Structured Review

Cell Signaling Technology Inc cleaved caspase 3
Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved <t>Caspase</t> 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P
Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 357 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/cleaved caspase 3/product/Cell Signaling Technology Inc
Average 99 stars, based on 357 article reviews
Price from $9.99 to $1999.99
cleaved caspase 3 - by Bioz Stars, 2020-10
99/100 stars

Images

1) Product Images from "SHP2 Inhibition Abrogates Adaptive Resistance to KRASG12C-Inhibition and Remodels the Tumor Microenvironment of KRAS-Mutant Tumors"

Article Title: SHP2 Inhibition Abrogates Adaptive Resistance to KRASG12C-Inhibition and Remodels the Tumor Microenvironment of KRAS-Mutant Tumors

Journal: bioRxiv

doi: 10.1101/2020.05.30.125138

Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved Caspase 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P
Figure Legend Snippet: Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved Caspase 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P

Techniques Used: Inhibition, In Vivo, Mouse Assay, Western Blot, Derivative Assay, Expressing, Staining

2) Product Images from "Direct therapeutic targeting of SWI/SNF induces epigenetic reprogramming and durable tumor regression in rhabdoid tumor"

Article Title: Direct therapeutic targeting of SWI/SNF induces epigenetic reprogramming and durable tumor regression in rhabdoid tumor

Journal: bioRxiv

doi: 10.1101/861484

Continuous infusion of mithramycin shows activity in an intramuscular rhabdoid tumor xenograft model. A:  Top, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 1 mg/kg/dose mithramycin intraperitoneal M, W, F for two weeks (green) relative to vehicle control (gray). Bottom, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 2.4 mg/kg of mithramycin (purple) or vehicle control (gray) administered continuously intraperitoneal over 72h. Most mice experienced a suppression or regression of tumor volume that persisted for more than 2 weeks following treatment. The shaded box indicates the duration of treatment. B:  Immunohistochemistry analysis recapitulates the biochemistry described in vitro. G401 tumor sections at 10X magnification stained for cleaved caspase 3 (CC3; apoptosis) or H3K27me3. A marked increase in CC3 correlates with H3K27me3 staining is seen only in mice treated with the continuous infusion schedule but not bolus dosing. C,D:  Prolonged durable response and cure of mice bearing G401 xenografts treated with  (C)  30mg/kg EC8042 administered continuously over 72-hours or  (D)  50 mg/kg EC8042 administered continuously over 144-hours. Treatment duration indicated by gray shaded box. Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text). E:  Bioluminescence imaging of G401 rhabdoid tumor xenografts correlates with caliper measurements in 7A and 7B. Two mice per treatment group were imaged (left) and quantified in the bar graph (right) . Error bars represent mean with SD. F:  Kaplan-Meier survival curves indicating extended survival for mice bearing established G401 xenografts treated with the 3-day or 7-day continuous infusions of EC8042 in (C) and (D). Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text).
Figure Legend Snippet: Continuous infusion of mithramycin shows activity in an intramuscular rhabdoid tumor xenograft model. A: Top, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 1 mg/kg/dose mithramycin intraperitoneal M, W, F for two weeks (green) relative to vehicle control (gray). Bottom, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 2.4 mg/kg of mithramycin (purple) or vehicle control (gray) administered continuously intraperitoneal over 72h. Most mice experienced a suppression or regression of tumor volume that persisted for more than 2 weeks following treatment. The shaded box indicates the duration of treatment. B: Immunohistochemistry analysis recapitulates the biochemistry described in vitro. G401 tumor sections at 10X magnification stained for cleaved caspase 3 (CC3; apoptosis) or H3K27me3. A marked increase in CC3 correlates with H3K27me3 staining is seen only in mice treated with the continuous infusion schedule but not bolus dosing. C,D: Prolonged durable response and cure of mice bearing G401 xenografts treated with (C) 30mg/kg EC8042 administered continuously over 72-hours or (D) 50 mg/kg EC8042 administered continuously over 144-hours. Treatment duration indicated by gray shaded box. Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text). E: Bioluminescence imaging of G401 rhabdoid tumor xenografts correlates with caliper measurements in 7A and 7B. Two mice per treatment group were imaged (left) and quantified in the bar graph (right) . Error bars represent mean with SD. F: Kaplan-Meier survival curves indicating extended survival for mice bearing established G401 xenografts treated with the 3-day or 7-day continuous infusions of EC8042 in (C) and (D). Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text).

Techniques Used: Activity Assay, Mouse Assay, Immunohistochemistry, In Vitro, Staining, Imaging

Mithramycin cellular sensitivity favors SWI/SNF mutant cancers. A:  Graph of IC50 as a function of cell line generated from a published screen of 445 agents in 63 sarcoma cell lines (  39 ). Cell lines with mutated or dysregulated SWI/SNF (green) cluster towards the right on the graph indicating these cell lines are more sensitive to mithramycin. B:  Table highlighting the SWI/SNF dysregulation status in the top 25 sarcoma cell lines from   Fig. 1A . Mutation status was confirmed in COSMIC (  49 ) or the DepMap ( https://depmap.org/portal/ ). C:  Dose response curves of rhabdoid tumor and Ewing sarcoma cell lines. RT cell lines (black) are sensitive to mithramycin treatment with a similar IC50 value as TC32 ES cells (grey). RT cell lines are not sensitive to three broadly-active chemotherapeutic agents: etoposide, doxorubicin or SN38. E:  Western blot showing concentration-dependent increase in H3K27me3 following exposure to 100nM, 50nM, 25nM mithramycin for 18h in BT12 cells relative to loading control (H3). E:  Mithramycin induces a concentration-dependent suppression of proliferation in BT12 cells. Cells were exposed to 25nM, 50nM, or 100nM MMA for 18h, replaced with drug- free media and monitored using live cell imaging. F:  Mithramycin induces morphological changes and apoptosis relative to control cells (solvent). BT12 cells treated with 25nM (left) or 100nM (right) mithramycin for 18h (top) or 48-hours (bottom) in the presence of cleaved caspase 3/7 reagent that fluoresces with caspase activation. G:  Mithramycin leads to H3K27me3 amplification in a time-dependent manner that precedes the induction of apoptosis as measured by the cleavage of PARP. Western blot lysates collected at 1h, 2h, 4h, 8h, 12h 16h, 18h of continuous 100nM mithramycin treatment. H:  Chromatin immunoprecipitation qPCR (ChIP-qPCR) of H3K27me3 at  MYT1  and  CCND1 . H3K27me3 occupancy is increased in a time-dependent manner.
Figure Legend Snippet: Mithramycin cellular sensitivity favors SWI/SNF mutant cancers. A: Graph of IC50 as a function of cell line generated from a published screen of 445 agents in 63 sarcoma cell lines ( 39 ). Cell lines with mutated or dysregulated SWI/SNF (green) cluster towards the right on the graph indicating these cell lines are more sensitive to mithramycin. B: Table highlighting the SWI/SNF dysregulation status in the top 25 sarcoma cell lines from Fig. 1A . Mutation status was confirmed in COSMIC ( 49 ) or the DepMap ( https://depmap.org/portal/ ). C: Dose response curves of rhabdoid tumor and Ewing sarcoma cell lines. RT cell lines (black) are sensitive to mithramycin treatment with a similar IC50 value as TC32 ES cells (grey). RT cell lines are not sensitive to three broadly-active chemotherapeutic agents: etoposide, doxorubicin or SN38. E: Western blot showing concentration-dependent increase in H3K27me3 following exposure to 100nM, 50nM, 25nM mithramycin for 18h in BT12 cells relative to loading control (H3). E: Mithramycin induces a concentration-dependent suppression of proliferation in BT12 cells. Cells were exposed to 25nM, 50nM, or 100nM MMA for 18h, replaced with drug- free media and monitored using live cell imaging. F: Mithramycin induces morphological changes and apoptosis relative to control cells (solvent). BT12 cells treated with 25nM (left) or 100nM (right) mithramycin for 18h (top) or 48-hours (bottom) in the presence of cleaved caspase 3/7 reagent that fluoresces with caspase activation. G: Mithramycin leads to H3K27me3 amplification in a time-dependent manner that precedes the induction of apoptosis as measured by the cleavage of PARP. Western blot lysates collected at 1h, 2h, 4h, 8h, 12h 16h, 18h of continuous 100nM mithramycin treatment. H: Chromatin immunoprecipitation qPCR (ChIP-qPCR) of H3K27me3 at MYT1 and CCND1 . H3K27me3 occupancy is increased in a time-dependent manner.

Techniques Used: Mutagenesis, Generated, Western Blot, Concentration Assay, Live Cell Imaging, Activation Assay, Amplification, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction

EC8042 induces a mixed phenotype that favors mesenchymal differentiation over apoptosis. A-D:  10X image of section of G401 treated tumors collected on day 4 (left) and day 8 (right) and stained for H3K27me3  (A) , H3K27ac  (B) , human mitochondria  (C) , and cleaved caspase 3  (D) . The sections compare vehicle to treatment started on day 1 with 30 mg/kg of EC8042 administered continuously for 72-hours (3-day pump). H3K27me3 increases and correlates with apoptosis (CC3) while H3K27ac decreases over time. H3K27ac and human mitochondrial staining decrease by day 4 compared with vehicle. Positive staining of H3K27me3 and CC3 is gone by day 8, as is human mitochondrial staining indicating no residual tumor.  E:  Immunohistochemistry analysis of H  E stains from G401 xenograft tumors on 1, 3, and 7-days after treatment with vehicle, 3-day EC8042 pump or 7-day EC8042 pump. EC8042 treated xenograft tumors exhibit evidence of mesenchymal differentiation compared to vehicle. microCT analysis of xenograft tumors on 7-days after treatment exhibit enhanced calcification compared to vehicle.
Figure Legend Snippet: EC8042 induces a mixed phenotype that favors mesenchymal differentiation over apoptosis. A-D: 10X image of section of G401 treated tumors collected on day 4 (left) and day 8 (right) and stained for H3K27me3 (A) , H3K27ac (B) , human mitochondria (C) , and cleaved caspase 3 (D) . The sections compare vehicle to treatment started on day 1 with 30 mg/kg of EC8042 administered continuously for 72-hours (3-day pump). H3K27me3 increases and correlates with apoptosis (CC3) while H3K27ac decreases over time. H3K27ac and human mitochondrial staining decrease by day 4 compared with vehicle. Positive staining of H3K27me3 and CC3 is gone by day 8, as is human mitochondrial staining indicating no residual tumor. E: Immunohistochemistry analysis of H E stains from G401 xenograft tumors on 1, 3, and 7-days after treatment with vehicle, 3-day EC8042 pump or 7-day EC8042 pump. EC8042 treated xenograft tumors exhibit evidence of mesenchymal differentiation compared to vehicle. microCT analysis of xenograft tumors on 7-days after treatment exhibit enhanced calcification compared to vehicle.

Techniques Used: Staining, Immunohistochemistry

3) Product Images from "Development of Aortic Valve Disease in Familial Hypercholesterolemic Swine: Implications for Elucidating Disease Etiology"

Article Title: Development of Aortic Valve Disease in Familial Hypercholesterolemic Swine: Implications for Elucidating Disease Etiology

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

doi: 10.1161/JAHA.115.002254

Representative histology of atherosclerotic lesions in the coronary arteries of adult  RFH  swine. A, Hematoxylin and eosin (H  E) staining and Movat's Pentachrome revealed wall thickening, fibrin accumulation, and plaque buildup in coronary arteries. B, α‐ SMA ‐positive cells were detected in the subendothelial region. C, Lipid oxidation was observed by immunostaining of oxApoB and malondialdehyde. D, Apoptotic cells were identified throughout the arteries by immunostaining for caspase 3. E, Leukocyte degranulation ( CD 107a) and the secretion of  MCP ‐1 were also observed in the intima, as expected in an atherosclerotic plaque. Scale bars: 400 μm (A and C); 200 μm (B, D, and E). Arrows indicate areas representative of positive staining. N=4 animals. MCP‐1 indicates monocyte chemoattractant protein‐1; oxApoB, oxidatively modified apolipoprotein B; RFH, Rapacz familial hypercholesterolemic; α‐SMA, alpha‐smooth muscle actin.
Figure Legend Snippet: Representative histology of atherosclerotic lesions in the coronary arteries of adult RFH swine. A, Hematoxylin and eosin (H E) staining and Movat's Pentachrome revealed wall thickening, fibrin accumulation, and plaque buildup in coronary arteries. B, α‐ SMA ‐positive cells were detected in the subendothelial region. C, Lipid oxidation was observed by immunostaining of oxApoB and malondialdehyde. D, Apoptotic cells were identified throughout the arteries by immunostaining for caspase 3. E, Leukocyte degranulation ( CD 107a) and the secretion of MCP ‐1 were also observed in the intima, as expected in an atherosclerotic plaque. Scale bars: 400 μm (A and C); 200 μm (B, D, and E). Arrows indicate areas representative of positive staining. N=4 animals. MCP‐1 indicates monocyte chemoattractant protein‐1; oxApoB, oxidatively modified apolipoprotein B; RFH, Rapacz familial hypercholesterolemic; α‐SMA, alpha‐smooth muscle actin.

Techniques Used: Staining, Immunostaining, Modification

4) Product Images from "Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions"

Article Title: Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions

Journal: Journal of Virology

doi: 10.1128/JVI.00070-12

Apoptotic effects of neomycin and antiapoptotic effects of angiogenin. (A) Serum-starved (8 h) HMVEC-d cells in 8-well chamber slides were either left untreated, treated with 1 μg/ml ANG, infected with 20 KSHV DNA/cell, or pretreated with 200 μM neomycin (N) for 1 h and then infected with the virus or treated with 1 μg/ml ANG. At 24 h, the slides were stained with anti-cleaved caspase-3 antibody and visualized under the fluorescence microscope. (B) HMVEC-d cells were treated similarly, and the samples were separated with a 10% gel and subjected to Western blotting for cleaved caspase-3.
Figure Legend Snippet: Apoptotic effects of neomycin and antiapoptotic effects of angiogenin. (A) Serum-starved (8 h) HMVEC-d cells in 8-well chamber slides were either left untreated, treated with 1 μg/ml ANG, infected with 20 KSHV DNA/cell, or pretreated with 200 μM neomycin (N) for 1 h and then infected with the virus or treated with 1 μg/ml ANG. At 24 h, the slides were stained with anti-cleaved caspase-3 antibody and visualized under the fluorescence microscope. (B) HMVEC-d cells were treated similarly, and the samples were separated with a 10% gel and subjected to Western blotting for cleaved caspase-3.

Techniques Used: Infection, Staining, Fluorescence, Microscopy, Western Blot

Reduction of cell survival upon angiogenin knockdown. (A) 293T cells were transfected with control sh-GFP and sh-Renilla (RL) plasmids or two sh-ANG (sh-ANG1 and sh-ANG2) plasmids, and ANG knockdown was checked by separating the lysates with a 12.5% gel and performing Western blotting for ANG. (B) ANG knockdown was also checked by real-time RT-PCR in sh-GFP- and sh-ANG1-transduced BCBL-1 cells. (C to E) BCBL-1 cells transduced with either sh-GFP or sh-ANG1 were subjected to Western blotting for p53, p-p53, p21, Bcl-2, Bax, caspase 3, and β-actin. Fold changes are indicated. (F) BCBL cells transduced with either sh-GFP or sh-ANG1 were stained with YO-PRO dye and analyzed by FACS for the percentage of live cells. Values shown represent the means ± standard deviations from three independent experiments. ***,  P
Figure Legend Snippet: Reduction of cell survival upon angiogenin knockdown. (A) 293T cells were transfected with control sh-GFP and sh-Renilla (RL) plasmids or two sh-ANG (sh-ANG1 and sh-ANG2) plasmids, and ANG knockdown was checked by separating the lysates with a 12.5% gel and performing Western blotting for ANG. (B) ANG knockdown was also checked by real-time RT-PCR in sh-GFP- and sh-ANG1-transduced BCBL-1 cells. (C to E) BCBL-1 cells transduced with either sh-GFP or sh-ANG1 were subjected to Western blotting for p53, p-p53, p21, Bcl-2, Bax, caspase 3, and β-actin. Fold changes are indicated. (F) BCBL cells transduced with either sh-GFP or sh-ANG1 were stained with YO-PRO dye and analyzed by FACS for the percentage of live cells. Values shown represent the means ± standard deviations from three independent experiments. ***, P

Techniques Used: Transfection, Western Blot, Quantitative RT-PCR, Transduction, Staining, FACS

5) Product Images from "IAP antagonists Birinapant and AT-406 efficiently synergise with either TRAIL, BRAF, or BCL-2 inhibitors to sensitise BRAFV600E colorectal tumour cells to apoptosis"

Article Title: IAP antagonists Birinapant and AT-406 efficiently synergise with either TRAIL, BRAF, or BCL-2 inhibitors to sensitise BRAFV600E colorectal tumour cells to apoptosis

