cell culture human foreskin fibroblast hs68 cells  (Sanyo)

 
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    Structured Review

    Sanyo cell culture human foreskin fibroblast hs68 cells
    The effect of decanal on UVB-induced collagen degradation in <t>Hs68</t> dermal fibroblasts. The collagen content was measured in the supernatant of Hs68 cells treated with either the vehicle or 25, 50 and 100 μM decanal for 24 h after UVB exposure. The final procollagen type I level was normalized to the total cellular protein content. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p
    Cell Culture Human Foreskin Fibroblast Hs68 Cells, supplied by Sanyo, used in various techniques. Bioz Stars score: 84/100, based on 101 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    cell culture human foreskin fibroblast hs68 cells - by Bioz Stars, 2020-08
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    Images

    1) Product Images from "Decanal Protects against UVB-Induced Photoaging in Human Dermal Fibroblasts via the cAMP Pathway"

    Article Title: Decanal Protects against UVB-Induced Photoaging in Human Dermal Fibroblasts via the cAMP Pathway

    Journal: Nutrients

    doi: 10.3390/nu12051214

    The effect of decanal on UVB-induced collagen degradation in Hs68 dermal fibroblasts. The collagen content was measured in the supernatant of Hs68 cells treated with either the vehicle or 25, 50 and 100 μM decanal for 24 h after UVB exposure. The final procollagen type I level was normalized to the total cellular protein content. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p
    Figure Legend Snippet: The effect of decanal on UVB-induced collagen degradation in Hs68 dermal fibroblasts. The collagen content was measured in the supernatant of Hs68 cells treated with either the vehicle or 25, 50 and 100 μM decanal for 24 h after UVB exposure. The final procollagen type I level was normalized to the total cellular protein content. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p

    Techniques Used:

    The effect of decanal on cell viability in Hs68 dermal fibroblasts. The cell viability of both ( A ) non-ultraviolet (UV) B-exposed and ( B ) UVB-exposed Hs68 cells was determined after incubation with either the vehicle or 25–200 μM decanal for 24 h. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p
    Figure Legend Snippet: The effect of decanal on cell viability in Hs68 dermal fibroblasts. The cell viability of both ( A ) non-ultraviolet (UV) B-exposed and ( B ) UVB-exposed Hs68 cells was determined after incubation with either the vehicle or 25–200 μM decanal for 24 h. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p

    Techniques Used: Incubation

    The effect of decanal on collagen synthesis via the cAMP pathway. Hs68 cells were treated with decanal or the vehicle and exposed to UVB. The SQ22536 or the vehicle was pretreated for 1 h before the decanal treatment. ( A ) Collagen contents; ( B ) mRNA expression of matrix metalloproteinase ( MMP) 1, MMP3 and MMP9 ; and ( C ) mRNA expression of collagen type I alpha 1 chain ( COL1A1 ), COL1A2 and COL3A1 were analyzed. The final procollagen type I level was normalized to the total cellular protein content. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p
    Figure Legend Snippet: The effect of decanal on collagen synthesis via the cAMP pathway. Hs68 cells were treated with decanal or the vehicle and exposed to UVB. The SQ22536 or the vehicle was pretreated for 1 h before the decanal treatment. ( A ) Collagen contents; ( B ) mRNA expression of matrix metalloproteinase ( MMP) 1, MMP3 and MMP9 ; and ( C ) mRNA expression of collagen type I alpha 1 chain ( COL1A1 ), COL1A2 and COL3A1 were analyzed. The final procollagen type I level was normalized to the total cellular protein content. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p

    Techniques Used: Expressing

    The effect of decanal on the cyclic adenosine monophosphate–protein kinase A (cAMP-PKA) pathway in Hs68 dermal fibroblasts. ( A ) The time course of decanal-induced intracellular cAMP levels was determined after incubation with either the vehicle or 50 μM decanal for 5, 15, 30, 45 and 60 min. ( B ) The protein expression of protein kinase A catalytic subunit (PKA Cα) was measured after incubation with 50 μM decanal for 30 min. Data are shown as the mean ± SEM ( n = 3). Significant differences between decanal and non-stimulated control groups are shown as * p
    Figure Legend Snippet: The effect of decanal on the cyclic adenosine monophosphate–protein kinase A (cAMP-PKA) pathway in Hs68 dermal fibroblasts. ( A ) The time course of decanal-induced intracellular cAMP levels was determined after incubation with either the vehicle or 50 μM decanal for 5, 15, 30, 45 and 60 min. ( B ) The protein expression of protein kinase A catalytic subunit (PKA Cα) was measured after incubation with 50 μM decanal for 30 min. Data are shown as the mean ± SEM ( n = 3). Significant differences between decanal and non-stimulated control groups are shown as * p

    Techniques Used: Incubation, Expressing

    The effect of decanal on the UVB-induced mitogen-activated protein kinase (MAPK) pathway in Hs68 dermal fibroblasts. Hs68 cells were treated with either decanal or the vehicle and exposed to UVB. ( A ) Phosphorylation of MAPK proteins (p38, c-Jun N-terminal kinase (JNK) and extracellular-signal-regulated kinase (ERK)) and ( B ) c-Jun and c-Fos (MAPK downstream molecules) were determined. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p
    Figure Legend Snippet: The effect of decanal on the UVB-induced mitogen-activated protein kinase (MAPK) pathway in Hs68 dermal fibroblasts. Hs68 cells were treated with either decanal or the vehicle and exposed to UVB. ( A ) Phosphorylation of MAPK proteins (p38, c-Jun N-terminal kinase (JNK) and extracellular-signal-regulated kinase (ERK)) and ( B ) c-Jun and c-Fos (MAPK downstream molecules) were determined. Data are shown as the mean ± SEM ( n = 3). Significant differences are indicated as * p

    Techniques Used:

    The effect of decanal on hyaluronic acid synthesis via the cAMP signaling pathway. Hs68 cells were treated with either decanal or the vehicle and exposed to UVB. Other samples were pretreated with either SQ22536 or the vehicle for 1 h before decanal treatment. ( A ) Hyaluronic acid concentrations and ( B ) the mRNA expression of HAS2 were examined. The final hyaluronic acid levels were normalized to the total cellular protein content. Data are shown as the mean ±SEM ( n = 3). Significant differences are indicated as * p
    Figure Legend Snippet: The effect of decanal on hyaluronic acid synthesis via the cAMP signaling pathway. Hs68 cells were treated with either decanal or the vehicle and exposed to UVB. Other samples were pretreated with either SQ22536 or the vehicle for 1 h before decanal treatment. ( A ) Hyaluronic acid concentrations and ( B ) the mRNA expression of HAS2 were examined. The final hyaluronic acid levels were normalized to the total cellular protein content. Data are shown as the mean ±SEM ( n = 3). Significant differences are indicated as * p

    Techniques Used: Expressing

    2) Product Images from "Suppression of MIF-induced neuronal apoptosis may underlie the therapeutic effects of effective components of Fufang Danshen in the treatment of Alzheimer's disease"

    Article Title: Suppression of MIF-induced neuronal apoptosis may underlie the therapeutic effects of effective components of Fufang Danshen in the treatment of Alzheimer's disease

    Journal: Acta Pharmacologica Sinica

    doi: 10.1038/aps.2017.210

    Effects of MIF on the growth of SH-SY5Y cells. (A) SH-SY5Y cells were treated with ISO-1 (10 μmol/L), LY294002 (10 μmol/L) and PS1145 (10 μmol/L) for 36 h, and cell proliferation was examined by MTT assay. (B) SH-SY5Y cells were
    Figure Legend Snippet: Effects of MIF on the growth of SH-SY5Y cells. (A) SH-SY5Y cells were treated with ISO-1 (10 μmol/L), LY294002 (10 μmol/L) and PS1145 (10 μmol/L) for 36 h, and cell proliferation was examined by MTT assay. (B) SH-SY5Y cells were

    Techniques Used: MTT Assay

    Effect of the FFDS components and their combination on MIF-induced apoptosis in SH-SY5Y cells using flow cytometry. (A) Effect of MIF on SH-SY5Y cells. After SH-SY5Y cells were treated with 50 ng/mL MIF for 24 h, the cell membrane permeability was increased,
    Figure Legend Snippet: Effect of the FFDS components and their combination on MIF-induced apoptosis in SH-SY5Y cells using flow cytometry. (A) Effect of MIF on SH-SY5Y cells. After SH-SY5Y cells were treated with 50 ng/mL MIF for 24 h, the cell membrane permeability was increased,

    Techniques Used: Flow Cytometry, Cytometry, Permeability

    Effect of the FFDS components and their combination on MIF-induced SH-SY5Y cell apoptosis. (A) Effect of MIF on SH-SY5Y cell growth ( *** P
    Figure Legend Snippet: Effect of the FFDS components and their combination on MIF-induced SH-SY5Y cell apoptosis. (A) Effect of MIF on SH-SY5Y cell growth ( *** P

    Techniques Used:

    3) Product Images from "Potential of Maintaining a Healthy Vaginal Environment by Two Lactobacillus Strains Isolated from Cocoa Fermentation"

    Article Title: Potential of Maintaining a Healthy Vaginal Environment by Two Lactobacillus Strains Isolated from Cocoa Fermentation

    Journal: BioMed Research International

    doi: 10.1155/2018/7571954

    Death of HMVII cells after 24 h of incubation with LAMP from M. hominis (MhLAMP) or M. genitalium (MgLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p
    Figure Legend Snippet: Death of HMVII cells after 24 h of incubation with LAMP from M. hominis (MhLAMP) or M. genitalium (MgLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p

    Techniques Used: Incubation

    Interaction of HMVII cells with 4 μ g/mL of U. parvum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UpLAMP. (c) HMVII with UpLAMP and L. fermentum FA4. (d) HMVII with UpLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with 4 μ g/mL of U. parvum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UpLAMP. (c) HMVII with UpLAMP and L. fermentum FA4. (d) HMVII with UpLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    Interaction of HMVII cells with 4 μ g/mL of M. hominis LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with MhLAMP. (c) HMVII with MhLAMP and L. fermentum FA4. (d) HMVII with MhLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with 4 μ g/mL of M. hominis LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with MhLAMP. (c) HMVII with MhLAMP and L. fermentum FA4. (d) HMVII with MhLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    Death of HMVII cells after 24 h of incubation with LAMP from U. parvum (UpLAMP) or U. urealyticum (UuLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p
    Figure Legend Snippet: Death of HMVII cells after 24 h of incubation with LAMP from U. parvum (UpLAMP) or U. urealyticum (UuLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p

    Techniques Used: Incubation

    Interaction of HMVII cells with U. urealyticum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UuLAMP. (c) HMVII with UuLAMP and L. fermentum FA4. (d) HMVII with UuLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with U. urealyticum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UuLAMP. (c) HMVII with UuLAMP and L. fermentum FA4. (d) HMVII with UuLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    4) Product Images from "NovelmiRNA-25 inhibits AMPD2 in peripheral blood mononuclear cells of patients with systemic lupus erythematosus and represents a promising novel biomarker"

    Article Title: NovelmiRNA-25 inhibits AMPD2 in peripheral blood mononuclear cells of patients with systemic lupus erythematosus and represents a promising novel biomarker

    Journal: Journal of Translational Medicine

    doi: 10.1186/s12967-018-1739-5

    NovelmiRNA-25 targets AMPD2 . a Predicted target sites in the AMPD2 3′UTR based on the NovelmiRNA-25 seed region. Mutations in the AMPD2 3′UTR generated missense sequences unable to pair with the NovelmiRNA-25 seed region. The structure of NovelmiRNA-25 bound to AMPD2 is depicted. b NovelmiRNA-25 and the AMPD2 3′UTR seeding region, highlighted in green, are highly conserved in mammals. c Activity of the luciferase gene linked to the 3′UTR of AMPD2 . pmirGLO firefly luciferase reporter plasmids with wild-type or mutated 3′UTR sequences of AMPD2 were transiently transfected into cells with NovelmiRNA-25 precursor or negative control and a Renilla luciferase reporter for normalization. Luciferase activities were measured after 48 h. The mean of the results from the cells transfected with pmirGLO control vector was set as 100%. Data are the mean and standard deviation (SD) of separate transfections (n = 3). d Downregulation of endogenous AMPD2 protein expression by NovelmiRNA-25. Western blotting of AMPD2 protein after transfection with negative control (NC) or NovelmiRNA-25 mimic in HEK293T cells. Expression levels were normalized to that of GAPDH; **P
    Figure Legend Snippet: NovelmiRNA-25 targets AMPD2 . a Predicted target sites in the AMPD2 3′UTR based on the NovelmiRNA-25 seed region. Mutations in the AMPD2 3′UTR generated missense sequences unable to pair with the NovelmiRNA-25 seed region. The structure of NovelmiRNA-25 bound to AMPD2 is depicted. b NovelmiRNA-25 and the AMPD2 3′UTR seeding region, highlighted in green, are highly conserved in mammals. c Activity of the luciferase gene linked to the 3′UTR of AMPD2 . pmirGLO firefly luciferase reporter plasmids with wild-type or mutated 3′UTR sequences of AMPD2 were transiently transfected into cells with NovelmiRNA-25 precursor or negative control and a Renilla luciferase reporter for normalization. Luciferase activities were measured after 48 h. The mean of the results from the cells transfected with pmirGLO control vector was set as 100%. Data are the mean and standard deviation (SD) of separate transfections (n = 3). d Downregulation of endogenous AMPD2 protein expression by NovelmiRNA-25. Western blotting of AMPD2 protein after transfection with negative control (NC) or NovelmiRNA-25 mimic in HEK293T cells. Expression levels were normalized to that of GAPDH; **P

    Techniques Used: Generated, Activity Assay, Luciferase, Transfection, Negative Control, Plasmid Preparation, Standard Deviation, Expressing, Western Blot

    5) Product Images from "Ultrasmall dopamine-coated nanogolds: preparation, characteristics, and CT imaging"

    Article Title: Ultrasmall dopamine-coated nanogolds: preparation, characteristics, and CT imaging

    Journal: Journal of Experimental Nanoscience

    doi: 10.1080/17458080.2015.1102343

    The cell viability results of (a) HeLa, (b) PC-3, and (c) IMR-90 cell lines when exposed to WDU AuNPs and WDU AuNPs@DPAs at different concentrations by MTT assay.
    Figure Legend Snippet: The cell viability results of (a) HeLa, (b) PC-3, and (c) IMR-90 cell lines when exposed to WDU AuNPs and WDU AuNPs@DPAs at different concentrations by MTT assay.

    Techniques Used: MTT Assay

    6) Product Images from "Characterization of the transcriptional and metabolic responses of pediatric high grade gliomas to mTOR-HIF-1α axis inhibition"

    Article Title: Characterization of the transcriptional and metabolic responses of pediatric high grade gliomas to mTOR-HIF-1α axis inhibition

    Journal: Oncotarget

    doi: 10.18632/oncotarget.16500

    Cell effects of rapamycin, irinotecan and the combination and impact on the mTOR-HIF-1/2α pathway in KNS42, SF188 and UW479 in both normoxic and hypoxic conditions The immunoblotting analyses of the protein expressions are presented for the cleaved PARP, phospho-H2AX and LC3B as markers of apoptosis, double strand breaks and autophagy, respectively, and for the phosphorylated forms of RICTOR, AKT, ERK and S6RP as markers of mTOR pathway and HIF-1/2α protein accumulations in Figure 5A . The Figure 5B is illustrated the quantification of western blot analyses (c-PARP/control ratio and LC3BI/LC3BII ratio). The Figure 5C is presented cleaved-caspase 3 western blot analysesfor each cell line and the significant gene set enrichment analyses testing apoptosis genes in SF188 and autophagy pathway in UW479.
    Figure Legend Snippet: Cell effects of rapamycin, irinotecan and the combination and impact on the mTOR-HIF-1/2α pathway in KNS42, SF188 and UW479 in both normoxic and hypoxic conditions The immunoblotting analyses of the protein expressions are presented for the cleaved PARP, phospho-H2AX and LC3B as markers of apoptosis, double strand breaks and autophagy, respectively, and for the phosphorylated forms of RICTOR, AKT, ERK and S6RP as markers of mTOR pathway and HIF-1/2α protein accumulations in Figure 5A . The Figure 5B is illustrated the quantification of western blot analyses (c-PARP/control ratio and LC3BI/LC3BII ratio). The Figure 5C is presented cleaved-caspase 3 western blot analysesfor each cell line and the significant gene set enrichment analyses testing apoptosis genes in SF188 and autophagy pathway in UW479.

    Techniques Used: Western Blot

    Impact of hypoxia (1% O2) on mTOR pathway activation, gene expression and proliferation in KNS42, SF188 and UW479 cell lines Figure 2A and 2B : Comparative immunoblotting analyses of mTOR pathway activation in KNS42, SF188, UW479 and the patient-derived cell line TC68 cultured in both normoxic and hypoxic conditions. Figure 2C, 2D, 2E, 2F, 2G, 2H : Comparative transcriptomic analyses. The relative expressions of HIF-1/2α and MYC are described in Figure 2C . Unsupervised analyses showing the top genes significantly upregulated or down-regulated in response to hypoxia (1% O2) comparatively to normoxia conditions are described in Figure 2D . Gene set enrichment analysis testing HIFs gene targets is shown in Figure 2E , MYC gene targets in Figure 2F and genes involved in cell cycle progression in Figure 2G . Figure 2H is presenting the impact of hypoxia on the proliferations of KNS42, SF188 and UW479, assessed by a colorimetric proliferation assay using crystal violet and the calculation of doubling time. In the Figure 2I , the effect on proliferation of siRNA transient transfection against HIF-1α, HIF-2α or both in SF188, KNS42 and UW479 cell lines in hypoxia. This effect was measured in triplicate and compared at 72 hours relatively to the control growth. *: p
    Figure Legend Snippet: Impact of hypoxia (1% O2) on mTOR pathway activation, gene expression and proliferation in KNS42, SF188 and UW479 cell lines Figure 2A and 2B : Comparative immunoblotting analyses of mTOR pathway activation in KNS42, SF188, UW479 and the patient-derived cell line TC68 cultured in both normoxic and hypoxic conditions. Figure 2C, 2D, 2E, 2F, 2G, 2H : Comparative transcriptomic analyses. The relative expressions of HIF-1/2α and MYC are described in Figure 2C . Unsupervised analyses showing the top genes significantly upregulated or down-regulated in response to hypoxia (1% O2) comparatively to normoxia conditions are described in Figure 2D . Gene set enrichment analysis testing HIFs gene targets is shown in Figure 2E , MYC gene targets in Figure 2F and genes involved in cell cycle progression in Figure 2G . Figure 2H is presenting the impact of hypoxia on the proliferations of KNS42, SF188 and UW479, assessed by a colorimetric proliferation assay using crystal violet and the calculation of doubling time. In the Figure 2I , the effect on proliferation of siRNA transient transfection against HIF-1α, HIF-2α or both in SF188, KNS42 and UW479 cell lines in hypoxia. This effect was measured in triplicate and compared at 72 hours relatively to the control growth. *: p

    Techniques Used: Activation Assay, Expressing, Derivative Assay, Cell Culture, Proliferation Assay, Transfection

    Efficacy of rapamycin, irinotecan and the combination of both in terms of anti-proliferative effect measured at 72 hours from treatment in normoxic and hypoxic conditions in the 3 cell lines and in TC68/TC35 This anti-proliferative effect was measured using a crystal violet colorimetric proliferation assay in the 3 cell lines and a trypan blue exclusion assay with automated counting of cells in TC68 and TC35. The 50% growth inhibition (GI50) values (95% CI) are reported for irinotecan in Figure 4A and for rapamycin in Figure 4B . Proliferation curves are reported in Figure 4C and combination indexes (CI) in Figure 4D . For TC68, the effect of drugs on neurosphere formation is described in Figure 4E (phase contrast microscopy) and the effect on proliferation in Figure 4F for TC68 and TC35.
    Figure Legend Snippet: Efficacy of rapamycin, irinotecan and the combination of both in terms of anti-proliferative effect measured at 72 hours from treatment in normoxic and hypoxic conditions in the 3 cell lines and in TC68/TC35 This anti-proliferative effect was measured using a crystal violet colorimetric proliferation assay in the 3 cell lines and a trypan blue exclusion assay with automated counting of cells in TC68 and TC35. The 50% growth inhibition (GI50) values (95% CI) are reported for irinotecan in Figure 4A and for rapamycin in Figure 4B . Proliferation curves are reported in Figure 4C and combination indexes (CI) in Figure 4D . For TC68, the effect of drugs on neurosphere formation is described in Figure 4E (phase contrast microscopy) and the effect on proliferation in Figure 4F for TC68 and TC35.

