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  • 94
    Thermo Fisher wortmannin
    TGF-β1 induces HMGA1 expression through PI3K signaling and ERK signaling in SW579 cells. (A) The effects of <t>wortmannin,</t> PD98059 and U0126 on the mRNA expression of HMGA1 induced by treatment with 5 ng/ml of TGF-β1 in SW579 cells. (B and C) The effects of wortmannin, PD98059 and U0126 on the expression of HMGA1 induced by treatment 5 ng/ml with TGF-β1 in SW579 cells. Fluorescence were gathered and analyzed with a fluorescence microscope (Olympus). * P
    Wortmannin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 148 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Thermo Fisher
    Average 94 stars, based on 148 article reviews
    Price from $9.99 to $1999.99
    wortmannin - by Bioz Stars, 2020-05
    94/100 stars
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    94
    Abcam wortmannin
    Microtopography and distribution of actin, PIP2, and PIP3 (A) Fibroblasts grown with microrod (between dashed lines) show strong focal adhesion formation (white arrow). (B, C) Fibroblasts on microtopography (PDMS, 400 kPa), show cells attached to micropost (top indicated by white line). Actin, red; nucleus, blue; PIP3, or PIP2 (B, C respectively), green (bar = 15 µm). PIP3 pattern is diffuse to cell periphery but PIP2 shows increased localization around microposts. (D) For actin dynamics, a stress fiber region of interest (red box) was bleached to approximately 40% initial intensity by a 488 nm laser at high power and observed during recovery in cells grown on flat 10 kPa, 400 kPa, or glass surfaces, microtopography, or on glass with neomycin or <t>wortmannin</t> treatment (bar = 15 µm). (E) Time course of recovery of fluorescence intensity determined kfrap kinetic constant was not significantly difference on different stiffnesses, microtopography, or with drug treatment. FRAP experiments in different conditions were conducted on cells from at least 3 separate cultures. Means ± SE.
    Wortmannin, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Abcam
    Average 94 stars, based on 33 article reviews
    Price from $9.99 to $1999.99
    wortmannin - by Bioz Stars, 2020-05
    94/100 stars
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    96
    Cell Signaling Technology Inc wortmannin
    PTEN -loss results in an increased cholesterol de novo synthesis. a Relative accumulation of Srebp2 , Hmgcr , Sqs and Hmgcs1 transcripts in prostate of wild type (WT) and Pten pc −/− mice. ( N = 10 per group). b Accumulation of cholesterol precursors, lanosterol, lathosterol, desmosterol and cholesterol in prostatic samples from wild type (WT) ( N = 4) and Pten pc −/− mice ( N = 5). c qPCR analysis of HMGCR , ABCA1 and FASN expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with T0901317 supplementation (1 µM). Experiments have been performed in three experimental replicates. d qPCR analysis of ABCA1 and HMGCR expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with mevalonate (500 µM). e qPCR analysis of HMGCR , ABCA1 and FASN expression in PC3 cells treated with LY294002 (20 µM) alone or with increasing amounts of mevalonate (100 µM, 500 µM, 1 mM, 10 mM). f Western blot analysis of SREBP2 cleaved form, HMGCR and SQLE respective accumulation in LNCaP cell line. AKTS473 is used to confirm <t>Wortmannin</t> and LY294002 treatment efficiency and β-ACTIN as a loading control. g Western blot analysis of HMGCR, SQLE and APOE in PC3 cells treated with LY294002 (20 µM) alone or in combination with mevalonate (500 µM). AKTS473 and AKT are used to confirm Wortmannin and LY294002 treatment efficiency and β-ACTIN as a loading control. h SREBP2 and LXR target genes expression in PC3 cells transfected with siSREBP2 or siGFP as control. Experiments have been performed in three experimental replicates. i Relative expression of Hmgcr , Abca1 and Fasn on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose) or Simvastatin (40 mg kg −1 ). j Immunofluorescence detection of FASN and ABCA1 on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose, N = 6) or Simvastatin (40 mg kg −1 , N = 6). Nuclei are stained using Hoescht ( blue ), scale bar 100 µm. k qPCR analysis of Abca1 and Fasn expression on MPEC Pten −/− or Pten −/− lxrαβ −/− treated with simvastatin alone (2,5 µM) or in combination with T0901317 (1 µM), N = 3 per group. All data are represented as mean ± SEM and statistical analyses were performed with the Student’s t -test; * p
    Wortmannin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 573 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Cell Signaling Technology Inc
    Average 96 stars, based on 573 article reviews
    Price from $9.99 to $1999.99
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    92
    Millipore wortmannin
    Level of plasma interleukin-6 and myeloperoxidase activity in the pancreas. A: Plasma interleukin (IL)-6 level after treatment with <t>wortmannin;</t> B: Myeloperoxidase (MPO) activity in pancreatic tissue after treatment with wortmannin. c P
    Wortmannin, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 7667 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Millipore
    Average 92 stars, based on 7667 article reviews
    Price from $9.99 to $1999.99
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    99
    Selleck Chemicals wortmannin
    Analysis of SOD activities (A), MDA contents (B) in the renal tissue, as well as plasma IL-10 (C) and TNF-α levels (D) in each groups. Electroacupuncture at bilaterally Zusanli and Neiguan acupoints significantly increased SOD activities, decreased MDA contents accompanied with increased levels of IL-10 and decreased TNF-α levels. Pretreatment with <t>wortmannin</t> significantly restrained the above effects of electroacupuncture in group ELW. Values were presented as mean ± SD (n = 10). The data were analyzed using one-way ANOVA and the Bonferroni test for multiple comparisons. Significant differences were indicated with an asterisk: *P
    Wortmannin, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 99/100, based on 191 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Selleck Chemicals
    Average 99 stars, based on 191 article reviews
    Price from $9.99 to $1999.99
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    95
    Tocris wortmannin
    Effects of the inhibitors on class I PI3K/Akt/mTOR axis signaling in canine C2 cells. Cells were treated with pan-class I PI3K inhibitor <t>Wortmannin</t> (W) at 1 μM and ZSTK474 (Z) at 1 μM, mTOR inhibitor Rapamycin (R) at 100 nM ( A ) and Akt inhibitor KP372-1 at 0, 150, 200 and 400 nM ( B ) for the indicated period of time. Whole cell lysates (comprising 50 μg total protein) were subjected to western blot with the indicated antibodies. β-actin was used as a loading control. N/A indicates data unavailable due to induction of apoptosis in all cells.
    Wortmannin, supplied by Tocris, used in various techniques. Bioz Stars score: 95/100, based on 220 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/Tocris
    Average 95 stars, based on 220 article reviews
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    85
    LKT Laboratories wortmannin
    Inhibition of Nrf2 activity by PI3K inhibitors. ARPE cells were treated with 10 μ M LY294002 for 16 hours ( A ) or <t>wortmannin</t> for 6 hours ( B ) at the indicated concentrations. The Nrf2 activity was measured by transient transfection with an ARE reporter
    Wortmannin, supplied by LKT Laboratories, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/wortmannin/product/LKT Laboratories
    Average 85 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    Image Search Results