Journal: BMC Cancer

doi: 10.1186/s12885-016-2606-5

Co-treatment of Birinapant with TRAIL can synergistically increase their efficiency and induce apoptosis in colorectal adenocarcinoma cells in 2D and 3D. 5a: Cell viability of cell line HT29 after co-treatment with the SMAC-mimetic Birinapant and the apoptotic agent TRAIL. Cells were either left untreated (ctr = control) or treated with Birinapant, TRAIL and their combination for 48 and 72 h and the % percentage cell viability was measured. The average of three independent experiments is presented. Columns = % percentage of cell viability, bars = SD. 5a-RKO: Respectively for cell line RKO (upper panel). To check for the reversibility of treatment effects, RKO cells were either left untreated (ctr = control), treated with either Birinapant, TRAIL or their combination for 48 h. Cells were then incubated with additional 48 h without treatments (NoT) and the % percentage cell viability was measured. The average of three independent experiments is presented. 5b-I. Protein levels of cIAP-1 and XIAP after 0.5, 1 and 5 μM AT-406 (lanes 3–5) or Birinapant (lanes 6–8) treatments respectively for 48 h, compared to control lanes 1 (no treatment) or 2 (DMSO treatment). 5b-II and 5b-III. Protein levels of PARP-1 and total Caspase-3 in HT29 cell line after co-treatment with SMAC-mimetic Birinapant and TRAIL for 48 and 72 h. Cells were either left untreated (lane 1) or treated with DMSO (lane 2), with 5 μM Birinapant (lane 3), with 10, 50 and 100 ng/mL TRAIL (lanes 4–6), or with 5 μM Birinapant and combination with TRAIL (lanes 7–9) for 48 and 72 h. Sensitive cell line HCT116 treated with TRAIL is used as positive control (lane 10). Using W. B., protein levels of PARP-1 (Figure 5b-II) and of total Caspase 3 (Figure 5b-III) were analyzed after the corresponding treatments. Data are representative for three independent experiments. 5b-I-RKO: Protein levels of cIAP-1 and XIAP after mono-treatments (lanes 3 and 4 for AT-406 and Birinapant respectively and lanes 5–7 for TRAIL 10, 50 and 100 ng/mL respectively) and co-treatments (lanes 8–13 as shown) for 24 h. Untreated (lane 1) and treated with DMSO (lane 2), cells are also presented. 5b-II-RKO and 5b-III-RKO: Protein levels of PARP-1 and total Caspase-3 after respective mono-treatments and co-treatments (same lines as 5-I-RKO). Data are representative of three independent experiments. 5C: Light microscopy images from HT29 culture after combined treatment with Birinapant and TRAIL. Detached (apoptotic) cells are shown in supernatant of co-treated groups for 48 and 72 h. Representative images. 5d: Confocal microscope images were taken after co-treatment with Birinapant, TRAIL and their combination. Nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images.. Scale bar: 9.9 μm. 5e. Light microscopy of three-dimensional culture of HT29 cells after co-treatment with Birinapant, TRAIL and their combinations in 3D for 6 days. Representative images. 5f: Confocal microscope images were taken after co-treatment with Birinapant, TRAIL and their combinations in 3D culture for 6 days. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 57.4 μm
Figure Legend Snippet: Co-treatment of Birinapant with TRAIL can synergistically increase their efficiency and induce apoptosis in colorectal adenocarcinoma cells in 2D and 3D. 5a: Cell viability of cell line HT29 after co-treatment with the SMAC-mimetic Birinapant and the apoptotic agent TRAIL. Cells were either left untreated (ctr = control) or treated with Birinapant, TRAIL and their combination for 48 and 72 h and the % percentage cell viability was measured. The average of three independent experiments is presented. Columns = % percentage of cell viability, bars = SD. 5a-RKO: Respectively for cell line RKO (upper panel). To check for the reversibility of treatment effects, RKO cells were either left untreated (ctr = control), treated with either Birinapant, TRAIL or their combination for 48 h. Cells were then incubated with additional 48 h without treatments (NoT) and the % percentage cell viability was measured. The average of three independent experiments is presented. 5b-I. Protein levels of cIAP-1 and XIAP after 0.5, 1 and 5 μM AT-406 (lanes 3–5) or Birinapant (lanes 6–8) treatments respectively for 48 h, compared to control lanes 1 (no treatment) or 2 (DMSO treatment). 5b-II and 5b-III. Protein levels of PARP-1 and total Caspase-3 in HT29 cell line after co-treatment with SMAC-mimetic Birinapant and TRAIL for 48 and 72 h. Cells were either left untreated (lane 1) or treated with DMSO (lane 2), with 5 μM Birinapant (lane 3), with 10, 50 and 100 ng/mL TRAIL (lanes 4–6), or with 5 μM Birinapant and combination with TRAIL (lanes 7–9) for 48 and 72 h. Sensitive cell line HCT116 treated with TRAIL is used as positive control (lane 10). Using W. B., protein levels of PARP-1 (Figure 5b-II) and of total Caspase 3 (Figure 5b-III) were analyzed after the corresponding treatments. Data are representative for three independent experiments. 5b-I-RKO: Protein levels of cIAP-1 and XIAP after mono-treatments (lanes 3 and 4 for AT-406 and Birinapant respectively and lanes 5–7 for TRAIL 10, 50 and 100 ng/mL respectively) and co-treatments (lanes 8–13 as shown) for 24 h. Untreated (lane 1) and treated with DMSO (lane 2), cells are also presented. 5b-II-RKO and 5b-III-RKO: Protein levels of PARP-1 and total Caspase-3 after respective mono-treatments and co-treatments (same lines as 5-I-RKO). Data are representative of three independent experiments. 5C: Light microscopy images from HT29 culture after combined treatment with Birinapant and TRAIL. Detached (apoptotic) cells are shown in supernatant of co-treated groups for 48 and 72 h. Representative images. 5d: Confocal microscope images were taken after co-treatment with Birinapant, TRAIL and their combination. Nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images.. Scale bar: 9.9 μm. 5e. Light microscopy of three-dimensional culture of HT29 cells after co-treatment with Birinapant, TRAIL and their combinations in 3D for 6 days. Representative images. 5f: Confocal microscope images were taken after co-treatment with Birinapant, TRAIL and their combinations in 3D culture for 6 days. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 57.4 μm

Techniques Used: Incubation, Positive Control, Light Microscopy, Microscopy, Staining

SMAC-mimetic AT-406 and TRAIL synergistically kill resistant tumour cells. 6a Cell viability of HT29 cell line after combined treatment with SMAC-mimetic AT-406 and the apoptotic agent TRAIL. Cells were either left untreated (ctr = control) or treated with AT-406 combined with TRAIL for 48 and 72 h and the % percentage cell viability was measured by SRB staining. The values are the average of three independent experiments and are presented as fold change of the absorbance of treated/untreated cells, for each condition. Columns = % percentage of cell viability, bars = SD. 6a-RKO: Respectively for cell line RKO. 6b. Confocal microscope images were taken after co-treatment with AT-406, TRAIL and their combinations for 48 and 72 h. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 9.9 μM
Figure Legend Snippet: SMAC-mimetic AT-406 and TRAIL synergistically kill resistant tumour cells. 6a Cell viability of HT29 cell line after combined treatment with SMAC-mimetic AT-406 and the apoptotic agent TRAIL. Cells were either left untreated (ctr = control) or treated with AT-406 combined with TRAIL for 48 and 72 h and the % percentage cell viability was measured by SRB staining. The values are the average of three independent experiments and are presented as fold change of the absorbance of treated/untreated cells, for each condition. Columns = % percentage of cell viability, bars = SD. 6a-RKO: Respectively for cell line RKO. 6b. Confocal microscope images were taken after co-treatment with AT-406, TRAIL and their combinations for 48 and 72 h. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 9.9 μM

Techniques Used: Sulforhodamine B Assay, Staining, Microscopy

Birinapant treatment results in reduced cell viability and appearance of apoptosis in selected colorectal adenocarcinoma cells. a : Cell viability of cell lines after treatment with SMAC-mimetics. Cells were either left untreated (ctr = control) or treated with 0.5 μM, 1 mM or 5 mM AT-406 and Birinapant for 48 h (1) and 72 h (2) and the % percentage cell viability was measured by SRB. Average of three independent experiments are presented as fold change of the absorbance of treated/untreated cells for each condition. Columns = % percentage of cell viability, bars = SD. b : Protein levels of XIAP, cIAP-1, PARP-1 and total caspase-3 in RKO and HCT116 were analysed by W.B. after treatment with 0.5, 1 and 5 μM SMAC-mimetics AT-406 (lanes 3–5) and Birinapant (lanes 6–8) for 48 and 72 h. Untreated (lane 1) or treated with DMSO cells (lane 2) were used as control. Proteins are quantified against α-Tubulin. Data are representative for three independent experiments. c - d : Confocal microscope images and Hoechst staining of RKO (2C) and HCT116 (2D) cell lines two-dimensional culture, after treatment with SMAC-mimetics AT-406 (A) and Birinapant (B). Crescent nuclei of RKO cells present after cell treatments are shown by arrows. Confocal microscope images were taken after treatment with SMAC-mimetics AT-406 and Birinapant in RKO (2c) and HCT116 (2d) 48 and 72 h. The nuclei were detected with HOECHST staining. Representative images are presented
Figure Legend Snippet: Birinapant treatment results in reduced cell viability and appearance of apoptosis in selected colorectal adenocarcinoma cells. a : Cell viability of cell lines after treatment with SMAC-mimetics. Cells were either left untreated (ctr = control) or treated with 0.5 μM, 1 mM or 5 mM AT-406 and Birinapant for 48 h (1) and 72 h (2) and the % percentage cell viability was measured by SRB. Average of three independent experiments are presented as fold change of the absorbance of treated/untreated cells for each condition. Columns = % percentage of cell viability, bars = SD. b : Protein levels of XIAP, cIAP-1, PARP-1 and total caspase-3 in RKO and HCT116 were analysed by W.B. after treatment with 0.5, 1 and 5 μM SMAC-mimetics AT-406 (lanes 3–5) and Birinapant (lanes 6–8) for 48 and 72 h. Untreated (lane 1) or treated with DMSO cells (lane 2) were used as control. Proteins are quantified against α-Tubulin. Data are representative for three independent experiments. c - d : Confocal microscope images and Hoechst staining of RKO (2C) and HCT116 (2D) cell lines two-dimensional culture, after treatment with SMAC-mimetics AT-406 (A) and Birinapant (B). Crescent nuclei of RKO cells present after cell treatments are shown by arrows. Confocal microscope images were taken after treatment with SMAC-mimetics AT-406 and Birinapant in RKO (2c) and HCT116 (2d) 48 and 72 h. The nuclei were detected with HOECHST staining. Representative images are presented

Techniques Used: Sulforhodamine B Assay, Microscopy, Staining

Pre-treatment with Birinapant and then co-treatment with Birinapant/ BRAF inhibitor PLX4720 synergistically induce apoptosis of colorectal adenocarcinoma cells in 2D and 3D. a : Cell viability after co-treatment with the SMAC-mimetics Birinapant or AT-406 in combination with the BRAF inhibitor PLX4720. Cells were either left untreated (ctr = control) or treated with 5 μM Birinapant, 5 μM AT-406 and 1 μM PLX4720 and their combinations for 48 and 72 h. The average of three independent experiments is presented as fold change of the absorbance of treated/untreated cells, for each condition. Columns = % percentage of cell viability, bars = SD. b : Cell viability after pre-treatment with the SMAC-mimetic Birinapant and then co-treatment with BRAF inhibitor PLX4720 and Birinapant. Cells were either left untreated (ctr = control) or treated with 0.5 or 1 μM Birinapant and 0.2 or 0.5 μM PLX4720. For the pre-treatment testing, cells were first incubated for 24 h with either 0.5 or 1 μM of Birinapant and then co-treated with 0.2 and 0.5 μM PLX4720 for another 24 or 48 h. c : Protein levels of PARP-1, total Caspase-3, XIAP, cIAP-1 and p-ERK1/2 in RKO by W.B., after pre-treatment with 0.5 or 1 μM Birinapant and then co-treatment with 0.2 or 0.5 μM PLX4720. Untreated cells were used as control. Proteins are quantified against α-Tubulin. Data are representative for three independent experiments. d : Light microscopy images from RKO culture after pre-treatment with Birinapant and then co-treatment with PLX4720. Detached cells are shown in supernatant of co-treated group. Several images were taken from untreated and treated with Birinapant (0.5 or 1 μM) and PLX4720 (0.2 or 0.5 μM) RKO cells while been cultured in 6-well plates for 48 and 72 h. Representative images are presented. e : Light microscopy of three-dimensional culture of RKO cells after co-treatment with 0.5 μM, 1 μM Birinapant and 0.2 μM, 0.5 μM PLX4720 and their combination in 3D culture for 6 d. Representative images. f : Confocal microscope images were taken after co-treatment with 0.5 μM, 1 μM Birinapant and 0.2 μM, 0.5 μM PLX4720 and combinations in 3D cultures for 6 days. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 20 μm
Figure Legend Snippet: Pre-treatment with Birinapant and then co-treatment with Birinapant/ BRAF inhibitor PLX4720 synergistically induce apoptosis of colorectal adenocarcinoma cells in 2D and 3D. a : Cell viability after co-treatment with the SMAC-mimetics Birinapant or AT-406 in combination with the BRAF inhibitor PLX4720. Cells were either left untreated (ctr = control) or treated with 5 μM Birinapant, 5 μM AT-406 and 1 μM PLX4720 and their combinations for 48 and 72 h. The average of three independent experiments is presented as fold change of the absorbance of treated/untreated cells, for each condition. Columns = % percentage of cell viability, bars = SD. b : Cell viability after pre-treatment with the SMAC-mimetic Birinapant and then co-treatment with BRAF inhibitor PLX4720 and Birinapant. Cells were either left untreated (ctr = control) or treated with 0.5 or 1 μM Birinapant and 0.2 or 0.5 μM PLX4720. For the pre-treatment testing, cells were first incubated for 24 h with either 0.5 or 1 μM of Birinapant and then co-treated with 0.2 and 0.5 μM PLX4720 for another 24 or 48 h. c : Protein levels of PARP-1, total Caspase-3, XIAP, cIAP-1 and p-ERK1/2 in RKO by W.B., after pre-treatment with 0.5 or 1 μM Birinapant and then co-treatment with 0.2 or 0.5 μM PLX4720. Untreated cells were used as control. Proteins are quantified against α-Tubulin. Data are representative for three independent experiments. d : Light microscopy images from RKO culture after pre-treatment with Birinapant and then co-treatment with PLX4720. Detached cells are shown in supernatant of co-treated group. Several images were taken from untreated and treated with Birinapant (0.5 or 1 μM) and PLX4720 (0.2 or 0.5 μM) RKO cells while been cultured in 6-well plates for 48 and 72 h. Representative images are presented. e : Light microscopy of three-dimensional culture of RKO cells after co-treatment with 0.5 μM, 1 μM Birinapant and 0.2 μM, 0.5 μM PLX4720 and their combination in 3D culture for 6 d. Representative images. f : Confocal microscope images were taken after co-treatment with 0.5 μM, 1 μM Birinapant and 0.2 μM, 0.5 μM PLX4720 and combinations in 3D cultures for 6 days. The nuclei were detected with HOECHST staining (blue color), cleaved Caspase-3 (green color). Representative images. Scale bar: 20 μm

Techniques Used: Incubation, Light Microscopy, Cell Culture, Microscopy, Staining

6) Product Images from "MMP-7 mediates cleavage of N-cadherin and promotes smooth muscle cell apoptosis"

Article Title: MMP-7 mediates cleavage of N-cadherin and promotes smooth muscle cell apoptosis

Journal: Cardiovascular Research

doi: 10.1093/cvr/cvq042

MMP-7 and apoptosis in atherosclerotic plaques. In situ zymography of coronary atherosclerotic plaque in the absence ( A ) and presence ( B ) of 10 nM MMP-7 inhibitor. Green colour indicates MMP activity and nuclei are stained blue with DAPI. Dual detection of cleaved caspase-3 (red) and MMP activity (green) in coronary plaque ( C ) and IMA ( D ). ( E ) High power image of marked area in ( C ), arrows indicate co-location of apoptosis and MMP activity. ( F ) Quantification of the percentage of apoptotic (ISEL positive) cells in atherosclerotic plaques from ApoE −/− /MMP-7 +/+ and ApoE −/− /MMP-7 −/− . * indicates a significant difference from ApoE −/− /MMP-7 +/+ . ISEL staining of atherosclerotic plaques from ApoE −/− /MMP-7 +/+ ( G ) and ApoE −/− /MMP-7 −/− ( H ) mice. ISEL positive apoptotic cells are green and nuclei are stained blue (DAPI).
Figure Legend Snippet: MMP-7 and apoptosis in atherosclerotic plaques. In situ zymography of coronary atherosclerotic plaque in the absence ( A ) and presence ( B ) of 10 nM MMP-7 inhibitor. Green colour indicates MMP activity and nuclei are stained blue with DAPI. Dual detection of cleaved caspase-3 (red) and MMP activity (green) in coronary plaque ( C ) and IMA ( D ). ( E ) High power image of marked area in ( C ), arrows indicate co-location of apoptosis and MMP activity. ( F ) Quantification of the percentage of apoptotic (ISEL positive) cells in atherosclerotic plaques from ApoE −/− /MMP-7 +/+ and ApoE −/− /MMP-7 −/− . * indicates a significant difference from ApoE −/− /MMP-7 +/+ . ISEL staining of atherosclerotic plaques from ApoE −/− /MMP-7 +/+ ( G ) and ApoE −/− /MMP-7 −/− ( H ) mice. ISEL positive apoptotic cells are green and nuclei are stained blue (DAPI).

Techniques Used: In Situ, Zymography, Activity Assay, Staining, Mouse Assay

N-cadherin, MMP-7, and apoptosis in human IMA and atherosclerotic plaques. Representative western blot for N-cadherin ( A ) and cleaved caspase-3 ( B ) in IMA and coronary and carotid atherosclerotic plaques, n = 3 per group. β-Tubulin is shown as loading control. Bar chart in ( A ) shows densitometric analysis of N-cadherin western blots. ( C ) MMP activity in IMA and atherosclerotic plaques, n = 15 and 20. * indicates a significant difference from IMA.
Figure Legend Snippet: N-cadherin, MMP-7, and apoptosis in human IMA and atherosclerotic plaques. Representative western blot for N-cadherin ( A ) and cleaved caspase-3 ( B ) in IMA and coronary and carotid atherosclerotic plaques, n = 3 per group. β-Tubulin is shown as loading control. Bar chart in ( A ) shows densitometric analysis of N-cadherin western blots. ( C ) MMP activity in IMA and atherosclerotic plaques, n = 15 and 20. * indicates a significant difference from IMA.

Techniques Used: Western Blot, Activity Assay

7) Product Images from "Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats"

Article Title: Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats

Journal: BMC Neuroscience

doi: 10.1186/1471-2202-12-12

Double staining with S-100 and cleaved caspase 3 in a control nerve (A) and in a repaired (B) sciatic nerve from the distal nerve segment . The staining indicated that cleaved caspase 3 stained cell nuclei (green) was associated with S-100 staining (red). DAPI-stained cells (blue) were used to localize cell nuclei. Length of bars 50 μm (A) and 25 μm (B).
Figure Legend Snippet: Double staining with S-100 and cleaved caspase 3 in a control nerve (A) and in a repaired (B) sciatic nerve from the distal nerve segment . The staining indicated that cleaved caspase 3 stained cell nuclei (green) was associated with S-100 staining (red). DAPI-stained cells (blue) were used to localize cell nuclei. Length of bars 50 μm (A) and 25 μm (B).

Techniques Used: Double Staining, Staining

Schematic drawing showing (A) the experimental design with immediate (I), delayed (D) and no nerve repairs (N and DN) and (B) the sites where p-ERK 1/2 and cleaved caspase 3 were analyzed .
Figure Legend Snippet: Schematic drawing showing (A) the experimental design with immediate (I), delayed (D) and no nerve repairs (N and DN) and (B) the sites where p-ERK 1/2 and cleaved caspase 3 were analyzed .

Techniques Used:

Results of immunocytochemical staining for ERK1/2 and cleaved caspase 3 analyzed in the distal nerve segment after immediate, delayed and no nerve repairs . Immunocytochemical staining for p-ERK1/2 (A) in longitudinal sections at the site of the lesion (a)-(d) [(a) group I (immediate repair), (b) group N (no repair), (c) group D (delayed repair), (d) group DN (delayed no repair)], and in the distal nerve segment (e)-(h) [(e) group I, (f) group N, (g) group D, (h) group DN]. Contralateral uninjured nerve is shown in (i). The scale bar represents 100 μm. The graphs show the percentages of immunopositive area for p-ERK1/2 (B) expressed as mean + SD. The stars indicate p-values *
Figure Legend Snippet: Results of immunocytochemical staining for ERK1/2 and cleaved caspase 3 analyzed in the distal nerve segment after immediate, delayed and no nerve repairs . Immunocytochemical staining for p-ERK1/2 (A) in longitudinal sections at the site of the lesion (a)-(d) [(a) group I (immediate repair), (b) group N (no repair), (c) group D (delayed repair), (d) group DN (delayed no repair)], and in the distal nerve segment (e)-(h) [(e) group I, (f) group N, (g) group D, (h) group DN]. Contralateral uninjured nerve is shown in (i). The scale bar represents 100 μm. The graphs show the percentages of immunopositive area for p-ERK1/2 (B) expressed as mean + SD. The stars indicate p-values *

Techniques Used: Staining

Graphs of regression analyses between length of neurofilaments (mm, dependent) and p-ERK 1/2 and cleaved caspase 3 (independent) at site of lesion and in distal nerve segment . For p-values and r 2 see Results.
Figure Legend Snippet: Graphs of regression analyses between length of neurofilaments (mm, dependent) and p-ERK 1/2 and cleaved caspase 3 (independent) at site of lesion and in distal nerve segment . For p-values and r 2 see Results.