    Techniques Used: Proliferation Assay, Trypan Blue Exclusion Assay, Inhibition, Microscopy

    7) Product Images from "CTCF and cohesin regulate chromatin loop stability with distinct dynamics"

    Article Title: CTCF and cohesin regulate chromatin loop stability with distinct dynamics

    Journal: eLife

    doi: 10.7554/eLife.25776

    Supplementary and control CTCF FRAP experiments. ( A ) Representative raw confocal microscopy images of H2B-Halo, C59 Halo-mCTCF and C32 Halo-hCTCF each labeled with 1 μM TMR just before, 1 s after, 10 s after and 4 min after bleaching a 1 μm circular spot (green circle). mES cells especially show significant movement on the minute time-scale and all movies were drift-corrected. ( B ) Photo-bleaching corrected FRAP curves measured at one frame per second in mESCs. C59 and C87 Halo-mCTCF show the behavior of endogenously tagged CTCF, and transiently transfected Halo-mCTCF expressed from a CMV-promoter is shown in black (OE: over-expressed). As can be clearly seen, the CTCF FRAP recovery is much faster when over-expressed. Error bars show standard error. ( C ) Photobleaching-corrected FRAP curves measured at one frame per second in human U2OS cells. Halo-3xNLS and H2B-Halo are controls for rapid and negligible recovery, respectively, demonstrating the validity our photobleaching- and drift-correction approaches. CTCF recovery in human U2OS cells is similar, albeit slightly slower than in mESCs. ( D ) Model-fitting of C87 and C59 Halo-mCTCF FRAP curves using a reaction dominant model (Materials and methods) for FRAP experiments at either 11 min and 0.5 Hz or 5 min and 1 Hz. The model-inferred residence time and the 95% confidence interval (CI) from the fitting is shown. ( E ) Dynamics of Halo-mCTCF (C59 mESCs) does not change between G1- and S/G2-phase of the cell cycle. G1 and S/G2 were distinguished using the Fucci system ( Figure 2—figure supplement 4 ). DOI: http://dx.doi.org/10.7554/eLife.25776.011
    Figure Legend Snippet: Supplementary and control CTCF FRAP experiments. ( A ) Representative raw confocal microscopy images of H2B-Halo, C59 Halo-mCTCF and C32 Halo-hCTCF each labeled with 1 μM TMR just before, 1 s after, 10 s after and 4 min after bleaching a 1 μm circular spot (green circle). mES cells especially show significant movement on the minute time-scale and all movies were drift-corrected. ( B ) Photo-bleaching corrected FRAP curves measured at one frame per second in mESCs. C59 and C87 Halo-mCTCF show the behavior of endogenously tagged CTCF, and transiently transfected Halo-mCTCF expressed from a CMV-promoter is shown in black (OE: over-expressed). As can be clearly seen, the CTCF FRAP recovery is much faster when over-expressed. Error bars show standard error. ( C ) Photobleaching-corrected FRAP curves measured at one frame per second in human U2OS cells. Halo-3xNLS and H2B-Halo are controls for rapid and negligible recovery, respectively, demonstrating the validity our photobleaching- and drift-correction approaches. CTCF recovery in human U2OS cells is similar, albeit slightly slower than in mESCs. ( D ) Model-fitting of C87 and C59 Halo-mCTCF FRAP curves using a reaction dominant model (Materials and methods) for FRAP experiments at either 11 min and 0.5 Hz or 5 min and 1 Hz. The model-inferred residence time and the 95% confidence interval (CI) from the fitting is shown. ( E ) Dynamics of Halo-mCTCF (C59 mESCs) does not change between G1- and S/G2-phase of the cell cycle. G1 and S/G2 were distinguished using the Fucci system ( Figure 2—figure supplement 4 ). DOI: http://dx.doi.org/10.7554/eLife.25776.011

    Techniques Used: Confocal Microscopy, Labeling, Transfection

    Tagging CTCF and Rad21 does not affect expression of key pluripotency genes or CTCF and Rad21 protein levels. ( A ) Expression of key mouse embryonic stem cell genes measured by qPCR was similar in wild-type (blue) and C59 (FLAG-Halo-mCTCF; mRad21-SNAP f -V5) (red) JM8.N4 mouse embryonic stem cells. ( B ) CTCF (red) and Rad21 (green) protein levels as measured by western blot and normalized to either H3 levels (solid bar) or TBP (hashed bar) was similar between wild-type and tagged mouse embryonic stem cells (WT, C87, C45, C59) and similar between wild-type and tagged human U2OS cells (WT, C32). Error bars show standard deviation among three replicates. DOI: http://dx.doi.org/10.7554/eLife.25776.006
    Figure Legend Snippet: Tagging CTCF and Rad21 does not affect expression of key pluripotency genes or CTCF and Rad21 protein levels. ( A ) Expression of key mouse embryonic stem cell genes measured by qPCR was similar in wild-type (blue) and C59 (FLAG-Halo-mCTCF; mRad21-SNAP f -V5) (red) JM8.N4 mouse embryonic stem cells. ( B ) CTCF (red) and Rad21 (green) protein levels as measured by western blot and normalized to either H3 levels (solid bar) or TBP (hashed bar) was similar between wild-type and tagged mouse embryonic stem cells (WT, C87, C45, C59) and similar between wild-type and tagged human U2OS cells (WT, C32). Error bars show standard deviation among three replicates. DOI: http://dx.doi.org/10.7554/eLife.25776.006

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Standard Deviation

    Supplementary stroboscopic paSMT experiments and controls. ( A ) Sketch illustrating stroboscopic paSMT. Sketch illustrating labeling Halo-tagged proteins, e.g. CTCF or Rad21, with PA-JF646. This dye remains dark until 405 nm activation, which converts it to regular fluorescent JF646. The advantage is that thousands of single-molecule trajectories can be recorded from a single cell at a density of ~0.5 fluorescent molecules per nucleus per frame, which makes tracking unambiguous, by using very low intensity 405 nm excitation. Since high 633 nm laser powers are used, most molecules bleach within 3–8 frames. We use PA-JF646 instead of PA-JF549 since the red-shifted 633 nm laser induces less photo-toxicity, although the displacement histograms were identical between PA-JF549 and PA-JF646. Moreover, we never record for more than 2 min per cell. We observed no visible signs of photo-toxicity after 2 min of paSMT. Below, sketch illustrating stroboscopic illumination. To minimize ‘motion-blurring’ of fast-diffusing molecules, we used pulsed 633 nm excitation with 1 ms pulses. The camera integration time was 4 ms + ~0.447 ms (frame-transfer mode) resulting in a frame rate of roughly 225 Hz. Below, raw microscopy images demonstrating that even fast-diffusing molecules can be imaged and tracked (red lines) at high-signal-to-background. ( B ) Overview of two-state dynamic displacement model. Full details are provided in the Materials and methods. Briefly, the model assumes molecules can exist in either a chromatin associating (specific and non-specific) state called ‘bound’ or in a free 3D diffusion state called ‘free’. A mathematical model describing how the distribution of displacements, r , depends on the time delay, fraction bound, diffusion constants, localization error and axial detection slice is shown below. Overall, the model contains three fitted parameters, which were estimated using least squares fitting to the raw displacements considering the first seven displacements (Δτ ~4.5 ms to 31.5 ms). For ease of visualization, we show displacement histograms in ( C–F ), but the fitting was performed on cumulative distribution functions (CDFs) to minimize binning artifacts. ( C–F ) displacement histograms for various cell lines all measured using the approach in ( A–B ). For ease of visualization, the displacement histograms are cut off at 1050 nm, but longer trajectories were included in the model fitting. ( C ) shows Halo-only and Halo-3xNLS in mESCs, which show negligible binding. Note, that most fast-diffusing molecules eventually move out of the focal plane. ( D ) shows various Halo-mCTCF constructs. C59 and C87 are endogenous Halo-mCTCF knock-ins. pL30-wt-Halo-mCTCF was transiently expressed using a weak promoter ( L30 ). Compared to Halo-mCTCF overexpressed by the strong CMV promoter, overexpressing Halo-mCTCF using a weak promoter ( L30 ) causes only a minor (10 percentage points; likely due to saturation of binding sites) underestimation of the fraction bound. Right, two transiently transfected Halo-mCTCF mutants: 11ZF-mut-Halo-mCTCF is a CTCF mutant with an essential His amino acid in all 11 zinc-fingers mutated to Arg, which should abolish specific DNA-binding. We used this mutant to estimate the non-specifically bound fraction. ΔZF-Halo-mCTCF has the entire 11-zinc-finger domain deleted. We used this mutant to verify that the zinc-finger domain solely is responsible for chromatin association. ( E ) H2B-Halo and Rad21 experiments in mESCs. We used H2B-halo as a control for a protein that is almost exclusively bound. Note that since we use an EF1a promoter to express H2B, which is not cell-cycle regulated, some H2B-molecules do show free diffusion. mRad21-Halo in S/G2 and G1 are also shown, as is transiently transfected wt-mRad21-Halo expressed using the low-expression promoter, L30. Even though mRad21-Halo is only weakly overexpressed, most molecules show free 3D diffusion. The overexpression artifact may be caused by the fact that without similar overexpression of Smc1, Smc3 and SA1/2, most Rad21 cannot form cohesin complexes. Finally, a Rad21-mutant (F601R, L605R, Q617K), which cannot form cohesin complexes was used to estimate the non-specifically bound Rad21 fractions. ( F ) Stroboscopic paSMT experiments in human U2OS cells. H2B-Halo and Halo-3xNLS were used as controls for mostly bound and free molecules and the same zinc-finger mutants as in mESCs were transiently transfected as control for non-specific chromatin association. Note that C32 Halo-hCTCF shows slightly more free diffusion than C59 and C87 in mESCs. DOI: http://dx.doi.org/10.7554/eLife.25776.017
    Figure Legend Snippet: Supplementary stroboscopic paSMT experiments and controls. ( A ) Sketch illustrating stroboscopic paSMT. Sketch illustrating labeling Halo-tagged proteins, e.g. CTCF or Rad21, with PA-JF646. This dye remains dark until 405 nm activation, which converts it to regular fluorescent JF646. The advantage is that thousands of single-molecule trajectories can be recorded from a single cell at a density of ~0.5 fluorescent molecules per nucleus per frame, which makes tracking unambiguous, by using very low intensity 405 nm excitation. Since high 633 nm laser powers are used, most molecules bleach within 3–8 frames. We use PA-JF646 instead of PA-JF549 since the red-shifted 633 nm laser induces less photo-toxicity, although the displacement histograms were identical between PA-JF549 and PA-JF646. Moreover, we never record for more than 2 min per cell. We observed no visible signs of photo-toxicity after 2 min of paSMT. Below, sketch illustrating stroboscopic illumination. To minimize ‘motion-blurring’ of fast-diffusing molecules, we used pulsed 633 nm excitation with 1 ms pulses. The camera integration time was 4 ms + ~0.447 ms (frame-transfer mode) resulting in a frame rate of roughly 225 Hz. Below, raw microscopy images demonstrating that even fast-diffusing molecules can be imaged and tracked (red lines) at high-signal-to-background. ( B ) Overview of two-state dynamic displacement model. Full details are provided in the Materials and methods. Briefly, the model assumes molecules can exist in either a chromatin associating (specific and non-specific) state called ‘bound’ or in a free 3D diffusion state called ‘free’. A mathematical model describing how the distribution of displacements, r , depends on the time delay, fraction bound, diffusion constants, localization error and axial detection slice is shown below. Overall, the model contains three fitted parameters, which were estimated using least squares fitting to the raw displacements considering the first seven displacements (Δτ ~4.5 ms to 31.5 ms). For ease of visualization, we show displacement histograms in ( C–F ), but the fitting was performed on cumulative distribution functions (CDFs) to minimize binning artifacts. ( C–F ) displacement histograms for various cell lines all measured using the approach in ( A–B ). For ease of visualization, the displacement histograms are cut off at 1050 nm, but longer trajectories were included in the model fitting. ( C ) shows Halo-only and Halo-3xNLS in mESCs, which show negligible binding. Note, that most fast-diffusing molecules eventually move out of the focal plane. ( D ) shows various Halo-mCTCF constructs. C59 and C87 are endogenous Halo-mCTCF knock-ins. pL30-wt-Halo-mCTCF was transiently expressed using a weak promoter ( L30 ). Compared to Halo-mCTCF overexpressed by the strong CMV promoter, overexpressing Halo-mCTCF using a weak promoter ( L30 ) causes only a minor (10 percentage points; likely due to saturation of binding sites) underestimation of the fraction bound. Right, two transiently transfected Halo-mCTCF mutants: 11ZF-mut-Halo-mCTCF is a CTCF mutant with an essential His amino acid in all 11 zinc-fingers mutated to Arg, which should abolish specific DNA-binding. We used this mutant to estimate the non-specifically bound fraction. ΔZF-Halo-mCTCF has the entire 11-zinc-finger domain deleted. We used this mutant to verify that the zinc-finger domain solely is responsible for chromatin association. ( E ) H2B-Halo and Rad21 experiments in mESCs. We used H2B-halo as a control for a protein that is almost exclusively bound. Note that since we use an EF1a promoter to express H2B, which is not cell-cycle regulated, some H2B-molecules do show free diffusion. mRad21-Halo in S/G2 and G1 are also shown, as is transiently transfected wt-mRad21-Halo expressed using the low-expression promoter, L30. Even though mRad21-Halo is only weakly overexpressed, most molecules show free 3D diffusion. The overexpression artifact may be caused by the fact that without similar overexpression of Smc1, Smc3 and SA1/2, most Rad21 cannot form cohesin complexes. Finally, a Rad21-mutant (F601R, L605R, Q617K), which cannot form cohesin complexes was used to estimate the non-specifically bound Rad21 fractions. ( F ) Stroboscopic paSMT experiments in human U2OS cells. H2B-Halo and Halo-3xNLS were used as controls for mostly bound and free molecules and the same zinc-finger mutants as in mESCs were transiently transfected as control for non-specific chromatin association. Note that C32 Halo-hCTCF shows slightly more free diffusion than C59 and C87 in mESCs. DOI: http://dx.doi.org/10.7554/eLife.25776.017

    Techniques Used: Labeling, Activation Assay, Mass Spectrometry, Microscopy, Diffusion-based Assay, Binding Assay, Construct, Transfection, Mutagenesis, Zinc-Fingers, Expressing, Over Expression

    Overview of super-resolution PALM approach and control experiments. ( A ) Representative super-resolution PALM reconstruction of Halo-mCTCF in C59 mouse embryonic stem cells. Left: full nucleus. Right: zoom-in on a 3 μm square before (top) and after (bottom) cluster assignment using a Bayesian clustering algorithm. ( B ) Representative super-resolution PALM reconstruction of mRad21-Halo in C45 mouse embryonic stem cells. Left: full nucleus. Right: zoom-in on a 3 μm square before (top) and after (bottom) cluster assignment using a Bayesian clustering algorithm. ( C ) Bar graphs showing fraction of molecules in clusters for different mES cell lines as inferred from Bayesian cluster assignments. Bar graphs show mean per 3 μm square and the error bars show the standard error. ( D ) Bar graphs showing cluster radii in clusters for different mES cell lines as inferred from Bayesian cluster assignments. Bar graphs show mean per 3 μm square and the error bars show the standard error. ( E ) Representative control for photo-blinking in apparent PALM clustering. U2OS C32 Halo-hCTCF was labeled with ~50:50 PA-JF549:PA-JF646 and imaged using two-color PALM. Clusters were assigned as in ( C–D ) and the fraction of CTCF molecules in each cluster labeled with JF549 and JF646 plotted. If PALM clustering was solely a photo-blinking artifact, clusters should be exclusively composed of either JF549 or JF646, whereas under ideal conditions the distribution should follow a binomial distribution. The observed distributions resemble the expected binomial distributions suggesting that most called clusters are not a photo-blinking artifact (Kullback-Leibler divergence ~0.3 bits). DOI: http://dx.doi.org/10.7554/eLife.25776.021
    Figure Legend Snippet: Overview of super-resolution PALM approach and control experiments. ( A ) Representative super-resolution PALM reconstruction of Halo-mCTCF in C59 mouse embryonic stem cells. Left: full nucleus. Right: zoom-in on a 3 μm square before (top) and after (bottom) cluster assignment using a Bayesian clustering algorithm. ( B ) Representative super-resolution PALM reconstruction of mRad21-Halo in C45 mouse embryonic stem cells. Left: full nucleus. Right: zoom-in on a 3 μm square before (top) and after (bottom) cluster assignment using a Bayesian clustering algorithm. ( C ) Bar graphs showing fraction of molecules in clusters for different mES cell lines as inferred from Bayesian cluster assignments. Bar graphs show mean per 3 μm square and the error bars show the standard error. ( D ) Bar graphs showing cluster radii in clusters for different mES cell lines as inferred from Bayesian cluster assignments. Bar graphs show mean per 3 μm square and the error bars show the standard error. ( E ) Representative control for photo-blinking in apparent PALM clustering. U2OS C32 Halo-hCTCF was labeled with ~50:50 PA-JF549:PA-JF646 and imaged using two-color PALM. Clusters were assigned as in ( C–D ) and the fraction of CTCF molecules in each cluster labeled with JF549 and JF646 plotted. If PALM clustering was solely a photo-blinking artifact, clusters should be exclusively composed of either JF549 or JF646, whereas under ideal conditions the distribution should follow a binomial distribution. The observed distributions resemble the expected binomial distributions suggesting that most called clusters are not a photo-blinking artifact (Kullback-Leibler divergence ~0.3 bits). DOI: http://dx.doi.org/10.7554/eLife.25776.021

    Techniques Used: Labeling

    8) Product Images from "Involvement of impaired autophagy and mitophagy in Neuro-2a cell damage under hypoxic and/or high-glucose conditions"

    Article Title: Involvement of impaired autophagy and mitophagy in Neuro-2a cell damage under hypoxic and/or high-glucose conditions

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-20162-1

    Autophagy impairment and autophagy induction under chronic hypoxia. ( a ) Neuro-2a cells were exposed to hypoxia and/or high glucose for 48 h, and the protein levels of P-AMPK, AMPK, P-mTOR and mTOR were measured by immunoblot. ( b ) The ImageJ densitometric analysis showed that chronic hypoxia upregulated the ratio of P-AMPK/AMPK and lowered the ratio of P-mTOR/ mTOR when compared with those under normoxia. High glucose decreased the ratio of P-AMPK/AMPK. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, # P
    Figure Legend Snippet: Autophagy impairment and autophagy induction under chronic hypoxia. ( a ) Neuro-2a cells were exposed to hypoxia and/or high glucose for 48 h, and the protein levels of P-AMPK, AMPK, P-mTOR and mTOR were measured by immunoblot. ( b ) The ImageJ densitometric analysis showed that chronic hypoxia upregulated the ratio of P-AMPK/AMPK and lowered the ratio of P-mTOR/ mTOR when compared with those under normoxia. High glucose decreased the ratio of P-AMPK/AMPK. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, # P

    Techniques Used:

    Accumulation of AVs under hypoxic stress. ( a ) Neuro-2a cells were submitted to hypoxia and/or high glucose for 48 h, and the protein levels of P62 and LC3B-II were measured by immunoblots, showing a time-dependent increase in levels of LC3-II and P62 under hypoxia compared with those under normoxia. In contrast, high glucose had no significant effect on their expression. ( b ) The ImageJ densitometric analysis of P62 and LC3B-II from the immunoblots is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: Accumulation of AVs under hypoxic stress. ( a ) Neuro-2a cells were submitted to hypoxia and/or high glucose for 48 h, and the protein levels of P62 and LC3B-II were measured by immunoblots, showing a time-dependent increase in levels of LC3-II and P62 under hypoxia compared with those under normoxia. In contrast, high glucose had no significant effect on their expression. ( b ) The ImageJ densitometric analysis of P62 and LC3B-II from the immunoblots is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: Western Blot, Expressing

    The accumulation of damaged mitochondria under hypoxia was independent of autophagy impairment. ( a ) Neuro-2a cells were submitted to hypoxia and/or high glucose for up to 42 h and then subjected to 10 nM BAF for an additional 6 h. Immunoblots were performed to determine the levels of BNIP3 and TOMM20. ( b ) The ImageJ densitometric analysis from immunoblots is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: The accumulation of damaged mitochondria under hypoxia was independent of autophagy impairment. ( a ) Neuro-2a cells were submitted to hypoxia and/or high glucose for up to 42 h and then subjected to 10 nM BAF for an additional 6 h. Immunoblots were performed to determine the levels of BNIP3 and TOMM20. ( b ) The ImageJ densitometric analysis from immunoblots is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: Western Blot

    The HIF-1α-BNIP3 pathway was activated during hypoxia. ( a ) Analysis of mitophagy-related protein expression in Neuro-2a cells by immunoblotting. Neuro-2a cells were exposed to hypoxia and/or high glucose for 48 h. Total cellular extracts were analysed by immunoblotting with antibodies against HIF-1α, BNIP3, PINK1 and TOMM20. ( b ) The ImageJ densitometric analysis is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: The HIF-1α-BNIP3 pathway was activated during hypoxia. ( a ) Analysis of mitophagy-related protein expression in Neuro-2a cells by immunoblotting. Neuro-2a cells were exposed to hypoxia and/or high glucose for 48 h. Total cellular extracts were analysed by immunoblotting with antibodies against HIF-1α, BNIP3, PINK1 and TOMM20. ( b ) The ImageJ densitometric analysis is shown. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: Expressing

    BNIP3-mediated mitophagy was insufficient during hypoxia. Neuro-2a cells were incubated for 48 h in each experiment, then fixed and immunostained with the LC3B, BNIP3, and COXIV antibodies. Cells were counterstained with DAPI to visualize nuclei. ( a ) Immunodetection of endogenous BNIP3 (green) colocalized with the mitochondrial marker COXIV (red), where it appeared as yellow. Scale bar: 50 μm. ( b ) Less colocalization of LC3B (green) and COXIV (red) was detected by fluorescence microscope. Scale bar: 50 μm. ( c ) Less colocalization of LC3B (green) and BNIP3 (red) was detected by fluorescence microscope. Scale bar: 50 μm.
    Figure Legend Snippet: BNIP3-mediated mitophagy was insufficient during hypoxia. Neuro-2a cells were incubated for 48 h in each experiment, then fixed and immunostained with the LC3B, BNIP3, and COXIV antibodies. Cells were counterstained with DAPI to visualize nuclei. ( a ) Immunodetection of endogenous BNIP3 (green) colocalized with the mitochondrial marker COXIV (red), where it appeared as yellow. Scale bar: 50 μm. ( b ) Less colocalization of LC3B (green) and COXIV (red) was detected by fluorescence microscope. Scale bar: 50 μm. ( c ) Less colocalization of LC3B (green) and BNIP3 (red) was detected by fluorescence microscope. Scale bar: 50 μm.