    TGF-β1 induces HMGA1 expression through PI3K signaling and ERK signaling in SW579 cells. (A) The effects of wortmannin, PD98059 and U0126 on the mRNA expression of HMGA1 induced by treatment with 5 ng/ml of TGF-β1 in SW579 cells. (B and C) The effects of wortmannin, PD98059 and U0126 on the expression of HMGA1 induced by treatment 5 ng/ml with TGF-β1 in SW579 cells. Fluorescence were gathered and analyzed with a fluorescence microscope (Olympus). * P

    Journal: International Journal of Oncology

    Article Title: TGF-β1 induces HMGA1 expression: The role of HMGA1 in thyroid cancer proliferation and invasion

    doi: 10.3892/ijo.2017.3958

    Figure Lengend Snippet: TGF-β1 induces HMGA1 expression through PI3K signaling and ERK signaling in SW579 cells. (A) The effects of wortmannin, PD98059 and U0126 on the mRNA expression of HMGA1 induced by treatment with 5 ng/ml of TGF-β1 in SW579 cells. (B and C) The effects of wortmannin, PD98059 and U0126 on the expression of HMGA1 induced by treatment 5 ng/ml with TGF-β1 in SW579 cells. Fluorescence were gathered and analyzed with a fluorescence microscope (Olympus). * P

    Article Snippet: RNA isolation and real-time RT-PCR The SW579 were treated with TGF β1 (0, 1, 2, 5 and 10 ng/ml) for 12 h or wortmannin (100 nM), PD98059 (20 µ M) and U0126 (25 µ M), and maintained in culture medium for 2 h. Total RNA of SW579 cells was extracted using TRIzol reagent (Invitrogen) and was reverse transcribed into cDNA using the first-strand synthesis kit (Gibco-BRL, Carlsbad, CA, USA).

    Techniques: Expressing, Fluorescence, Microscopy

    TGF-β1 enhances the promoter activity of HMGA1 in SW579 cells. (A) The effects of TGF-β1 on HMGA1 promoter activity. HMGA1 promoter, control vectors or pGL4.1 vectors were transfected into SW579 cells, and treated with the indicated concentrations of TGF-β1. (B) The effects of 5 ng/ml TGF-β1 on HMGA1 promoter activity. HMGA1 promoter, control vectors or pGL4.1 vectors were transfected into SW579 cells, and treated with 5 ng/ml TGF-β1 for the indicated time. (C) The effects of 5 ng/ml TGF-β1 on HMGA1 promoter activity with wortmannin (100 nM), PD98059 (20 µ M) or U0126 (25 µ M) in SW579 cells, * P

    Journal: International Journal of Oncology

    Article Title: TGF-β1 induces HMGA1 expression: The role of HMGA1 in thyroid cancer proliferation and invasion

    doi: 10.3892/ijo.2017.3958

    Figure Lengend Snippet: TGF-β1 enhances the promoter activity of HMGA1 in SW579 cells. (A) The effects of TGF-β1 on HMGA1 promoter activity. HMGA1 promoter, control vectors or pGL4.1 vectors were transfected into SW579 cells, and treated with the indicated concentrations of TGF-β1. (B) The effects of 5 ng/ml TGF-β1 on HMGA1 promoter activity. HMGA1 promoter, control vectors or pGL4.1 vectors were transfected into SW579 cells, and treated with 5 ng/ml TGF-β1 for the indicated time. (C) The effects of 5 ng/ml TGF-β1 on HMGA1 promoter activity with wortmannin (100 nM), PD98059 (20 µ M) or U0126 (25 µ M) in SW579 cells, * P

    Article Snippet: RNA isolation and real-time RT-PCR The SW579 were treated with TGF β1 (0, 1, 2, 5 and 10 ng/ml) for 12 h or wortmannin (100 nM), PD98059 (20 µ M) and U0126 (25 µ M), and maintained in culture medium for 2 h. Total RNA of SW579 cells was extracted using TRIzol reagent (Invitrogen) and was reverse transcribed into cDNA using the first-strand synthesis kit (Gibco-BRL, Carlsbad, CA, USA).