Techniques Used:

8) Product Images from "Synthetic lethality of glutaminolysis inhibition, autophagy inactivation and asparagine depletion in colon cancer"

Article Title: Synthetic lethality of glutaminolysis inhibition, autophagy inactivation and asparagine depletion in colon cancer

Journal: Oncotarget

doi: 10.18632/oncotarget.16844

Inactivation of autophagy enhanced growth inhibition induced by glutamine starvation (A) The effect of autophagy inhibitor-CQ (10μM) on cell viability in the presence or absence of glutamine for 72h was determined by MTS assay. (B) and (C) The effect of autophagy inhibitor-CQ (10μM) on cell apoptosis in the presence or absence of glutamine for 72h was assessed using flow cytometry (B) and Western blotting detection of cleaved caspase 3 and PARP cleavage (for 48h) (C) . (D) MTS assay in SW620 cells as in (A) . (E) and (F) Apoptosis detection in SW620 cells as in (B) and (C) , respectively. All experiments were repeated for 3 times and the representative results were shown. The asterisks indicate statistical difference (p
Figure Legend Snippet: Inactivation of autophagy enhanced growth inhibition induced by glutamine starvation (A) The effect of autophagy inhibitor-CQ (10μM) on cell viability in the presence or absence of glutamine for 72h was determined by MTS assay. (B) and (C) The effect of autophagy inhibitor-CQ (10μM) on cell apoptosis in the presence or absence of glutamine for 72h was assessed using flow cytometry (B) and Western blotting detection of cleaved caspase 3 and PARP cleavage (for 48h) (C) . (D) MTS assay in SW620 cells as in (A) . (E) and (F) Apoptosis detection in SW620 cells as in (B) and (C) , respectively. All experiments were repeated for 3 times and the representative results were shown. The asterisks indicate statistical difference (p

Techniques Used: Inhibition, MTS Assay, Flow Cytometry, Cytometry, Western Blot

9) Product Images from "Characterization of injury in isolated rat proximal tubules during cold incubation and rewarming"

Article Title: Characterization of injury in isolated rat proximal tubules during cold incubation and rewarming

Journal: PLoS ONE

doi: 10.1371/journal.pone.0180553

Caspase 3 cleavage of isolated renal proximal tubules following rewarming after 48 h of cold incubation. Isolated tubules were incubated at 4°C in chloride-rich solution 1 or its chloride-poor counterpart solution 2 each in the absence and presence of the iron chelators deferoxamine (0.5 mM) and LK 614 (20 μM) for 48 h. Tubule suspensions were rewarmed in extra-cellular buffer at 37°C for two hours. Total protein extracts of renal proximal tubules were prepared using RIPA buffer and equal amounts of protein (40 μg) were separated by SDS-PAGE and transferred onto nitrocellulose membrane. Full-length and cleaved caspase 3 was assessed in tubules of four preparations using a rabbit antibody against caspase 3. (A) Representative Western Blot for caspase 3 and cleaved caspase 3. (B) Loading control. Equal amounts of total protein (8 μg) of the protein extracts were separated by SDS-PAGE and coomassie staining was performed to check for comparable loading. (C) Quantification of cleaved caspase 3 (expressed as percentage of the respective signal of non-stored control tubules; n = 4). ** p
Figure Legend Snippet: Caspase 3 cleavage of isolated renal proximal tubules following rewarming after 48 h of cold incubation. Isolated tubules were incubated at 4°C in chloride-rich solution 1 or its chloride-poor counterpart solution 2 each in the absence and presence of the iron chelators deferoxamine (0.5 mM) and LK 614 (20 μM) for 48 h. Tubule suspensions were rewarmed in extra-cellular buffer at 37°C for two hours. Total protein extracts of renal proximal tubules were prepared using RIPA buffer and equal amounts of protein (40 μg) were separated by SDS-PAGE and transferred onto nitrocellulose membrane. Full-length and cleaved caspase 3 was assessed in tubules of four preparations using a rabbit antibody against caspase 3. (A) Representative Western Blot for caspase 3 and cleaved caspase 3. (B) Loading control. Equal amounts of total protein (8 μg) of the protein extracts were separated by SDS-PAGE and coomassie staining was performed to check for comparable loading. (C) Quantification of cleaved caspase 3 (expressed as percentage of the respective signal of non-stored control tubules; n = 4). ** p

Techniques Used: Isolation, Incubation, SDS Page, Western Blot, Staining

10) Product Images from "Hepatocyte apoptosis is tumor promoting in murine nonalcoholic steatohepatitis"

Article Title: Hepatocyte apoptosis is tumor promoting in murine nonalcoholic steatohepatitis

Journal: Cell Death & Disease

doi: 10.1038/s41419-020-2283-9

Mcl1 deficiency in hepatocytes promotes tumor development in mice fed the FFC diet for 10 months. Mcl1 ∆hep mice and control littermates (WT) were placed on standard chow or FFC diet for 10 months. Blood and livers were harvested at the end of the study. a Body weight at the end of the feeding study. b Liver weight as percentage of total body weight. c , d Liver injury was assessed by serum AST and ALT activity. e Presence of tumors was assesses by histology. Chow-WT n = 9 mice; Chow-Mcl1 ∆hep n = 21 mice; FFC-WT n = 14 mice; FFC-Mcl1 ∆hep n = 18 mice. f Macroscopic view of livers (scale bar 5 mm). g Representative images of H E-stained liver sections of FFC-fed Mcl1 ∆hep mice. Left and middle image: tumors lack distinct borders; middle and right image: tumors show nodule-in-nodule appearance (scale bar 0.5 mm or 2.5 mm as indicated). h , i Liver sections of FFC-fed Mcl1 ∆hep mice were stained for H E, glutamine synthetase, collagen IV, GP73, β-catenin, cleaved caspase 3 (red arrows) and Ki67 and representative images are shown. Dotted line marks tumor (t) border (scale bar 50 μm). j Real-time qPCR analysis of genes known to be dysregulated in liver cancer. P value calculated for differences found between tumors of Mcl1 ∆hep mice fed chow vs FFC diet. Bars represent mean ± SEM. a , b Chow-WT n = 5 mice; Chow-Mcl1 ∆hep n = 13 mice; FFC-WT n = 14 mice; FFC-Mcl1 ∆hep n = 18 mice; c , d Chow-WT n = 5 mice; Chow-Mcl1 ∆hep n = 10 mice; FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 13 mice; ** p
Figure Legend Snippet: Mcl1 deficiency in hepatocytes promotes tumor development in mice fed the FFC diet for 10 months. Mcl1 ∆hep mice and control littermates (WT) were placed on standard chow or FFC diet for 10 months. Blood and livers were harvested at the end of the study. a Body weight at the end of the feeding study. b Liver weight as percentage of total body weight. c , d Liver injury was assessed by serum AST and ALT activity. e Presence of tumors was assesses by histology. Chow-WT n = 9 mice; Chow-Mcl1 ∆hep n = 21 mice; FFC-WT n = 14 mice; FFC-Mcl1 ∆hep n = 18 mice. f Macroscopic view of livers (scale bar 5 mm). g Representative images of H E-stained liver sections of FFC-fed Mcl1 ∆hep mice. Left and middle image: tumors lack distinct borders; middle and right image: tumors show nodule-in-nodule appearance (scale bar 0.5 mm or 2.5 mm as indicated). h , i Liver sections of FFC-fed Mcl1 ∆hep mice were stained for H E, glutamine synthetase, collagen IV, GP73, β-catenin, cleaved caspase 3 (red arrows) and Ki67 and representative images are shown. Dotted line marks tumor (t) border (scale bar 50 μm). j Real-time qPCR analysis of genes known to be dysregulated in liver cancer. P value calculated for differences found between tumors of Mcl1 ∆hep mice fed chow vs FFC diet. Bars represent mean ± SEM. a , b Chow-WT n = 5 mice; Chow-Mcl1 ∆hep n = 13 mice; FFC-WT n = 14 mice; FFC-Mcl1 ∆hep n = 18 mice; c , d Chow-WT n = 5 mice; Chow-Mcl1 ∆hep n = 10 mice; FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 13 mice; ** p

Techniques Used: Mouse Assay, AST Assay, Activity Assay, Staining, Real-time Polymerase Chain Reaction

Mcl1 deficiency in hepatocytes exacerbates liver injury induced by the FFC diet. Mcl1 ∆hep mice and control littermates (WT) were placed on standard chow or FFC diet for 4 months. Blood and livers were harvested at the end of the study. a Body weight at the end of the feeding study. b Liver weight as percentage of total body weight. c Liver triglycerides were measured in tissue homogenates using biochemical assay. d , e Liver injury was assessed by serum AST and ALT activity. f Hepatic apoptosis was assessed by immunohistochemistry and counting cleaved caspase 3-positive cells per 20× fields. n = 5 mice/group. g NAFLD activity score. FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 7 mice; h Representative images of H E staining (arrows point to inflammatory foci), Sudan red staining for lipids, and immunohistochemistry for cleaved caspase 3 (arrowheads) in liver tissue samples (scale bar 50 μm). Bars represent mean ± SEM. a – e Chow-WT n = 9 mice; Chow-Mcl1 ∆hep n = 5 mice; FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 7 mice; *** p
Figure Legend Snippet: Mcl1 deficiency in hepatocytes exacerbates liver injury induced by the FFC diet. Mcl1 ∆hep mice and control littermates (WT) were placed on standard chow or FFC diet for 4 months. Blood and livers were harvested at the end of the study. a Body weight at the end of the feeding study. b Liver weight as percentage of total body weight. c Liver triglycerides were measured in tissue homogenates using biochemical assay. d , e Liver injury was assessed by serum AST and ALT activity. f Hepatic apoptosis was assessed by immunohistochemistry and counting cleaved caspase 3-positive cells per 20× fields. n = 5 mice/group. g NAFLD activity score. FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 7 mice; h Representative images of H E staining (arrows point to inflammatory foci), Sudan red staining for lipids, and immunohistochemistry for cleaved caspase 3 (arrowheads) in liver tissue samples (scale bar 50 μm). Bars represent mean ± SEM. a – e Chow-WT n = 9 mice; Chow-Mcl1 ∆hep n = 5 mice; FFC-WT n = 12 mice; FFC-Mcl1 ∆hep n = 7 mice; *** p

Techniques Used: Mouse Assay, AST Assay, Activity Assay, Immunohistochemistry, Staining

11) Product Images from "PARP Inhibitors Synergize with Gemcitabine by Potentiating DNA Damage in Non-Small Cell Lung Cancer"

Article Title: PARP Inhibitors Synergize with Gemcitabine by Potentiating DNA Damage in Non-Small Cell Lung Cancer

Journal: International journal of cancer

doi: 10.1002/ijc.31770

Induction of apoptosis in NSCLC cell lines by treatment with gemcitabine and BMN673 alone or combined. (a) Western blot showing changes in cleaved caspase-3 protein expression in H23 and SK-MES-1 cells after 48 hours of treatment with gemcitabine (GEM; 5 nM), BMN673 (50 nM), both agents, or DMSO vehicle control. (b) Quantification of changes in relative cleaved caspase-3 protein expression in H23 and SK-MES-1 cells after 48 hours of treatment with gemcitabine (5 nM), BMN673 (50 nM), both agents combined, or DMSO vehicle control. (c) Analysis of apoptosis in H23 and SK-MES-1 cells treated with BMN673 (50 nM), gemcitabine (5 nM), or both for 48 hours. The percentage of apoptotic cells was determined using Annexin V/propidium iodide staining. (d) Quantification of flow cytometry experiments from 2c.
Figure Legend Snippet: Induction of apoptosis in NSCLC cell lines by treatment with gemcitabine and BMN673 alone or combined. (a) Western blot showing changes in cleaved caspase-3 protein expression in H23 and SK-MES-1 cells after 48 hours of treatment with gemcitabine (GEM; 5 nM), BMN673 (50 nM), both agents, or DMSO vehicle control. (b) Quantification of changes in relative cleaved caspase-3 protein expression in H23 and SK-MES-1 cells after 48 hours of treatment with gemcitabine (5 nM), BMN673 (50 nM), both agents combined, or DMSO vehicle control. (c) Analysis of apoptosis in H23 and SK-MES-1 cells treated with BMN673 (50 nM), gemcitabine (5 nM), or both for 48 hours. The percentage of apoptotic cells was determined using Annexin V/propidium iodide staining. (d) Quantification of flow cytometry experiments from 2c.

Techniques Used: Western Blot, Expressing, Staining, Flow Cytometry

12) Product Images from "Vinyl chloride dysregulates metabolic homeostasis and enhances diet‐induced liver injury in mice"

Article Title: Vinyl chloride dysregulates metabolic homeostasis and enhances diet‐induced liver injury in mice

Journal: Hepatology Communications

doi: 10.1002/hep4.1151

Effect of VC on apoptosis. (A) Representative western blots and densitometric analysis for whole liver caspase‐3 protein are shown. (B) TUNEL staining was performed as described in Materials and Methods for the 12‐week samples. Representative photomicrographs are shown (magnification ×200). (C) TUNEL‐positive hepatocytes and NPCs were quantified as described in Materials and Methods and are expressed as TUNEL‐positive cells per 1,000 hepatocytes. a P
Figure Legend Snippet: Effect of VC on apoptosis. (A) Representative western blots and densitometric analysis for whole liver caspase‐3 protein are shown. (B) TUNEL staining was performed as described in Materials and Methods for the 12‐week samples. Representative photomicrographs are shown (magnification ×200). (C) TUNEL‐positive hepatocytes and NPCs were quantified as described in Materials and Methods and are expressed as TUNEL‐positive cells per 1,000 hepatocytes. a P

Techniques Used: Western Blot, TUNEL Assay, Staining

13) Product Images from "Downregulation of microRNA-21 inhibited radiation-resistance of esophageal squamous cell carcinoma"

Article Title: Downregulation of microRNA-21 inhibited radiation-resistance of esophageal squamous cell carcinoma

Journal: Cancer Cell International

doi: 10.1186/s12935-018-0502-6

Expression of cleaved-caspase-3 and cleaved-PARP in miR-21 mimics and inhibitor transfected-TE-1 cells after exposure of 6 Gy for 48 h. After transfection with miR-21 mimics and inhibitor, cells were irradiated with 6 Gy for 48 h. And then, the cells were collected and the levels of cleaved-caspase-3 and cleaved-PARP were detected by western blot
Figure Legend Snippet: Expression of cleaved-caspase-3 and cleaved-PARP in miR-21 mimics and inhibitor transfected-TE-1 cells after exposure of 6 Gy for 48 h. After transfection with miR-21 mimics and inhibitor, cells were irradiated with 6 Gy for 48 h. And then, the cells were collected and the levels of cleaved-caspase-3 and cleaved-PARP were detected by western blot

Techniques Used: Expressing, Transfection, Irradiation, Western Blot

14) Product Images from "The cryoprotectant trehalose could inhibit ERS-induced apoptosis by activating autophagy in cryoprotected rat valves"

Article Title: The cryoprotectant trehalose could inhibit ERS-induced apoptosis by activating autophagy in cryoprotected rat valves

Journal: PLoS ONE

doi: 10.1371/journal.pone.0194078

Effect of trehalose on apoptosis and ERS in cryopreserved valve cells. (A and B) Western blotting using a cleaved caspase-3 antibody on cryopreserved valve cells treated with trehalose (0.1 mol/L) or DMSO for 16 weeks. (C, D and E) Western blotting using GRP78 and CHOP antibodies on cryopreserved valves treated with trehalose (0.1 mol/L) for 16 weeks. Data are expressed as the mean ± SD ( n = 20 per group). ** P
Figure Legend Snippet: Effect of trehalose on apoptosis and ERS in cryopreserved valve cells. (A and B) Western blotting using a cleaved caspase-3 antibody on cryopreserved valve cells treated with trehalose (0.1 mol/L) or DMSO for 16 weeks. (C, D and E) Western blotting using GRP78 and CHOP antibodies on cryopreserved valves treated with trehalose (0.1 mol/L) for 16 weeks. Data are expressed as the mean ± SD ( n = 20 per group). ** P

Techniques Used: Western Blot

15) Product Images from "LRH-1 mitigates intestinal inflammatory disease by maintaining epithelial homeostasis and cell survival"

Article Title: LRH-1 mitigates intestinal inflammatory disease by maintaining epithelial homeostasis and cell survival

Journal: Nature Communications

doi: 10.1038/s41467-018-06137-w

In vivo rescue of Lrh1 IEC-KO mice by hLRH-1 reverses cell death. a LRH-1 protein levels in hLrh1 IEC-Flex enteroids detected by anti-LRH-1 antibody with arrowheads indicating migration of human (blue) or mouse (black) LRH-1 before or after addition of 4OHT, which eliminates mLRH-1 and promotes hLRH-1 expression; protein extracts were isolated 72 h later. b Relative levels of mLRH-1 and a downstream LRH-1 target gene, Ctrb1 in wild-type ( Lrh1 fl/fl ), Lrh1 IEC- KO , and hLrh1 IEC-Flex enteroids, with values normalized to wild type set at 1.0. Generation of hLrh1 IEC-Flex is described in Methods. For all panels, data were generated from three independent wells of enteroids (~50 organoids per well) done in triplicate. c Percentage of cell death in hLrh1 IEC-Flex enteroids with TNFα (10 ng/ml, 40 h) after eliminating mLRH-1 (gray) and expressing hLRH-1 (blue) by addition of 4OHT for 48 h. Data are also shown for treated Lrh1 fl/fl enteroids (black). All values are normalized to five independent wells of untreated Lrh1 fl/fl enteroids, which is taken to be 0%. d Immunofluorescence of wild-type ( Lrh1 fl/fl ), Lrh1 IEC-KO , and hLrh1 IEC-Flex ileum from adult male mice treated with two consecutive injections of tamoxifen. Staining for activated Casp3 (red) and CD44 (green), which marks intestinal epithelial crypt cells, is shown at lower (first column) and higher (second column) magnification. The appearance of apoptotic cells is indicated in the crypt region as well as the villus (white arrowheads and dashed white line) in Lrh1 IEC-KO ileum; some signal is also observed after expressing hLRH-1 in hLrh1 IEC-Flex . Scale bars = 50 μm. N = 2 per genotype. For panels b and c error bars are SEM with statistical analyses determined by Student’s unpaired t test, two tailed with p values of * p =
Figure Legend Snippet: In vivo rescue of Lrh1 IEC-KO mice by hLRH-1 reverses cell death. a LRH-1 protein levels in hLrh1 IEC-Flex enteroids detected by anti-LRH-1 antibody with arrowheads indicating migration of human (blue) or mouse (black) LRH-1 before or after addition of 4OHT, which eliminates mLRH-1 and promotes hLRH-1 expression; protein extracts were isolated 72 h later. b Relative levels of mLRH-1 and a downstream LRH-1 target gene, Ctrb1 in wild-type ( Lrh1 fl/fl ), Lrh1 IEC- KO , and hLrh1 IEC-Flex enteroids, with values normalized to wild type set at 1.0. Generation of hLrh1 IEC-Flex is described in Methods. For all panels, data were generated from three independent wells of enteroids (~50 organoids per well) done in triplicate. c Percentage of cell death in hLrh1 IEC-Flex enteroids with TNFα (10 ng/ml, 40 h) after eliminating mLRH-1 (gray) and expressing hLRH-1 (blue) by addition of 4OHT for 48 h. Data are also shown for treated Lrh1 fl/fl enteroids (black). All values are normalized to five independent wells of untreated Lrh1 fl/fl enteroids, which is taken to be 0%. d Immunofluorescence of wild-type ( Lrh1 fl/fl ), Lrh1 IEC-KO , and hLrh1 IEC-Flex ileum from adult male mice treated with two consecutive injections of tamoxifen. Staining for activated Casp3 (red) and CD44 (green), which marks intestinal epithelial crypt cells, is shown at lower (first column) and higher (second column) magnification. The appearance of apoptotic cells is indicated in the crypt region as well as the villus (white arrowheads and dashed white line) in Lrh1 IEC-KO ileum; some signal is also observed after expressing hLRH-1 in hLrh1 IEC-Flex . Scale bars = 50 μm. N = 2 per genotype. For panels b and c error bars are SEM with statistical analyses determined by Student’s unpaired t test, two tailed with p values of * p =