    Techniques Used: Incubation, Immunodetection, Marker, Fluorescence, Microscopy

    Inhibition of autophagy degradation induced cell death. ( a ) An MTS assay was used to access Neuro-2a cell proliferation under hypoxia and/or normoxia in the presence or absence of BAF. Treatment with BAF induced a decreased OD value under normoxia but did not further change the OD value under hypoxia. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). ( b ) Immunoblots were performed to determine the level of the cleaved caspase-3 protein in Neuro-2a cells. BAF treatment induced an increase in the level of cleaved caspase-3 protein under normoxia but did not further change the expression of cleaved caspase-3 under hypoxia. The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: Inhibition of autophagy degradation induced cell death. ( a ) An MTS assay was used to access Neuro-2a cell proliferation under hypoxia and/or normoxia in the presence or absence of BAF. Treatment with BAF induced a decreased OD value under normoxia but did not further change the OD value under hypoxia. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). ( b ) Immunoblots were performed to determine the level of the cleaved caspase-3 protein in Neuro-2a cells. BAF treatment induced an increase in the level of cleaved caspase-3 protein under normoxia but did not further change the expression of cleaved caspase-3 under hypoxia. The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: Inhibition, MTS Assay, Western Blot, Expressing

    Effect of chronic hypoxia and/or high glucose on Neuro-2a cell viability. ( a ) Measurement of Neuro-2a cell proliferation. Neuro-2a cells were exposed to hypoxia and/or high glucose for up to 48 h. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). Cell proliferation was determined by the MTS assay. ( b , c ) Western blot analysis of cleaved caspase-3 expression in Neuro-2a cells. The ImageJ densitometric analysis of the cleaved caspase-3 from immunoblots is shown. The results were expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: Effect of chronic hypoxia and/or high glucose on Neuro-2a cell viability. ( a ) Measurement of Neuro-2a cell proliferation. Neuro-2a cells were exposed to hypoxia and/or high glucose for up to 48 h. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). Cell proliferation was determined by the MTS assay. ( b , c ) Western blot analysis of cleaved caspase-3 expression in Neuro-2a cells. The ImageJ densitometric analysis of the cleaved caspase-3 from immunoblots is shown. The results were expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: MTS Assay, Western Blot, Expressing

    Chronic hypoxia triggered a loss of mitochondrial potential and destruction of the mitochondrial structure. ( a ) Neuro-2a cells were subjected to hypoxia and/or high glucose for up to 48 h. The mitochondria potential was measured by JC1 staining. Quantitative analysis by flow cytometry revealed that chronic hypoxia lowered the ratio of the red and green fluorescence, while high glucose did not change it with or without hypoxia. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p
    Figure Legend Snippet: Chronic hypoxia triggered a loss of mitochondrial potential and destruction of the mitochondrial structure. ( a ) Neuro-2a cells were subjected to hypoxia and/or high glucose for up to 48 h. The mitochondria potential was measured by JC1 staining. Quantitative analysis by flow cytometry revealed that chronic hypoxia lowered the ratio of the red and green fluorescence, while high glucose did not change it with or without hypoxia. “C” represented control conditions (25 mM glucose), “G” represented high glucose conditions (75 mM glucose). The results are expressed as the mean ± SEM from three independent experiments. Two-way ANOVA, ** p

    Techniques Used: Staining, Flow Cytometry, Cytometry, Fluorescence

    9) Product Images from "Anti-inflammatory activity of a sulfated polysaccharide isolated from an enzymatic digest of brown seaweed Sargassum horneri in RAW 264.7 cells"

    Article Title: Anti-inflammatory activity of a sulfated polysaccharide isolated from an enzymatic digest of brown seaweed Sargassum horneri in RAW 264.7 cells

    Journal: Nutrition Research and Practice

    doi: 10.4162/nrp.2017.11.1.3

    Effects of crude polysaccharides from the Celluclast enzyme digest (CCP) on LPS-induced iNOS and COX-2 protein expression in RAW 264.7 cells. Cells (1 × 10 5  cells/mL) were treated with indicated concentrations of CCP (25, 50, or 100 µg/mL) for 1 h before LPS (1 µg/mL) treatment for 24 h. Cell lysates (40 µg) were resolved by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against iNOS and COX-2. (A) The proteins were then visualized by ECL. (B) The intensity of the bands was measured by ImageJ software. Relative amounts of iNOS and COX-2 compared to β-actin and the density ratio represented the relative intensity of each band against that of the β-actin. The gel shown is a representative of the results from three separate experiments. The inhibitory effect of Effects of the crude polysaccharides from the CCP on the production of pro-inflammatory cytokines in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells (1 × 10 5 ) were stimulated with LPS (1 µg/mL) for 24 h with or without CCP. Supernatants were collected, and TNF-α (C) and IL-1β (D) levels in the culture supernatant were determined by ELISA according to the manufacturer's instructions. Data points and bars represent the arithmetic means ± SD (n = 3). Each data point represents the mean ± SEM ( * P
    Figure Legend Snippet: Effects of crude polysaccharides from the Celluclast enzyme digest (CCP) on LPS-induced iNOS and COX-2 protein expression in RAW 264.7 cells. Cells (1 × 10 5 cells/mL) were treated with indicated concentrations of CCP (25, 50, or 100 µg/mL) for 1 h before LPS (1 µg/mL) treatment for 24 h. Cell lysates (40 µg) were resolved by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against iNOS and COX-2. (A) The proteins were then visualized by ECL. (B) The intensity of the bands was measured by ImageJ software. Relative amounts of iNOS and COX-2 compared to β-actin and the density ratio represented the relative intensity of each band against that of the β-actin. The gel shown is a representative of the results from three separate experiments. The inhibitory effect of Effects of the crude polysaccharides from the CCP on the production of pro-inflammatory cytokines in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells (1 × 10 5 ) were stimulated with LPS (1 µg/mL) for 24 h with or without CCP. Supernatants were collected, and TNF-α (C) and IL-1β (D) levels in the culture supernatant were determined by ELISA according to the manufacturer's instructions. Data points and bars represent the arithmetic means ± SD (n = 3). Each data point represents the mean ± SEM ( * P

    Techniques Used: Expressing, SDS Page, Software, Enzyme-linked Immunosorbent Assay

    (A) Cytotoxicity of crude polysaccharides (CP) on RAW 264.7 cells in the presence of LPS. The viability of cells without sample and LPS has been taken as reference (100%). (B) Dose-dependent inhibition of NO production by CPs in LPS-stimulated RAW264.7 macrophages. The level of NO production is expressed as percentages of that of the group treated with LPS alone. (C) The inhibitory effects of the crude polysaccharides from the Celluclast enzyme digest (CCP) on the production of PGE 2  in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells (1 × 10 5 ) were stimulated with LPS (1 µg/mL) for 24-h with or without CCP. Supernatants were collected and levels of PGE 2  in the culture supernatant were determined by ELISA according to the manufacturer's instructions. Data points and bars represent the arithmetic means ± SD (n = 3). Each data point represents the mean ± SD ( * P
    Figure Legend Snippet: (A) Cytotoxicity of crude polysaccharides (CP) on RAW 264.7 cells in the presence of LPS. The viability of cells without sample and LPS has been taken as reference (100%). (B) Dose-dependent inhibition of NO production by CPs in LPS-stimulated RAW264.7 macrophages. The level of NO production is expressed as percentages of that of the group treated with LPS alone. (C) The inhibitory effects of the crude polysaccharides from the Celluclast enzyme digest (CCP) on the production of PGE 2 in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells (1 × 10 5 ) were stimulated with LPS (1 µg/mL) for 24-h with or without CCP. Supernatants were collected and levels of PGE 2 in the culture supernatant were determined by ELISA according to the manufacturer's instructions. Data points and bars represent the arithmetic means ± SD (n = 3). Each data point represents the mean ± SD ( * P

    Techniques Used: Inhibition, Enzyme-linked Immunosorbent Assay

    Inhibitory effects of the crude polysaccharides from the Celluclast enzyme digest (CCP) on NF-κB and MAPK activation in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells were treated with LPS (1 µg/mL) or CCP (100 µg/mL) for 10 or 20 min. (A) and (B) Cytosolic cell lysates, (C) and (D) nuclear protein extracts (40 µg) were resolved by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against NF-κB p50 and p65. The proteins were then visualized by ECL. (E) and (F) Effects of CCP on MAPK activation induced by LPS in RAW 264.7 cells. RAW 264.7 cells were treated with LPS (1 µg/mL) or CCP (100 µg/mL) for 10 or 20 min. Total protein (40 µg) was separated by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against ERK and p38. The proteins were then visualized by ECL. The intensity of the bands was measured by ImageJ software. Relative amounts of density ratio represented the relative intensity of each band against that of the standard protein. The results shown are representative of those obtained from three independent experiments. Each data point represents the mean ± SEM ( * P
    Figure Legend Snippet: Inhibitory effects of the crude polysaccharides from the Celluclast enzyme digest (CCP) on NF-κB and MAPK activation in LPS-stimulated RAW 264.7 cells. RAW 264.7 cells were treated with LPS (1 µg/mL) or CCP (100 µg/mL) for 10 or 20 min. (A) and (B) Cytosolic cell lysates, (C) and (D) nuclear protein extracts (40 µg) were resolved by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against NF-κB p50 and p65. The proteins were then visualized by ECL. (E) and (F) Effects of CCP on MAPK activation induced by LPS in RAW 264.7 cells. RAW 264.7 cells were treated with LPS (1 µg/mL) or CCP (100 µg/mL) for 10 or 20 min. Total protein (40 µg) was separated by 10% SDS-PAGE, transferred to nitrocellulose membranes, and probed with antibodies against ERK and p38. The proteins were then visualized by ECL. The intensity of the bands was measured by ImageJ software. Relative amounts of density ratio represented the relative intensity of each band against that of the standard protein. The results shown are representative of those obtained from three independent experiments. Each data point represents the mean ± SEM ( * P

    Techniques Used: Activation Assay, SDS Page, Software

    10) Product Images from "CRISPR/Cas9 gene editing for the creation of an MGAT1-deficient CHO cell line to control HIV-1 vaccine glycosylation"

    Article Title: CRISPR/Cas9 gene editing for the creation of an MGAT1-deficient CHO cell line to control HIV-1 vaccine glycosylation

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.2005817

    Flow chart of MGAT1 gene editing and cell line selection strategy. (A) A plasmid containing the Cas9 nuclease, tracrRNA, and a gRNA sequence was electroporated into suspension adapted CHO-S cells. (B) Twenty-four hours following transfection, the cells were distributed into 96-well tissue culture plates at a density of 0.5 cells/well. (C) Between 12 and 15 days later, wells with 20% or greater confluency were transferred to 24-well plates. (D) After 5 days of growth in 24-well plates, a 0.2-mL aliquot was removed from each well, and cells were tested for the ability to bind fluorescein-labeled GNA. (E) GNA-binding cells were then expanded to shake flasks, and cell lines were transiently transfected with a gene encoding A244-rgp120. The cell culture supernatants were then collected after 5 days and tested for binding of gp120 to the prototypic glycan-dependent, broadly neutralizing monoclonal antibody PG9. This representative plot (F) is shown for demonstrative process purposes only. A detailed plot of this data is show in Fig 4A . (G) The gene encoding MGAT1 was sequenced from GNA-binding cell lines that exhibited robust growth and the ability to secrete PG9-binding gp120. The specific mutations induced by NHEJR were determined by Sanger sequencing. Cas9, CRISPR-associated protein 9; CHO, Chinese hamster ovary; FIA, fluorescence immunoassay; GNA, Galanthus nivalis lectin; gRNA, guide RNA; MGAT1, Mannosyl (Alpha-1,3-)-Glycoprotein Beta-1,2-N-Acetylglucosaminyltransferase; NHEJR, nonhomologous end joining repair; rgp120, recombinant gp120; tracrRNA, trans -activating CRISPR RNA.
    Figure Legend Snippet: Flow chart of MGAT1 gene editing and cell line selection strategy. (A) A plasmid containing the Cas9 nuclease, tracrRNA, and a gRNA sequence was electroporated into suspension adapted CHO-S cells. (B) Twenty-four hours following transfection, the cells were distributed into 96-well tissue culture plates at a density of 0.5 cells/well. (C) Between 12 and 15 days later, wells with 20% or greater confluency were transferred to 24-well plates. (D) After 5 days of growth in 24-well plates, a 0.2-mL aliquot was removed from each well, and cells were tested for the ability to bind fluorescein-labeled GNA. (E) GNA-binding cells were then expanded to shake flasks, and cell lines were transiently transfected with a gene encoding A244-rgp120. The cell culture supernatants were then collected after 5 days and tested for binding of gp120 to the prototypic glycan-dependent, broadly neutralizing monoclonal antibody PG9. This representative plot (F) is shown for demonstrative process purposes only. A detailed plot of this data is show in Fig 4A . (G) The gene encoding MGAT1 was sequenced from GNA-binding cell lines that exhibited robust growth and the ability to secrete PG9-binding gp120. The specific mutations induced by NHEJR were determined by Sanger sequencing. Cas9, CRISPR-associated protein 9; CHO, Chinese hamster ovary; FIA, fluorescence immunoassay; GNA, Galanthus nivalis lectin; gRNA, guide RNA; MGAT1, Mannosyl (Alpha-1,3-)-Glycoprotein Beta-1,2-N-Acetylglucosaminyltransferase; NHEJR, nonhomologous end joining repair; rgp120, recombinant gp120; tracrRNA, trans -activating CRISPR RNA.

    Techniques Used: Flow Cytometry, Selection, Plasmid Preparation, Sequencing, Transfection, Labeling, Binding Assay, Cell Culture, CRISPR, Fluorescence, Recombinant

    11) Product Images from "Absorptive interactions of concurrent oral administration of (+)-catechin and puerarin in rats and the underlying mechanisms"

    Article Title: Absorptive interactions of concurrent oral administration of (+)-catechin and puerarin in rats and the underlying mechanisms

    Journal: Acta Pharmacologica Sinica

    doi: 10.1038/aps.2015.164

    Cell transport of (+)-C. (A) The transport amount of (+)-C (400 μmol/L) across Caco-2 cell monolayers over time. (B) Concentration-dependent transport rate of (+)-C across Caco-2 cell monolayers.
    Figure Legend Snippet: Cell transport of (+)-C. (A) The transport amount of (+)-C (400 μmol/L) across Caco-2 cell monolayers over time. (B) Concentration-dependent transport rate of (+)-C across Caco-2 cell monolayers.

    Techniques Used: Concentration Assay

    (+)-C and Pue transport across Caco-2 cell monolayers
    Figure Legend Snippet: (+)-C and Pue transport across Caco-2 cell monolayers

    Techniques Used:

    (+)-C and Pue transport across Caco-2 cell monolayers
    Figure Legend Snippet: (+)-C and Pue transport across Caco-2 cell monolayers

    Techniques Used:

    12) Product Images from "Ultrasmall dopamine-coated nanogolds: preparation, characteristics, and CT imaging"

    Article Title: Ultrasmall dopamine-coated nanogolds: preparation, characteristics, and CT imaging

    Journal: Journal of Experimental Nanoscience

    doi: 10.1080/17458080.2015.1102343

    The cell viability results of (a) HeLa, (b) PC-3, and (c) IMR-90 cell lines when exposed to WDU AuNPs and WDU AuNPs@DPAs at different concentrations by MTT assay.
    Figure Legend Snippet: The cell viability results of (a) HeLa, (b) PC-3, and (c) IMR-90 cell lines when exposed to WDU AuNPs and WDU AuNPs@DPAs at different concentrations by MTT assay.

    Techniques Used: MTT Assay

    13) Product Images from "Lipid-like Nanoparticles for Small Interfering RNA Delivery to Endothelial Cells"

    Article Title: Lipid-like Nanoparticles for Small Interfering RNA Delivery to Endothelial Cells

    Journal: Advanced functional materials

    doi: 10.1002/adfm.200900519

    Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and serum-deprived (endothelial basal medium-2 (EBM-2) with no serum and growth factors) condition. (A) RT-PCR for SHP-1, KDR/Flk-1, and eNOS of siRNA-transfected HUVECs at
    Figure Legend Snippet: Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and serum-deprived (endothelial basal medium-2 (EBM-2) with no serum and growth factors) condition. (A) RT-PCR for SHP-1, KDR/Flk-1, and eNOS of siRNA-transfected HUVECs at

    Techniques Used: Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction

    14) Product Images from "Anti-inflammatory and anti-genotoxic activity of branched chain amino acids (BCAA) in lipopolysaccharide (LPS) stimulated RAW 264.7 macrophages"

    Article Title: Anti-inflammatory and anti-genotoxic activity of branched chain amino acids (BCAA) in lipopolysaccharide (LPS) stimulated RAW 264.7 macrophages

    Journal: Food Science and Biotechnology

    doi: 10.1007/s10068-017-0165-4

    Effect of BCAAs on iNOS mRNA expression in LPS-stimulated RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments.  a–d Different letters  among samples indicate significant differences by Duncan’s multiple range test ( p
    Figure Legend Snippet: Effect of BCAAs on iNOS mRNA expression in LPS-stimulated RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments. a–d Different letters among samples indicate significant differences by Duncan’s multiple range test ( p

    Techniques Used: Expressing, Standard Deviation

    Antigenotoxic effect of 400 μM BCAAs on 200 μM H 2 O 2  induced DNA damage in RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine.  NC  negative control; PC: 200 μM H 2 O 2  treated positive control. The data are expressed as the mean ± standard deviation (SD) of three separate experiments.  a–c Different letters  among samples indicate significant differences by Duncan’s multiple range test ( p
    Figure Legend Snippet: Antigenotoxic effect of 400 μM BCAAs on 200 μM H 2 O 2 induced DNA damage in RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. NC negative control; PC: 200 μM H 2 O 2 treated positive control. The data are expressed as the mean ± standard deviation (SD) of three separate experiments. a–c Different letters among samples indicate significant differences by Duncan’s multiple range test ( p

    Techniques Used: Negative Control, Positive Control, Standard Deviation

    Effect of BCAAs on expression of IL-6 ( A ) and COX-2 ( B ) mRNA in LPS-induced RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments.  a,b Different letters  among samples indicate significant differences by Duncan’s multiple range test ( p
    Figure Legend Snippet: Effect of BCAAs on expression of IL-6 ( A ) and COX-2 ( B ) mRNA in LPS-induced RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments. a,b Different letters among samples indicate significant differences by Duncan’s multiple range test ( p

    Techniques Used: Expressing, Standard Deviation

    Effect of BCAAs on cell viability ( A ) and NO production ( B ) in LPS-induced RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments.  a–i Different letters  among samples indicate significant differences by Duncan’s multiple range test ( p
    Figure Legend Snippet: Effect of BCAAs on cell viability ( A ) and NO production ( B ) in LPS-induced RAW 264.7 macrophages. Val, valine; Ile, isoleucine; Lue, leucine. The data are expressed as the mean ± standard deviation (SD) of three separate experiments. a–i Different letters among samples indicate significant differences by Duncan’s multiple range test ( p

    Techniques Used: Standard Deviation

    Comet images of RAW 264.7 macrophages treated with BCAAs.  A  Negative control;  B  200 μM H 2 O 2  treated positive control;  C  Valine 400 μM + 200 μM H 2 O 2 ;  D  Isoleucine 400 μM + 200 μM H 2 O 2 ;  E  Leucine 400 μM + 200 μM H 2 O 2
    Figure Legend Snippet: Comet images of RAW 264.7 macrophages treated with BCAAs. A Negative control; B 200 μM H 2 O 2 treated positive control; C Valine 400 μM + 200 μM H 2 O 2 ; D Isoleucine 400 μM + 200 μM H 2 O 2 ; E Leucine 400 μM + 200 μM H 2 O 2