    Techniques: Activity Assay, Transfection

    ERK1/2 and AKT signaling pathways are involved in the upregulation of VEGF expression induced by IGF1. The upregulation of HIF-1α and GPER protein expression observed treating CAFs ( a ) and SKBR3 cells ( b ) with 100 ng/mL IGF1 for 8 hours is abolished in the presence of 10 μM IGF1R inhibitor AG1024 (AG), 10 μM MEK inhibitor PD98059 (PD) and 100 nM PI3K inhibitor Wortmannin (WM). The activation of ERK1/2 and AKT (Ser 473) is prevented in CAFs ( c , e ) and SKBR3 cells ( d , f ) treated for 60 minutes with 100 ng/mL IGF1, alone and in combination with AG (10 μM), PD (10 μM) and WM (100 nM). ERK2, AKT and β-actin serve as loading control, as indicated. g VEGF protein expression in CAFs treated with 100 ng/mL IGF1 for 8 hours, alone and in combination with AG (10 μM), PD (10 μM) and WM (100 nM), as evidenced by immunfluoerscence experiment. VEGF accumulation is shown by the green signal , nuclei are stained by DAPI ( blue signal ), bar scale 100 μM. Results shown are representative of two independent experiments. Evaluation of luciferase activity in SKBR3 cells infected with a HRE reporter construct (SKBR3-HRE-luc) ( h ), and in SKBR3 cells transiently transfected with a GPER (pGPER) ( i ) or a VEGF (pVEGF) promoter construct ( j ) and treated with 100 ng/mL IGF1 for 18 h in the presence of AG, PD and WM. Luciferase activity was normalized to the internal transfection control. Results are expressed as the % change of normalized RLU values relative to vehicle-treated cells. Each data point represents the mean ± SEM of two independent experiments performed in triplicate. (**) p

    Journal: Breast Cancer Research : BCR

    Article Title: GPER mediates the angiocrine actions induced by IGF1 through the HIF-1α/VEGF pathway in the breast tumor microenvironment

    doi: 10.1186/s13058-017-0923-5

    Figure Lengend Snippet: ERK1/2 and AKT signaling pathways are involved in the upregulation of VEGF expression induced by IGF1. The upregulation of HIF-1α and GPER protein expression observed treating CAFs ( a ) and SKBR3 cells ( b ) with 100 ng/mL IGF1 for 8 hours is abolished in the presence of 10 μM IGF1R inhibitor AG1024 (AG), 10 μM MEK inhibitor PD98059 (PD) and 100 nM PI3K inhibitor Wortmannin (WM). The activation of ERK1/2 and AKT (Ser 473) is prevented in CAFs ( c , e ) and SKBR3 cells ( d , f ) treated for 60 minutes with 100 ng/mL IGF1, alone and in combination with AG (10 μM), PD (10 μM) and WM (100 nM). ERK2, AKT and β-actin serve as loading control, as indicated. g VEGF protein expression in CAFs treated with 100 ng/mL IGF1 for 8 hours, alone and in combination with AG (10 μM), PD (10 μM) and WM (100 nM), as evidenced by immunfluoerscence experiment. VEGF accumulation is shown by the green signal , nuclei are stained by DAPI ( blue signal ), bar scale 100 μM. Results shown are representative of two independent experiments. Evaluation of luciferase activity in SKBR3 cells infected with a HRE reporter construct (SKBR3-HRE-luc) ( h ), and in SKBR3 cells transiently transfected with a GPER (pGPER) ( i ) or a VEGF (pVEGF) promoter construct ( j ) and treated with 100 ng/mL IGF1 for 18 h in the presence of AG, PD and WM. Luciferase activity was normalized to the internal transfection control. Results are expressed as the % change of normalized RLU values relative to vehicle-treated cells. Each data point represents the mean ± SEM of two independent experiments performed in triplicate. (**) p

    Article Snippet: Wortmannin (WM) was acquired from Life Technologies Ltd. SU5416 was supplied by Generon Ltd. Human VEGF was purchased from Peprotech EC: Limited.

    Techniques: Expressing, Activation Assay, Staining, Luciferase, Activity Assay, Infection, Construct, Transfection

    A. ERα phosphorylation induced by EGF through the PI3K/MAPK kinase pathway. Representative Western blot showing the phosphorylation status of ERα (ser118 and ser 167), ERK1/2, pAKT and pS6K1 in MCF-7 cells in controls and after treatment for 30 min with EGF. Endogenous ERK1/2, ERα and Δ-actin are the loading controls. The above phosphorylation Status of controls and EGF-induced parameters were also assessed following pretreatment of cells with MEK in inhibitor, U0126 (10μM) or Wortmannin (0.5 μM)(Wort) or both inhibitors for 2 h prior to the addition of EGF. B. Lack of participation of JAK2 in MAPK and PI3K pathways induced by EGF. Western blot showing pER1/2, and pAKT phosphorylation of cells cultured in presence or absence of EGF for 1 h following preincubation with or without JAK2 inhibitor AG-490 for 2 h. Δ actin was used as loading control. C. EGF-induced up-regulation of hPIII transcriptional activity abolished by ERK1/2 and PI3K inhibitors. Cells transfected with hPIII promoter construct or PGL2 vector (control) were pre-incubated with inhibitors U0126 or Wortmanin (Wort) for 2 h prior to addition of buffer in controls or the EGF stimulus and further incubated for 16 h for evaluation of promoter activity. Asterisks (*) indicate Statistically significant changes between EGF untreated and treated groups (Student t -test; P