Techniques Used: In Vivo, Mouse Assay, Migration, Expressing, Isolation, Generated, Immunofluorescence, Staining, Two Tailed Test

Rescue of Lrh1 IEC-KO mouse organoids by hLRH-1. a Ribbon diagrams of mouse (left) and human (middle) LRH-1 ligand-binding pocket highlighting species-specific structural features of salt bridge (dotted black line and red arrowhead) versus coordination of phospholipid ligand (red stick; dotted black lines). Model of hLRH-1 pocket mutant (right) showing placement of pocket-obscuring residues (gold). b AAV8-directed GFP expression in organoids after 24 h (BF brightfield, GFP fluorescence, white arrowhead indicates representative GFP + cell). Nuclear hLRH-1 expression 4 d post-infection detected with anti-Flag. Scale bar = 100 μm. c Titration of hLRH-1 protein by infectious titer of AAV8-hLRH1 (3.3×10 10 − 4.1×10 9 genome copies) after 4 d (left); mock infection is without virus. LRH-1 detected by anti-LRH-1 (upper panel) or anti-FLAG (middle panel). NS: nonspecific band. Western blot for AAV-hLRH1 and AAV-PM detected by FLAG antibody from Lrh1 fl/fl organoids infected with equal titer of AAV (right). d Casp3 signal in untreated Lrh1 IEC-KO organoid crypts infected with either AAV-GFP or AAV-hLRH1 with Casp3 + cells indicated (white arrowheads). Scale bar = 50 μm. e Expression of active Casp3 protein in Lrh1 IEC-KO organoids infected with AAV-GFP or AAV-hLRH1 for 72 h prior to TNFα (10 ng/ml, 40 h). f Percent cell death in Lrh1 IEC-KO organoids infected with mock (gray), AAV-GFP (green), AAV-hLRH1 (blue), or AAV-hPM (hLRH-1 pocket mutant; light blue) for 72 h, and then treated with TNFα (10 ng/ml, 40 h). g Fold change by RT-qPCR in Lrh1 fl/fl (black) and Lrh1 IEC-KO (gray) organoids, or in Lrh1 IEC-KO organoids subsequently infected with AAV-GFP (green) or AAV-hLRH1 (blue) for 72 h. h Uptake of fluorescent dextran in Lrh1 IEC-KO organoids infected with AAV-GFP or AAV-hLRH1 for 72 h followed by TNFα (10 ng/ml, 40 h) as per Fig. 1h . Scale bar = 100 μm. i Viability of TNFα-exposed Lrh1 fl/fl organoids (20 ng/ml, 40 h) and 5-FU (5 μg/ml, 24 h), respectively, overexpressing hLRH-1 by approximately two times endogenous levels. Control organoids were infected with AAV-GFP (black bar). For panels f − i error bars are SEM with statistical analyses as per Fig. 1 with * p =
Figure Legend Snippet: Rescue of Lrh1 IEC-KO mouse organoids by hLRH-1. a Ribbon diagrams of mouse (left) and human (middle) LRH-1 ligand-binding pocket highlighting species-specific structural features of salt bridge (dotted black line and red arrowhead) versus coordination of phospholipid ligand (red stick; dotted black lines). Model of hLRH-1 pocket mutant (right) showing placement of pocket-obscuring residues (gold). b AAV8-directed GFP expression in organoids after 24 h (BF brightfield, GFP fluorescence, white arrowhead indicates representative GFP + cell). Nuclear hLRH-1 expression 4 d post-infection detected with anti-Flag. Scale bar = 100 μm. c Titration of hLRH-1 protein by infectious titer of AAV8-hLRH1 (3.3×10 10 − 4.1×10 9 genome copies) after 4 d (left); mock infection is without virus. LRH-1 detected by anti-LRH-1 (upper panel) or anti-FLAG (middle panel). NS: nonspecific band. Western blot for AAV-hLRH1 and AAV-PM detected by FLAG antibody from Lrh1 fl/fl organoids infected with equal titer of AAV (right). d Casp3 signal in untreated Lrh1 IEC-KO organoid crypts infected with either AAV-GFP or AAV-hLRH1 with Casp3 + cells indicated (white arrowheads). Scale bar = 50 μm. e Expression of active Casp3 protein in Lrh1 IEC-KO organoids infected with AAV-GFP or AAV-hLRH1 for 72 h prior to TNFα (10 ng/ml, 40 h). f Percent cell death in Lrh1 IEC-KO organoids infected with mock (gray), AAV-GFP (green), AAV-hLRH1 (blue), or AAV-hPM (hLRH-1 pocket mutant; light blue) for 72 h, and then treated with TNFα (10 ng/ml, 40 h). g Fold change by RT-qPCR in Lrh1 fl/fl (black) and Lrh1 IEC-KO (gray) organoids, or in Lrh1 IEC-KO organoids subsequently infected with AAV-GFP (green) or AAV-hLRH1 (blue) for 72 h. h Uptake of fluorescent dextran in Lrh1 IEC-KO organoids infected with AAV-GFP or AAV-hLRH1 for 72 h followed by TNFα (10 ng/ml, 40 h) as per Fig. 1h . Scale bar = 100 μm. i Viability of TNFα-exposed Lrh1 fl/fl organoids (20 ng/ml, 40 h) and 5-FU (5 μg/ml, 24 h), respectively, overexpressing hLRH-1 by approximately two times endogenous levels. Control organoids were infected with AAV-GFP (black bar). For panels f − i error bars are SEM with statistical analyses as per Fig. 1 with * p =

Techniques Used: Ligand Binding Assay, Mutagenesis, Expressing, Fluorescence, Infection, Titration, Western Blot, Quantitative RT-PCR

16) Product Images from "Cell death‐inducing DFF45‐like effector C gene silencing alleviates pulmonary vascular remodeling in a type 2 diabetic rat model"

Article Title: Cell death‐inducing DFF45‐like effector C gene silencing alleviates pulmonary vascular remodeling in a type 2 diabetic rat model

Journal: Journal of Diabetes Investigation

doi: 10.1111/jdi.12768

(a) Cleaved caspase‐3 and cleaved poly (adenosine diphosphate‐ribose) polymerase ( PARP ) in lung tissue measured by western blotting. (b) Cleaved caspase‐3 (** P
Figure Legend Snippet: (a) Cleaved caspase‐3 and cleaved poly (adenosine diphosphate‐ribose) polymerase ( PARP ) in lung tissue measured by western blotting. (b) Cleaved caspase‐3 (** P

Techniques Used: Western Blot

17) Product Images from "A phase II clinical trial of the Aurora and angiogenic kinase inhibitor ENMD-2076 for previously treated, advanced, or metastatic triple-negative breast cancer"

Article Title: A phase II clinical trial of the Aurora and angiogenic kinase inhibitor ENMD-2076 for previously treated, advanced, or metastatic triple-negative breast cancer

Journal: Breast Cancer Research : BCR

doi: 10.1186/s13058-018-1014-y

Effects of ENMD-2076 on pharmacodynamic markers in serial tumor biopsies obtained in a subset of patients. a Paired samples were available for eight patients at baseline prior to dosing (C1D1) and postdose on days 14–16 (C1D15). An additional sample was obtained in one patient who experienced stable disease for ten cycles followed by progression (end of treatment [EOT]). Tissue was analyzed by IHC for Ki-67 as a marker of cellular proliferation. b Nonresponder. Staining was performed as in panel a . Note that there is no decrease in proliferation or increase in apoptosis in the nonresponder following ENMD-2076 treatment. Samples were analyzed by IHC for CD34 expression as a marker of microvessel density. Changes were independent of tumor response and clinical benefit to ENMD-2076 treatment. Patients 01-005, 01-030, 02-004, 02-006, 02-012, and 02-027 had progressive disease (PD) at first imaging assessment following two cycles; 01-028 had stable disease (SD) for ten cycles; and 01-031 had SD for four cycles. c Immunoflurorescence analysis of tumor biopsies for 4′,6-diamidino-2-phenylindole (DAPI), p53, and p73 in a patient who had stable disease by Response Evaluation Criteria in Solid Tumors (RECIST version 1.1) after two cycles of treatment and then progressed after cycle 3. Patient has a p53 mutation R273S. Note an increase in p53 and p73 following treatment, which is consistent with preclinical findings in patient-derived tumor xenograft models. IHC images from 01-028 responder. Cleaved caspase 3, Ki-67, and CD34 on serial tumor biopsies were used to assess apoptosis, proliferation, and microvessel density, respectively, in a patient responding to ENMD-2076 treatment with prolonged stable disease for ten cycles. Biopsies were obtained prior to treatment, 15 days after treatment, and at the time of disease progression day 280. Formalin-fixed, paraffin-embedded tissue sections were stained with the indicated antibodies, and representative images were obtained at × 20 magnification. Note an increase in cleaved caspase 3 and a decrease in Ki-67 and CD34 in the posttreatment biopsy. At the time of disease progression, these changes were reversed. Changes in ( d ) Ki-67 and ( e ) CD34 (microvessel density) in serial tumor biopsies. Baseline and day 15 samples were available for eight patients. An additional sample was obtained from one patient at the time of progression following prolonged stable disease. Ki-67 and CD34 were assessed using IHC. SA-β-gal Senescence-associated β-galactosidase
Figure Legend Snippet: Effects of ENMD-2076 on pharmacodynamic markers in serial tumor biopsies obtained in a subset of patients. a Paired samples were available for eight patients at baseline prior to dosing (C1D1) and postdose on days 14–16 (C1D15). An additional sample was obtained in one patient who experienced stable disease for ten cycles followed by progression (end of treatment [EOT]). Tissue was analyzed by IHC for Ki-67 as a marker of cellular proliferation. b Nonresponder. Staining was performed as in panel a . Note that there is no decrease in proliferation or increase in apoptosis in the nonresponder following ENMD-2076 treatment. Samples were analyzed by IHC for CD34 expression as a marker of microvessel density. Changes were independent of tumor response and clinical benefit to ENMD-2076 treatment. Patients 01-005, 01-030, 02-004, 02-006, 02-012, and 02-027 had progressive disease (PD) at first imaging assessment following two cycles; 01-028 had stable disease (SD) for ten cycles; and 01-031 had SD for four cycles. c Immunoflurorescence analysis of tumor biopsies for 4′,6-diamidino-2-phenylindole (DAPI), p53, and p73 in a patient who had stable disease by Response Evaluation Criteria in Solid Tumors (RECIST version 1.1) after two cycles of treatment and then progressed after cycle 3. Patient has a p53 mutation R273S. Note an increase in p53 and p73 following treatment, which is consistent with preclinical findings in patient-derived tumor xenograft models. IHC images from 01-028 responder. Cleaved caspase 3, Ki-67, and CD34 on serial tumor biopsies were used to assess apoptosis, proliferation, and microvessel density, respectively, in a patient responding to ENMD-2076 treatment with prolonged stable disease for ten cycles. Biopsies were obtained prior to treatment, 15 days after treatment, and at the time of disease progression day 280. Formalin-fixed, paraffin-embedded tissue sections were stained with the indicated antibodies, and representative images were obtained at × 20 magnification. Note an increase in cleaved caspase 3 and a decrease in Ki-67 and CD34 in the posttreatment biopsy. At the time of disease progression, these changes were reversed. Changes in ( d ) Ki-67 and ( e ) CD34 (microvessel density) in serial tumor biopsies. Baseline and day 15 samples were available for eight patients. An additional sample was obtained from one patient at the time of progression following prolonged stable disease. Ki-67 and CD34 were assessed using IHC. SA-β-gal Senescence-associated β-galactosidase

Techniques Used: Immunohistochemistry, Marker, Staining, Expressing, Imaging, Mutagenesis, Derivative Assay, Formalin-fixed Paraffin-Embedded

18) Product Images from "Moderate hypothermia inhibits microglial activation after traumatic brain injury by modulating autophagy/apoptosis and the MyD88-dependent TLR4 signaling pathway"

Article Title: Moderate hypothermia inhibits microglial activation after traumatic brain injury by modulating autophagy/apoptosis and the MyD88-dependent TLR4 signaling pathway

Journal: Journal of Neuroinflammation

doi: 10.1186/s12974-018-1315-1

Immunofluorescence analysis of cleaved caspase 3 (red) and Iba-1 (green) from the injured cortex and ipsilateral hippocampus. a Immunohistochemical staining of cleaved caspase 3 and Iba-1 from the injured cortex and ipsilateral hippocampus. Arrows indicate co-localization of cleaved caspase 3 and Iba-1 (magnification, × 630). b Number of cleaved caspase 3-positive microglia in the injured cortex and ipsilateral hippocampus. At least 10 randomly selected microscopic fields were used for counting. Data in the bar graphs represent mean ± SD. **** P
Figure Legend Snippet: Immunofluorescence analysis of cleaved caspase 3 (red) and Iba-1 (green) from the injured cortex and ipsilateral hippocampus. a Immunohistochemical staining of cleaved caspase 3 and Iba-1 from the injured cortex and ipsilateral hippocampus. Arrows indicate co-localization of cleaved caspase 3 and Iba-1 (magnification, × 630). b Number of cleaved caspase 3-positive microglia in the injured cortex and ipsilateral hippocampus. At least 10 randomly selected microscopic fields were used for counting. Data in the bar graphs represent mean ± SD. **** P

Techniques Used: Immunofluorescence, Immunohistochemistry, Staining

Immunofluorescence analysis and Western blotting of cleaved caspase 3 expression in the injured cortex and ipsilateral hippocampus. a Number of cleaved caspase 3-positive cells in the injured cortex and ipsilateral hippocampus. Data in the bar graphs represent mean ± SD. **** P
Figure Legend Snippet: Immunofluorescence analysis and Western blotting of cleaved caspase 3 expression in the injured cortex and ipsilateral hippocampus. a Number of cleaved caspase 3-positive cells in the injured cortex and ipsilateral hippocampus. Data in the bar graphs represent mean ± SD. **** P

Techniques Used: Immunofluorescence, Western Blot, Expressing

19) Product Images from "The Role of IL-6RA in UHMWPE Promotes Proliferation in Fibro-Like Synovial Cells"

Article Title: The Role of IL-6RA in UHMWPE Promotes Proliferation in Fibro-Like Synovial Cells

Journal: BioMed Research International

doi: 10.1155/2018/3928915

Effects of UHMWPE on apoptosis of FLS cells. FLS were incubated with 0-1.0 g/L UHMWPE for 0, 1, and 7 d or incubated with 1.0 g/L UHMWPE for 0, 1, and 7 d. Apoptotic and necrotic cell populations were analyzed by flow cytometry; quantitative analysis of apoptotic cells after UHMWPE treatment, n=5 (a and b). The expression of apoptosis-related proteins in FLS cells was detected by Western blot assay and the changes of Bcl-2, Bax, caspase-3, and cleaved-caspase-3 were statistically analyzed, n=5 (c). Data was obtained from three independent experiments; results were shown as means ± standard deviation. ∗ p
Figure Legend Snippet: Effects of UHMWPE on apoptosis of FLS cells. FLS were incubated with 0-1.0 g/L UHMWPE for 0, 1, and 7 d or incubated with 1.0 g/L UHMWPE for 0, 1, and 7 d. Apoptotic and necrotic cell populations were analyzed by flow cytometry; quantitative analysis of apoptotic cells after UHMWPE treatment, n=5 (a and b). The expression of apoptosis-related proteins in FLS cells was detected by Western blot assay and the changes of Bcl-2, Bax, caspase-3, and cleaved-caspase-3 were statistically analyzed, n=5 (c). Data was obtained from three independent experiments; results were shown as means ± standard deviation. ∗ p

Techniques Used: Incubation, Flow Cytometry, Cytometry, Expressing, Western Blot, Standard Deviation

The antagonistic effect of IL-6RA on FLS cells treated with UHMWPE for 7 d. The experiment was divided into 3 groups, including UHMWPE (1 g/L), UHMWPE (1 g/L) + IL-6RA (50 μ g/ml), and IL-6RA(50 μ g/ml). Apoptotic rate was detected by flow cytometry after treatment IL-6RA, n=5 (a). The expression levels of apoptosis-related proteins including Bax, Bcl2, caspase-3, and cleaved-caspase-3 were analyzed by Western blot after treatment IL-6RA, n=5 (b). Data was obtained from three independent experiments; results were shown as means ± standard deviation. ∗ p
Figure Legend Snippet: The antagonistic effect of IL-6RA on FLS cells treated with UHMWPE for 7 d. The experiment was divided into 3 groups, including UHMWPE (1 g/L), UHMWPE (1 g/L) + IL-6RA (50 μ g/ml), and IL-6RA(50 μ g/ml). Apoptotic rate was detected by flow cytometry after treatment IL-6RA, n=5 (a). The expression levels of apoptosis-related proteins including Bax, Bcl2, caspase-3, and cleaved-caspase-3 were analyzed by Western blot after treatment IL-6RA, n=5 (b). Data was obtained from three independent experiments; results were shown as means ± standard deviation. ∗ p

Techniques Used: Flow Cytometry, Cytometry, Expressing, Western Blot, Standard Deviation

20) Product Images from "Excess Hydrocortisone Hampers Placental Nutrient Uptake Disrupting Cellular Metabolism"

Article Title: Excess Hydrocortisone Hampers Placental Nutrient Uptake Disrupting Cellular Metabolism

Journal: BioMed Research International

doi: 10.1155/2018/5106174

Assessment of hydrocortisone-induced cellular death by apoptosis in healthy-term placental explants. (a) Representative microphotographs of TUNEL assay to determine apoptosis in each experimental condition; cells undergoing apoptosis present fluorescent green nuclei. (b) Quantification of the % of TUNEL positive cells per condition. (c) Active caspases 3 and 7 as measured by luminometry, using the Caspase Glo Assay. (d) Cleaved caspase 3 as determined by western blot and normalized by actin levels. Values are presented as means ± SEM for 5 independent experiments. ∗ p≤0.05; ∗∗ p≤0.01; ∗∗∗ p≤0.001 relative to control.
Figure Legend Snippet: Assessment of hydrocortisone-induced cellular death by apoptosis in healthy-term placental explants. (a) Representative microphotographs of TUNEL assay to determine apoptosis in each experimental condition; cells undergoing apoptosis present fluorescent green nuclei. (b) Quantification of the % of TUNEL positive cells per condition. (c) Active caspases 3 and 7 as measured by luminometry, using the Caspase Glo Assay. (d) Cleaved caspase 3 as determined by western blot and normalized by actin levels. Values are presented as means ± SEM for 5 independent experiments. ∗ p≤0.05; ∗∗ p≤0.01; ∗∗∗ p≤0.001 relative to control.