    Techniques Used: Negative Control, Positive Control

    15) Product Images from "Iron and Iron Regulatory Proteins in Amoeboid Microglial Cells Are Linked to Oligodendrocyte Death in Hypoxic Neonatal Rat Periventricular White Matter through Production of Proinflammatory Cytokines and Reactive Oxygen/Nitrogen Species"

    Article Title: Iron and Iron Regulatory Proteins in Amoeboid Microglial Cells Are Linked to Oligodendrocyte Death in Hypoxic Neonatal Rat Periventricular White Matter through Production of Proinflammatory Cytokines and Reactive Oxygen/Nitrogen Species

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.2250-11.2011

    A , Bar graph in A shows significant changes in the total iron content in microglial culture. Note the reduction of iron in hypoxic microglial cells when treated with deferoxamine (H+D). Significant differences in total iron level between control (C), hypoxic (H), and deferoxamine treated groups are expressed as follows: * p
    Figure Legend Snippet: A , Bar graph in A shows significant changes in the total iron content in microglial culture. Note the reduction of iron in hypoxic microglial cells when treated with deferoxamine (H+D). Significant differences in total iron level between control (C), hypoxic (H), and deferoxamine treated groups are expressed as follows: * p

    Techniques Used:

    A , Bar graph represents the glutathione content in primary oligodendrocytes in control group (C), oligodendrocytes treated with conditioned medium from control microglia (CCM), oligodendrocytes treated with deferoxamine (DCM), hypoxic oligodendrocytes (H), oligodendrocytes treated with conditioned medium from hypoxic microglia (HCM), and with conditioned medium from deferoxamine treated hypoxic microglial cells (DHCM). B , Bar graph represents the MDA concentrations in oligodendrocyte cultures in all six groups. Significant differences between the groups are expressed as follows: * p
    Figure Legend Snippet: A , Bar graph represents the glutathione content in primary oligodendrocytes in control group (C), oligodendrocytes treated with conditioned medium from control microglia (CCM), oligodendrocytes treated with deferoxamine (DCM), hypoxic oligodendrocytes (H), oligodendrocytes treated with conditioned medium from hypoxic microglia (HCM), and with conditioned medium from deferoxamine treated hypoxic microglial cells (DHCM). B , Bar graph represents the MDA concentrations in oligodendrocyte cultures in all six groups. Significant differences between the groups are expressed as follows: * p

    Techniques Used: Multiple Displacement Amplification

    TUNEL assay of oligodendrocytes showing a significant increase in apoptosis following hypoxia and on treatment with conditioned medium from microglial cells subjected to hypoxia. A shows the confocal images of DAPI-stained nucleus in all of the groups, B shows the confocal images of cells that are TUNEL positive in all of the groups, and C shows the colocalization of DAPI- and TUNEL-positive cells. Bar graph in D shows the percentage of cells that are TUNEL positive in each group (C, control; CCM, treated with conditioned medium control microglia; DCM, treated with deferoxamine; H, hypoxia; HCM, treated with conditioned medium from hypoxic microglia; DHCM, treated with conditioned medium from hypoxic microglial cells plus deferoxamine). Scale bars, 50 μm. Significant differences between the various groups are expressed as follows: * p
    Figure Legend Snippet: TUNEL assay of oligodendrocytes showing a significant increase in apoptosis following hypoxia and on treatment with conditioned medium from microglial cells subjected to hypoxia. A shows the confocal images of DAPI-stained nucleus in all of the groups, B shows the confocal images of cells that are TUNEL positive in all of the groups, and C shows the colocalization of DAPI- and TUNEL-positive cells. Bar graph in D shows the percentage of cells that are TUNEL positive in each group (C, control; CCM, treated with conditioned medium control microglia; DCM, treated with deferoxamine; H, hypoxia; HCM, treated with conditioned medium from hypoxic microglia; DHCM, treated with conditioned medium from hypoxic microglial cells plus deferoxamine). Scale bars, 50 μm. Significant differences between the various groups are expressed as follows: * p

    Techniques Used: TUNEL Assay, Staining

    16) Product Images from "Innovative biodegradable poly(L-lactide)/collagen/hydroxyapatite composite fibrous scaffolds promote osteoblastic proliferation and differentiation"

    Article Title: Innovative biodegradable poly(L-lactide)/collagen/hydroxyapatite composite fibrous scaffolds promote osteoblastic proliferation and differentiation

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S146679

    The effects of different electrospun scaffolds on the cell differentiation of MC3T3-E1 cells. Notes: ( A ) Quantification of the mineralized matrix of cells cultured on different nanofiber scaffolds; ( B ) the mineralized matrix nodules of cells cultured on different nanofiber scaffolds stained by ARS: (a) control without differentiation medium, (b) control with differentiation medium, (c) PLLA, (d) PLLA/HA, (e) PLLA/Coll, (f) PLLA/Coll/HA; ( C ) the expression of mRNA for OCN, Runx-2, and BMP-2 in cells cultured on different nanofiber scaffolds. Data are expressed as mean ± standard deviation from three independent experiments (n=6). ** P
    Figure Legend Snippet: The effects of different electrospun scaffolds on the cell differentiation of MC3T3-E1 cells. Notes: ( A ) Quantification of the mineralized matrix of cells cultured on different nanofiber scaffolds; ( B ) the mineralized matrix nodules of cells cultured on different nanofiber scaffolds stained by ARS: (a) control without differentiation medium, (b) control with differentiation medium, (c) PLLA, (d) PLLA/HA, (e) PLLA/Coll, (f) PLLA/Coll/HA; ( C ) the expression of mRNA for OCN, Runx-2, and BMP-2 in cells cultured on different nanofiber scaffolds. Data are expressed as mean ± standard deviation from three independent experiments (n=6). ** P

    Techniques Used: Cell Differentiation, Cell Culture, Staining, Expressing, Standard Deviation

    The effects of different electrospun scaffolds on the viability and ALP activity of MC3T3-E1 cells. Notes: ( A ) The viability of cells cultured on different electrospun scaffolds for 1, 3, and 7 days; ( B ) fluorescence microscopy of cells cultured on different electrospun scaffolds for 1 and 7 days; ( C ) ALP activity of cells cultured on different electrospun scaffolds. The blue dots indicate the DNA of live cells stained with DAPI. The green color indicates the cytoskeleton of cells stained with actin green. Data are expressed as mean ± standard deviation from three independent experiments (n=6). ** P
    Figure Legend Snippet: The effects of different electrospun scaffolds on the viability and ALP activity of MC3T3-E1 cells. Notes: ( A ) The viability of cells cultured on different electrospun scaffolds for 1, 3, and 7 days; ( B ) fluorescence microscopy of cells cultured on different electrospun scaffolds for 1 and 7 days; ( C ) ALP activity of cells cultured on different electrospun scaffolds. The blue dots indicate the DNA of live cells stained with DAPI. The green color indicates the cytoskeleton of cells stained with actin green. Data are expressed as mean ± standard deviation from three independent experiments (n=6). ** P

    Techniques Used: ALP Assay, Activity Assay, Cell Culture, Fluorescence, Microscopy, Staining, Standard Deviation

    17) Product Images from "Voltage-gated ion channels mediate the electrotaxis of glioblastoma cells in a hybrid PMMA/PDMS microdevice"

    Article Title: Voltage-gated ion channels mediate the electrotaxis of glioblastoma cells in a hybrid PMMA/PDMS microdevice

    Journal: bioRxiv

    doi: 10.1101/2020.02.14.948638

    The electrotactic directedness of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; ‡ indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; * indicates P
    Figure Legend Snippet: The electrotactic directedness of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; ‡ indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; * indicates P

    Techniques Used: Inhibition, Adsorption, Cell Culture

    Phase contrast images of T98G and U-251MG cells before and after 6 hours 300 V m −1 stimulation. The electric field is from left to right. Only T98G cells demonstrate prominent perpendicular allignment after electrical stimulation.
    Figure Legend Snippet: Phase contrast images of T98G and U-251MG cells before and after 6 hours 300 V m −1 stimulation. The electric field is from left to right. Only T98G cells demonstrate prominent perpendicular allignment after electrical stimulation.

    Techniques Used:

    Time series plot of orientation in electrically stimulated glioblastoma cells. T98G cell results are indicated in closed symbols. U-251MG cell results are indicated in open symbols. Only T98G cells demonstrate prominent perpendicular alignment after electrical stimulation. The dashed lines indicate the 95% confidence interval for respective groups.
    Figure Legend Snippet: Time series plot of orientation in electrically stimulated glioblastoma cells. T98G cell results are indicated in closed symbols. U-251MG cell results are indicated in open symbols. Only T98G cells demonstrate prominent perpendicular alignment after electrical stimulation. The dashed lines indicate the 95% confidence interval for respective groups.

    Techniques Used:

    Phase contrast images of T98G U-251MG cells under EF stimulations in different microenvironments after 6 hours. The scale bars represent 100 μ m. 4-AP: 4-aminopyridine. T98G cells demonstrate perpendicular alignment after electric field stimulation in cell culture media with or without FBS. However, without calcium and FBS in the cell culture medium, T98G cells’ viability decreases. Furthermore, in cell culture medium supplemented with 4 mM 4-AP, T98G also lose viability and start to detach from the substrate. These trends were not observed in the U-251MG cells, suggesting heterogeneity between cell lines.
    Figure Legend Snippet: Phase contrast images of T98G U-251MG cells under EF stimulations in different microenvironments after 6 hours. The scale bars represent 100 μ m. 4-AP: 4-aminopyridine. T98G cells demonstrate perpendicular alignment after electric field stimulation in cell culture media with or without FBS. However, without calcium and FBS in the cell culture medium, T98G cells’ viability decreases. Furthermore, in cell culture medium supplemented with 4 mM 4-AP, T98G also lose viability and start to detach from the substrate. These trends were not observed in the U-251MG cells, suggesting heterogeneity between cell lines.

    Techniques Used: Cell Culture

    The electrotaxis of T98G U-251MG glioblastoma cells in dependence of the extracellular serum and calcium by varying the medium recipe. (a) The electrotaxis directedness; (b) The electrotaxis speed; n.s. indicates not significant; **** indicate P
    Figure Legend Snippet: The electrotaxis of T98G U-251MG glioblastoma cells in dependence of the extracellular serum and calcium by varying the medium recipe. (a) The electrotaxis directedness; (b) The electrotaxis speed; n.s. indicates not significant; **** indicate P

    Techniques Used:

    Video clip showing the electrotaxis of U-251MG glioblastoma cells suppressed with 200 nM Agatoxin IVA on P/Q-type VGCC under 300 V m −1 dcEF for six hours and the respective tracking results using Usiigaci software.
    Figure Legend Snippet: Video clip showing the electrotaxis of U-251MG glioblastoma cells suppressed with 200 nM Agatoxin IVA on P/Q-type VGCC under 300 V m −1 dcEF for six hours and the respective tracking results using Usiigaci software.

    Techniques Used: Cross-linking Immunoprecipitation, Software

    Phase contrast images of T98G U-251MG cells on various ECMs. Scale bar: 100 μ m.
    Figure Legend Snippet: Phase contrast images of T98G U-251MG cells on various ECMs. Scale bar: 100 μ m.

    Techniques Used:

    The electrotaxis of T98G U-251MG glioblastoma cells on various ECMs after 6 hours under 300 V m −1 EF stimulation. (a) The electrotaxis directedness; (b) The electrotaxis speed; † indicates the electrotaxis groups tested against those without EF stimulation; All other groups are statistically compared to their respective controls on Geltrex™ ECM; n.s. indicates not significant; * indicates P
    Figure Legend Snippet: The electrotaxis of T98G U-251MG glioblastoma cells on various ECMs after 6 hours under 300 V m −1 EF stimulation. (a) The electrotaxis directedness; (b) The electrotaxis speed; † indicates the electrotaxis groups tested against those without EF stimulation; All other groups are statistically compared to their respective controls on Geltrex™ ECM; n.s. indicates not significant; * indicates P

    Techniques Used:

    The electrotaxis of T98G U-251MG glioblastoma cells in dependence of extracellular calcium by varying concentration of calcium chelators, EDTA and EGTA. (a) The electrotaxis directedness; (b) The electrotaxis speed; † indicate the electrotaxis groups tested against those without EF stimulation; All groups are statistically compared to controls in cell culture media with 10% FBS; n.s. indicates not significant; **** indicate P
    Figure Legend Snippet: The electrotaxis of T98G U-251MG glioblastoma cells in dependence of extracellular calcium by varying concentration of calcium chelators, EDTA and EGTA. (a) The electrotaxis directedness; (b) The electrotaxis speed; † indicate the electrotaxis groups tested against those without EF stimulation; All groups are statistically compared to controls in cell culture media with 10% FBS; n.s. indicates not significant; **** indicate P

    Techniques Used: Concentration Assay, Cell Culture

    Video clip showing the electrotaxis of U-251MG glioblastoma cells suppressed with 4-AP on A-type VGKC under 300 V m −1 dcEF for six hours and the respective tracking results using Usiigaci software.
    Figure Legend Snippet: Video clip showing the electrotaxis of U-251MG glioblastoma cells suppressed with 4-AP on A-type VGKC under 300 V m −1 dcEF for six hours and the respective tracking results using Usiigaci software.

    Techniques Used: Cross-linking Immunoprecipitation, Software

    The results of U-251MG cell seeding inside microchannels by various methods. (a, d, g, j) In tip loading method, cells are introduced by using gravitational flow with micropipette tips. The cells can flocculate inside the tips and in microchannels as illustrated in (d). The microscopy image of seeded cells is shown in (g) and magnified in (j); (b, e, h, k) In tip injection method, cells are injected into the channels and tips are removed. The small hydrostatic pressure differences between the inlet/outlet (shown as Δh) will contribute to hydrodynamic flow and disturb the cell distribution, causing non-uniform cell distribution and aggregates as shown in (e). The microscopy image of seeded cells is shown in (h) and magnified in (k); (c, f, i, l) In our pressure-balanced sub-merged cell seeding method, the hydrostatic pressure difference is eliminated. The injected cells remain uniform through-out the channel as shown in (f). The microscopy image of seeded cells is shown in (i) and magnified in (l). The uniform and sparse cell seeding method is suitable for many different applications from single cell tracking, ensembled cell studies to cell assembly. The scale bars in (g, h, i) represent 500 μ m. The scale bars in (j, k, l) represent 200 μ m.
    Figure Legend Snippet: The results of U-251MG cell seeding inside microchannels by various methods. (a, d, g, j) In tip loading method, cells are introduced by using gravitational flow with micropipette tips. The cells can flocculate inside the tips and in microchannels as illustrated in (d). The microscopy image of seeded cells is shown in (g) and magnified in (j); (b, e, h, k) In tip injection method, cells are injected into the channels and tips are removed. The small hydrostatic pressure differences between the inlet/outlet (shown as Δh) will contribute to hydrodynamic flow and disturb the cell distribution, causing non-uniform cell distribution and aggregates as shown in (e). The microscopy image of seeded cells is shown in (h) and magnified in (k); (c, f, i, l) In our pressure-balanced sub-merged cell seeding method, the hydrostatic pressure difference is eliminated. The injected cells remain uniform through-out the channel as shown in (f). The microscopy image of seeded cells is shown in (i) and magnified in (l). The uniform and sparse cell seeding method is suitable for many different applications from single cell tracking, ensembled cell studies to cell assembly. The scale bars in (g, h, i) represent 500 μ m. The scale bars in (j, k, l) represent 200 μ m.

    Techniques Used: Microscopy, Injection, Single Cell Tracking

    The electrotactic speed of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; † indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; ** indicate P
    Figure Legend Snippet: The electrotactic speed of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; † indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; ** indicate P

    Techniques Used: Inhibition, Adsorption, Cell Culture

    Video clip showing the random migration of U-251MG glioblastoma cells for six hours and the respective tracking results using Usiigaci software.
    Figure Legend Snippet: Video clip showing the random migration of U-251MG glioblastoma cells for six hours and the respective tracking results using Usiigaci software.

    Techniques Used: Cross-linking Immunoprecipitation, Migration, Software

    The electrotaxis of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on L-type (50 μ M Gadolinium [Gd] and Nicardipine) and N-type VGCCs (Conotoxin GVIA). a) The electrotactic directedness of the two cells. (b) The electrotactic speed of the two cells. ‡ indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; **** indicate P
    Figure Legend Snippet: The electrotaxis of T98G U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 hours with pharmacological inhibition on L-type (50 μ M Gadolinium [Gd] and Nicardipine) and N-type VGCCs (Conotoxin GVIA). a) The electrotactic directedness of the two cells. (b) The electrotactic speed of the two cells. ‡ indicate the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; **** indicate P

    Techniques Used: Inhibition, Adsorption, Cell Culture

    18) Product Images from "Best of Both Worlds: Simultaneous High-Light and Shade-Tolerance Adaptations within Individual Leaves of the Living Stone Lithops aucampiae"

    Article Title: Best of Both Worlds: Simultaneous High-Light and Shade-Tolerance Adaptations within Individual Leaves of the Living Stone Lithops aucampiae

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0075671

    Anatomy of Lithops aucampiae A i). Un-earthed plant, lateral view; ii) Plant face and iii) vertical section through leaf transect. Bar = 10 mm B longitudinal leaf transect showing epidermis under crossed-polarised light ( i, ii, iii ), UV ( iv, v, vi ) and white light ( vii, viii, ix ). Scale bar = 50 µm.
    Figure Legend Snippet: Anatomy of Lithops aucampiae A i). Un-earthed plant, lateral view; ii) Plant face and iii) vertical section through leaf transect. Bar = 10 mm B longitudinal leaf transect showing epidermis under crossed-polarised light ( i, ii, iii ), UV ( iv, v, vi ) and white light ( vii, viii, ix ). Scale bar = 50 µm.

    Techniques Used:

    19) Product Images from "MicroRNA-186-5p is expressed highly in ethanol-induced cardiomyocytes and regulates apoptosis via the target gene XIAP"

    Article Title: MicroRNA-186-5p is expressed highly in ethanol-induced cardiomyocytes and regulates apoptosis via the target gene XIAP

    Journal: Molecular Medicine Reports

    doi: 10.3892/mmr.2019.9953

    RT-qPCR and western blotting detect the expression levels of miR-186-5p or XIAP in cardiomyocytes following transfection with miR-186 mimic/inhibitor or XIAP plasmid. (A) RT-qPCR determined that the mRNA expression levels of miR-186-5p in AC16 cardiomyocytes increased following transfection with miR-186-5p mimic, while decreasing following transfection with miR-186-5p inhibitor. *P
    Figure Legend Snippet: RT-qPCR and western blotting detect the expression levels of miR-186-5p or XIAP in cardiomyocytes following transfection with miR-186 mimic/inhibitor or XIAP plasmid. (A) RT-qPCR determined that the mRNA expression levels of miR-186-5p in AC16 cardiomyocytes increased following transfection with miR-186-5p mimic, while decreasing following transfection with miR-186-5p inhibitor. *P

    Techniques Used: Quantitative RT-PCR, Western Blot, Expressing, Transfection, Plasmid Preparation

    Western blotting and reverse-transcription quantitative polymerase chain reaction to detect the expression levels of XIAP in ethanol-treated AC16 cardiomyocytes. (A) Protein expression levels of XIAP in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol. The 0 mmol/l treatment was the control group, and the 200 and 800 mmol/l were the experimental groups. The protein levels of XIAP differed between the experimental groups and the control group, decreasing in the experimental groups. (B) Protein expression levels of XIAP in 800 mmol/l ethanol-treated AC16 cardiomyocytes following different time interventions. The 0 h treatment was the control group, and the 24 and 48 h groups were the experimental ones. The protein expression levels of XIAP differed between the experimental groups and the control group, decreasing following ethanol treatment. (C) mRNA expression levels of XIAP in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol, and (D) mRNA expression levels of XIAP in 800 mmol/l ethanol-treated cardiomyocytes after different time interventions. The 0 mmol/l group was the control group, and 200 and 800 mmol/l were the experimental groups. The expression levels of XIAP differed between the experimental groups and the control group, decreasing in the experimental groups with increasing ethanol concentrations and longer treatment times. *P
    Figure Legend Snippet: Western blotting and reverse-transcription quantitative polymerase chain reaction to detect the expression levels of XIAP in ethanol-treated AC16 cardiomyocytes. (A) Protein expression levels of XIAP in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol. The 0 mmol/l treatment was the control group, and the 200 and 800 mmol/l were the experimental groups. The protein levels of XIAP differed between the experimental groups and the control group, decreasing in the experimental groups. (B) Protein expression levels of XIAP in 800 mmol/l ethanol-treated AC16 cardiomyocytes following different time interventions. The 0 h treatment was the control group, and the 24 and 48 h groups were the experimental ones. The protein expression levels of XIAP differed between the experimental groups and the control group, decreasing following ethanol treatment. (C) mRNA expression levels of XIAP in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol, and (D) mRNA expression levels of XIAP in 800 mmol/l ethanol-treated cardiomyocytes after different time interventions. The 0 mmol/l group was the control group, and 200 and 800 mmol/l were the experimental groups. The expression levels of XIAP differed between the experimental groups and the control group, decreasing in the experimental groups with increasing ethanol concentrations and longer treatment times. *P

    Techniques Used: Western Blot, Real-time Polymerase Chain Reaction, Expressing

    Western blotting and reverse-transcription quantitative polymerase chain reaction to detect the expression levels of XIAP in cardiomyocytes transfected with miR-186 mimic/inhibitor. (A) The protein expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes decreased following transfection with miR-186-5p mimic into cells. (B) The protein expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased following transfection with miR-186-5p inhibitor into cells. (C) The mRNA expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes decreased following transfection miR-186-5p mimic into cells. *P
    Figure Legend Snippet: Western blotting and reverse-transcription quantitative polymerase chain reaction to detect the expression levels of XIAP in cardiomyocytes transfected with miR-186 mimic/inhibitor. (A) The protein expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes decreased following transfection with miR-186-5p mimic into cells. (B) The protein expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased following transfection with miR-186-5p inhibitor into cells. (C) The mRNA expression levels of XIAP in ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes decreased following transfection miR-186-5p mimic into cells. *P

    Techniques Used: Western Blot, Real-time Polymerase Chain Reaction, Expressing, Transfection

    Flow cytometry analyses of the apoptosis levels following ethanol treatment. Q4-1, necrotic cells and mechanically injured cells; Q4-2, late-stage apoptotic cells; Q4-3, living cells; Q4-4, early-stage apoptotic cells. The 0 mmol/l (24 or 48 h) groups were the control group, while 200 mmol/l (24 or 48 h) and 800 mmol/l (24 or 48 h) groups were the experimental groups. The results demonstrated that the level of apoptosis in AC16 cardiomyocytes increased following ethanol treatment compared with the control groups, and this was dependent on ethanol concentration and length of ethanol exposure. PE, phycoerythrin.
    Figure Legend Snippet: Flow cytometry analyses of the apoptosis levels following ethanol treatment. Q4-1, necrotic cells and mechanically injured cells; Q4-2, late-stage apoptotic cells; Q4-3, living cells; Q4-4, early-stage apoptotic cells. The 0 mmol/l (24 or 48 h) groups were the control group, while 200 mmol/l (24 or 48 h) and 800 mmol/l (24 or 48 h) groups were the experimental groups. The results demonstrated that the level of apoptosis in AC16 cardiomyocytes increased following ethanol treatment compared with the control groups, and this was dependent on ethanol concentration and length of ethanol exposure. PE, phycoerythrin.