    Journal: Oncotarget

    Article Title: Role of EGF/ERBB1 in the transcriptional regulation of the prolactin receptor independent of estrogen and prolactin in breast cancer cells

    doi: 10.18632/oncotarget.11579

    Figure Lengend Snippet: A. ERα phosphorylation induced by EGF through the PI3K/MAPK kinase pathway. Representative Western blot showing the phosphorylation status of ERα (ser118 and ser 167), ERK1/2, pAKT and pS6K1 in MCF-7 cells in controls and after treatment for 30 min with EGF. Endogenous ERK1/2, ERα and Δ-actin are the loading controls. The above phosphorylation Status of controls and EGF-induced parameters were also assessed following pretreatment of cells with MEK in inhibitor, U0126 (10μM) or Wortmannin (0.5 μM)(Wort) or both inhibitors for 2 h prior to the addition of EGF. B. Lack of participation of JAK2 in MAPK and PI3K pathways induced by EGF. Western blot showing pER1/2, and pAKT phosphorylation of cells cultured in presence or absence of EGF for 1 h following preincubation with or without JAK2 inhibitor AG-490 for 2 h. Δ actin was used as loading control. C. EGF-induced up-regulation of hPIII transcriptional activity abolished by ERK1/2 and PI3K inhibitors. Cells transfected with hPIII promoter construct or PGL2 vector (control) were pre-incubated with inhibitors U0126 or Wortmanin (Wort) for 2 h prior to addition of buffer in controls or the EGF stimulus and further incubated for 16 h for evaluation of promoter activity. Asterisks (*) indicate Statistically significant changes between EGF untreated and treated groups (Student t -test; P

    Article Snippet: Western blot analysis Whole cell lysates from MCF-7 cells cultured in presence and absence of EGF (100 ng/ml) or treated with either or both U0126 and Wortmannin inhibitors were extracted using RIPA lysis buffer (Thermo Scientific, Rockford, IL) in presence of 1x protease and Phosphatase inhibitor cocktail (Thermo Scientific).

    Techniques: Western Blot, Cell Culture, Activity Assay, Transfection, Construct, Plasmid Preparation, Incubation

    Microtopography and distribution of actin, PIP2, and PIP3 (A) Fibroblasts grown with microrod (between dashed lines) show strong focal adhesion formation (white arrow). (B, C) Fibroblasts on microtopography (PDMS, 400 kPa), show cells attached to micropost (top indicated by white line). Actin, red; nucleus, blue; PIP3, or PIP2 (B, C respectively), green (bar = 15 µm). PIP3 pattern is diffuse to cell periphery but PIP2 shows increased localization around microposts. (D) For actin dynamics, a stress fiber region of interest (red box) was bleached to approximately 40% initial intensity by a 488 nm laser at high power and observed during recovery in cells grown on flat 10 kPa, 400 kPa, or glass surfaces, microtopography, or on glass with neomycin or wortmannin treatment (bar = 15 µm). (E) Time course of recovery of fluorescence intensity determined kfrap kinetic constant was not significantly difference on different stiffnesses, microtopography, or with drug treatment. FRAP experiments in different conditions were conducted on cells from at least 3 separate cultures. Means ± SE.

    Journal: Journal of cellular physiology

    Article Title: Lipid signaling affects primary fibroblast collective migration and anchorage in response to stiffness and microtopography

    doi: 10.1002/jcp.26236

    Figure Lengend Snippet: Microtopography and distribution of actin, PIP2, and PIP3 (A) Fibroblasts grown with microrod (between dashed lines) show strong focal adhesion formation (white arrow). (B, C) Fibroblasts on microtopography (PDMS, 400 kPa), show cells attached to micropost (top indicated by white line). Actin, red; nucleus, blue; PIP3, or PIP2 (B, C respectively), green (bar = 15 µm). PIP3 pattern is diffuse to cell periphery but PIP2 shows increased localization around microposts. (D) For actin dynamics, a stress fiber region of interest (red box) was bleached to approximately 40% initial intensity by a 488 nm laser at high power and observed during recovery in cells grown on flat 10 kPa, 400 kPa, or glass surfaces, microtopography, or on glass with neomycin or wortmannin treatment (bar = 15 µm). (E) Time course of recovery of fluorescence intensity determined kfrap kinetic constant was not significantly difference on different stiffnesses, microtopography, or with drug treatment. FRAP experiments in different conditions were conducted on cells from at least 3 separate cultures. Means ± SE.

    Article Snippet: Fibroblasts grown on 10kPa, 100kPa, 400 kPa, or glass and treated with neomycin or wortmannin were fixed in 10% formalin at 7 hours after barrier removal, probed for focal adhesions with an antibody to phospho-paxillin [Y113] (Abcam, cat. ab32084, Cambridge, MA) in a 1% BSA, 0.1% Tween-20 solution.