Techniques Used: TUNEL Assay, Caspase-Glo Assay, Western Blot

21) Product Images from "Effects of Several Cosmetic Preservatives on ROS-Dependent Apoptosis of Rat Neural Progenitor Cells"

Article Title: Effects of Several Cosmetic Preservatives on ROS-Dependent Apoptosis of Rat Neural Progenitor Cells

Journal: Biomolecules & Therapeutics

doi: 10.4062/biomolther.2017.221

Apoptotic effect of preservatives on NPCs. The apoptotic protein expression was measured at 24 h exposure after treatment with NPCs. Cleaved caspase-3 and cleaved caspase-8 were quantified by densitometric procedure reflected as bar graphs. Values are expressed as the mean ± SEM (N=5). * p
Figure Legend Snippet: Apoptotic effect of preservatives on NPCs. The apoptotic protein expression was measured at 24 h exposure after treatment with NPCs. Cleaved caspase-3 and cleaved caspase-8 were quantified by densitometric procedure reflected as bar graphs. Values are expressed as the mean ± SEM (N=5). * p

Techniques Used: Expressing

22) Product Images from "Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain"

Article Title: Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain

Journal: Pediatric research

doi: 10.1038/pr.2018.49

Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.
Figure Legend Snippet: Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.

Techniques Used: Expressing, Western Blot

Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.
Figure Legend Snippet: Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.

Techniques Used: Expressing, Western Blot, Staining

23) Product Images from "Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain"

Article Title: Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain

Journal: Pediatric research

doi: 10.1038/pr.2018.49

Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.
Figure Legend Snippet: Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.

Techniques Used: Expressing, Western Blot

Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.
Figure Legend Snippet: Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.

Techniques Used: Expressing, Western Blot, Staining

24) Product Images from "Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain"

Article Title: Up-regulation of cholesterol 24-hydroxylase following hypoxia-ischemia in neonatal mouse brain

Journal: Pediatric research

doi: 10.1038/pr.2018.49

Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.
Figure Legend Snippet: Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr and 24hr after neonatal HI. A. Protein expression of cleaved caspase-3 in the cortices was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to an internal control (IC). Sham vs. HI, *p=0.0112 at 6hr; *p=0.0027 at 24hr; *p=0.0228 at 72hr (n=5–6 for sham animals, n=7–10 for HI animals from 6–72hr). B. Correlation of the serum levels of 24S-HC with the expression of cleaved caspase-3 at 6hr (top) and at 24hr (bottom). Sample number, R 2 and p values are shown in the graphs.

Techniques Used: Expressing, Western Blot

Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.
Figure Legend Snippet: Up-regulation of CYP46A1 with a concomitant increase of 24S-HC in the ipsilateral cortex after neonatal HI. A. Protein expression of CYP46A1 was measured by western blotting at the indicated time points and presented as the OD ratio to β-actin and normalized to the values of sham 0hr (graph on the right, sham vs. HI: * p= 0.0339 at 6hr; * p=0.0026 at 24hr; n=5–6 for sham animals, n=6–12 for HI animals at 6–72hr). B. Immunofluorescent staining with anti-CYP46A1 antibody at 24hr after HI showed enhanced expression in the ipsilateral cortex than that in the contralateral side and in the sham animals (left panel s). CYP46A1 staining in the ipsi-cortex was largely co-localized with the expression of cleaved caspase-3 (middle panels). C. Increased production of 24S-HC (ng/mg tissue wet weight) in the ipsilateral cortex at 6hr and 24hr after HI (sham vs. HI: *p= 0.0167 at 6hr; *p=0.0085 at 24hr; n=3–6 for sham animals, n=5–7 for HI animals at 6–72hr). The time course of the changes in 24S-HC in the sham-, contra- and ipsilateral cortex is shown on the right. At 6hr, 24S-HC in the ipsilateral cortex was higher than that in the contralateral side (ipsi- vs. contra-: *p=0.0493). #: Significant difference between the HI ipsi- and the sham animals only, but not between the HI ipsi- and HI contralateral hemisphere.

Techniques Used: Expressing, Western Blot, Staining

25) Product Images from "Valproic acid enhances the viability of random pattern skin flaps: involvement of enhancing angiogenesis and inhibiting oxidative stress and apoptosis"

Article Title: Valproic acid enhances the viability of random pattern skin flaps: involvement of enhancing angiogenesis and inhibiting oxidative stress and apoptosis

Journal: Drug Design, Development and Therapy

doi: 10.2147/DDDT.S186222

VPA reduced apoptosis in random skin flaps. Notes: ( A ) Cleaved CASP3 expression in each group as assessed by IHC (original magnification ×200). ( B ) Protein expression of cleaved CASP3 in each group as assessed by Western blot analysis. The gels have been run under the same experimental conditions, and cropped blots are used here. ( C ) The optical density values of cleaved CASP3. ( D ) Densitometry result of cleaved CASP3 protein expression in the two groups. Values are expressed as mean ± SEM, n=6 per group. * P
Figure Legend Snippet: VPA reduced apoptosis in random skin flaps. Notes: ( A ) Cleaved CASP3 expression in each group as assessed by IHC (original magnification ×200). ( B ) Protein expression of cleaved CASP3 in each group as assessed by Western blot analysis. The gels have been run under the same experimental conditions, and cropped blots are used here. ( C ) The optical density values of cleaved CASP3. ( D ) Densitometry result of cleaved CASP3 protein expression in the two groups. Values are expressed as mean ± SEM, n=6 per group. * P

Techniques Used: Expressing, Immunohistochemistry, Western Blot

26) Product Images from "Icotinib inhibits the proliferation of hepatocellular carcinoma cells in vitro and in vivo dependently on EGFR activation and PDL1 expression"

Article Title: Icotinib inhibits the proliferation of hepatocellular carcinoma cells in vitro and in vivo dependently on EGFR activation and PDL1 expression

Journal: OncoTargets and therapy

doi: 10.2147/OTT.S179844

Effect of icotinib on expression of C-caspase 3, BCL2, p-EGFR, and PDL1 in SMMC7721 and Huh7 cells. Notes:  ( A ) Representative Western blot images; ( B ) quantification analysis of protein expression by ImageJ. ** P
Figure Legend Snippet: Effect of icotinib on expression of C-caspase 3, BCL2, p-EGFR, and PDL1 in SMMC7721 and Huh7 cells. Notes: ( A ) Representative Western blot images; ( B ) quantification analysis of protein expression by ImageJ. ** P

Techniques Used: Expressing, Western Blot

27) Product Images from "The HDAC Inhibitor, SAHA, Combined with Cisplatin Synergistically Induces Apoptosis in Alpha-fetoprotein-producing Hepatoid Adenocarcinoma Cells"

Article Title: The HDAC Inhibitor, SAHA, Combined with Cisplatin Synergistically Induces Apoptosis in Alpha-fetoprotein-producing Hepatoid Adenocarcinoma Cells

Journal: Acta Histochemica et Cytochemica

doi: 10.1267/ahc.18044

Effect of cisplatin and SAHA on induction of apoptotic cell death in VAT-39 cells. ( A ) Immunohistochemical localization of cleaved caspase-3 in 5 μM cisplatin and 2 μM SAHA-treated VAT-39 cells. ( B ) Counts of cleaved caspase-3-positive cells are shown in the bar graph. ( C ) Western blot analysis detected double bands of cleaved caspase-3 (17 kDa and 19 kDa). Isolated proteins (20 μg) were subjected to SDS-PAGE. β-actin (42 kDa) was used as a loading control. ( D ) Cell death was examined by TUNEL assay using the Mebstain apoptosis TUNEL kit. Arrows indicate TUNEL-positive cells among cisplatin and SAHA-treated cells. *** P
Figure Legend Snippet: Effect of cisplatin and SAHA on induction of apoptotic cell death in VAT-39 cells. ( A ) Immunohistochemical localization of cleaved caspase-3 in 5 μM cisplatin and 2 μM SAHA-treated VAT-39 cells. ( B ) Counts of cleaved caspase-3-positive cells are shown in the bar graph. ( C ) Western blot analysis detected double bands of cleaved caspase-3 (17 kDa and 19 kDa). Isolated proteins (20 μg) were subjected to SDS-PAGE. β-actin (42 kDa) was used as a loading control. ( D ) Cell death was examined by TUNEL assay using the Mebstain apoptosis TUNEL kit. Arrows indicate TUNEL-positive cells among cisplatin and SAHA-treated cells. *** P

Techniques Used: Immunohistochemistry, Western Blot, Isolation, SDS Page, TUNEL Assay

28) Product Images from "Development of a fluorescence-based cellular apoptosis reporter"

Article Title: Development of a fluorescence-based cellular apoptosis reporter

Journal: Methods and Applications in Fluorescence

doi: 10.1088/2050-6120/aae6f8

Cellular distribution of NGD6 in SCC cell line. Representative confocal images of SCC NGD6 cells untreated or treated with 250 nM staurosporine for 24 h (STS). Blue = nuclei stained with Hoechst; Green = cellular distribution of GFP construct; Red = Cleaved caspase-3 immunofluorescence; Merge = overlay of channels. Scale bars = 10 μ m.
Figure Legend Snippet: Cellular distribution of NGD6 in SCC cell line. Representative confocal images of SCC NGD6 cells untreated or treated with 250 nM staurosporine for 24 h (STS). Blue = nuclei stained with Hoechst; Green = cellular distribution of GFP construct; Red = Cleaved caspase-3 immunofluorescence; Merge = overlay of channels. Scale bars = 10 μ m.

Techniques Used: Staining, Construct, Immunofluorescence

Activation of caspase-3 in 4T1 and SCC cells. (A) Cell lysates from 4T1 cells treated with increasing concentrations of doxorubicin for 24 h, or (B) 4T1 cells treated with 4 μ M doxorubicin for given time periods, or (C) SCC cells treated with increasing concentrations of staursporine for 24 h, or (D) SCC cells treated with 250 nM staurosporine for given time periods, were subjected to western blot analysis using cleaved caspase-3 and PARP antibodies. All membranes were stripped and re-probed with β -actin as a loading control.
Figure Legend Snippet: Activation of caspase-3 in 4T1 and SCC cells. (A) Cell lysates from 4T1 cells treated with increasing concentrations of doxorubicin for 24 h, or (B) 4T1 cells treated with 4 μ M doxorubicin for given time periods, or (C) SCC cells treated with increasing concentrations of staursporine for 24 h, or (D) SCC cells treated with 250 nM staurosporine for given time periods, were subjected to western blot analysis using cleaved caspase-3 and PARP antibodies. All membranes were stripped and re-probed with β -actin as a loading control.

Techniques Used: Activation Assay, Western Blot

Design and generation of pCasFSwitch. (A) pCasFSwitch is a fluorescent reporter construct designed to identify cells undergoing caspase-3 mediated apoptosis by a switch in a GFP signal from the plasma membrane of the cell to the nucleus. Schematic representations of the constructs showing the caspase-3 cleavage site in red. Probe nomenclature shown beside each schematic. (B) pCasFSwitch was constructed from pEGFP-N1. Step 1: insertion of two or three nuclear localization sequences (NLS) at the N-terminal of GFP, to generate p2NLS-GFP and p3NLS-GFP respectively. White arrows indicate nuclear GFP, and white arrowheads show cytosolic GFP. Step 2: insertion of plasma membrane localization sequences (PLS) at the C-terminal of GFP, using the hexalysine stretch employed by K-Ras4B, or the hypervariable domain of H-Ras, yielding the intermediate constructs pNLS-GFP-6KPLS and pNLS-GFP-HRPLS respectively. Step3: insertion of the caspase-3 cleavage domain between GFP and the PLS to generate the final constructs, pNLS-GFP-DEVDG-6KPLS (NGD6) and pNLS-GFP-DEVDG-HRPLS (NGDH). Functionality testing for each of the constructs was carried out by transient transfection into HEK293T cells and analysis by confocal microscopy. Representative images for each of the intermediate constructs is shown below the appropriate construct schematic. Blue = nuclei stained with Hoechst; Green = cellular distribution of the GFP construct. Merge = overlay of blue and green channels superimposed on DIC image. Scale bars = 10 μ m.
Figure Legend Snippet: Design and generation of pCasFSwitch. (A) pCasFSwitch is a fluorescent reporter construct designed to identify cells undergoing caspase-3 mediated apoptosis by a switch in a GFP signal from the plasma membrane of the cell to the nucleus. Schematic representations of the constructs showing the caspase-3 cleavage site in red. Probe nomenclature shown beside each schematic. (B) pCasFSwitch was constructed from pEGFP-N1. Step 1: insertion of two or three nuclear localization sequences (NLS) at the N-terminal of GFP, to generate p2NLS-GFP and p3NLS-GFP respectively. White arrows indicate nuclear GFP, and white arrowheads show cytosolic GFP. Step 2: insertion of plasma membrane localization sequences (PLS) at the C-terminal of GFP, using the hexalysine stretch employed by K-Ras4B, or the hypervariable domain of H-Ras, yielding the intermediate constructs pNLS-GFP-6KPLS and pNLS-GFP-HRPLS respectively. Step3: insertion of the caspase-3 cleavage domain between GFP and the PLS to generate the final constructs, pNLS-GFP-DEVDG-6KPLS (NGD6) and pNLS-GFP-DEVDG-HRPLS (NGDH). Functionality testing for each of the constructs was carried out by transient transfection into HEK293T cells and analysis by confocal microscopy. Representative images for each of the intermediate constructs is shown below the appropriate construct schematic. Blue = nuclei stained with Hoechst; Green = cellular distribution of the GFP construct. Merge = overlay of blue and green channels superimposed on DIC image. Scale bars = 10 μ m.

Techniques Used: Construct, Transfection, Confocal Microscopy, Staining

Cleavage analysis of NGD6 in SCC cell line. (A) Schematic representation of caspase-3 mediated cleavage of NGD6 showing the un-cleaved and cleaved structures. Probe nomenclature and molecular weight shown beside each schematic. (B) Lysates from; SCC cells, SCC NGD6 cells left untreated or treated with 250 nM staurosporine for 24 h (STS), and untreated SCC NGD6 cells incubated with recombinant caspase-3 (C3) or Cathepsin B (CB) for given time periods (as indicated), were subjected to western blot analysis using GFP, cleaved caspase-3 and PARP antibodies. All membranes were stripped and re-probed with β -actin as a loading control.
Figure Legend Snippet: Cleavage analysis of NGD6 in SCC cell line. (A) Schematic representation of caspase-3 mediated cleavage of NGD6 showing the un-cleaved and cleaved structures. Probe nomenclature and molecular weight shown beside each schematic. (B) Lysates from; SCC cells, SCC NGD6 cells left untreated or treated with 250 nM staurosporine for 24 h (STS), and untreated SCC NGD6 cells incubated with recombinant caspase-3 (C3) or Cathepsin B (CB) for given time periods (as indicated), were subjected to western blot analysis using GFP, cleaved caspase-3 and PARP antibodies. All membranes were stripped and re-probed with β -actin as a loading control.

Techniques Used: Molecular Weight, Incubation, Recombinant, Western Blot

Cellular distribution of NGN6 in SCC cell line. Representative confocal images of SCC NGN6 cells untreated or treated with 250 nM staurosporine for 24 h (STS). Blue = nuclei stained with Hoechst; Green = cellular distribution of GFP construct; Red = Cleaved caspase-3 immunofluorescence; Merge = overlay of both channels. Scale bars = 10 μ m.
Figure Legend Snippet: Cellular distribution of NGN6 in SCC cell line. Representative confocal images of SCC NGN6 cells untreated or treated with 250 nM staurosporine for 24 h (STS). Blue = nuclei stained with Hoechst; Green = cellular distribution of GFP construct; Red = Cleaved caspase-3 immunofluorescence; Merge = overlay of both channels. Scale bars = 10 μ m.

Techniques Used: Staining, Construct, Immunofluorescence

29) Product Images from "Magnetic multiwalled carbon nanotubes with controlled release of epirubicin: an intravesical instillation system for bladder cancer"

Article Title: Magnetic multiwalled carbon nanotubes with controlled release of epirubicin: an intravesical instillation system for bladder cancer

Journal: International Journal of Nanomedicine

doi: 10.2147/IJN.S189688

Immunohistochemical staining of exemplary tissue sections of bladders in different groups and statistical analysis of the IOD/area data. Notes:  ( A ) Representative light-microscopy sections of BCL2, BAX, and cleaved caspase 3-stained bladder tissue; ( B ) statistical analysis of IOD/area data of bladder-tissue sections for BCL2, BAX, and cleaved caspase 3. * P
Figure Legend Snippet: Immunohistochemical staining of exemplary tissue sections of bladders in different groups and statistical analysis of the IOD/area data. Notes: ( A ) Representative light-microscopy sections of BCL2, BAX, and cleaved caspase 3-stained bladder tissue; ( B ) statistical analysis of IOD/area data of bladder-tissue sections for BCL2, BAX, and cleaved caspase 3. * P

Techniques Used: Immunohistochemistry, Staining, Light Microscopy

30) Product Images from "Farnesol, a Fungal Quorum-Sensing Molecule Triggers Apoptosis in Human Oral Squamous Carcinoma Cells"

Article Title: Farnesol, a Fungal Quorum-Sensing Molecule Triggers Apoptosis in Human Oral Squamous Carcinoma Cells

Journal: Neoplasia (New York, N.Y.)

doi:

Western blot analysis: Assessment of OSCC 9 cell protein expression, all after 48 hours of treatment, demonstrating (A) a decrease in survivin expression and (B) an increase in cleaved-caspase 3 and cleaved-caspase 9 expression in cells exposed to synthetic 30- to 60- µ M farnesol (F30, F60) or  C. albicans  spent culture media grown for 48 to 72 hours (CA48h, CA72h), without a change in expression with the non-farnesol-producing  C. albicans  spent culture media grown for 24 to 72 hours (NF24, NF48h, NF72h). The restoration of caspase expression to baseline levels is seen in the presence of 48 hours of combined DAG and farnesol (DAG-F60) treatment. Actin was used as the loading control.
Figure Legend Snippet: Western blot analysis: Assessment of OSCC 9 cell protein expression, all after 48 hours of treatment, demonstrating (A) a decrease in survivin expression and (B) an increase in cleaved-caspase 3 and cleaved-caspase 9 expression in cells exposed to synthetic 30- to 60- µ M farnesol (F30, F60) or C. albicans spent culture media grown for 48 to 72 hours (CA48h, CA72h), without a change in expression with the non-farnesol-producing C. albicans spent culture media grown for 24 to 72 hours (NF24, NF48h, NF72h). The restoration of caspase expression to baseline levels is seen in the presence of 48 hours of combined DAG and farnesol (DAG-F60) treatment. Actin was used as the loading control.