    Techniques Used: Flow Cytometry, Cytometry, Concentration Assay

    Transfecting miR-186-5p mimic affects the levels of ethanol-induced apoptosis of cardiomyocytes. Q2-1, necrotic cells and mechanically injured cells; Q2-2, late-stage apoptotic cells; Q2-3, living cells; Q2-4, early-stage apoptotic cells. The apoptosis levels of ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased, while the apoptosis levels further increased following transfection with the miR-186-5p mimic plasmid. miR, microRNA; FITC, fluorescein isothiocyanate; PE, phycoerythrin.
    Figure Legend Snippet: Transfecting miR-186-5p mimic affects the levels of ethanol-induced apoptosis of cardiomyocytes. Q2-1, necrotic cells and mechanically injured cells; Q2-2, late-stage apoptotic cells; Q2-3, living cells; Q2-4, early-stage apoptotic cells. The apoptosis levels of ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased, while the apoptosis levels further increased following transfection with the miR-186-5p mimic plasmid. miR, microRNA; FITC, fluorescein isothiocyanate; PE, phycoerythrin.

    Techniques Used: Transfection, Plasmid Preparation

    Reverse-transcription quantitative polymerase chain reaction detects the mRNA expression levels of miR-186-5p in ethanol-treated AC16 cardiomyocytes. (A) The mRNA expression levels of miR-186-5p in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol. The 0 mmol/l treatment was the control group, and 200 and 800 mmol/l were the experimental groups. The mRNA expression levels of miR-186-5p differed between the experimental groups and the control group, increasing with higher concentrations of ethanol. (B) The mRNA expression levels of miR-186-5p in 800 mmol/l ethanol-treated AC16 cardiomyocytes following different time interventions. The 0 h was the control group, and 24 and 48 h were the experimental groups. The mRNA expression levels of miR-186-5p also differed between each experimental group and the control, and increasing with longer exposures to ethanol. *P
    Figure Legend Snippet: Reverse-transcription quantitative polymerase chain reaction detects the mRNA expression levels of miR-186-5p in ethanol-treated AC16 cardiomyocytes. (A) The mRNA expression levels of miR-186-5p in ethanol-treated AC16 cardiomyocytes after 24 h intervention with different concentrations of ethanol. The 0 mmol/l treatment was the control group, and 200 and 800 mmol/l were the experimental groups. The mRNA expression levels of miR-186-5p differed between the experimental groups and the control group, increasing with higher concentrations of ethanol. (B) The mRNA expression levels of miR-186-5p in 800 mmol/l ethanol-treated AC16 cardiomyocytes following different time interventions. The 0 h was the control group, and 24 and 48 h were the experimental groups. The mRNA expression levels of miR-186-5p also differed between each experimental group and the control, and increasing with longer exposures to ethanol. *P

    Techniques Used: Real-time Polymerase Chain Reaction, Expressing

    Transfecting the XIAP plasmid reduces the levels of ethanol-induced apoptosis of cardiomyocytes. Q2-1, necrotic cells and mechanically injured cells; Q2-2, late-stage apoptotic cells. Q2-3, living cells; Q2-4, early-stage apoptotic cells. The apoptosis levels of ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased, but the apoptosis levels decreased following transfection with the XIAP plasmid. XIAP, X-linked inhibitor of apoptosis protein; FITC, fluorescein isothiocyanate; PE, phycoerythrin.
    Figure Legend Snippet: Transfecting the XIAP plasmid reduces the levels of ethanol-induced apoptosis of cardiomyocytes. Q2-1, necrotic cells and mechanically injured cells; Q2-2, late-stage apoptotic cells. Q2-3, living cells; Q2-4, early-stage apoptotic cells. The apoptosis levels of ethanol (800 mmol/l, 24 h)-treated AC16 cardiomyocytes increased, but the apoptosis levels decreased following transfection with the XIAP plasmid. XIAP, X-linked inhibitor of apoptosis protein; FITC, fluorescein isothiocyanate; PE, phycoerythrin.

    Techniques Used: Plasmid Preparation, Transfection

    20) Product Images from "The role of MglA for adaptation to oxidative stress of Francisella tularensis LVS"

    Article Title: The role of MglA for adaptation to oxidative stress of Francisella tularensis LVS

    Journal: BMC Microbiology

    doi: 10.1186/1471-2180-12-14

    Analysis of oxidized proteins by the Oxyblot assay . Relative amounts of oxidized proteins in LVS, Δ mglA , or FUU301 during growth in an aerobic or microaerobic environment. Similar results were seen in two additional experiments. The first well of each preparation contained 2.5 ng of protein and the following wells two-fold dilutions thereof. Controls contain non-derivatized samples, and demonstrate the specificity of the antibodies used for detection of oxidative damage.
    Figure Legend Snippet: Analysis of oxidized proteins by the Oxyblot assay . Relative amounts of oxidized proteins in LVS, Δ mglA , or FUU301 during growth in an aerobic or microaerobic environment. Similar results were seen in two additional experiments. The first well of each preparation contained 2.5 ng of protein and the following wells two-fold dilutions thereof. Controls contain non-derivatized samples, and demonstrate the specificity of the antibodies used for detection of oxidative damage.

    Techniques Used:

    Growth of LVS (squares) and Δ mglA (triangles) in CDM in an aerobic (closed symbols) or microaerobic (open symbols) milieu . The diagram shows one representative experiment and similar results were seen in three additional experiments. The error bars represent the standard error of means and are included for all strains but are small for some data points and are therefore not visible in the diagram.
    Figure Legend Snippet: Growth of LVS (squares) and Δ mglA (triangles) in CDM in an aerobic (closed symbols) or microaerobic (open symbols) milieu . The diagram shows one representative experiment and similar results were seen in three additional experiments. The error bars represent the standard error of means and are included for all strains but are small for some data points and are therefore not visible in the diagram.

    Techniques Used:

    21) Product Images from "Evaluation of Bufadienolides as the Main Antitumor Components in Cinobufacin Injection for Liver and Gastric Cancer Therapy"

    Article Title: Evaluation of Bufadienolides as the Main Antitumor Components in Cinobufacin Injection for Liver and Gastric Cancer Therapy

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0169141

    Evaluation of antitumor effects of different Cinobufacin components. Growth inhibition effects of different concentrations of bufadienolides (Buf), alkaloids (Alk), nucleosides (Nuc), and peptides (Pep) extracted from Cinobufacin on BEL-7402 (A) and BGC-823 cells (B). Cells was monitored for 48hrs after adding different Cinobufacin extracts.
    Figure Legend Snippet: Evaluation of antitumor effects of different Cinobufacin components. Growth inhibition effects of different concentrations of bufadienolides (Buf), alkaloids (Alk), nucleosides (Nuc), and peptides (Pep) extracted from Cinobufacin on BEL-7402 (A) and BGC-823 cells (B). Cells was monitored for 48hrs after adding different Cinobufacin extracts.

    Techniques Used: Inhibition

    22) Product Images from "Anti-Interleukin-22-Neutralizing Antibody Attenuates Angiotensin II-Induced Cardiac Hypertrophy in Mice"

    Article Title: Anti-Interleukin-22-Neutralizing Antibody Attenuates Angiotensin II-Induced Cardiac Hypertrophy in Mice

    Journal: Mediators of Inflammation

    doi: 10.1155/2017/5635929

    Effect of IL-22 nAb on cardiac hypertrophy in vitro. H9C2 cells were treated with saline + PBS, saline + IL-22 nAb, angiotensin II + PBS, and angiotensin II + IL-22 nAb. n > 50 cells for each group. ∗ p
    Figure Legend Snippet: Effect of IL-22 nAb on cardiac hypertrophy in vitro. H9C2 cells were treated with saline + PBS, saline + IL-22 nAb, angiotensin II + PBS, and angiotensin II + IL-22 nAb. n > 50 cells for each group. ∗ p

    Techniques Used: In Vitro

    23) Product Images from "Potential of Maintaining a Healthy Vaginal Environment by Two Lactobacillus Strains Isolated from Cocoa Fermentation"

    Article Title: Potential of Maintaining a Healthy Vaginal Environment by Two Lactobacillus Strains Isolated from Cocoa Fermentation

    Journal: BioMed Research International

    doi: 10.1155/2018/7571954

    Death of HMVII cells after 24 h of incubation with LAMP from M. hominis (MhLAMP) or M. genitalium (MgLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p
    Figure Legend Snippet: Death of HMVII cells after 24 h of incubation with LAMP from M. hominis (MhLAMP) or M. genitalium (MgLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p

    Techniques Used: Incubation

    Death of HMVII cells after 24 h of incubation with LAMP from U. parvum (UpLAMP) or U. urealyticum (UuLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p
    Figure Legend Snippet: Death of HMVII cells after 24 h of incubation with LAMP from U. parvum (UpLAMP) or U. urealyticum (UuLAMP) with and without L. plantarum PA3 or L. fermentum FA4. ∗p

    Techniques Used: Incubation

    Interaction of HMVII cells with 4 μ g/mL of M. hominis LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with MhLAMP. (c) HMVII with MhLAMP and L. fermentum FA4. (d) HMVII with MhLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with 4 μ g/mL of M. hominis LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with MhLAMP. (c) HMVII with MhLAMP and L. fermentum FA4. (d) HMVII with MhLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    Interaction of HMVII cells with U. urealyticum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UuLAMP. (c) HMVII with UuLAMP and L. fermentum FA4. (d) HMVII with UuLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with U. urealyticum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UuLAMP. (c) HMVII with UuLAMP and L. fermentum FA4. (d) HMVII with UuLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    Interaction of HMVII cells with 4 μ g/mL of U. parvum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UpLAMP. (c) HMVII with UpLAMP and L. fermentum FA4. (d) HMVII with UpLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).
    Figure Legend Snippet: Interaction of HMVII cells with 4 μ g/mL of U. parvum LAMP with and without L. fermentum FA4 or L. plantarum PA3. (a) Control (HMVII cells alone). (b) HMVII with UpLAMP. (c) HMVII with UpLAMP and L. fermentum FA4. (d) HMVII with UpLAMP and L. plantarum PA3. Green arrows indicate intact HMVII cells, red arrows indicate HMVII cells with altered morphology, and yellow arrows indicate lactobacilli adhered to whole cells (scanning electron microscopy, ×2500).

    Techniques Used: Electron Microscopy

    24) Product Images from "Liver epithelial cells proliferate under hypoxia and protect the liver from ischemic injury via expression of HIF-1 alpha target genes"

    Article Title: Liver epithelial cells proliferate under hypoxia and protect the liver from ischemic injury via expression of HIF-1 alpha target genes

    Journal: Surgery

    doi: 10.1016/j.surg.2012.03.003

    Changes in the partial oxygen pressure in the medium during hypoxic condition of 2% O 2 , 5% CO 2 , balance N 2 in a multigas incubator at 37°C ( n = 4 each).
    Figure Legend Snippet: Changes in the partial oxygen pressure in the medium during hypoxic condition of 2% O 2 , 5% CO 2 , balance N 2 in a multigas incubator at 37°C ( n = 4 each).

    Techniques Used:

    25) Product Images from "Physioxia: a more effective approach for culturing human adipose-derived stem cells for cell transplantation"

    Article Title: Physioxia: a more effective approach for culturing human adipose-derived stem cells for cell transplantation

    Journal: Stem Cell Research & Therapy

    doi: 10.1186/s13287-018-0891-4

    Physioxia improved ASC survivability under ischemic conditions. ASCs (1 × 10 4 ) were seeded onto 96-well plates and incubated in four hostile environments for 24 h: ( a ) ischemic model, 1% O 2 , pH 6.4 and 0.56 μM glucose; ( b ) hypoxic model, 1% O 2 , pH 7.4 and 5.6 μM glucose; ( c ) acidic model, 20% O 2 , pH 6.4 and 5.6 μM glucose; ( d ) nutrient-depleted model, 20% O 2 , pH 7.4 and 0.56 μM glucose. ( A ) Fluorescent images showing the cell death rate by live/dead cell staining. The cell death rate was obtained by the ratio of dead cells to total cells. Three fields were quantified. ( B ) The cell survival rate was detected by WST-8 presented as the ratio of OD 24 to OD 0 . Data are presented as the mean ± SD, * P
    Figure Legend Snippet: Physioxia improved ASC survivability under ischemic conditions. ASCs (1 × 10 4 ) were seeded onto 96-well plates and incubated in four hostile environments for 24 h: ( a ) ischemic model, 1% O 2 , pH 6.4 and 0.56 μM glucose; ( b ) hypoxic model, 1% O 2 , pH 7.4 and 5.6 μM glucose; ( c ) acidic model, 20% O 2 , pH 6.4 and 5.6 μM glucose; ( d ) nutrient-depleted model, 20% O 2 , pH 7.4 and 0.56 μM glucose. ( A ) Fluorescent images showing the cell death rate by live/dead cell staining. The cell death rate was obtained by the ratio of dead cells to total cells. Three fields were quantified. ( B ) The cell survival rate was detected by WST-8 presented as the ratio of OD 24 to OD 0 . Data are presented as the mean ± SD, * P

    Techniques Used: Incubation, Staining

    Physioxia increased ASC survivability in vivo. After mixing with 80 μL of fibrin gel, 1 × 10 6 P-ASCs or H-ASCs were subcutaneously transplanted into the dorsum of nude mice. The implants were extracted after 24, 48, and 72 h. a TUNEL assay was used to stain the nucleus of dead cells. The black arrows indicate dead cells. b The TUNEL + cell rate was determined by the ratio of TUNEL + cells versus total cells. Three fields were quantified. Data are presented as the mean ± SD, * P
    Figure Legend Snippet: Physioxia increased ASC survivability in vivo. After mixing with 80 μL of fibrin gel, 1 × 10 6 P-ASCs or H-ASCs were subcutaneously transplanted into the dorsum of nude mice. The implants were extracted after 24, 48, and 72 h. a TUNEL assay was used to stain the nucleus of dead cells. The black arrows indicate dead cells. b The TUNEL + cell rate was determined by the ratio of TUNEL + cells versus total cells. Three fields were quantified. Data are presented as the mean ± SD, * P

    Techniques Used: In Vivo, Mouse Assay, TUNEL Assay, Staining

    Characterization of P-ASCs and H-ASCs. Human ASCs were cultured under physioxia (2% O 2 , P-ASCs) or hyperoxia (20% O 2 , H-ASCs) for the entire in vitro period until assayed at passage 3. a Flow cytometry was applied to demonstrate the immunophenotype of P-ASCs and H-ASCs. b Morphology of ASCs cultured in α-modified Eagle’s medium (α-MEM) with 10% fetal bovine serum (FBS). c After inducing adipogenesis for 7 days, lipid clusters were stained by oil red O. d Oil red O staining was quantified by the ratio of oil red O + area to total image area for three fields. Data are presented as the mean ± SD, * P > 0.05. e Western blot of hypoxia-inducible factor 1 (HIF-1) and β-actin (reference gene) expression. Scale bar = 50 μm. ASCs adipose-derived stem cells, H-ASCs hyperoxia ASCs, P-ASCs physioxia ASCs
    Figure Legend Snippet: Characterization of P-ASCs and H-ASCs. Human ASCs were cultured under physioxia (2% O 2 , P-ASCs) or hyperoxia (20% O 2 , H-ASCs) for the entire in vitro period until assayed at passage 3. a Flow cytometry was applied to demonstrate the immunophenotype of P-ASCs and H-ASCs. b Morphology of ASCs cultured in α-modified Eagle’s medium (α-MEM) with 10% fetal bovine serum (FBS). c After inducing adipogenesis for 7 days, lipid clusters were stained by oil red O. d Oil red O staining was quantified by the ratio of oil red O + area to total image area for three fields. Data are presented as the mean ± SD, * P > 0.05. e Western blot of hypoxia-inducible factor 1 (HIF-1) and β-actin (reference gene) expression. Scale bar = 50 μm. ASCs adipose-derived stem cells, H-ASCs hyperoxia ASCs, P-ASCs physioxia ASCs

    Techniques Used: Cell Culture, In Vitro, Flow Cytometry, Cytometry, Modification, Staining, Western Blot, Expressing, Derivative Assay

    Physioxia enhanced ASC proliferation and migration through ROS upregulation. a The proliferation of P-ASCs and H-ASCs measured by WST-8 and cell doubling curves. b and d P-ASCs were treated with 100 μM BHA to inhibit ROS, as detected by flow cytometry. The relative MFI was quantified by the ratio of the MFI for P-ASCs and P-ASCs (BHA) to that of H-ASCs. c The proliferation of P-ASCs, H-ASCs and P-ASCs (BHA) measured by WST-8 and cell doubling curves. e Transwell assays were used for determining cell migration, and the migrated cells were stained by 0.1% crystal violet. f The crystal violet in migrated cells was extracted by 10% acetic acid, and the optical density values were determined. The cell doubling curve was produced by dividing the cell number by 10 4 and then transforming the values to log 2 . Data are presented as the mean ± SD, * P
    Figure Legend Snippet: Physioxia enhanced ASC proliferation and migration through ROS upregulation. a The proliferation of P-ASCs and H-ASCs measured by WST-8 and cell doubling curves. b and d P-ASCs were treated with 100 μM BHA to inhibit ROS, as detected by flow cytometry. The relative MFI was quantified by the ratio of the MFI for P-ASCs and P-ASCs (BHA) to that of H-ASCs. c The proliferation of P-ASCs, H-ASCs and P-ASCs (BHA) measured by WST-8 and cell doubling curves. e Transwell assays were used for determining cell migration, and the migrated cells were stained by 0.1% crystal violet. f The crystal violet in migrated cells was extracted by 10% acetic acid, and the optical density values were determined. The cell doubling curve was produced by dividing the cell number by 10 4 and then transforming the values to log 2 . Data are presented as the mean ± SD, * P