    Techniques: Fluorescence

    Focal adhesion and actin stress fibers in leading lamella of migratory fibroblasts with varying stiffness and altered PIP2 level Fibroblasts were grown on PAA substrate of 10 kPa or 100kPa, or glass ( > 1 GPa) stiffness and viewed with a fluorescent microscope. (A) Untreated fibroblasts (left panels), neomycin treatment (central panels), and wortmannin treatment (right panels) have similar cytoskeletal structure with actin stress fibers (red) ending in focal adhesions (green). Actin, red; nucleus, blue; focal adhesion, paxillin, green (bar = 10 µm). (B) Diagram to show sampling method for the distance between the focal adhesion and the lamella membrane by 5 white lines per cell. (C) Histogram with focal adhesion distance from leading edge membrane to show variation with stiffness and drug treatment. Means ± SE; ** P

    Journal: Journal of cellular physiology

    Article Title: Lipid signaling affects primary fibroblast collective migration and anchorage in response to stiffness and microtopography

    doi: 10.1002/jcp.26236

    Figure Lengend Snippet: Focal adhesion and actin stress fibers in leading lamella of migratory fibroblasts with varying stiffness and altered PIP2 level Fibroblasts were grown on PAA substrate of 10 kPa or 100kPa, or glass ( > 1 GPa) stiffness and viewed with a fluorescent microscope. (A) Untreated fibroblasts (left panels), neomycin treatment (central panels), and wortmannin treatment (right panels) have similar cytoskeletal structure with actin stress fibers (red) ending in focal adhesions (green). Actin, red; nucleus, blue; focal adhesion, paxillin, green (bar = 10 µm). (B) Diagram to show sampling method for the distance between the focal adhesion and the lamella membrane by 5 white lines per cell. (C) Histogram with focal adhesion distance from leading edge membrane to show variation with stiffness and drug treatment. Means ± SE; ** P

    Article Snippet: Fibroblasts grown on 10kPa, 100kPa, 400 kPa, or glass and treated with neomycin or wortmannin were fixed in 10% formalin at 7 hours after barrier removal, probed for focal adhesions with an antibody to phospho-paxillin [Y113] (Abcam, cat. ab32084, Cambridge, MA) in a 1% BSA, 0.1% Tween-20 solution.

    Techniques: Microscopy, Sampling

    Localization of actin, lamellipodin, PIP2, and PIP3 with varying stiffness and altered PIP2 level Fibroblasts grown on glass and viewed with a fluorescent microscope shown untreated (left panels), with neomycin treatment (central panels), and with wortmannin treatment (right panels). Actin, red; nucleus, blue; lamellipodin, PIP2, and PIP3 green with their respective antibodies (bar = 10 µm). (A) Lamellipodin is distributed on the leading edge membrane under all conditions. (B) PIP3 antibody shows diffuse pattern with elevation at lamella under all conditions. (C) PIP2 antibody shows localization near the leading edge of the fibroblast. Significantly, neomycin-treated cells show even greater localization at lamellar edge. (D) A PIP2 biosensor (green) transfected living fibroblasts grown on glass and shows a distribution similar to the PIP2 antibody detection. (E, F) Histogram to show the ratio of intensity of PIP3 or PIP2, respectively from ten microns near the leading edge to ten microns in the cell interior. PIP3 shows no difference between treatments. Ratio of lamellar to interior PIP2 is significantly higher in neomycin-treated cells compared to untreated. Means ± SE; * P

    Journal: Journal of cellular physiology

    Article Title: Lipid signaling affects primary fibroblast collective migration and anchorage in response to stiffness and microtopography

    doi: 10.1002/jcp.26236

    Figure Lengend Snippet: Localization of actin, lamellipodin, PIP2, and PIP3 with varying stiffness and altered PIP2 level Fibroblasts grown on glass and viewed with a fluorescent microscope shown untreated (left panels), with neomycin treatment (central panels), and with wortmannin treatment (right panels). Actin, red; nucleus, blue; lamellipodin, PIP2, and PIP3 green with their respective antibodies (bar = 10 µm). (A) Lamellipodin is distributed on the leading edge membrane under all conditions. (B) PIP3 antibody shows diffuse pattern with elevation at lamella under all conditions. (C) PIP2 antibody shows localization near the leading edge of the fibroblast. Significantly, neomycin-treated cells show even greater localization at lamellar edge. (D) A PIP2 biosensor (green) transfected living fibroblasts grown on glass and shows a distribution similar to the PIP2 antibody detection. (E, F) Histogram to show the ratio of intensity of PIP3 or PIP2, respectively from ten microns near the leading edge to ten microns in the cell interior. PIP3 shows no difference between treatments. Ratio of lamellar to interior PIP2 is significantly higher in neomycin-treated cells compared to untreated. Means ± SE; * P

    Article Snippet: Fibroblasts grown on 10kPa, 100kPa, 400 kPa, or glass and treated with neomycin or wortmannin were fixed in 10% formalin at 7 hours after barrier removal, probed for focal adhesions with an antibody to phospho-paxillin [Y113] (Abcam, cat. ab32084, Cambridge, MA) in a 1% BSA, 0.1% Tween-20 solution.