Techniques Used: Western Blot, Expressing

31) Product Images from "p27 Protein Protects Metabolically Stressed Cardiomyocytes from Apoptosis by Promoting Autophagy *"

Article Title: p27 Protein Protects Metabolically Stressed Cardiomyocytes from Apoptosis by Promoting Autophagy *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M113.542795

Knockdown of p27 suppresses autophagy and increases apoptosis in glucose-deprived ( GD ) cardiomyocytes. A , Western blot shows lysates from cardiomyocytes treated with and without shRNA to knock down p27 versus control shRNA ( NT ) in the presence or absence of glucose. B , shRNAp27 reduced p27 and LC3-II levels and increased p62 levels. Apoptosis marker cleaved caspase 3 was increased in cardiomyocytes undergoing p27 knockdown. C , representative photomicrographs show cardiomyocytes fixed and stained for LC3 ( green ), α-actinin ( red ), and nuclei ( blue ). D , histogram shows the number of LC3 + dots/cell. E , TUNEL-positive nuclei are quantified. *, p
Figure Legend Snippet: Knockdown of p27 suppresses autophagy and increases apoptosis in glucose-deprived ( GD ) cardiomyocytes. A , Western blot shows lysates from cardiomyocytes treated with and without shRNA to knock down p27 versus control shRNA ( NT ) in the presence or absence of glucose. B , shRNAp27 reduced p27 and LC3-II levels and increased p62 levels. Apoptosis marker cleaved caspase 3 was increased in cardiomyocytes undergoing p27 knockdown. C , representative photomicrographs show cardiomyocytes fixed and stained for LC3 ( green ), α-actinin ( red ), and nuclei ( blue ). D , histogram shows the number of LC3 + dots/cell. E , TUNEL-positive nuclei are quantified. *, p

Techniques Used: Western Blot, shRNA, Marker, Staining, TUNEL Assay

TAT-p27-reduced cardiomyocyte apoptosis following glucose deprivation is autophagy-dependent. Glucose-deprived ( GD ) cardiomyocytes were treated with TAT-p27 in the presence or absence of autophagy flux inhibitor Baf-A1. A and B , TAT-p27-treated glucose-deprived cardiomyocytes showed increased LC3-II levels and reduced p62 and caspase 3 cleavage compared with TAT-β-Gal ( lanes 3 versus 1 ). However, addition of Baf-A1 increased apoptosis. *, p
Figure Legend Snippet: TAT-p27-reduced cardiomyocyte apoptosis following glucose deprivation is autophagy-dependent. Glucose-deprived ( GD ) cardiomyocytes were treated with TAT-p27 in the presence or absence of autophagy flux inhibitor Baf-A1. A and B , TAT-p27-treated glucose-deprived cardiomyocytes showed increased LC3-II levels and reduced p62 and caspase 3 cleavage compared with TAT-β-Gal ( lanes 3 versus 1 ). However, addition of Baf-A1 increased apoptosis. *, p

Techniques Used:

TAT-p27 reduced apoptosis post-MI by promoting autophagy. A and B , homogenates from mice undergoing sham and LAD ligation surgery, with the latter receiving treatments with TAT-p27 or TAT-β-Gal alone or in combination with the autophagy inhibitor CQ, were probed for LC3-II, p62, and cleaved caspase 3. β-Tubulin served as loading control. Compared with sham-operated controls, post-MI groups revealed higher levels of LC3-II and cleaved caspase 3 levels (*, p
Figure Legend Snippet: TAT-p27 reduced apoptosis post-MI by promoting autophagy. A and B , homogenates from mice undergoing sham and LAD ligation surgery, with the latter receiving treatments with TAT-p27 or TAT-β-Gal alone or in combination with the autophagy inhibitor CQ, were probed for LC3-II, p62, and cleaved caspase 3. β-Tubulin served as loading control. Compared with sham-operated controls, post-MI groups revealed higher levels of LC3-II and cleaved caspase 3 levels (*, p

Techniques Used: Mouse Assay, Ligation

TAT-p27 reduced glucose deprivation-induced apoptosis by promoting autophagy. TAT-p27-reduced apoptosis in glucose-deprived cardiomyocytes was abolished by 3-MA or CQ. A and B , 3-MA blocks the initial autophagy step, reducing LC3-II activation and increasing cleaved caspase 3 levels even in the presence of TAT-p27. *, p
Figure Legend Snippet: TAT-p27 reduced glucose deprivation-induced apoptosis by promoting autophagy. TAT-p27-reduced apoptosis in glucose-deprived cardiomyocytes was abolished by 3-MA or CQ. A and B , 3-MA blocks the initial autophagy step, reducing LC3-II activation and increasing cleaved caspase 3 levels even in the presence of TAT-p27. *, p

Techniques Used: Activation Assay

32) Product Images from "REV3L confers chemoresistance to cisplatin in human gliomas: The potential of its RNAi for synergistic therapy"

Article Title: REV3L confers chemoresistance to cisplatin in human gliomas: The potential of its RNAi for synergistic therapy

Journal: Neuro-Oncology

doi: 10.1215/15228517-2009-015

REV3L knockdown could efficiently synergize with cisplatin to induce glioma cell apoptosis via the mitochondria-mediated apop-totic pathway. (A) Time-dependent expression of cleaved caspase-3 after exposure to a single dose of cisplatin (1 μmol/l)
Figure Legend Snippet: REV3L knockdown could efficiently synergize with cisplatin to induce glioma cell apoptosis via the mitochondria-mediated apop-totic pathway. (A) Time-dependent expression of cleaved caspase-3 after exposure to a single dose of cisplatin (1 μmol/l)

Techniques Used: Expressing

REV3L overexpression conferred resistance to DNA cross-linking agents via inhibition of cisplatin-induced cell death of mitochondria-mediated apoptotic pathway. (A) The expression of cleaved caspase-3 and cytochrome c after exposure to two doses of cisplatin
Figure Legend Snippet: REV3L overexpression conferred resistance to DNA cross-linking agents via inhibition of cisplatin-induced cell death of mitochondria-mediated apoptotic pathway. (A) The expression of cleaved caspase-3 and cytochrome c after exposure to two doses of cisplatin

Techniques Used: Over Expression, Inhibition, Expressing

33) Product Images from "Proteomic Analysis of Shear Stress-Mediated Protection from TNF-alpha in Endothelial Cells"

Article Title: Proteomic Analysis of Shear Stress-Mediated Protection from TNF-alpha in Endothelial Cells

Journal: Microcirculation (New York, N.Y. : 1994)

doi: 10.1111/j.1549-8719.2010.00031.x

A, Quantitative densitometry comparing cleaved caspase 3 and poly-ADP-Ribose-Polymerase (PARP) protein levels in non-treated control cells, cells treated with 10ng/ml of TNF-alpha for 12hrs, cells exposed to shear for 18hrs (10 dynes/cm 2 ), and cells first
Figure Legend Snippet: A, Quantitative densitometry comparing cleaved caspase 3 and poly-ADP-Ribose-Polymerase (PARP) protein levels in non-treated control cells, cells treated with 10ng/ml of TNF-alpha for 12hrs, cells exposed to shear for 18hrs (10 dynes/cm 2 ), and cells first

Techniques Used:

34) Product Images from "MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats"

Article Title: MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats

Journal: Experimental neurology

doi: 10.1016/j.expneurol.2015.04.016

The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

35) Product Images from "MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats"

Article Title: MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats

Journal: Experimental neurology

doi: 10.1016/j.expneurol.2015.04.016

The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

36) Product Images from "MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats"

Article Title: MFGE8/Integrin β3 Pathway Alleviates Apoptosis and Inflammation in Early Brain Injury after Subarachnoid Hemorrhage in Rats

Journal: Experimental neurology

doi: 10.1016/j.expneurol.2015.04.016

The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The adverse effects of silencing endogenous MFGE8 by siRNA at 24 hours after SAH Administration of MFGE8 siRNA decreased the modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E), IL-1β (F) showed that MFGE8 siRNA decreased the protein level of endogenous MFGE8 and upregulated Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: Integrinβ3 siRNA reversed the protective effects of rhMFGE8 treatment at 24 hours after SAH Integrinβ3 siRNA decreased Modified Garcia Score (A) and Beam Balance Testing Score (B) in the presence of rhMFGE8 treatment. Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that integrinβ3 siRNA decreased the protein level of MFGE8 and attenuated Capase3 and IL-1β expressions in the presence of rhMFGE8 treatment. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P
Figure Legend Snippet: The protective effects of rhMFGE8 treatment at 24 hours after SAH Administration of rhMFGE8 increased the Modified Garcia Score (A) and Beam Balance Testing Score (B). Representative Western blots bands (C) and quantitative analysis of MFGE8 (D), CC3 (E) and IL-1β (F) showed that rhMFGE8 treatment increased the protein level of MFGE8 and decreased Capase3 and IL-1β expressions. MFGE8: Milk fat globule-epidermal growth factor-factor 8; rhMFGE8: Recombinant human milk fat globule-epidermal growth factor-factor 8; CC3: Cleaved caspase 3; n=6 for each group; * P

Techniques Used: Modification, Western Blot, Recombinant

37) Product Images from "Tropism of and Innate Immune Responses to the Novel Human Betacoronavirus Lineage C Virus in Human Ex Vivo Respiratory Organ Cultures"

Article Title: Tropism of and Innate Immune Responses to the Novel Human Betacoronavirus Lineage C Virus in Human Ex Vivo Respiratory Organ Cultures

Journal: Journal of Virology

doi: 10.1128/JVI.00009-13

Apoptotic cells identified in human lung tissue ex vivo culture upon HCoV-EMC and SARS-CoV infection. (A to C) Ex vivo culture of lung tissue mock infected (A) or infected with HCoV-EMC (B) or SARS-CoV (C) at 48 hpi. The reddish brown stain identifies the presence of cleaved caspase 3. (D and E) Costaining of HCoV-EMC (D) and SARS-CoV (E) antigen (pink stain) with cleaved caspase 3 (reddish brown stain).
Figure Legend Snippet: Apoptotic cells identified in human lung tissue ex vivo culture upon HCoV-EMC and SARS-CoV infection. (A to C) Ex vivo culture of lung tissue mock infected (A) or infected with HCoV-EMC (B) or SARS-CoV (C) at 48 hpi. The reddish brown stain identifies the presence of cleaved caspase 3. (D and E) Costaining of HCoV-EMC (D) and SARS-CoV (E) antigen (pink stain) with cleaved caspase 3 (reddish brown stain).

Techniques Used: Ex Vivo, Infection, Staining

38) Product Images from "Sensitivity of staurosporine-induced differentiated RGC-5 cells to homocysteine"

Article Title: Sensitivity of staurosporine-induced differentiated RGC-5 cells to homocysteine

Journal: Current eye research

doi: 10.3109/02713680903421194

TUNEL analysis and immunodetection of cleaved caspase-3 in undifferentiated RGC-5 cells treated with various doses of homocysteine for 18 h (A) RGC-5 cells were seeded onto coverslips and processed for TUNEL analysis with either no treatment, or treatment with 1 mM homocysteine, 5 mM homocysteine, and 10 mM homocysteine. TUNEL positive cells (arrows) are indicated by bright white (green when in color) fluorescence; Hoescht 33258 staining (light gray (blue when in color) fluorescence) allows visualization of all cell nuclei. Scale bar denotes 25 µm. (B) DIC images of cells treated as described in panel A. (C) Protein isolated from cells were subjected to SDS-PAGE, transferred to a nitrocellulose membrane, and probed with an antibody against cleaved caspase-3. β-actin served as the loading control. Lanes: (1) RGC-5 cells, no treatment; (2) RGC-5 cells + 1 mM homocysteine; (3) RGC-5 cells + 5 mM homocysteine; (4) RGC-5 cells + 10 mM homocysteine. β-actin: M r ~ 45 kDa, pro-form caspase-3: M r ~ 32 kDa, cleaved form caspase-3: M r ~ 17 kDa.
Figure Legend Snippet: TUNEL analysis and immunodetection of cleaved caspase-3 in undifferentiated RGC-5 cells treated with various doses of homocysteine for 18 h (A) RGC-5 cells were seeded onto coverslips and processed for TUNEL analysis with either no treatment, or treatment with 1 mM homocysteine, 5 mM homocysteine, and 10 mM homocysteine. TUNEL positive cells (arrows) are indicated by bright white (green when in color) fluorescence; Hoescht 33258 staining (light gray (blue when in color) fluorescence) allows visualization of all cell nuclei. Scale bar denotes 25 µm. (B) DIC images of cells treated as described in panel A. (C) Protein isolated from cells were subjected to SDS-PAGE, transferred to a nitrocellulose membrane, and probed with an antibody against cleaved caspase-3. β-actin served as the loading control. Lanes: (1) RGC-5 cells, no treatment; (2) RGC-5 cells + 1 mM homocysteine; (3) RGC-5 cells + 5 mM homocysteine; (4) RGC-5 cells + 10 mM homocysteine. β-actin: M r ~ 45 kDa, pro-form caspase-3: M r ~ 32 kDa, cleaved form caspase-3: M r ~ 17 kDa.

Techniques Used: TUNEL Assay, Immunodetection, Fluorescence, Staining, Isolation, SDS Page

Immunodetection of cleaved caspase-3 in undifferentiated and staurosporine-differentiated RGC-5 cells treated with homocysteine, glutamate or X:XO for 18 h Protein was isolated from cells and subjected to SDS-PAGE, transferred to a nitrocellulose membrane, and probed with an antibody against cleaved caspase-3. β-actin served as the loading control. β-actin: M r ~ 45 kDa, pro-form caspase-3: M r ~ 32 kDa, cleaved form caspase-3: M r ~ 17 kDa. Lanes: (1) RGC-5 cells, no staurosporine differentiation; (2) RGC-5 cells + staurosporine, (3) RGC-5 cells + 1 mM homocysteine, (4) RGC-5 cells + 1 mM glutamate, (5) RGC-5 cells + 25 µM xanthine:10 mU/ml xanthine oxidase (6) RGC-5 cells + staurosporine + 1 mM homocysteine, (7) RGC-5 cells + staurosporine + 1 mM glutamate, (8) RGC-5 cells + staurosporine + 25 µM xanthine:10 mU/ml xanthine oxidase.
Figure Legend Snippet: Immunodetection of cleaved caspase-3 in undifferentiated and staurosporine-differentiated RGC-5 cells treated with homocysteine, glutamate or X:XO for 18 h Protein was isolated from cells and subjected to SDS-PAGE, transferred to a nitrocellulose membrane, and probed with an antibody against cleaved caspase-3. β-actin served as the loading control. β-actin: M r ~ 45 kDa, pro-form caspase-3: M r ~ 32 kDa, cleaved form caspase-3: M r ~ 17 kDa. Lanes: (1) RGC-5 cells, no staurosporine differentiation; (2) RGC-5 cells + staurosporine, (3) RGC-5 cells + 1 mM homocysteine, (4) RGC-5 cells + 1 mM glutamate, (5) RGC-5 cells + 25 µM xanthine:10 mU/ml xanthine oxidase (6) RGC-5 cells + staurosporine + 1 mM homocysteine, (7) RGC-5 cells + staurosporine + 1 mM glutamate, (8) RGC-5 cells + staurosporine + 25 µM xanthine:10 mU/ml xanthine oxidase.

Techniques Used: Immunodetection, Isolation, SDS Page

39) Product Images from "Reovirus Activates Transforming Growth Factor ? and Bone Morphogenetic Protein Signaling Pathways in the Central Nervous System That Contribute to Neuronal Survival following Infection ▿"

Article Title: Reovirus Activates Transforming Growth Factor ? and Bone Morphogenetic Protein Signaling Pathways in the Central Nervous System That Contribute to Neuronal Survival following Infection ▿

Journal: Journal of Virology

doi: 10.1128/JVI.02433-08

Inhibition of reovirus-induced TGF-β signal activation increases apoptosis in vivo. (A) Whole-brain lysates from mock-infected and reovirus-infected (1,000 PFU, i.c. injection) Swiss Webster pups sacrificed at day 8 postinfection following daily intraperitoneal treatment with TGF-βRI inhibitor (TGF-βRI Inh III) or vehicle control were resolved on 10% polyacrylamide gels, transferred to nitrocellulose membranes, and probed with anti-pSMAD3, anti-cleaved caspase 3 (cl-Casp3), anti-cleaved PARP (cl-PARP), and anti-β-actin. Immunoblots are representative of three individual replicates per treatment group from multiple litters. (B to D) Immunohistochemistry staining of the same treatment groups revealed no differences in viral antigen (green) or in the distribution of cleaved caspase 3 (red) in animals treated with vehicle control or TGF-βRI inhibitor. Images shown are of the cingulate cortex. Original magnification, ×400.
Figure Legend Snippet: Inhibition of reovirus-induced TGF-β signal activation increases apoptosis in vivo. (A) Whole-brain lysates from mock-infected and reovirus-infected (1,000 PFU, i.c. injection) Swiss Webster pups sacrificed at day 8 postinfection following daily intraperitoneal treatment with TGF-βRI inhibitor (TGF-βRI Inh III) or vehicle control were resolved on 10% polyacrylamide gels, transferred to nitrocellulose membranes, and probed with anti-pSMAD3, anti-cleaved caspase 3 (cl-Casp3), anti-cleaved PARP (cl-PARP), and anti-β-actin. Immunoblots are representative of three individual replicates per treatment group from multiple litters. (B to D) Immunohistochemistry staining of the same treatment groups revealed no differences in viral antigen (green) or in the distribution of cleaved caspase 3 (red) in animals treated with vehicle control or TGF-βRI inhibitor. Images shown are of the cingulate cortex. Original magnification, ×400.