    Techniques Used: Migration, Flow Cytometry, Cytometry, Staining, Produced

    Physioxia promoted angiogenic ability of ASCs. ASCs (2 × 10 4 ) were seeded onto 96-well plates coated with 50 μL of Matrigel and cultured for 6 h. a Mesh-like structures resulting from tube formation assay. b , c and d Total mesh, branching length, and junction values per field of view were quantified by ImageJ. Five fields were quantified. e Expression levels of mRNA encoding VEGF, VEGFR2, and vWF as measured by qRT-PCR. Data are presented as the mean ± SD, * P
    Figure Legend Snippet: Physioxia promoted angiogenic ability of ASCs. ASCs (2 × 10 4 ) were seeded onto 96-well plates coated with 50 μL of Matrigel and cultured for 6 h. a Mesh-like structures resulting from tube formation assay. b , c and d Total mesh, branching length, and junction values per field of view were quantified by ImageJ. Five fields were quantified. e Expression levels of mRNA encoding VEGF, VEGFR2, and vWF as measured by qRT-PCR. Data are presented as the mean ± SD, * P

    Techniques Used: Cell Culture, Tube Formation Assay, Expressing, Quantitative RT-PCR

    Physioxia inhibited ASC senescence and apoptosis. a Microscopy images of senescent cells shown by SA-β-Gal staining. b SA-β-Gal staining results were quantified by the ratio of SA-β-Gal + area to the total image area for three fields. c Cell apoptosis measured by flow cytometry using annexin V-FITC/PI double staining. Q1-UL, mechanical error; Q1-UR, late apoptotic or necrotic cells; Q1-LL, viable cells; Q1-LR, early apoptotic cells. d The ratio of viable cells acquired from Q1-LL. Data are presented as the mean ± SD, * P
    Figure Legend Snippet: Physioxia inhibited ASC senescence and apoptosis. a Microscopy images of senescent cells shown by SA-β-Gal staining. b SA-β-Gal staining results were quantified by the ratio of SA-β-Gal + area to the total image area for three fields. c Cell apoptosis measured by flow cytometry using annexin V-FITC/PI double staining. Q1-UL, mechanical error; Q1-UR, late apoptotic or necrotic cells; Q1-LL, viable cells; Q1-LR, early apoptotic cells. d The ratio of viable cells acquired from Q1-LL. Data are presented as the mean ± SD, * P

    Techniques Used: Microscopy, Staining, Flow Cytometry, Cytometry, Double Staining

    26) Product Images from "Zinc inhibits the reproductive toxicity of Zearalenone in immortalized murine ovarian granular KK-1 cells"

    Article Title: Zinc inhibits the reproductive toxicity of Zearalenone in immortalized murine ovarian granular KK-1 cells

    Journal: Scientific Reports

    doi: 10.1038/srep14277

    Zinc inhibits ZEA-induced oxidative stress. The effects of ZnSO 4 , GZn and TPEN on ZEA-induced ROS production ( A ) and the MDA increase ( B ). The ROS production is expressed by the fluorescence intensity of DCFH-DA, while the level of MDA is expressed as the ratio to the protein concentration. The effects of 25 μM ZnSO 4 and GZn on the mRNA expression of Mt2 ( C ) and Sod1 ( E ) at different time points (0, 1, 6, 12, 24 h). The effects of 24 h of treatment with ZnSO 4 , GZn, or ZEA on the Mt2 ( D ), Sod1 ( F ) and Mtf1 ( G ) levels were also examined. KK-1 cells were treated as described in the legend for Fig. 1 . The gene expression was assessed by real-time PCR after RNA isolation and reverse transcription. The cycle threshold (Ct) values of triplicate samples were averaged, and the mRNA expression relative to the control group was normalized to that of the housekeeping gene. The values are the means ± SD of three independent experiments. Different characters indicate that there was a significant difference between the compared groups ( p
    Figure Legend Snippet: Zinc inhibits ZEA-induced oxidative stress. The effects of ZnSO 4 , GZn and TPEN on ZEA-induced ROS production ( A ) and the MDA increase ( B ). The ROS production is expressed by the fluorescence intensity of DCFH-DA, while the level of MDA is expressed as the ratio to the protein concentration. The effects of 25 μM ZnSO 4 and GZn on the mRNA expression of Mt2 ( C ) and Sod1 ( E ) at different time points (0, 1, 6, 12, 24 h). The effects of 24 h of treatment with ZnSO 4 , GZn, or ZEA on the Mt2 ( D ), Sod1 ( F ) and Mtf1 ( G ) levels were also examined. KK-1 cells were treated as described in the legend for Fig. 1 . The gene expression was assessed by real-time PCR after RNA isolation and reverse transcription. The cycle threshold (Ct) values of triplicate samples were averaged, and the mRNA expression relative to the control group was normalized to that of the housekeeping gene. The values are the means ± SD of three independent experiments. Different characters indicate that there was a significant difference between the compared groups ( p

    Techniques Used: Multiple Displacement Amplification, Fluorescence, Protein Concentration, Expressing, Real-time Polymerase Chain Reaction, Isolation

    Cytotoxicity of ZnSO 4 , GZn, and TPEN. KK-1 cells were incubated with various concentrations of ZnSO 4 (0–150 μM), GZn (0–150 μM), or TPEN (0–20 μM) for 24 h. The effects of ZnSO 4 ( A ), GZn ( B ) and TPEN ( C ) on the cell viability were assessed by a CCK-8 assay. The effects of a 24-h coincubation with zinc and ZEA on the cell viability ( D ): C, 0.05% DMSO (v/v in DMEM); Zn, 25 μM ZnSO 4 ; GZn, 25 μM GZn; HZn, 12.5 μM ZnSO 4 and 12.5 μM GZn; ZEA, 20 μM ZEA; T, 2.5 μM TPEN; Z + Zn, 25 μM ZnSO 4 and 20 μM ZEA; Z + GZn, 25 μM GZn and 20 μM ZEA; HZn + ZEA, 12.5 μM ZnSO 4 , 12.5 μM GZn and 20 μM ZEA; Z + T, 20 μM ZEA and 2.5 μM TPEN. The changes in the intracellular zinc concentration after treatment with ZnSO 4 , GZn or TPEN treatments ( E ). KK-1 cells were incubated with 0.05% DMSO (v/v in DMEM), 25 μM ZnSO 4 , 25 μM GZn or 2.5 μM TPEN. The cellular zinc concentration was determined using the FACSCalibur instrument at different time points (0, 1, 6, 12, 24 h) as described in the Materials and methods. The effects of ZnSO 4 and GZn on the mRNA expression of zinc transporters Slc30a1 ( F ) and Slc39a1 ( G ). KK-1 cells were incubated with 0.05% DMSO (v/v in DMEM), 25 μM ZnSO 4 or 25 μM GZn for different time periods (0, 1, 6, 12, 24 h). The values are the means ± SD of three independent experiments. The characters indicate significant differences between the compared groups ( p
    Figure Legend Snippet: Cytotoxicity of ZnSO 4 , GZn, and TPEN. KK-1 cells were incubated with various concentrations of ZnSO 4 (0–150 μM), GZn (0–150 μM), or TPEN (0–20 μM) for 24 h. The effects of ZnSO 4 ( A ), GZn ( B ) and TPEN ( C ) on the cell viability were assessed by a CCK-8 assay. The effects of a 24-h coincubation with zinc and ZEA on the cell viability ( D ): C, 0.05% DMSO (v/v in DMEM); Zn, 25 μM ZnSO 4 ; GZn, 25 μM GZn; HZn, 12.5 μM ZnSO 4 and 12.5 μM GZn; ZEA, 20 μM ZEA; T, 2.5 μM TPEN; Z + Zn, 25 μM ZnSO 4 and 20 μM ZEA; Z + GZn, 25 μM GZn and 20 μM ZEA; HZn + ZEA, 12.5 μM ZnSO 4 , 12.5 μM GZn and 20 μM ZEA; Z + T, 20 μM ZEA and 2.5 μM TPEN. The changes in the intracellular zinc concentration after treatment with ZnSO 4 , GZn or TPEN treatments ( E ). KK-1 cells were incubated with 0.05% DMSO (v/v in DMEM), 25 μM ZnSO 4 , 25 μM GZn or 2.5 μM TPEN. The cellular zinc concentration was determined using the FACSCalibur instrument at different time points (0, 1, 6, 12, 24 h) as described in the Materials and methods. The effects of ZnSO 4 and GZn on the mRNA expression of zinc transporters Slc30a1 ( F ) and Slc39a1 ( G ). KK-1 cells were incubated with 0.05% DMSO (v/v in DMEM), 25 μM ZnSO 4 or 25 μM GZn for different time periods (0, 1, 6, 12, 24 h). The values are the means ± SD of three independent experiments. The characters indicate significant differences between the compared groups ( p

    Techniques Used: Incubation, CCK-8 Assay, Concentration Assay, Expressing

    Zinc inhibits ZEA-induced cell apoptosis. The effects of ZnSO 4 , GZn, and ZEA on the Δψm ( A ) and the percentage of cell apoptosis ( B ). The Δψm is expressed as the fluorescence intensity ratio of the red over the green staining. The effects of ZnSO 4 , GZn, and ZEA on the mRNA expression of Bax ( C ), Casp3 ( D ) and Casp9 ( E ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate significant differences between the compared groups ( p
    Figure Legend Snippet: Zinc inhibits ZEA-induced cell apoptosis. The effects of ZnSO 4 , GZn, and ZEA on the Δψm ( A ) and the percentage of cell apoptosis ( B ). The Δψm is expressed as the fluorescence intensity ratio of the red over the green staining. The effects of ZnSO 4 , GZn, and ZEA on the mRNA expression of Bax ( C ), Casp3 ( D ) and Casp9 ( E ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate significant differences between the compared groups ( p

    Techniques Used: Fluorescence, Staining, Expressing

    The effects of zinc were validated at the protein level by Western blotting. The effects of ZnSO 4, GZn, and ZEA on the protein expression of Bax ( A ) and Casp9 ( B ). The results of the Western blot analysis and the relative expression of each protein are shown. The effects of ZnSO 4 , GZn, and ZEA on the cell cycle was detected by a FACSCalibur instrument. The results are shown in ( C ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate that there was a significant difference between the compared groups ( p
    Figure Legend Snippet: The effects of zinc were validated at the protein level by Western blotting. The effects of ZnSO 4, GZn, and ZEA on the protein expression of Bax ( A ) and Casp9 ( B ). The results of the Western blot analysis and the relative expression of each protein are shown. The effects of ZnSO 4 , GZn, and ZEA on the cell cycle was detected by a FACSCalibur instrument. The results are shown in ( C ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate that there was a significant difference between the compared groups ( p

    Techniques Used: Western Blot, Expressing

    Zinc regulates steroidogenic enzymes and promotes estrogen production. The effects of ZnSO 4 , GZn, and ZEA on the transcription of Star ( A ), Cyp11a1 ( B ), Cyp17a1 ( C ) and Hsd3b1 ( D ) are shown. The effects of ZnSO 4 , GZn, and ZEA on the protein expression of these steroidogenic enzymes were determined by Western blotting, and the results and relative expression of Star ( F ) and Cyp11a1 ( G ) are shown. The effects of ZnSO 4 , GZn, and ZEA on the estrogen production were determined by RIA ( E ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate significant differences between the compared groups ( p
    Figure Legend Snippet: Zinc regulates steroidogenic enzymes and promotes estrogen production. The effects of ZnSO 4 , GZn, and ZEA on the transcription of Star ( A ), Cyp11a1 ( B ), Cyp17a1 ( C ) and Hsd3b1 ( D ) are shown. The effects of ZnSO 4 , GZn, and ZEA on the protein expression of these steroidogenic enzymes were determined by Western blotting, and the results and relative expression of Star ( F ) and Cyp11a1 ( G ) are shown. The effects of ZnSO 4 , GZn, and ZEA on the estrogen production were determined by RIA ( E ). KK-1 cells were treated as described in the legend for Fig. 1 . The values are the means ± SD of three independent experiments. Different characters indicate significant differences between the compared groups ( p

    Techniques Used: Expressing, Western Blot

    27) Product Images from "Heparin-binding epidermal growth factor-like growth factor is a potent regulator of invasion activity in oral squamous cell carcinoma"

    Article Title: Heparin-binding epidermal growth factor-like growth factor is a potent regulator of invasion activity in oral squamous cell carcinoma

    Journal: Oncology Reports

    doi: 10.3892/or.2011.1616

    Shedding of HB-EGF upregulates MMP-9 expression through EGFR receptor activation. (A) The TACE inhibitor TAPI-2 reduced MMP-9 mRNA levels in HSC3 cells. (B) The EGFR inhibitor AG1478 downregulated MMP-9 mRNA levels in HSC3 cells
    Figure Legend Snippet: Shedding of HB-EGF upregulates MMP-9 expression through EGFR receptor activation. (A) The TACE inhibitor TAPI-2 reduced MMP-9 mRNA levels in HSC3 cells. (B) The EGFR inhibitor AG1478 downregulated MMP-9 mRNA levels in HSC3 cells

    Techniques Used: Expressing, Activation Assay

    Expression levels of EGF family mRNA in HSC3 cells. Amphiregulin (AREG), epigen (EPHN), epiregulin (EREG), HB-EGF (HBEGF), TGF-α (TGF), betacellulin (BTC), EGF and β-actin mRNA expression levels in HSC3 cells were analyzed by RT-qPCR.
    Figure Legend Snippet: Expression levels of EGF family mRNA in HSC3 cells. Amphiregulin (AREG), epigen (EPHN), epiregulin (EREG), HB-EGF (HBEGF), TGF-α (TGF), betacellulin (BTC), EGF and β-actin mRNA expression levels in HSC3 cells were analyzed by RT-qPCR.

    Techniques Used: Expressing, Quantitative RT-PCR

    Knockdown of heparin-binding epidermal growth factor-like growth factor (HB-EGF) affects invasion activity in HSC3 cells. (A) Expression of HB-EGF mRNA in HSC3 cells treated with siRNAs was determined by real-time RT-PCR. The amount of HB-EGF mRNA was significantly reduced in HB-EGF siRNA-treated HSC3 cells compared with that in control no-siRNA or scrambled-siRNA-treated HSC3 cells. (B) Effect of HB-EGF on HSC3 cell proliferation. The MTT assay showed similar proliferation rates among cells treated with HB-EGF siRNA (closed circle), no-siRNA (triangle), and scrambled-siRNA (open circle). (C) Invasion activity of HB-EGF siRNA-transfected HSC3 cells as determined by the Matrigel invasion assay. The invasion index represents the ratio of cells migrating through a Matrigel-coated membrane/cells migrating through a control non-coated membrane. No-siRNA and scrambled-siRNA transfectants were highly invasive, whereas the invasiveness of HB-EGF siRNA-transfected HSC3 cells was significantly lower.
    Figure Legend Snippet: Knockdown of heparin-binding epidermal growth factor-like growth factor (HB-EGF) affects invasion activity in HSC3 cells. (A) Expression of HB-EGF mRNA in HSC3 cells treated with siRNAs was determined by real-time RT-PCR. The amount of HB-EGF mRNA was significantly reduced in HB-EGF siRNA-treated HSC3 cells compared with that in control no-siRNA or scrambled-siRNA-treated HSC3 cells. (B) Effect of HB-EGF on HSC3 cell proliferation. The MTT assay showed similar proliferation rates among cells treated with HB-EGF siRNA (closed circle), no-siRNA (triangle), and scrambled-siRNA (open circle). (C) Invasion activity of HB-EGF siRNA-transfected HSC3 cells as determined by the Matrigel invasion assay. The invasion index represents the ratio of cells migrating through a Matrigel-coated membrane/cells migrating through a control non-coated membrane. No-siRNA and scrambled-siRNA transfectants were highly invasive, whereas the invasiveness of HB-EGF siRNA-transfected HSC3 cells was significantly lower.

    Techniques Used: Binding Assay, Activity Assay, Expressing, Quantitative RT-PCR, MTT Assay, Transfection, Invasion Assay

    Heparin-binding epidermal growth factor-like growth factor (HB-EGF) upregulates MMP-9 expression. (A) MMP-9 mRNA expression and collagenase activity in siRNA-treated HSC3 cells was determined by RT-PCR and zymography, respectively. The MMP-9 mRNA level was reduced in HB-EGF siRNA-treated cells compared with the levels in control no-siRNA and scrambled-siRNA-treated HSC3 cells. (B) The addition of s-HB-EGF to the culture medium upregulated MMP-9 mRNA levels in HSC3 cells.
    Figure Legend Snippet: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) upregulates MMP-9 expression. (A) MMP-9 mRNA expression and collagenase activity in siRNA-treated HSC3 cells was determined by RT-PCR and zymography, respectively. The MMP-9 mRNA level was reduced in HB-EGF siRNA-treated cells compared with the levels in control no-siRNA and scrambled-siRNA-treated HSC3 cells. (B) The addition of s-HB-EGF to the culture medium upregulated MMP-9 mRNA levels in HSC3 cells.

    Techniques Used: Binding Assay, Expressing, Activity Assay, Reverse Transcription Polymerase Chain Reaction, Zymography

    28) Product Images from "Delivery of Small Interfering RNA for Inhibition of Endothelial Cell Apoptosis by Hypoxia and Serum Deprivation"

    Article Title: Delivery of Small Interfering RNA for Inhibition of Endothelial Cell Apoptosis by Hypoxia and Serum Deprivation

    Journal: Biochemical and biophysical research communications

    doi: 10.1016/j.bbrc.2008.08.123

    Silencing of target gene (TNFR-1 and SHP-1) expression by siRNA transfection. (A) RT-PCR of HUVECs for each target at 2 days after transfection with TNFR-1, SHP-1, or GFP siRNA. Quantitative real-time PCR of siRNA-transfected HUVECs (n=3) for (B) TNFR-1
    Figure Legend Snippet: Silencing of target gene (TNFR-1 and SHP-1) expression by siRNA transfection. (A) RT-PCR of HUVECs for each target at 2 days after transfection with TNFR-1, SHP-1, or GFP siRNA. Quantitative real-time PCR of siRNA-transfected HUVECs (n=3) for (B) TNFR-1

    Techniques Used: Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction

    Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. (A) RT-PCR for various molecules of siRNA-transfected HUVECs retrieved at days
    Figure Legend Snippet: Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. (A) RT-PCR for various molecules of siRNA-transfected HUVECs retrieved at days

    Techniques Used: Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction

    Apoptotic activity of siRNA-transfected HUVECs cultured under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. (A) TUNEL staining of siRNA-transfected HUVECs at 2 days after culture (×100).
    Figure Legend Snippet: Apoptotic activity of siRNA-transfected HUVECs cultured under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. (A) TUNEL staining of siRNA-transfected HUVECs at 2 days after culture (×100).

    Techniques Used: Activity Assay, Transfection, Cell Culture, TUNEL Assay, Staining

    Capillary formation by siRNA-transfected HUVECs cultured for 2 days under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. When cultured onto GFR-Matrigel, HUVECs transfected with siRNAs
    Figure Legend Snippet: Capillary formation by siRNA-transfected HUVECs cultured for 2 days under hypoxic (1% oxygen) and survival factor-deprived (EBM-2 basal medium with no serum and growth factors) conditions. When cultured onto GFR-Matrigel, HUVECs transfected with siRNAs

    Techniques Used: Transfection, Cell Culture

    29) Product Images from "Crocus sativus L. (Saffron) Stigma Aqueous Extract Induces Apoptosis in Alveolar Human Lung Cancer Cells through Caspase-Dependent Pathways Activation"

    Article Title: Crocus sativus L. (Saffron) Stigma Aqueous Extract Induces Apoptosis in Alveolar Human Lung Cancer Cells through Caspase-Dependent Pathways Activation

    Journal: BioMed Research International

    doi: 10.1155/2013/417928

    Comparison of cytotoxicty effect of saffron on cell viability of the A549 cells. Morphological changes of cells after 48-hour treatment with different concentrations of saffron (200, 400 μ g/mL), (a) untreated A549 cells (b) the A549 cells treated with 200 ( μ g/mL) saffron and (c) the A549 cells treated with 400 ( μ g/mL) saffron.
    Figure Legend Snippet: Comparison of cytotoxicty effect of saffron on cell viability of the A549 cells. Morphological changes of cells after 48-hour treatment with different concentrations of saffron (200, 400 μ g/mL), (a) untreated A549 cells (b) the A549 cells treated with 200 ( μ g/mL) saffron and (c) the A549 cells treated with 400 ( μ g/mL) saffron.

    Techniques Used:

    Nuclear changes of saffron-treated A549 cells. A549 cells seeded on chamber slides were treated with media or different saffron concentrations. Suspension of the A549 cells was incubated with media (a), 200 μ g/mL of saffron (b), and 400 μ g/mL of saffron (c) for 48 hours at 37°C to induce apoptosis. Cells were then labeled with FAM-VAD-FMK for 60 minutes at 37°C. Cells were washed, and then Hoechst stain was added and incubated for 5 minutes. Wet-mount slides were prepared and the photos of the same cells were taken and superimposed. Caspase activity (green) was detected using a band pass filter (excitation at 488 nm, emission at 520 nm). Nuclear staining by Hoechst stain (blue) was revealed using a UV filter (excitation at 365 nm, emission at 480 nm). In these pictures, the yellow arrows represent the nuclear breakdown and chromatin condensation of A549 cells induced by saffron.
    Figure Legend Snippet: Nuclear changes of saffron-treated A549 cells. A549 cells seeded on chamber slides were treated with media or different saffron concentrations. Suspension of the A549 cells was incubated with media (a), 200 μ g/mL of saffron (b), and 400 μ g/mL of saffron (c) for 48 hours at 37°C to induce apoptosis. Cells were then labeled with FAM-VAD-FMK for 60 minutes at 37°C. Cells were washed, and then Hoechst stain was added and incubated for 5 minutes. Wet-mount slides were prepared and the photos of the same cells were taken and superimposed. Caspase activity (green) was detected using a band pass filter (excitation at 488 nm, emission at 520 nm). Nuclear staining by Hoechst stain (blue) was revealed using a UV filter (excitation at 365 nm, emission at 480 nm). In these pictures, the yellow arrows represent the nuclear breakdown and chromatin condensation of A549 cells induced by saffron.