    Techniques: Microscopy, Transfection

    Fibroblast migration in a wound closure model is regulated by substrate stiffness and PIP2 availability Fibroblasts were grown on flat surfaces with stiffness varying from 10 kPa to over 1 GPa (glass) and subjected to treatment with drugs that reduce PIP2availability (neomycin, 500 µM) or increase PIP2 via PI3K inhibition (wortmannin, 1 µM). Phase images of cells grown on glass at (A) one hour after barrier removal (t1) and (B) at 7 hours after barrier removal (t7) to show distance migrated over time (bar = 250 µm). (C) Migration velocity was significantly highest on the softest surface (10 kPa) of untreated fibroblast compared to 400 kPa (+ in black bars, P

    Journal: Journal of cellular physiology

    Article Title: Lipid signaling affects primary fibroblast collective migration and anchorage in response to stiffness and microtopography

    doi: 10.1002/jcp.26236

    Figure Lengend Snippet: Fibroblast migration in a wound closure model is regulated by substrate stiffness and PIP2 availability Fibroblasts were grown on flat surfaces with stiffness varying from 10 kPa to over 1 GPa (glass) and subjected to treatment with drugs that reduce PIP2availability (neomycin, 500 µM) or increase PIP2 via PI3K inhibition (wortmannin, 1 µM). Phase images of cells grown on glass at (A) one hour after barrier removal (t1) and (B) at 7 hours after barrier removal (t7) to show distance migrated over time (bar = 250 µm). (C) Migration velocity was significantly highest on the softest surface (10 kPa) of untreated fibroblast compared to 400 kPa (+ in black bars, P

    Article Snippet: Fibroblasts grown on 10kPa, 100kPa, 400 kPa, or glass and treated with neomycin or wortmannin were fixed in 10% formalin at 7 hours after barrier removal, probed for focal adhesions with an antibody to phospho-paxillin [Y113] (Abcam, cat. ab32084, Cambridge, MA) in a 1% BSA, 0.1% Tween-20 solution.

    Techniques: Migration, Inhibition

    PTEN -loss results in an increased cholesterol de novo synthesis. a Relative accumulation of Srebp2 , Hmgcr , Sqs and Hmgcs1 transcripts in prostate of wild type (WT) and Pten pc −/− mice. ( N = 10 per group). b Accumulation of cholesterol precursors, lanosterol, lathosterol, desmosterol and cholesterol in prostatic samples from wild type (WT) ( N = 4) and Pten pc −/− mice ( N = 5). c qPCR analysis of HMGCR , ABCA1 and FASN expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with T0901317 supplementation (1 µM). Experiments have been performed in three experimental replicates. d qPCR analysis of ABCA1 and HMGCR expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with mevalonate (500 µM). e qPCR analysis of HMGCR , ABCA1 and FASN expression in PC3 cells treated with LY294002 (20 µM) alone or with increasing amounts of mevalonate (100 µM, 500 µM, 1 mM, 10 mM). f Western blot analysis of SREBP2 cleaved form, HMGCR and SQLE respective accumulation in LNCaP cell line. AKTS473 is used to confirm Wortmannin and LY294002 treatment efficiency and β-ACTIN as a loading control. g Western blot analysis of HMGCR, SQLE and APOE in PC3 cells treated with LY294002 (20 µM) alone or in combination with mevalonate (500 µM). AKTS473 and AKT are used to confirm Wortmannin and LY294002 treatment efficiency and β-ACTIN as a loading control. h SREBP2 and LXR target genes expression in PC3 cells transfected with siSREBP2 or siGFP as control. Experiments have been performed in three experimental replicates. i Relative expression of Hmgcr , Abca1 and Fasn on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose) or Simvastatin (40 mg kg −1 ). j Immunofluorescence detection of FASN and ABCA1 on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose, N = 6) or Simvastatin (40 mg kg −1 , N = 6). Nuclei are stained using Hoescht ( blue ), scale bar 100 µm. k qPCR analysis of Abca1 and Fasn expression on MPEC Pten −/− or Pten −/− lxrαβ −/− treated with simvastatin alone (2,5 µM) or in combination with T0901317 (1 µM), N = 3 per group. All data are represented as mean ± SEM and statistical analyses were performed with the Student’s t -test; * p

    Journal: Nature Communications

    Article Title: Liver X receptors constrain tumor development and metastasis dissemination in PTEN-deficient prostate cancer

    doi: 10.1038/s41467-017-00508-5

    Figure Lengend Snippet: PTEN -loss results in an increased cholesterol de novo synthesis. a Relative accumulation of Srebp2 , Hmgcr , Sqs and Hmgcs1 transcripts in prostate of wild type (WT) and Pten pc −/− mice. ( N = 10 per group). b Accumulation of cholesterol precursors, lanosterol, lathosterol, desmosterol and cholesterol in prostatic samples from wild type (WT) ( N = 4) and Pten pc −/− mice ( N = 5). c qPCR analysis of HMGCR , ABCA1 and FASN expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with T0901317 supplementation (1 µM). Experiments have been performed in three experimental replicates. d qPCR analysis of ABCA1 and HMGCR expression in LNCaP cells treated with Simvastatin (2,5 µM) alone or with mevalonate (500 µM). e qPCR analysis of HMGCR , ABCA1 and FASN expression in PC3 cells treated with LY294002 (20 µM) alone or with increasing amounts of mevalonate (100 µM, 500 µM, 1 mM, 10 mM). f Western blot analysis of SREBP2 cleaved form, HMGCR and SQLE respective accumulation in LNCaP cell line. AKTS473 is used to confirm Wortmannin and LY294002 treatment efficiency and β-ACTIN as a loading control. g Western blot analysis of HMGCR, SQLE and APOE in PC3 cells treated with LY294002 (20 µM) alone or in combination with mevalonate (500 µM). AKTS473 and AKT are used to confirm Wortmannin and LY294002 treatment efficiency and β-ACTIN as a loading control. h SREBP2 and LXR target genes expression in PC3 cells transfected with siSREBP2 or siGFP as control. Experiments have been performed in three experimental replicates. i Relative expression of Hmgcr , Abca1 and Fasn on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose) or Simvastatin (40 mg kg −1 ). j Immunofluorescence detection of FASN and ABCA1 on prostate samples from wild type mice and Pten pc −/− mice treated with vehicle (methylcellulose, N = 6) or Simvastatin (40 mg kg −1 , N = 6). Nuclei are stained using Hoescht ( blue ), scale bar 100 µm. k qPCR analysis of Abca1 and Fasn expression on MPEC Pten −/− or Pten −/− lxrαβ −/− treated with simvastatin alone (2,5 µM) or in combination with T0901317 (1 µM), N = 3 per group. All data are represented as mean ± SEM and statistical analyses were performed with the Student’s t -test; * p