Techniques Used: Inhibition, Activation Assay, In Vivo, Infection, Injection, Western Blot, Immunohistochemistry, Staining

BMP6 ligand treatment of reovirus-infected MCCs prevents apoptosis. (A to C) Mock-infected (A) or reovirus-infected (B and C) (MOI, 100) primary MCCs were treated daily with vehicle control (B) or BMP6 ligand (C) and analyzed at day 4 postinfection using immunocytochemistry with antibodies to cleaved caspase 3 (green) and microtubule-associated protein 2 (red). Nuclear staining (Hoechst) is shown in blue. (D) Blinded cell counts of anti-cleaved caspase 3 (anti-cl-caspase 3)-positive cells in three replicate experiments show a threefold ( P
Figure Legend Snippet: BMP6 ligand treatment of reovirus-infected MCCs prevents apoptosis. (A to C) Mock-infected (A) or reovirus-infected (B and C) (MOI, 100) primary MCCs were treated daily with vehicle control (B) or BMP6 ligand (C) and analyzed at day 4 postinfection using immunocytochemistry with antibodies to cleaved caspase 3 (green) and microtubule-associated protein 2 (red). Nuclear staining (Hoechst) is shown in blue. (D) Blinded cell counts of anti-cleaved caspase 3 (anti-cl-caspase 3)-positive cells in three replicate experiments show a threefold ( P

Techniques Used: Infection, Immunocytochemistry, Staining

40) Product Images from "Role of ?-Adrenergic Receptor Regulation of TNF-? and Insulin Signaling in Retinal M?ller Cells"

Article Title: Role of ?-Adrenergic Receptor Regulation of TNF-? and Insulin Signaling in Retinal M?ller Cells

Journal: Investigative Ophthalmology & Visual Science

doi: 10.1167/iovs.11-8631

( A ) ELISA showed a significant increase in caspase-3 in samples treated with ICI 118551 (a β-1-adrenergic receptor antagonist) compared with untreated samples. Caspase-3 levels were significantly decreased in samples treated with CGP 20712A (a
Figure Legend Snippet: ( A ) ELISA showed a significant increase in caspase-3 in samples treated with ICI 118551 (a β-1-adrenergic receptor antagonist) compared with untreated samples. Caspase-3 levels were significantly decreased in samples treated with CGP 20712A (a

Techniques Used: Enzyme-linked Immunosorbent Assay

ELISA of cleaved caspase-3 in rat Müller cells showed significantly increased apoptosis in high glucose compared with low glucose. Treatment with isoproterenol significantly decreased caspase-3 activity after 24 hours of treatment of Müller
Figure Legend Snippet: ELISA of cleaved caspase-3 in rat Müller cells showed significantly increased apoptosis in high glucose compared with low glucose. Treatment with isoproterenol significantly decreased caspase-3 activity after 24 hours of treatment of Müller

Techniques Used: Enzyme-linked Immunosorbent Assay, Activity Assay

Related Articles

Incubation:

Article Title: Cytoplasmic liver kinase B1 promotes the growth of human lung adenocarcinoma by enhancing autophagy, et al. Cytoplasmic liver kinase B1 promotes the growth of human lung adenocarcinoma by enhancing autophagy
Article Snippet: .. After being blocked with 5% fat‐free milk powder in TBST, the membranes were incubated with primary antibodies against Beclin1 (ePR1733Y, 1:800 dilution; Abcam, Cambridge, UK), LC3 (L7543, 1:1000 dilution; Sigma), Atg4 (D62C10, 1:1000 dilution; Cell Signaling Technology), Atg7 (D12B11, 1:1000 dilution; Cell Signaling Technology), Atg12 (D88H11, 1:1000 dilution; Cell Signaling Technology), cleaved caspase‐3 (5A1E, 1:1000 dilution; Cell Signaling Technology), caspase‐3 (8G10, 1:1000 dilution; Cell Signaling Technology), cleaved poly(ADP‐ribose) polymerase (PARP) (19F4, 1:1000 dilution; Cell Signaling Technology), PARP (46D11, 1:1000 dilution; Cell Signaling Technology), H3 (1B1B2, 1:1000 dilution; Cell Signaling Technology), and β‐actin (sc‐130301, 1:1000 dilution; Santa Cruz Biotechnology, Santa Cruz, CA, USA) at 4°C overnight. .. After being washed, the bound antibodies were detected with HRP‐conjugated secondary antibodies and visualized using the enhanced chemiluminescent reagents (Millipore).

other:

Article Title: Dexmedetomidine Ameliorates Acute Stress-Induced Kidney Injury by Attenuating Oxidative Stress and Apoptosis through Inhibition of the ROS/JNK Signaling Pathway
Article Snippet: Therefore, the site of kidney injury was determined by measuring localization of cleaved caspase 3 in kidney tissue.

Article Title: Dexmedetomidine Ameliorates Acute Stress-Induced Kidney Injury by Attenuating Oxidative Stress and Apoptosis through Inhibition of the ROS/JNK Signaling Pathway
Article Snippet: As shown in Figures and , cleaved caspase 3 was primarily expressed in renal tubules.

Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 88
    Cell Signaling Technology Inc whole mount cleaved caspase 3 staining
    p53 25,26,53,54/+  Embryos Exhibit Additional Features of CHARGE Syndrome and p53-Dependent Cellular Responses (a)  Double outlet right ventricle (DORV) in E13.5 p53 25,26,53,54/+  heart (50%, n=6). Top: Main pulmonary artery (MPA) connects via pulmonary valve (PV) to right ventricle (RV) in both control and p53 25,26,53,54/+  embryo. Bottom: Aorta (Ao) in control embryo connects to left ventricle (LV) via aortic valve (AV) Φ . Aorta in p53 25,26,53,54/+  embryo connects to RV via AV*. (b)  Abnormal atrioventricular cushions in E13.5 p53 25,26,53,54/+  heart (75%, n=4) fail to elongateinto mature mitral (mv, arrowhead) and tricuspid (tv, arrow) valves. RA: right atrium; LA: left atrium.  (c)  E13.5  p53 25,26,53,54/+  kidneys are smaller (79%), with fewer average glomeruli (13 vs. 3; n=5; arrows), than controls. (d) p53 25,26,53,54/+  embryonic phenotypes observed in CHARGE (+present, −absent).  (e)  Left: Cleaved-caspase 3 (CC3; Top) and p53 (Bottom) immunohistochemistry in E15.5 retinas. Arrows: CC3-positive cells. Right: CC3-positive cells per retinal area. ***p-value=0.007; one-tailed Welsh’s t-test (n=5).  (f)  BrdU immunofluorescence in E9.5 Pax3 +  NCCs (delineated by green-dotted line;  Extended-Data Fig. 6c ). Right: Percentage BrdU-positive cells per total Pax3 +  NCCs ***p-value=0.004 one-tailed Student’s t-test (n=4).
    Whole Mount Cleaved Caspase 3 Staining, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 88/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/whole mount cleaved caspase 3 staining/product/Cell Signaling Technology Inc
    Average 88 stars, based on 9 article reviews
    Price from $9.99 to $1999.99
    whole mount cleaved caspase 3 staining - by Bioz Stars, 2020-10
    88/100 stars
      Buy from Supplier

    92
    Cell Signaling Technology Inc cleaved caspase 3
    Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved <t>Caspase</t> 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 92/100, based on 357 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cleaved caspase 3/product/Cell Signaling Technology Inc
    Average 92 stars, based on 357 article reviews
    Price from $9.99 to $1999.99
    cleaved caspase 3 - by Bioz Stars, 2020-10
    92/100 stars
      Buy from Supplier

    85
    Cell Signaling Technology Inc 17 kda cleaved caspase 3 forms
    AAV2 induction of apoptosis in MDA-MB-468 cells results in PARP cleavage following activation of caspases of the intrinsic pathway . MDA-MB-468 monolayer cell cultures were synchronized in G1, followed by infection with AAV2. Cell pellets were collected each day over a 7 day period. Cells were passaged 1:2 on day 2 and day 5. Detection of caspases and their cleavage/activation was performed by Western blotting. Total protein extracts were prepared as described previously [ 48 ]. Sixty micrograms of total protein extracts from AAV2 infected and control MDA-MB-468 cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect the 35 <t>kDa</t> pro-caspase form of <t>caspase-3,</t> proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology). To detect the 35 kDa pro-caspase form of caspase-6, proteins were resolved in a 10% SDS-PAGE gel and to detect the 15 kDa cleaved form of caspase-6, proteins were resolved in a 15% SDS-PAGE gel and detected with a rabbit polyclonal antibody (Cell Signaling Technology). To detect both the pro- and cleaved forms of caspase-7, caspase-8 and caspase-9, proteins were resolved in a 10% SDS-PAGE gel. The 35 kDa pro-caspase form and the 30 kDa and 20 kDa cleaved forms of caspase-7 was detected with a mouse monoclonal antibody (Cell Signaling). Caspase-8 was detected with a mouse monoclonal antibody (Alexis Biochemicals). The 47 kDa pro-caspase and 35 kDa cleaved of caspase-9 was detected with a rabbit polyclonal antibody (Cell Signaling). To detect the pro- (116 kDa) and cleaved- (89 kDa) forms of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).
    17 Kda Cleaved Caspase 3 Forms, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/17 kda cleaved caspase 3 forms/product/Cell Signaling Technology Inc
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    17 kda cleaved caspase 3 forms - by Bioz Stars, 2020-10
    85/100 stars
      Buy from Supplier

    Image Search Results


    p53 25,26,53,54/+  Embryos Exhibit Additional Features of CHARGE Syndrome and p53-Dependent Cellular Responses (a)  Double outlet right ventricle (DORV) in E13.5 p53 25,26,53,54/+  heart (50%, n=6). Top: Main pulmonary artery (MPA) connects via pulmonary valve (PV) to right ventricle (RV) in both control and p53 25,26,53,54/+  embryo. Bottom: Aorta (Ao) in control embryo connects to left ventricle (LV) via aortic valve (AV) Φ . Aorta in p53 25,26,53,54/+  embryo connects to RV via AV*. (b)  Abnormal atrioventricular cushions in E13.5 p53 25,26,53,54/+  heart (75%, n=4) fail to elongateinto mature mitral (mv, arrowhead) and tricuspid (tv, arrow) valves. RA: right atrium; LA: left atrium.  (c)  E13.5  p53 25,26,53,54/+  kidneys are smaller (79%), with fewer average glomeruli (13 vs. 3; n=5; arrows), than controls. (d) p53 25,26,53,54/+  embryonic phenotypes observed in CHARGE (+present, −absent).  (e)  Left: Cleaved-caspase 3 (CC3; Top) and p53 (Bottom) immunohistochemistry in E15.5 retinas. Arrows: CC3-positive cells. Right: CC3-positive cells per retinal area. ***p-value=0.007; one-tailed Welsh’s t-test (n=5).  (f)  BrdU immunofluorescence in E9.5 Pax3 +  NCCs (delineated by green-dotted line;  Extended-Data Fig. 6c ). Right: Percentage BrdU-positive cells per total Pax3 +  NCCs ***p-value=0.004 one-tailed Student’s t-test (n=4).

    Journal: Nature

    Article Title: Inappropriate p53 Activation During Development Induces Features of CHARGE Syndrome

    doi: 10.1038/nature13585

    Figure Lengend Snippet: p53 25,26,53,54/+ Embryos Exhibit Additional Features of CHARGE Syndrome and p53-Dependent Cellular Responses (a) Double outlet right ventricle (DORV) in E13.5 p53 25,26,53,54/+ heart (50%, n=6). Top: Main pulmonary artery (MPA) connects via pulmonary valve (PV) to right ventricle (RV) in both control and p53 25,26,53,54/+ embryo. Bottom: Aorta (Ao) in control embryo connects to left ventricle (LV) via aortic valve (AV) Φ . Aorta in p53 25,26,53,54/+ embryo connects to RV via AV*. (b) Abnormal atrioventricular cushions in E13.5 p53 25,26,53,54/+ heart (75%, n=4) fail to elongateinto mature mitral (mv, arrowhead) and tricuspid (tv, arrow) valves. RA: right atrium; LA: left atrium. (c) E13.5 p53 25,26,53,54/+ kidneys are smaller (79%), with fewer average glomeruli (13 vs. 3; n=5; arrows), than controls. (d) p53 25,26,53,54/+ embryonic phenotypes observed in CHARGE (+present, −absent). (e) Left: Cleaved-caspase 3 (CC3; Top) and p53 (Bottom) immunohistochemistry in E15.5 retinas. Arrows: CC3-positive cells. Right: CC3-positive cells per retinal area. ***p-value=0.007; one-tailed Welsh’s t-test (n=5). (f) BrdU immunofluorescence in E9.5 Pax3 + NCCs (delineated by green-dotted line; Extended-Data Fig. 6c ). Right: Percentage BrdU-positive cells per total Pax3 + NCCs ***p-value=0.004 one-tailed Student’s t-test (n=4).

    Article Snippet: Whole-mount cleaved-caspase 3 staining was performed as described with anti-cleaved-caspase 3 antibody (Cell Signaling #9664) and developed with DAB (Vector Labs).

    Techniques: Immunohistochemistry, One-tailed Test, Immunofluorescence

    p53 25,26,53,54/+  Embryo Tissues Display Increased Apoptosis and Decreased Proliferation (a)  Left: Immunofluorescence for Phospho-Histone H3 (red) in the retina of E13.5 control and  p53 25,26,53,54/+ embryos. Right: Quantification of Phospho-Histone H3 positive cells per retina area relative to littermate controls. **p-value=0.006 by one-tailed Welsh’s t-test (n=4). ( b ) Left: Immunohistochemistry for cleaved-caspase 3 (CC3) in thymi of control (left) and  p53 25,26,53,54/+  (right) embryos. Inset: close-up image of cleaved-caspase 3 positive region. Right: Quantification of CC3-positive cells per thymic area. *p-value=0.02 by one-tailed Student’s t-test (n=4).  (c)  Immunofluorescence for Pax3 (green) in neural crest cells of E9.5 control and p53 25,26,53,54/+  embryos was used to identify neural crest cells in   Figure 2f .  (d) Left: Immunofluorescence for cleaved-caspase 3 (CC3, red) and Pax3 (green) in neural crest cells of E9.5 control and  p53 25,26,53,54/+ embryos.  p53 25,26,53,54/+  embryos have more apoptotic (red) neural crest cells, as determined by Pax3-positive staining (green), compared to control littermates. Right: Quantification of CC3 positive cells per total neural crest cell number. p-value=0.14 by one-tailed Student’s t-test (n=4). (e)  Left: Immunofluorescence for cleaved-caspase 3 (CC3, red) in otic vesicle of E9.5 control and  p53 25,26,53,54/+  embryos. Right: Quantification of CC3 positive cells per total cell number. *p-value=0.03 by one-tailed Student’s t-test (n=3). ( f ) Whole-mount cleaved-caspase 3 staining in E8.5 control and p53 25,26,53,54/+  embryos reveals enhanced apoptosis in the neuroepithelium of  p53 25,26,53,54/+  embryos (right) but not in controls (left). Close-up shows magnification of the caudal neuroepithelium (bottom). Arrows indicate cleaved-caspase 3 positive regions.

    Journal: Nature

    Article Title: Inappropriate p53 Activation During Development Induces Features of CHARGE Syndrome

    doi: 10.1038/nature13585

    Figure Lengend Snippet: p53 25,26,53,54/+ Embryo Tissues Display Increased Apoptosis and Decreased Proliferation (a) Left: Immunofluorescence for Phospho-Histone H3 (red) in the retina of E13.5 control and p53 25,26,53,54/+ embryos. Right: Quantification of Phospho-Histone H3 positive cells per retina area relative to littermate controls. **p-value=0.006 by one-tailed Welsh’s t-test (n=4). ( b ) Left: Immunohistochemistry for cleaved-caspase 3 (CC3) in thymi of control (left) and p53 25,26,53,54/+ (right) embryos. Inset: close-up image of cleaved-caspase 3 positive region. Right: Quantification of CC3-positive cells per thymic area. *p-value=0.02 by one-tailed Student’s t-test (n=4). (c) Immunofluorescence for Pax3 (green) in neural crest cells of E9.5 control and p53 25,26,53,54/+ embryos was used to identify neural crest cells in Figure 2f . (d) Left: Immunofluorescence for cleaved-caspase 3 (CC3, red) and Pax3 (green) in neural crest cells of E9.5 control and p53 25,26,53,54/+ embryos. p53 25,26,53,54/+ embryos have more apoptotic (red) neural crest cells, as determined by Pax3-positive staining (green), compared to control littermates. Right: Quantification of CC3 positive cells per total neural crest cell number. p-value=0.14 by one-tailed Student’s t-test (n=4). (e) Left: Immunofluorescence for cleaved-caspase 3 (CC3, red) in otic vesicle of E9.5 control and p53 25,26,53,54/+ embryos. Right: Quantification of CC3 positive cells per total cell number. *p-value=0.03 by one-tailed Student’s t-test (n=3). ( f ) Whole-mount cleaved-caspase 3 staining in E8.5 control and p53 25,26,53,54/+ embryos reveals enhanced apoptosis in the neuroepithelium of p53 25,26,53,54/+ embryos (right) but not in controls (left). Close-up shows magnification of the caudal neuroepithelium (bottom). Arrows indicate cleaved-caspase 3 positive regions.

    Article Snippet: Whole-mount cleaved-caspase 3 staining was performed as described with anti-cleaved-caspase 3 antibody (Cell Signaling #9664) and developed with DAB (Vector Labs).

    Techniques: Immunofluorescence, One-tailed Test, Immunohistochemistry, Staining

    Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved Caspase 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P

    Journal: bioRxiv

    Article Title: SHP2 Inhibition Abrogates Adaptive Resistance to KRASG12C-Inhibition and Remodels the Tumor Microenvironment of KRAS-Mutant Tumors

    doi: 10.1101/2020.05.30.125138

    Figure Lengend Snippet: Combined ARS1620/SHP2 inhibition is highly efficacious in PDAC models in vivo . A , Pancreas tumors were established in syngeneic mice by orthotopic injections of KCP cells, and 14 days later, mice were treated with vehicle, SHP099, ARS1620 or both drugs (Combo), as depicted. Tumor weight was quantified in a cohort at Day 0 (baseline) and in treated mice at Day 10. B , Immunoblots of KCP-derived tumor lysates showing effects of the indicated treatments on KRAS G12C -GTP, pERK, and DUSP6 levels. C , ERK-dependent gene expression, assessed by RNAseq, in KCP tumors treated for 3 days, as indicated in A (colors indicate log2FC). D-E , Time-dependent increase in RTK (D) and RTK ligands (E) gene expression in KCP-derived orthotopic tumors after vehicle, SHP099, ARS1620 and Combo treatment at Day 3, determined by RNAseq (colors represent log2FC). F , H E, Masson Trichome, CD31, pERK, Ki67 and cleaved Caspase 3 staining and quantification in KCP tumor sections from mice after 10 days of treatment, as indicated. G , KCP tumors were established in syngeneic mice and allowed to grow to much larger size before treatments were initiated, as depicted in the scheme. Tumor weight was quantified in one cohort before treatment, in another cohort after 12 days of treatment, and after drug withdrawal, at Day 27, as indicated. H , Kaplan-Meier curve of KCP tumor-bearing mice after withdrawal of the indicated drugs (top). Tumor growth curve after withdrawal of indicated treatment at day 12 (bottom). H , Response of sub-cutaneous NY53 patient-derived xenograft to treatment with vehicle, SHP099, ARS1620 or both drugs. For all experiments, drug doses were: SHP099 (75 mg/kg body weight, daily), ARS1620 (200 mg/kg body weight, daily) or both drugs (daily). Data represent mean ± SD; *P

    Article Snippet: IHC for pERK (Cell Signaling, 4370), CD31 (Cell Signaling, D8V9E), Cleaved Caspase 3 (Cell Signaling, D3E9), Ki67 (Spring Biosciences, SP6), αSMA (Abcam, ab5694) was performed on sections from paraformaldehyde-fixed tumors.

    Techniques: Inhibition, In Vivo, Mouse Assay, Western Blot, Derivative Assay, Expressing, Staining

    Continuous infusion of mithramycin shows activity in an intramuscular rhabdoid tumor xenograft model. A:  Top, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 1 mg/kg/dose mithramycin intraperitoneal M, W, F for two weeks (green) relative to vehicle control (gray). Bottom, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 2.4 mg/kg of mithramycin (purple) or vehicle control (gray) administered continuously intraperitoneal over 72h. Most mice experienced a suppression or regression of tumor volume that persisted for more than 2 weeks following treatment. The shaded box indicates the duration of treatment. B:  Immunohistochemistry analysis recapitulates the biochemistry described in vitro. G401 tumor sections at 10X magnification stained for cleaved caspase 3 (CC3; apoptosis) or H3K27me3. A marked increase in CC3 correlates with H3K27me3 staining is seen only in mice treated with the continuous infusion schedule but not bolus dosing. C,D:  Prolonged durable response and cure of mice bearing G401 xenografts treated with  (C)  30mg/kg EC8042 administered continuously over 72-hours or  (D)  50 mg/kg EC8042 administered continuously over 144-hours. Treatment duration indicated by gray shaded box. Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text). E:  Bioluminescence imaging of G401 rhabdoid tumor xenografts correlates with caliper measurements in 7A and 7B. Two mice per treatment group were imaged (left) and quantified in the bar graph (right) . Error bars represent mean with SD. F:  Kaplan-Meier survival curves indicating extended survival for mice bearing established G401 xenografts treated with the 3-day or 7-day continuous infusions of EC8042 in (C) and (D). Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text).