    Techniques Used: Incubation, Labeling, Staining, Activity Assay

    The A549 cells were treated with media, noninduced cells (a) or saffron extract 200 μ g/mL (b) or 400 μ g/mL (c) induced cells, for 48 hours. Cells were labeled with FAMVAD-FMK for 1 hour, washed, and analyzed. Caspase activity was detected using a BD FACSCalibur flow cytometer.
    Figure Legend Snippet: The A549 cells were treated with media, noninduced cells (a) or saffron extract 200 μ g/mL (b) or 400 μ g/mL (c) induced cells, for 48 hours. Cells were labeled with FAMVAD-FMK for 1 hour, washed, and analyzed. Caspase activity was detected using a BD FACSCalibur flow cytometer.

    Techniques Used: Labeling, Activity Assay, Flow Cytometry, Cytometry

    Exposure of the A549 cells for 24, 48, and 72 h with saffron. Cells were treated with different concentrations of saffron extract (100, 200, 400, and 800 μ g/mL) for 24 (a), 48 (b), and 72 (c) hours. Viability was quantitated by MTT assay. Results are mean ± SEM. The asterisks are indicators of statistical difference obtained separately at different time points compared to their controls shown in figure as * P
    Figure Legend Snippet: Exposure of the A549 cells for 24, 48, and 72 h with saffron. Cells were treated with different concentrations of saffron extract (100, 200, 400, and 800 μ g/mL) for 24 (a), 48 (b), and 72 (c) hours. Viability was quantitated by MTT assay. Results are mean ± SEM. The asterisks are indicators of statistical difference obtained separately at different time points compared to their controls shown in figure as * P

    Techniques Used: MTT Assay

    Effect of saffron on exponentially growing A549 cells. Scattergrams of LSC measurements representing the cells stained with FAM-VAD-FMK (green fluorescence) and PI (red fluorescence). (A) Control cells exponentially growing in standard condition. (B) Cells exposed to 200 μ g/mL saffron for 48 h during exponential growth. (C) Cells exposed to 400 μ g/mL saffron for 48 h during exponential growth. Each scatterplot is divided into four compartments depending on the cell status as revealed by green versus red fluorescence. a: live cells; b: early apoptotic cells; c: late apoptotic cells; d: secondary necrotic cells.
    Figure Legend Snippet: Effect of saffron on exponentially growing A549 cells. Scattergrams of LSC measurements representing the cells stained with FAM-VAD-FMK (green fluorescence) and PI (red fluorescence). (A) Control cells exponentially growing in standard condition. (B) Cells exposed to 200 μ g/mL saffron for 48 h during exponential growth. (C) Cells exposed to 400 μ g/mL saffron for 48 h during exponential growth. Each scatterplot is divided into four compartments depending on the cell status as revealed by green versus red fluorescence. a: live cells; b: early apoptotic cells; c: late apoptotic cells; d: secondary necrotic cells.

    Techniques Used: Staining, Fluorescence

    30) Product Images from "Hypoxia differentially regulated CXCR4 and CXCR7 signaling in colon cancer"

    Article Title: Hypoxia differentially regulated CXCR4 and CXCR7 signaling in colon cancer

    Journal: Molecular Cancer

    doi: 10.1186/1476-4598-13-58

    Activation of Akt and Erk oncogenic pathways by stimulation of CXCR4/CXCL12 axis in SW480 cell line. Effect of CXCL12 stimulation on Akt and Erk oncogenic pathways. SW480 cells were stimulated with CXCL12 at concentration of 0.5 nM. The cells cultivated either in normoxia or in hypoxia (1%O2) and previously starved for 4 h in a serum-free medium before adding CXCL12, were evaluated for protein kinase phosphorylation.
    Figure Legend Snippet: Activation of Akt and Erk oncogenic pathways by stimulation of CXCR4/CXCL12 axis in SW480 cell line. Effect of CXCL12 stimulation on Akt and Erk oncogenic pathways. SW480 cells were stimulated with CXCL12 at concentration of 0.5 nM. The cells cultivated either in normoxia or in hypoxia (1%O2) and previously starved for 4 h in a serum-free medium before adding CXCL12, were evaluated for protein kinase phosphorylation.

    Techniques Used: Activation Assay, Concentration Assay

    CXCR4 expression in different colon cell lines cultured in hypoxia. (A) Transcript CXCR4 expression in normoxia (20% O2) and hypoxia (1% O2) in SW480, HCT116 and HT29 cell lines. (B) Transcript CXCR7 expression in normoxia (20% O2) and in hypoxia (3% and 1% O2) in SW480 cell line. Trancript HIF-1α (C) and CXCR4 (D) expression after siRNA #1 or #2 anti-HIF-1α transfection; * p = 0.015; ** p = 0.001.
    Figure Legend Snippet: CXCR4 expression in different colon cell lines cultured in hypoxia. (A) Transcript CXCR4 expression in normoxia (20% O2) and hypoxia (1% O2) in SW480, HCT116 and HT29 cell lines. (B) Transcript CXCR7 expression in normoxia (20% O2) and in hypoxia (3% and 1% O2) in SW480 cell line. Trancript HIF-1α (C) and CXCR4 (D) expression after siRNA #1 or #2 anti-HIF-1α transfection; * p = 0.015; ** p = 0.001.

    Techniques Used: Expressing, Cell Culture, Transfection

    Regulation of CXCR4 and CXCR7 protein expression at the cell membrane using flow cytometry. (A) CXCR4 and CXCR7 expression at the cell membrane of SW480, HCT116, and HT29, in normoxia (20% O2) and hypoxia (1% O 2 ). (B) CXCR4 and CXCR7 expression in SW480 cells after siRNA anti-HIF-1α or anti-CXCR4 in hypoxia. (C) Cell membrane CXCR4 protein expression after transient passage in hypoxia. SW480, HCT116 and HT29 cells were cultured 24 hours in 3% hypoxia and further maintained for 8, 24 and 48 hours in normoxia.
    Figure Legend Snippet: Regulation of CXCR4 and CXCR7 protein expression at the cell membrane using flow cytometry. (A) CXCR4 and CXCR7 expression at the cell membrane of SW480, HCT116, and HT29, in normoxia (20% O2) and hypoxia (1% O 2 ). (B) CXCR4 and CXCR7 expression in SW480 cells after siRNA anti-HIF-1α or anti-CXCR4 in hypoxia. (C) Cell membrane CXCR4 protein expression after transient passage in hypoxia. SW480, HCT116 and HT29 cells were cultured 24 hours in 3% hypoxia and further maintained for 8, 24 and 48 hours in normoxia.

    Techniques Used: Expressing, Flow Cytometry, Cytometry, Cell Culture

    Impact of CXCR4/CXCL12 axis inhibition in SW480 cell line. (A) Effect of CXCR4/CXCL12 axis inhibition on Akt and Erk phosphorylation. Cells are treated with CXCR4 and CXCR7 siRNAs in hypoxia (1% O 2 ) in presence of 0.5 nM CXCL12 for 15 minutes with or without the CXCR4 antagonist AMD 3100 (10 μM). (B) SW480 cell migration in hypoxia (3% and 1% O2) using a Boyden chamber assay. After 24 h incubation, cells remaining on the upper chamber are mechanically removed. The cells that have migrated to the lower chamber are counted after staining with fluorescent dye (DAPI or 4',6-diamidino-2-phenylindole). Quantification was performed by microscope counting five random fields for each chamber (* p
    Figure Legend Snippet: Impact of CXCR4/CXCL12 axis inhibition in SW480 cell line. (A) Effect of CXCR4/CXCL12 axis inhibition on Akt and Erk phosphorylation. Cells are treated with CXCR4 and CXCR7 siRNAs in hypoxia (1% O 2 ) in presence of 0.5 nM CXCL12 for 15 minutes with or without the CXCR4 antagonist AMD 3100 (10 μM). (B) SW480 cell migration in hypoxia (3% and 1% O2) using a Boyden chamber assay. After 24 h incubation, cells remaining on the upper chamber are mechanically removed. The cells that have migrated to the lower chamber are counted after staining with fluorescent dye (DAPI or 4',6-diamidino-2-phenylindole). Quantification was performed by microscope counting five random fields for each chamber (* p

    Techniques Used: Inhibition, Migration, Boyden Chamber Assay, Incubation, Staining, Microscopy

    31) Product Images from "Exosomes Secreted from CXCR4 Overexpressing Mesenchymal Stem Cells Promote Cardioprotection via Akt Signaling Pathway following Myocardial Infarction"

    Article Title: Exosomes Secreted from CXCR4 Overexpressing Mesenchymal Stem Cells Promote Cardioprotection via Akt Signaling Pathway following Myocardial Infarction

    Journal: Stem Cells International

    doi: 10.1155/2015/659890

    The in vitro angiogenic properties of MSC CR4 -derived exosomes. (a) Induction of tube formation by MSC-derived exosomes in HUVEC. (b) Quantitative data of total branch points in various groups. (c) VEGF expression in various groups was analyzed by qPCR. * P
    Figure Legend Snippet: The in vitro angiogenic properties of MSC CR4 -derived exosomes. (a) Induction of tube formation by MSC-derived exosomes in HUVEC. (b) Quantitative data of total branch points in various groups. (c) VEGF expression in various groups was analyzed by qPCR. * P

    Techniques Used: In Vitro, Derivative Assay, Expressing, Real-time Polymerase Chain Reaction

    Expression of CXCR4 in MSC and MSC-derived exosomes. (a) CXCR4 expression was identified by Western blotting in MSC from various groups with quantitative analysis. ((b) and (c)) The expression of CXCR4 in MSC-derived exosome was detected by Western blotting and quantitative analysis. All values expressed as mean ± SEM. * P
    Figure Legend Snippet: Expression of CXCR4 in MSC and MSC-derived exosomes. (a) CXCR4 expression was identified by Western blotting in MSC from various groups with quantitative analysis. ((b) and (c)) The expression of CXCR4 in MSC-derived exosome was detected by Western blotting and quantitative analysis. All values expressed as mean ± SEM. * P

    Techniques Used: Expressing, Derivative Assay, Western Blot

    Schematic depiction of functional exosomes secreted from MSC overexpressing CXCR4 for activation of signaling pathways for restoration of LV function after MI. In exosome generating cells, CXCR4 are selectively incorporated into the intraluminal vesicles (IVLs) of multivesicular endosomes (MVEs) from the plasma membrane. Then, by fusing with the plasma membrane, MVEs release CXCR4-enriched exosome into the extracellular milieu. CXCR4-enriched exosomes can bind to the plasma membrane of the targeting cells. These recruited CXCR4-enriched exosomes may either fuse directly with the plasma membrane or fuse with delimiting membrane of an endocytic compartment. Then, released CXCR4 can be delivered into the membrane or cytosol of the targeting cell which contributes to SDF-1 α /CXCR4 interaction. In infarcted heart tissue, increased binding of SDF-1 α to CXCR4 activates G-protein-coupled receptor kinases, which activate a cascade of signaling pathways in cells. Pretreatment of stem/progenitor cells with CXCR4-enriched exosome can reduce MI-induced cell death and promote angiogenesis through PI3K/Akt signaling pathway activation, thereby enhancing heart function improvement.
    Figure Legend Snippet: Schematic depiction of functional exosomes secreted from MSC overexpressing CXCR4 for activation of signaling pathways for restoration of LV function after MI. In exosome generating cells, CXCR4 are selectively incorporated into the intraluminal vesicles (IVLs) of multivesicular endosomes (MVEs) from the plasma membrane. Then, by fusing with the plasma membrane, MVEs release CXCR4-enriched exosome into the extracellular milieu. CXCR4-enriched exosomes can bind to the plasma membrane of the targeting cells. These recruited CXCR4-enriched exosomes may either fuse directly with the plasma membrane or fuse with delimiting membrane of an endocytic compartment. Then, released CXCR4 can be delivered into the membrane or cytosol of the targeting cell which contributes to SDF-1 α /CXCR4 interaction. In infarcted heart tissue, increased binding of SDF-1 α to CXCR4 activates G-protein-coupled receptor kinases, which activate a cascade of signaling pathways in cells. Pretreatment of stem/progenitor cells with CXCR4-enriched exosome can reduce MI-induced cell death and promote angiogenesis through PI3K/Akt signaling pathway activation, thereby enhancing heart function improvement.

    Techniques Used: Functional Assay, Activation Assay, Binding Assay

    The effect of MSC CR4 -derived exosomes on apoptosis. (a) TUNEL staining for CM apoptosis under hypoxic condition. CM labeled with cardiac specific antibody, sarcomeric α -actinin (red). Nuclei were labeled with DAPI (blue) and TUNEL positive nuclei (green). Original magnification ×200. (b) Quantitative data for the numbers of TUNEL positive nuclei in various experimental groups. All values expressed as mean ± SEM. * P
    Figure Legend Snippet: The effect of MSC CR4 -derived exosomes on apoptosis. (a) TUNEL staining for CM apoptosis under hypoxic condition. CM labeled with cardiac specific antibody, sarcomeric α -actinin (red). Nuclei were labeled with DAPI (blue) and TUNEL positive nuclei (green). Original magnification ×200. (b) Quantitative data for the numbers of TUNEL positive nuclei in various experimental groups. All values expressed as mean ± SEM. * P

    Techniques Used: Derivative Assay, TUNEL Assay, Staining, Labeling

    Characterization of MSC-derived exosomes. (a) Transmission electron microscopy (TEM) micrograph of exosomes purified from MSC. Isolated exosomes showed spherical and membrane encapsulated with the diameters varying between 40 and 90 nm. (b) Representative dynamic light scattering (DLS) number distribution measurement of isolated exosome population demonstrates a single peak (~90 nm diameter) indicating they are free of contamination. (c) Expression of specific exosomal markers, CD9 and CD63, in exosomes was identified by Western blotting. Exo Ctrl , exosomes isolated from MSC transfected with null lentivirus (MSC Ctrl ); Exo CR4 , exosomes isolated from MSC transfected with lentivirus overexpressing CXCR4 (MSC CR4 ); Exo siCR4 , exosomes isolated from MSC transfected with lentivirus siRNA against CXCR4 gene (MSC siCR4 ); Exo + GW4860, exosomes isolated from MSC with additional sphingomyelinase inhibitor GW4860. (d) Uptake of exosomes by neonatal cardiomyocytes (CM). CM were stained by cardiac specific antibody, sarcomeric α -actinin (red). MSC CR4 -derived exosomes were labeled with PKH67 (green) and incubated with CM for 48 hr.
    Figure Legend Snippet: Characterization of MSC-derived exosomes. (a) Transmission electron microscopy (TEM) micrograph of exosomes purified from MSC. Isolated exosomes showed spherical and membrane encapsulated with the diameters varying between 40 and 90 nm. (b) Representative dynamic light scattering (DLS) number distribution measurement of isolated exosome population demonstrates a single peak (~90 nm diameter) indicating they are free of contamination. (c) Expression of specific exosomal markers, CD9 and CD63, in exosomes was identified by Western blotting. Exo Ctrl , exosomes isolated from MSC transfected with null lentivirus (MSC Ctrl ); Exo CR4 , exosomes isolated from MSC transfected with lentivirus overexpressing CXCR4 (MSC CR4 ); Exo siCR4 , exosomes isolated from MSC transfected with lentivirus siRNA against CXCR4 gene (MSC siCR4 ); Exo + GW4860, exosomes isolated from MSC with additional sphingomyelinase inhibitor GW4860. (d) Uptake of exosomes by neonatal cardiomyocytes (CM). CM were stained by cardiac specific antibody, sarcomeric α -actinin (red). MSC CR4 -derived exosomes were labeled with PKH67 (green) and incubated with CM for 48 hr.

    Techniques Used: Derivative Assay, Transmission Assay, Electron Microscopy, Transmission Electron Microscopy, Purification, Isolation, Expressing, Western Blot, Transfection, Staining, Labeling, Incubation

    32) Product Images from "Immunosuppression of breast cancer cells mediated by transforming growth factor-β in exosomes from cancer cells"

    Article Title: Immunosuppression of breast cancer cells mediated by transforming growth factor-β in exosomes from cancer cells

    Journal: Oncology Letters

    doi: 10.3892/ol.2015.3841

    Exosomes were collected from MDA-MB-231 and BT-474 cells cultured under normoxic or hypoxic conditions and lysed with 1% Nonidet P-40. Following SDS-PAGE, immunoblotting and exposure to detection reagents, relative densitometry was performed, which revealed
    Figure Legend Snippet: Exosomes were collected from MDA-MB-231 and BT-474 cells cultured under normoxic or hypoxic conditions and lysed with 1% Nonidet P-40. Following SDS-PAGE, immunoblotting and exposure to detection reagents, relative densitometry was performed, which revealed

    Techniques Used: Multiple Displacement Amplification, Cell Culture, SDS Page

    33) Product Images from "Luciferase-Specific Coelenterazine Analogues for Optical Contamination-Free Bioassays"

    Article Title: Luciferase-Specific Coelenterazine Analogues for Optical Contamination-Free Bioassays

    Journal: Scientific Reports

    doi: 10.1038/s41598-017-00955-6

    Biased selectivity of the synthetic luciferins to marine luciferases. ( a ) The relative selectivity of representative synthetic luciferins according to luciferases in cell lysates. ( b ) The corresponding live-cell images on a 6-channel microslide, demonstrating the relative luciferase-selectivity of the selected coelenterazine (CTZ) analogues in living mammalian cells. The optical intensity ranges of 6piOH-2H-CTZ, 6etOH-CTZ, and 6piOH-CTZ were 1024–2261, 512–2214, 512–2841 RLU, respectively. ( c ) The time course of the optical intensities after substrate injection to the microslide channels growing live cells. The bioluminescence exhibited clearly distinctive time courses according to the chemical structures of the synthetic luciferins and the kinds of marine luciferases. The blue- and red-marked lines indicate the intensity profiles of RLuc8.6-535 and ALuc16 by time, respectively. Greatly enhancing features in the optical intensity by time were observed with ALuc16. Abbreviations: GLuc, Gaussia princeps luciferase; RLuc8.6-535, Renilla reniformis luciferase 8.6-535; ALuc16, artificial luciferase 16; CLuc, Cypridina luciferase; FLuc, firefly luciferase; CBgreen, click beetle luciferase green.
    Figure Legend Snippet: Biased selectivity of the synthetic luciferins to marine luciferases. ( a ) The relative selectivity of representative synthetic luciferins according to luciferases in cell lysates. ( b ) The corresponding live-cell images on a 6-channel microslide, demonstrating the relative luciferase-selectivity of the selected coelenterazine (CTZ) analogues in living mammalian cells. The optical intensity ranges of 6piOH-2H-CTZ, 6etOH-CTZ, and 6piOH-CTZ were 1024–2261, 512–2214, 512–2841 RLU, respectively. ( c ) The time course of the optical intensities after substrate injection to the microslide channels growing live cells. The bioluminescence exhibited clearly distinctive time courses according to the chemical structures of the synthetic luciferins and the kinds of marine luciferases. The blue- and red-marked lines indicate the intensity profiles of RLuc8.6-535 and ALuc16 by time, respectively. Greatly enhancing features in the optical intensity by time were observed with ALuc16. Abbreviations: GLuc, Gaussia princeps luciferase; RLuc8.6-535, Renilla reniformis luciferase 8.6-535; ALuc16, artificial luciferase 16; CLuc, Cypridina luciferase; FLuc, firefly luciferase; CBgreen, click beetle luciferase green.

    Techniques Used: Luciferase, Injection

    34) Product Images from "Immunomodulatory Polysaccharide from Chlorophytum borivilianum Roots"

    Article Title: Immunomodulatory Polysaccharide from Chlorophytum borivilianum Roots

    Journal: Evidence-based Complementary and Alternative Medicine : eCAM

    doi: 10.1093/ecam/neq012

    IC 50 values of C. borivilianum aqueous extract and fractions against P388 cells.
    Figure Legend Snippet: IC 50 values of C. borivilianum aqueous extract and fractions against P388 cells.