    Article Snippet: Simvastatin (S6196), Doxycycline (D9891) and Vitamine E (T3376) were purchased from Sigma-Aldrich, Wortmannin (#9951) and LY294002 (#9901) from Cell Signaling Technology.

    Techniques: Mouse Assay, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Transfection, Immunofluorescence, Staining

    PTEN -status controls LXR activities in PCa cells through PI3K/AKT pathway. a Relative expression levels of Abca1 , Abcg1 , Idol , Fasn and ( b ) PTEN, AKTS473, AKT, FASN and β-ACTIN protein accumulation levels in MPECs (mouse prostatic epithelial cells) wild-type ( + / + ) and Pten −/− . c Immunofluorescence of PTEN–GFP and ABCA1 in LNCaP-PTEN inducible cell line treated with DMSO or Doxycycline (25 µM). Nuclei are stained using Hoescht ( blue ). Scale bar : 100 µm. d Quantitative measurement of ABCA1-positive cells has been performed. e Relative expression of LXR target genes in LNCaP-PTEN-inducible cell line treated with DMSO or Doxycycline (25 µM). f , g Relative gene expression analysis and protein accumulation of LXR targets in LNCaP cells transfected with PTEN and PTENC124A expression construct vs. empty vector. h , i Relative expression and protein accumulations of LXR targets in LNCaP cells treated with PI3K inhibitors Wortmannin (0.5 µM) or LY294002 (20 µM). j Relative accumulation of ABCA1 , ABCG1 , FASN and IDOL in LNCaP cells transfected with AKTd/n (dominant-negative) expression vector. k Immunofluorescence against AKTS473 and ABCA1 in DU145 transfected with myrAKT or p110CAAX (dominant-positive) expression vectors. Nuclei are stained using Hoescht ( blue ). Scale bar : 100 µm. l Relative accumulation of ABCA1 , ABCG1 , FASN and IDOL in DU145 transfected with myrAKT of p110CAAX expression vectors. m Luciferase activity measurement in MEF Pten −/− transfected with 3xLXREtk-Luc reporter construct or tk-Luc construct as a control and with PTEN and PTENC124A expression construct versus empty vector. n Luciferase activity measurement in MEF + / + , Lxr α −/− , Lxr β −/− or Lxr αβ −/− transfected with 3xLXREtk-Luc reporter construct and expression vector encoding myrAKT. T0901317 (1 µM) treatment has been performed as a control. MEF Lxr αβ −/− were rescued using LXRα and LXRβ expression vectors. o Abca1 and Fasn relative expression in MPEC Pten −/− and Pten −/− lxrαβ −/− treated with Wortmannin (0.5 µM), LY294002 (20 µM) and/or T0901317 (1 µM). For whole experiments, the results represent the means ± SEM of three independent experiments; * p

    Journal: Nature Communications

    Article Title: Liver X receptors constrain tumor development and metastasis dissemination in PTEN-deficient prostate cancer

    doi: 10.1038/s41467-017-00508-5

    Figure Lengend Snippet: PTEN -status controls LXR activities in PCa cells through PI3K/AKT pathway. a Relative expression levels of Abca1 , Abcg1 , Idol , Fasn and ( b ) PTEN, AKTS473, AKT, FASN and β-ACTIN protein accumulation levels in MPECs (mouse prostatic epithelial cells) wild-type ( + / + ) and Pten −/− . c Immunofluorescence of PTEN–GFP and ABCA1 in LNCaP-PTEN inducible cell line treated with DMSO or Doxycycline (25 µM). Nuclei are stained using Hoescht ( blue ). Scale bar : 100 µm. d Quantitative measurement of ABCA1-positive cells has been performed. e Relative expression of LXR target genes in LNCaP-PTEN-inducible cell line treated with DMSO or Doxycycline (25 µM). f , g Relative gene expression analysis and protein accumulation of LXR targets in LNCaP cells transfected with PTEN and PTENC124A expression construct vs. empty vector. h , i Relative expression and protein accumulations of LXR targets in LNCaP cells treated with PI3K inhibitors Wortmannin (0.5 µM) or LY294002 (20 µM). j Relative accumulation of ABCA1 , ABCG1 , FASN and IDOL in LNCaP cells transfected with AKTd/n (dominant-negative) expression vector. k Immunofluorescence against AKTS473 and ABCA1 in DU145 transfected with myrAKT or p110CAAX (dominant-positive) expression vectors. Nuclei are stained using Hoescht ( blue ). Scale bar : 100 µm. l Relative accumulation of ABCA1 , ABCG1 , FASN and IDOL in DU145 transfected with myrAKT of p110CAAX expression vectors. m Luciferase activity measurement in MEF Pten −/− transfected with 3xLXREtk-Luc reporter construct or tk-Luc construct as a control and with PTEN and PTENC124A expression construct versus empty vector. n Luciferase activity measurement in MEF + / + , Lxr α −/− , Lxr β −/− or Lxr αβ −/− transfected with 3xLXREtk-Luc reporter construct and expression vector encoding myrAKT. T0901317 (1 µM) treatment has been performed as a control. MEF Lxr αβ −/− were rescued using LXRα and LXRβ expression vectors. o Abca1 and Fasn relative expression in MPEC Pten −/− and Pten −/− lxrαβ −/− treated with Wortmannin (0.5 µM), LY294002 (20 µM) and/or T0901317 (1 µM). For whole experiments, the results represent the means ± SEM of three independent experiments; * p