    Journal: bioRxiv

    Article Title: Direct therapeutic targeting of SWI/SNF induces epigenetic reprogramming and durable tumor regression in rhabdoid tumor

    doi: 10.1101/861484

    Figure Lengend Snippet: Continuous infusion of mithramycin shows activity in an intramuscular rhabdoid tumor xenograft model. A: Top, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 1 mg/kg/dose mithramycin intraperitoneal M, W, F for two weeks (green) relative to vehicle control (gray). Bottom, Spaghetti plot showing tumor volumes of individual tumors in mice bearing G401 xenografts treated with 2.4 mg/kg of mithramycin (purple) or vehicle control (gray) administered continuously intraperitoneal over 72h. Most mice experienced a suppression or regression of tumor volume that persisted for more than 2 weeks following treatment. The shaded box indicates the duration of treatment. B: Immunohistochemistry analysis recapitulates the biochemistry described in vitro. G401 tumor sections at 10X magnification stained for cleaved caspase 3 (CC3; apoptosis) or H3K27me3. A marked increase in CC3 correlates with H3K27me3 staining is seen only in mice treated with the continuous infusion schedule but not bolus dosing. C,D: Prolonged durable response and cure of mice bearing G401 xenografts treated with (C) 30mg/kg EC8042 administered continuously over 72-hours or (D) 50 mg/kg EC8042 administered continuously over 144-hours. Treatment duration indicated by gray shaded box. Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text). E: Bioluminescence imaging of G401 rhabdoid tumor xenografts correlates with caliper measurements in 7A and 7B. Two mice per treatment group were imaged (left) and quantified in the bar graph (right) . Error bars represent mean with SD. F: Kaplan-Meier survival curves indicating extended survival for mice bearing established G401 xenografts treated with the 3-day or 7-day continuous infusions of EC8042 in (C) and (D). Asterisks indicate an animal sacrificed due to unknown causes not related to tumor progression or drug toxicity (see text).

    Article Snippet: Tissue was incubated with H3K27me3 (Abcam, 1:250), Cleaved Caspase-3 (Cell Signaling, 1:250), H3K27ac (Abcam, 1:1250), or human mitochondria (Abcam, 1:800) washed and then secondary antibody (Envision+System HRP labelled polymer Anti-Rabbit, Dako 1:100).

    Techniques: Activity Assay, Mouse Assay, Immunohistochemistry, In Vitro, Staining, Imaging

    Mithramycin cellular sensitivity favors SWI/SNF mutant cancers. A:  Graph of IC50 as a function of cell line generated from a published screen of 445 agents in 63 sarcoma cell lines (  39 ). Cell lines with mutated or dysregulated SWI/SNF (green) cluster towards the right on the graph indicating these cell lines are more sensitive to mithramycin. B:  Table highlighting the SWI/SNF dysregulation status in the top 25 sarcoma cell lines from   Fig. 1A . Mutation status was confirmed in COSMIC (  49 ) or the DepMap ( https://depmap.org/portal/ ). C:  Dose response curves of rhabdoid tumor and Ewing sarcoma cell lines. RT cell lines (black) are sensitive to mithramycin treatment with a similar IC50 value as TC32 ES cells (grey). RT cell lines are not sensitive to three broadly-active chemotherapeutic agents: etoposide, doxorubicin or SN38. E:  Western blot showing concentration-dependent increase in H3K27me3 following exposure to 100nM, 50nM, 25nM mithramycin for 18h in BT12 cells relative to loading control (H3). E:  Mithramycin induces a concentration-dependent suppression of proliferation in BT12 cells. Cells were exposed to 25nM, 50nM, or 100nM MMA for 18h, replaced with drug- free media and monitored using live cell imaging. F:  Mithramycin induces morphological changes and apoptosis relative to control cells (solvent). BT12 cells treated with 25nM (left) or 100nM (right) mithramycin for 18h (top) or 48-hours (bottom) in the presence of cleaved caspase 3/7 reagent that fluoresces with caspase activation. G:  Mithramycin leads to H3K27me3 amplification in a time-dependent manner that precedes the induction of apoptosis as measured by the cleavage of PARP. Western blot lysates collected at 1h, 2h, 4h, 8h, 12h 16h, 18h of continuous 100nM mithramycin treatment. H:  Chromatin immunoprecipitation qPCR (ChIP-qPCR) of H3K27me3 at  MYT1  and  CCND1 . H3K27me3 occupancy is increased in a time-dependent manner.

    Journal: bioRxiv

    Article Title: Direct therapeutic targeting of SWI/SNF induces epigenetic reprogramming and durable tumor regression in rhabdoid tumor

    doi: 10.1101/861484

    Figure Lengend Snippet: Mithramycin cellular sensitivity favors SWI/SNF mutant cancers. A: Graph of IC50 as a function of cell line generated from a published screen of 445 agents in 63 sarcoma cell lines ( 39 ). Cell lines with mutated or dysregulated SWI/SNF (green) cluster towards the right on the graph indicating these cell lines are more sensitive to mithramycin. B: Table highlighting the SWI/SNF dysregulation status in the top 25 sarcoma cell lines from Fig. 1A . Mutation status was confirmed in COSMIC ( 49 ) or the DepMap ( https://depmap.org/portal/ ). C: Dose response curves of rhabdoid tumor and Ewing sarcoma cell lines. RT cell lines (black) are sensitive to mithramycin treatment with a similar IC50 value as TC32 ES cells (grey). RT cell lines are not sensitive to three broadly-active chemotherapeutic agents: etoposide, doxorubicin or SN38. E: Western blot showing concentration-dependent increase in H3K27me3 following exposure to 100nM, 50nM, 25nM mithramycin for 18h in BT12 cells relative to loading control (H3). E: Mithramycin induces a concentration-dependent suppression of proliferation in BT12 cells. Cells were exposed to 25nM, 50nM, or 100nM MMA for 18h, replaced with drug- free media and monitored using live cell imaging. F: Mithramycin induces morphological changes and apoptosis relative to control cells (solvent). BT12 cells treated with 25nM (left) or 100nM (right) mithramycin for 18h (top) or 48-hours (bottom) in the presence of cleaved caspase 3/7 reagent that fluoresces with caspase activation. G: Mithramycin leads to H3K27me3 amplification in a time-dependent manner that precedes the induction of apoptosis as measured by the cleavage of PARP. Western blot lysates collected at 1h, 2h, 4h, 8h, 12h 16h, 18h of continuous 100nM mithramycin treatment. H: Chromatin immunoprecipitation qPCR (ChIP-qPCR) of H3K27me3 at MYT1 and CCND1 . H3K27me3 occupancy is increased in a time-dependent manner.

    Article Snippet: Tissue was incubated with H3K27me3 (Abcam, 1:250), Cleaved Caspase-3 (Cell Signaling, 1:250), H3K27ac (Abcam, 1:1250), or human mitochondria (Abcam, 1:800) washed and then secondary antibody (Envision+System HRP labelled polymer Anti-Rabbit, Dako 1:100).

    Techniques: Mutagenesis, Generated, Western Blot, Concentration Assay, Live Cell Imaging, Activation Assay, Amplification, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction

    EC8042 induces a mixed phenotype that favors mesenchymal differentiation over apoptosis. A-D:  10X image of section of G401 treated tumors collected on day 4 (left) and day 8 (right) and stained for H3K27me3  (A) , H3K27ac  (B) , human mitochondria  (C) , and cleaved caspase 3  (D) . The sections compare vehicle to treatment started on day 1 with 30 mg/kg of EC8042 administered continuously for 72-hours (3-day pump). H3K27me3 increases and correlates with apoptosis (CC3) while H3K27ac decreases over time. H3K27ac and human mitochondrial staining decrease by day 4 compared with vehicle. Positive staining of H3K27me3 and CC3 is gone by day 8, as is human mitochondrial staining indicating no residual tumor.  E:  Immunohistochemistry analysis of H  E stains from G401 xenograft tumors on 1, 3, and 7-days after treatment with vehicle, 3-day EC8042 pump or 7-day EC8042 pump. EC8042 treated xenograft tumors exhibit evidence of mesenchymal differentiation compared to vehicle. microCT analysis of xenograft tumors on 7-days after treatment exhibit enhanced calcification compared to vehicle.

    Journal: bioRxiv

    Article Title: Direct therapeutic targeting of SWI/SNF induces epigenetic reprogramming and durable tumor regression in rhabdoid tumor

    doi: 10.1101/861484

    Figure Lengend Snippet: EC8042 induces a mixed phenotype that favors mesenchymal differentiation over apoptosis. A-D: 10X image of section of G401 treated tumors collected on day 4 (left) and day 8 (right) and stained for H3K27me3 (A) , H3K27ac (B) , human mitochondria (C) , and cleaved caspase 3 (D) . The sections compare vehicle to treatment started on day 1 with 30 mg/kg of EC8042 administered continuously for 72-hours (3-day pump). H3K27me3 increases and correlates with apoptosis (CC3) while H3K27ac decreases over time. H3K27ac and human mitochondrial staining decrease by day 4 compared with vehicle. Positive staining of H3K27me3 and CC3 is gone by day 8, as is human mitochondrial staining indicating no residual tumor. E: Immunohistochemistry analysis of H E stains from G401 xenograft tumors on 1, 3, and 7-days after treatment with vehicle, 3-day EC8042 pump or 7-day EC8042 pump. EC8042 treated xenograft tumors exhibit evidence of mesenchymal differentiation compared to vehicle. microCT analysis of xenograft tumors on 7-days after treatment exhibit enhanced calcification compared to vehicle.

    Article Snippet: Tissue was incubated with H3K27me3 (Abcam, 1:250), Cleaved Caspase-3 (Cell Signaling, 1:250), H3K27ac (Abcam, 1:1250), or human mitochondria (Abcam, 1:800) washed and then secondary antibody (Envision+System HRP labelled polymer Anti-Rabbit, Dako 1:100).

    Techniques: Staining, Immunohistochemistry

    AAV2 induction of apoptosis in MDA-MB-468 cells results in PARP cleavage following activation of caspases of the intrinsic pathway . MDA-MB-468 monolayer cell cultures were synchronized in G1, followed by infection with AAV2. Cell pellets were collected each day over a 7 day period. Cells were passaged 1:2 on day 2 and day 5. Detection of caspases and their cleavage/activation was performed by Western blotting. Total protein extracts were prepared as described previously [ 48 ]. Sixty micrograms of total protein extracts from AAV2 infected and control MDA-MB-468 cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology). To detect the 35 kDa pro-caspase form of caspase-6, proteins were resolved in a 10% SDS-PAGE gel and to detect the 15 kDa cleaved form of caspase-6, proteins were resolved in a 15% SDS-PAGE gel and detected with a rabbit polyclonal antibody (Cell Signaling Technology). To detect both the pro- and cleaved forms of caspase-7, caspase-8 and caspase-9, proteins were resolved in a 10% SDS-PAGE gel. The 35 kDa pro-caspase form and the 30 kDa and 20 kDa cleaved forms of caspase-7 was detected with a mouse monoclonal antibody (Cell Signaling). Caspase-8 was detected with a mouse monoclonal antibody (Alexis Biochemicals). The 47 kDa pro-caspase and 35 kDa cleaved of caspase-9 was detected with a rabbit polyclonal antibody (Cell Signaling). To detect the pro- (116 kDa) and cleaved- (89 kDa) forms of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Journal: Molecular Cancer

    Article Title: Adeno-associated virus type 2 infection activates caspase dependent and independent apoptosis in multiple breast cancer lines but not in normal mammary epithelial cells

    doi: 10.1186/1476-4598-10-97

    Figure Lengend Snippet: AAV2 induction of apoptosis in MDA-MB-468 cells results in PARP cleavage following activation of caspases of the intrinsic pathway . MDA-MB-468 monolayer cell cultures were synchronized in G1, followed by infection with AAV2. Cell pellets were collected each day over a 7 day period. Cells were passaged 1:2 on day 2 and day 5. Detection of caspases and their cleavage/activation was performed by Western blotting. Total protein extracts were prepared as described previously [ 48 ]. Sixty micrograms of total protein extracts from AAV2 infected and control MDA-MB-468 cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology). To detect the 35 kDa pro-caspase form of caspase-6, proteins were resolved in a 10% SDS-PAGE gel and to detect the 15 kDa cleaved form of caspase-6, proteins were resolved in a 15% SDS-PAGE gel and detected with a rabbit polyclonal antibody (Cell Signaling Technology). To detect both the pro- and cleaved forms of caspase-7, caspase-8 and caspase-9, proteins were resolved in a 10% SDS-PAGE gel. The 35 kDa pro-caspase form and the 30 kDa and 20 kDa cleaved forms of caspase-7 was detected with a mouse monoclonal antibody (Cell Signaling). Caspase-8 was detected with a mouse monoclonal antibody (Alexis Biochemicals). The 47 kDa pro-caspase and 35 kDa cleaved of caspase-9 was detected with a rabbit polyclonal antibody (Cell Signaling). To detect the pro- (116 kDa) and cleaved- (89 kDa) forms of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Article Snippet: To detect the 19 kDa and 17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology) and used at a dilution of 1:1000.

    Techniques: Multiple Displacement Amplification, Activation Assay, Infection, Western Blot, Polyacrylamide Gel Electrophoresis, SDS Page

    AAV2 induction of apoptosis/cell death in MDA-MB-231 cells results in activation of caspases of both the intrinsic and extrinsic pathways but not PARP cleavage . Left panel: MDA-MB-231 monolayer cell cultures were synchronized in G1, followed by infection with AAV2. Cell pellets were collected each day over a 7 day period. Cells were passaged 1:2 on day 2 and day 5. Detection of caspases and their cleavage/activation was performed by Western blotting. Total protein extracts were prepared as described previously [ 48 ]. Sixty micrograms of total protein extracts from AAV2 infected and control MDA-MB-468 cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology). To detect the 35 kDa pro-caspase form of caspase-6, proteins were resolved in a 10% SDS-PAGE gel and to detect the 15 kDa cleaved form of caspase-6, proteins were resolved in a 15% SDS-PAGE gel and detected with a rabbit polyclonal antibody (Cell Signaling Technology). To detect both the pro- and cleaved forms of caspase-7, caspase-8 and caspase-9, proteins were resolved in a 10% SDS-PAGE gel. The 35 kDa pro-caspase form and the 30 kDa/20 kDa cleaved forms of caspase-7 was detected with a mouse monoclonal antibody (Cell Signaling). The pro-caspase and cleaved 28 kDa form of caspase-8 was detected with a mouse monoclonal antibody (Alexis Biochemicals). The 47 kDa pro-caspase and 37 kDa/35 kDa cleaved forms of caspase-9 were detected with a rabbit polyclonal antibody (Cell Signaling). To detect the pro- (116 kDa) form of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Journal: Molecular Cancer

    Article Title: Adeno-associated virus type 2 infection activates caspase dependent and independent apoptosis in multiple breast cancer lines but not in normal mammary epithelial cells

    doi: 10.1186/1476-4598-10-97

    Figure Lengend Snippet: AAV2 induction of apoptosis/cell death in MDA-MB-231 cells results in activation of caspases of both the intrinsic and extrinsic pathways but not PARP cleavage . Left panel: MDA-MB-231 monolayer cell cultures were synchronized in G1, followed by infection with AAV2. Cell pellets were collected each day over a 7 day period. Cells were passaged 1:2 on day 2 and day 5. Detection of caspases and their cleavage/activation was performed by Western blotting. Total protein extracts were prepared as described previously [ 48 ]. Sixty micrograms of total protein extracts from AAV2 infected and control MDA-MB-468 cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology). To detect the 35 kDa pro-caspase form of caspase-6, proteins were resolved in a 10% SDS-PAGE gel and to detect the 15 kDa cleaved form of caspase-6, proteins were resolved in a 15% SDS-PAGE gel and detected with a rabbit polyclonal antibody (Cell Signaling Technology). To detect both the pro- and cleaved forms of caspase-7, caspase-8 and caspase-9, proteins were resolved in a 10% SDS-PAGE gel. The 35 kDa pro-caspase form and the 30 kDa/20 kDa cleaved forms of caspase-7 was detected with a mouse monoclonal antibody (Cell Signaling). The pro-caspase and cleaved 28 kDa form of caspase-8 was detected with a mouse monoclonal antibody (Alexis Biochemicals). The 47 kDa pro-caspase and 37 kDa/35 kDa cleaved forms of caspase-9 were detected with a rabbit polyclonal antibody (Cell Signaling). To detect the pro- (116 kDa) form of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Article Snippet: To detect the 19 kDa and 17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology) and used at a dilution of 1:1000.

    Techniques: Multiple Displacement Amplification, Activation Assay, Infection, Western Blot, Polyacrylamide Gel Electrophoresis, SDS Page

    AAV2 infection of nHMECs does not result in activation of caspases of either the intrinsic and extrinsic pathways of apoptosis . nHMEC monolayer cultures were synchronized in G1 as described, followed by infection with AAV2. Cell pellets were collected each day over a 5 day period. Cells were passaged 1:2 on day 2. Detection of caspases and their cleavage/activation was performed by Western blotting. Sixty micrograms of total protein extracts from AAV2 infected and control nHMEC cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect caspase-8 proteins were resolved in a 10% SDS-PAGE gel and detected with a mouse monoclonal antibody (Alexis Biochemicals). To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the pro- (116 kDa) form of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Journal: Molecular Cancer

    Article Title: Adeno-associated virus type 2 infection activates caspase dependent and independent apoptosis in multiple breast cancer lines but not in normal mammary epithelial cells

    doi: 10.1186/1476-4598-10-97

    Figure Lengend Snippet: AAV2 infection of nHMECs does not result in activation of caspases of either the intrinsic and extrinsic pathways of apoptosis . nHMEC monolayer cultures were synchronized in G1 as described, followed by infection with AAV2. Cell pellets were collected each day over a 5 day period. Cells were passaged 1:2 on day 2. Detection of caspases and their cleavage/activation was performed by Western blotting. Sixty micrograms of total protein extracts from AAV2 infected and control nHMEC cells were resolved in SDS-polyacrylamide gel electrophoresis (PAGE) gels. To detect caspase-8 proteins were resolved in a 10% SDS-PAGE gel and detected with a mouse monoclonal antibody (Alexis Biochemicals). To detect the 35 kDa pro-caspase form of caspase-3, proteins were resolved in a 10% SDS-PAGE gel and detected with caspase-3 rabbit monoclonal antibody (Cell Signaling Technology). To detect the pro- (116 kDa) form of PARP, proteins were resolved in a 7.5% SDS-PAGE gel and detected with a rabbit monoclonal antibody (Cell Signaling).

    Article Snippet: To detect the 19 kDa and 17-kDa cleaved caspase-3 forms, proteins were resolved in a 15% SDS-PAGE gel and detected with rabbit polyclonal antibody against cleaved caspase-3 (Cell Signaling Technology) and used at a dilution of 1:1000.

    Techniques: Infection, Activation Assay, Western Blot, Polyacrylamide Gel Electrophoresis, SDS Page