    Techniques Used:

    35) Product Images from "Lipid-like Nanoparticles for Small Interfering RNA Delivery to Endothelial Cells"

    Article Title: Lipid-like Nanoparticles for Small Interfering RNA Delivery to Endothelial Cells

    Journal: Advanced functional materials

    doi: 10.1002/adfm.200900519

    Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and serum-deprived (endothelial basal medium-2 (EBM-2) with no serum and growth factors) condition. (A) RT-PCR for SHP-1, KDR/Flk-1, and eNOS of siRNA-transfected HUVECs at
    Figure Legend Snippet: Gene expression profiles in siRNA-transfected HUVECs under hypoxic (1% oxygen) and serum-deprived (endothelial basal medium-2 (EBM-2) with no serum and growth factors) condition. (A) RT-PCR for SHP-1, KDR/Flk-1, and eNOS of siRNA-transfected HUVECs at

    Techniques Used: Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction

    Screening of lipidoids for siRNA delivery to HUVECs. (A) GAPDH activity of HUVECs (n=4) at 2 days after siGAPDH transfection. The % reduction in GAPDH activity following transfection with siRNA-lipidoid complexes at various lipidoid/siRNA weight ratios
    Figure Legend Snippet: Screening of lipidoids for siRNA delivery to HUVECs. (A) GAPDH activity of HUVECs (n=4) at 2 days after siGAPDH transfection. The % reduction in GAPDH activity following transfection with siRNA-lipidoid complexes at various lipidoid/siRNA weight ratios

    Techniques Used: Activity Assay, Transfection

    Silencing of SHP-1 by siSHP-1 delivery in HUVECs. RT-PCR and quantitative real-time PCR (n=3) to examine SHP-1 expression in HUVECs at 2 days after siRNA transfection using NA114 and Lipofectamine 2000 under culture condition with (A) 2% FBS and (B) 10%
    Figure Legend Snippet: Silencing of SHP-1 by siSHP-1 delivery in HUVECs. RT-PCR and quantitative real-time PCR (n=3) to examine SHP-1 expression in HUVECs at 2 days after siRNA transfection using NA114 and Lipofectamine 2000 under culture condition with (A) 2% FBS and (B) 10%

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Expressing, Transfection

    Inhibition of apoptotic activity in siSHP-1-transfected HUVECs cultured under hypoxic (1% oxygen) and serum-deprived (EBM-2 with no serum and growth factors) condition. (A) TUNEL staining of siRNA-transfected HUVECs and capillary formation (arrows) by
    Figure Legend Snippet: Inhibition of apoptotic activity in siSHP-1-transfected HUVECs cultured under hypoxic (1% oxygen) and serum-deprived (EBM-2 with no serum and growth factors) condition. (A) TUNEL staining of siRNA-transfected HUVECs and capillary formation (arrows) by

    Techniques Used: Inhibition, Activity Assay, Transfection, Cell Culture, TUNEL Assay, Staining

    36) Product Images from "Voltage-gated ion channels mediate the electrotaxis of glioblastoma cells in a hybrid PMMA/PDMS microdevice"

    Article Title: Voltage-gated ion channels mediate the electrotaxis of glioblastoma cells in a hybrid PMMA/PDMS microdevice

    Journal: APL Bioengineering

    doi: 10.1063/5.0004893

    The results of U-251MG cell seeding inside microchannels by various methods. [Left column: (a), (d), (g), and (j)]. In the tip loading method, cells are introduced by using gravitational flow with micropipette tips. The cells can flocculate inside the tips and in microchannels as illustrated in (d). The microscopy image of seeded cells is shown in (g) and magnified in (j); [Middle column: (b), (e), (h), and (k)] in the tip injection method, cells are injected into the channels and tips are removed. The small hydrostatic pressure differences between the inlet/outlet (shown as Δh) will contribute to hydrodynamic flow and disturb the cell distribution, causing non-uniform cell distribution and aggregates as shown in (e). The microscopy image of seeded cells is shown in (h) and magnified in (k); [Right column: (c), (f), (i), and (l)] in our pressure-balanced submerged cell seeding method, the hydrostatic pressure difference is eliminated. The injected cells remain uniform throughout the channel as shown in (f). The microscopy image of seeded cells is shown in (i) and magnified in (l). The uniform and sparse cell seeding method is suitable for different applications such as single cell tracking, ensembled cell studies, and cell assembly. The scale bars in [(g), (h), and (i)] represent 500 μ m. The scale bars in [(j), (k), and (l)] represent 200 μ m.
    Figure Legend Snippet: The results of U-251MG cell seeding inside microchannels by various methods. [Left column: (a), (d), (g), and (j)]. In the tip loading method, cells are introduced by using gravitational flow with micropipette tips. The cells can flocculate inside the tips and in microchannels as illustrated in (d). The microscopy image of seeded cells is shown in (g) and magnified in (j); [Middle column: (b), (e), (h), and (k)] in the tip injection method, cells are injected into the channels and tips are removed. The small hydrostatic pressure differences between the inlet/outlet (shown as Δh) will contribute to hydrodynamic flow and disturb the cell distribution, causing non-uniform cell distribution and aggregates as shown in (e). The microscopy image of seeded cells is shown in (h) and magnified in (k); [Right column: (c), (f), (i), and (l)] in our pressure-balanced submerged cell seeding method, the hydrostatic pressure difference is eliminated. The injected cells remain uniform throughout the channel as shown in (f). The microscopy image of seeded cells is shown in (i) and magnified in (l). The uniform and sparse cell seeding method is suitable for different applications such as single cell tracking, ensembled cell studies, and cell assembly. The scale bars in [(g), (h), and (i)] represent 500 μ m. The scale bars in [(j), (k), and (l)] represent 200 μ m.

    Techniques Used: Microscopy, Injection, Single Cell Tracking

    Inhibitory effects of selected ion channels on glioblastoma cell electrotaxis. (a) The electrotactic directedness of T98G and U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 h with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; ‡ indicates the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; * indicates P
    Figure Legend Snippet: Inhibitory effects of selected ion channels on glioblastoma cell electrotaxis. (a) The electrotactic directedness of T98G and U-251MG glioblastoma cells under 300 V m −1 dcEF after 6 h with pharmacological inhibition on various ion channels. † indicates the electrotaxis tested against those without EF stimulation; ‡ indicates the electrotaxis group tested against those with cytochrome C which prevents adsorption of short peptides to experimental apparatus; All other groups are statistically compared to their respective controls in cell culture media with 10% FBS; n.s. indicates not significant; * indicates P

    Techniques Used: Inhibition, Adsorption, Cell Culture

    The electrotaxis and electro-alignment of T98G and U-251MG glioblastoma cells in HMEFC. (a) The electrotaxis directedness of T98G cells in four different electric field strengths (EFSs); (b) the electrotaxis directedness of U-251MG cells in four different EFSs; [(c) and (d)] The rose plots showing the frequency of T98G (c) and U-251MG (d) electrotaxis with and without electric field. The r-vectors displaying the direction tendencies of each group are indicated by arrows; P values in Mardia–Watson–Wheeler tests are shown; (e) phase contrast images of T98G and (f) U-251MG cells before and after 6 h under 300 V m −1 stimulation; the electric field is applied from left to right. Only T98G cells demonstrate prominent perpendicular alignment under the electrical stimulation.
    Figure Legend Snippet: The electrotaxis and electro-alignment of T98G and U-251MG glioblastoma cells in HMEFC. (a) The electrotaxis directedness of T98G cells in four different electric field strengths (EFSs); (b) the electrotaxis directedness of U-251MG cells in four different EFSs; [(c) and (d)] The rose plots showing the frequency of T98G (c) and U-251MG (d) electrotaxis with and without electric field. The r-vectors displaying the direction tendencies of each group are indicated by arrows; P values in Mardia–Watson–Wheeler tests are shown; (e) phase contrast images of T98G and (f) U-251MG cells before and after 6 h under 300 V m −1 stimulation; the electric field is applied from left to right. Only T98G cells demonstrate prominent perpendicular alignment under the electrical stimulation.

    Techniques Used:

    37) Product Images from "Peptidoglycan ld-Carboxypeptidase Pgp2 Influences Campylobacter jejuni Helical Cell Shape and Pathogenic Properties and Provides the Substrate for the dl-Carboxypeptidase Pgp1 *"

    Article Title: Peptidoglycan ld-Carboxypeptidase Pgp2 Influences Campylobacter jejuni Helical Cell Shape and Pathogenic Properties and Provides the Substrate for the dl-Carboxypeptidase Pgp1 *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M113.491829

    The effect of pgp2 deletion on host-related phenotypes. A and B , PG isolated from the Δ pgp2 mutant shows reduced human Nod1 activation and no change in human Nod2 stimulation. To assay the change in the ability of C. jejuni Δ pgp2 PG to activate Nod proteins, human embryonic kidney cells (HEK293T) were co-transfected with either 0.2 or 2 μg of C. jejuni 81-176, Δ pgp2 , or Δ pgp2c PG, vectors for a nuclear factor-κB (NF-κB) luciferase reporter, and either Nod1 ( A ) or Nod2 ( B ). Nod activation was determined by measuring the activity of a NF-κB luciferase reporter in comparison with the non-stimulated ( NS ) negative control. Positive controls used were tripeptide l -Ala-γ- d -Glu- meso -DAP ( TriDAP ) and MDP. Data represent the mean ± S.E. of four independent experiments. C , invasion and intracellular survival ability of the Δ pgp2 strain in the INT407 epithelial cell line was assessed by a gentamicin ( Gm ) protection assay and showed no change in comparison with wild type. Gentamicin was added 3 h postinfection with the C. jejuni wild-type 81-176 and Δ pgp2 strains. After 2 h, the gentamicin was washed off, and the cells were incubated with fresh MEM containing 3% FBS and a low dose of gentamicin. cfu were determined for each well by lysing the cells with water and plating the dilutions onto MH-trimethoprim-vancomycin plates. Data represent the mean ± S.E. of three independent experiments. D , pgp2 deletion has no effect on IL-8 secretion in the INT407 epithelial cell line. ELISA was used to quantify IL-8 levels secreted by uninfected INT407 epithelial cell lines and cells infected for 8 and 24 h with C. jejuni wild-type 81-176, Δ pgp2, and Δ pgp2 c strains. Data represent the mean ± S.E. ( error bars ) of three independent experiments. E , the Δ pgp2 mutant strain is defective for chick colonization. Each point represents the log cfu/g cecal contents of an individual chick 7 days postcolonization with 10 4 cfu of the indicated C. jejuni strains. The geometric mean is denoted by a black bar . *, statistically significant difference using the unpaired Student's t test, with *, **, and *** indicating p
    Figure Legend Snippet: The effect of pgp2 deletion on host-related phenotypes. A and B , PG isolated from the Δ pgp2 mutant shows reduced human Nod1 activation and no change in human Nod2 stimulation. To assay the change in the ability of C. jejuni Δ pgp2 PG to activate Nod proteins, human embryonic kidney cells (HEK293T) were co-transfected with either 0.2 or 2 μg of C. jejuni 81-176, Δ pgp2 , or Δ pgp2c PG, vectors for a nuclear factor-κB (NF-κB) luciferase reporter, and either Nod1 ( A ) or Nod2 ( B ). Nod activation was determined by measuring the activity of a NF-κB luciferase reporter in comparison with the non-stimulated ( NS ) negative control. Positive controls used were tripeptide l -Ala-γ- d -Glu- meso -DAP ( TriDAP ) and MDP. Data represent the mean ± S.E. of four independent experiments. C , invasion and intracellular survival ability of the Δ pgp2 strain in the INT407 epithelial cell line was assessed by a gentamicin ( Gm ) protection assay and showed no change in comparison with wild type. Gentamicin was added 3 h postinfection with the C. jejuni wild-type 81-176 and Δ pgp2 strains. After 2 h, the gentamicin was washed off, and the cells were incubated with fresh MEM containing 3% FBS and a low dose of gentamicin. cfu were determined for each well by lysing the cells with water and plating the dilutions onto MH-trimethoprim-vancomycin plates. Data represent the mean ± S.E. of three independent experiments. D , pgp2 deletion has no effect on IL-8 secretion in the INT407 epithelial cell line. ELISA was used to quantify IL-8 levels secreted by uninfected INT407 epithelial cell lines and cells infected for 8 and 24 h with C. jejuni wild-type 81-176, Δ pgp2, and Δ pgp2 c strains. Data represent the mean ± S.E. ( error bars ) of three independent experiments. E , the Δ pgp2 mutant strain is defective for chick colonization. Each point represents the log cfu/g cecal contents of an individual chick 7 days postcolonization with 10 4 cfu of the indicated C. jejuni strains. The geometric mean is denoted by a black bar . *, statistically significant difference using the unpaired Student's t test, with *, **, and *** indicating p

    Techniques Used: Isolation, Mutagenesis, Activation Assay, Transfection, Luciferase, Activity Assay, Negative Control, Incubation, Enzyme-linked Immunosorbent Assay, Infection

    38) Product Images from "Isolation, characterization, and in silico, in vitro and in vivo antiulcer studies of isoimperatorin crystallized from Ostericum koreanum"

    Article Title: Isolation, characterization, and in silico, in vitro and in vivo antiulcer studies of isoimperatorin crystallized from Ostericum koreanum

    Journal: Pharmaceutical Biology

    doi: 10.1080/13880209.2016.1257641

    Ortep view of the crystal structure of isoimperatorin.
    Figure Legend Snippet: Ortep view of the crystal structure of isoimperatorin.

    Techniques Used:

    Interactions between isoimperatorin and Jack bean urease (PDB ID 3LA4) at the active site, generated using Discovery studio 2.1.0. The light silver colour shows the backbone of the urease protein in solid ribbon format. Carbon and oxygen atoms of the ligand molecule are shown in green and red, respectively. The interacting residues are shown in blue, while the dotted lines indicate the binding distances (Å).
    Figure Legend Snippet: Interactions between isoimperatorin and Jack bean urease (PDB ID 3LA4) at the active site, generated using Discovery studio 2.1.0. The light silver colour shows the backbone of the urease protein in solid ribbon format. Carbon and oxygen atoms of the ligand molecule are shown in green and red, respectively. The interacting residues are shown in blue, while the dotted lines indicate the binding distances (Å).

    Techniques Used: Generated, Binding Assay

    Effect of isoimperatorin treatment on the viability of rabbit articular chondrocytes. Chondrocytes were treated with the indicated concentrations (0.0–737.74 μM) of isoimperatorin for 24 h. Photographs of chondrocytes (A) were taken with a phase-contrast microscope (200× magnification). Cell viability was estimated using the methyl thiazole tetrazolium (MTT) assay. The graphical data (B) are expressed as mean ± standard error of mean.
    Figure Legend Snippet: Effect of isoimperatorin treatment on the viability of rabbit articular chondrocytes. Chondrocytes were treated with the indicated concentrations (0.0–737.74 μM) of isoimperatorin for 24 h. Photographs of chondrocytes (A) were taken with a phase-contrast microscope (200× magnification). Cell viability was estimated using the methyl thiazole tetrazolium (MTT) assay. The graphical data (B) are expressed as mean ± standard error of mean.

    Techniques Used: Microscopy, MTT Assay

    Chemical structure of isoimperatorin.
    Figure Legend Snippet: Chemical structure of isoimperatorin.

    Techniques Used:

    Effect of isoimperatorin on ulcer index for ulcers induced by ethanol, indomethacin, or pylorus ligation. Numbers indicate the % protection with respect to the control groups of different ulcer models. Bars represent the mean ± SEM of each group ( n = 5). One-way analysis of variance (ANOVA) was employed and significant differences between control groups and groups treated with isoimperatorin are represented by * p
    Figure Legend Snippet: Effect of isoimperatorin on ulcer index for ulcers induced by ethanol, indomethacin, or pylorus ligation. Numbers indicate the % protection with respect to the control groups of different ulcer models. Bars represent the mean ± SEM of each group ( n = 5). One-way analysis of variance (ANOVA) was employed and significant differences between control groups and groups treated with isoimperatorin are represented by * p

    Techniques Used: Ligation

    Effect of isoimperatorin on the dose-dependent expression of type II collagen in primary chondrocytes. After the treatment of chondrocytes with indicated concentrations of isoimperatorin for 24 h, (A) type II collagen expression was detected using Western blot analysis. (B) Isoimperatorin increased the expression of type II collagen in primary chondrocytes in the presence or absence of IL-1β. GAPDH was used as the loading control.
    Figure Legend Snippet: Effect of isoimperatorin on the dose-dependent expression of type II collagen in primary chondrocytes. After the treatment of chondrocytes with indicated concentrations of isoimperatorin for 24 h, (A) type II collagen expression was detected using Western blot analysis. (B) Isoimperatorin increased the expression of type II collagen in primary chondrocytes in the presence or absence of IL-1β. GAPDH was used as the loading control.

    Techniques Used: Expressing, Western Blot

    39) Product Images from "Puerarin attenuates the inflammatory response and apoptosis in LPS-stimulated cardiomyocytes"

    Article Title: Puerarin attenuates the inflammatory response and apoptosis in LPS-stimulated cardiomyocytes

    Journal: Experimental and Therapeutic Medicine

    doi: 10.3892/etm.2015.2910

    Effect of puerarin on mRNA expression levels of inflammatory mediators (A) IL-1β and (B) TNF-α in H9c2 cardiomyocytes. Puerarin decreased the LPS (1 µg/ml)-induced expression of IL-1β and TNF-α in a concentration-dependent manner. *P
    Figure Legend Snippet: Effect of puerarin on mRNA expression levels of inflammatory mediators (A) IL-1β and (B) TNF-α in H9c2 cardiomyocytes. Puerarin decreased the LPS (1 µg/ml)-induced expression of IL-1β and TNF-α in a concentration-dependent manner. *P

    Techniques Used: Expressing, Concentration Assay

    Effect of puerarin on apoptosis. (A and B) TUNEL staining results revealed that Pue (40 µM) attenuated apoptosis in H9c2 cardiomyocytes following stimulation with LPS (1 µg/ml) for 48 h. (A) Electron microscopy of H9c2 cardiomyocytes (original magnification, ×400). (B) Quantitative results of TUNEL staining. *P
    Figure Legend Snippet: Effect of puerarin on apoptosis. (A and B) TUNEL staining results revealed that Pue (40 µM) attenuated apoptosis in H9c2 cardiomyocytes following stimulation with LPS (1 µg/ml) for 48 h. (A) Electron microscopy of H9c2 cardiomyocytes (original magnification, ×400). (B) Quantitative results of TUNEL staining. *P

    Techniques Used: TUNEL Assay, Staining, Electron Microscopy

    40) Product Images from "Cartilage storage at 4 °C with regular culture medium replacement benefits chondrocyte viability of osteochondral grafts in vitro"

    Article Title: Cartilage storage at 4 °C with regular culture medium replacement benefits chondrocyte viability of osteochondral grafts in vitro

    Journal: Cell and Tissue Banking

    doi: 10.1007/s10561-016-9556-7

    EB-FDA fluorescence staining of chondrocytes at days 7, 21 and 35 in different storage conditions. Chondrocytes emitting green (alive) and red (dead) fluorescence are shown at each time point. Blue dots represent impurities. At day 7, dead cell numbers were low in all groups. Viable chondrocyte numbers were significantly higher in Group A1 compared with the other three groups. On day 21, there were more dead cells than on day 7 for all groups. On day 35, dead cell numbers continued to increase in all groups, with the highest numbers in Group B1 and Group B2. (Color figure online)
    Figure Legend Snippet: EB-FDA fluorescence staining of chondrocytes at days 7, 21 and 35 in different storage conditions. Chondrocytes emitting green (alive) and red (dead) fluorescence are shown at each time point. Blue dots represent impurities. At day 7, dead cell numbers were low in all groups. Viable chondrocyte numbers were significantly higher in Group A1 compared with the other three groups. On day 21, there were more dead cells than on day 7 for all groups. On day 35, dead cell numbers continued to increase in all groups, with the highest numbers in Group B1 and Group B2. (Color figure online)

    Techniques Used: Fluorescence, Staining

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    Article Snippet: .. Cell culture and transfection HEK293T cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) at 37 °C in a humidified atmosphere containing 5% CO2 (SANYO, MCO-175, Osaka prefecture, Japan). .. For cell transfection, HEK293T cells were seeded into 24-well plates at a density of 2 × 104 cells/well.

    Article Title: Decanal Protects against UVB-Induced Photoaging in Human Dermal Fibroblasts via the cAMP Pathway
    Article Snippet: .. Cell Culture Human foreskin fibroblast Hs68 cells were incubated in high-glucose Dulbecco’s modified Eagle medium supplemented with 10% (v/v ) fetal bovine serum and 1% 100 U/mL penicillin-streptomycin at 37 °C in a 5% CO2 incubator (Sanyo, Osaka, Japan). .. UV Irradiation We used the UV cross-linker (Model CL-1000M; UVP, Upland, CA, USA) for UVB (280–320 nm) irradiation.

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  • 93
    Sanyo co2 incubator
    Co2 Incubator, supplied by Sanyo, used in various techniques. Bioz Stars score: 93/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/co2 incubator/product/Sanyo
    Average 93 stars, based on 95 article reviews
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    co2 incubator - by Bioz Stars, 2020-08
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