    Article Snippet: Simvastatin (S6196), Doxycycline (D9891) and Vitamine E (T3376) were purchased from Sigma-Aldrich, Wortmannin (#9951) and LY294002 (#9901) from Cell Signaling Technology.

    Techniques: Expressing, Immunofluorescence, Staining, Transfection, Construct, Plasmid Preparation, Dominant Negative Mutation, Luciferase, Activity Assay

    Level of plasma interleukin-6 and myeloperoxidase activity in the pancreas. A: Plasma interleukin (IL)-6 level after treatment with wortmannin; B: Myeloperoxidase (MPO) activity in pancreatic tissue after treatment with wortmannin. c P

    Journal: World Journal of Gastroenterology : WJG

    Article Title: H2S mitigates severe acute pancreatitis through the PI3K/AKT-NF-κB pathway in vivo

    doi: 10.3748/wjg.v21.i15.4555

    Figure Lengend Snippet: Level of plasma interleukin-6 and myeloperoxidase activity in the pancreas. A: Plasma interleukin (IL)-6 level after treatment with wortmannin; B: Myeloperoxidase (MPO) activity in pancreatic tissue after treatment with wortmannin. c P

    Article Snippet: In the wortmannin (W) + SAP, 5 mg/kg NaHS + W + SAP, and 100 mg/kg NaHS + W + SAP groups, the animals were treated as described for the SAP group, with 5 mg/kg NaHS + SAP and 100 mg/kg NaHS + SAP, except that wortmannin (1.4 mg/kg, Sigma)[ ], a PI3 K inhibitor, was dissolved in dimethyl sulfoxide (DMSO) and administered 30 min before the first L-Arg injection.

    Techniques: Activity Assay

    Phosphoinositide-3-kinase (PI3K) and p-Akt protein in rat lung. Top , Representative Western blots are shown with the PI3/K, p-Akt and β-actin labeled band. Bottom , Densitometry of all Western blot results from rat lungs of the 6 groups. Data are reported as means±SD for n=6 animals per group. LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. **P

    Journal: Brazilian Journal of Medical and Biological Research

    Article Title: Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway

    doi: 10.1590/1414-431X20143949

    Figure Lengend Snippet: Phosphoinositide-3-kinase (PI3K) and p-Akt protein in rat lung. Top , Representative Western blots are shown with the PI3/K, p-Akt and β-actin labeled band. Bottom , Densitometry of all Western blot results from rat lungs of the 6 groups. Data are reported as means±SD for n=6 animals per group. LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. **P

    Article Snippet: Thirty-six rats were randomly divided into 6 groups as follows: control group (saline only), propofol group (20 mg·kg−1 ·h−1 propofol for 2 h, iv ; Fresenius Kabi, China) , wortmannin group (0.6 mg/kg wortmannin, iv ; Sigma Aldrich, USA), LPS group (5 mg/kgEscherichia coli B55:5, iv ; Sigma Aldrich), propofol+LPS group (20 mg·kg−1 ·h−1 propofol was administered 1 h before LPS, and followed by infusion of 20 mg·kg−1 ·h−1 propofol for 1 h), and wortmannin+propofol+LPS group (0.6 mg/kg wortmannin was administered 30 min before infusion of propofol, and followed by the same protocol as the propofol+LPS group).

    Techniques: Western Blot, Labeling

    The lung wet to dry weight (W/D) ratio, and concentration of proteins, TNF-α and IL-6 in bronchoalveolar lavage fluid (BALF). Data are reported as means±SD for n=8 animals per group. Data are representative of triplicate experiments with similar results. TNF-α: tumor necrosis factor-α; IL-6: interleukin-6; LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. **P

    Journal: Brazilian Journal of Medical and Biological Research

    Article Title: Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway

    doi: 10.1590/1414-431X20143949

    Figure Lengend Snippet: The lung wet to dry weight (W/D) ratio, and concentration of proteins, TNF-α and IL-6 in bronchoalveolar lavage fluid (BALF). Data are reported as means±SD for n=8 animals per group. Data are representative of triplicate experiments with similar results. TNF-α: tumor necrosis factor-α; IL-6: interleukin-6; LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. **P

    Article Snippet: Thirty-six rats were randomly divided into 6 groups as follows: control group (saline only), propofol group (20 mg·kg−1 ·h−1 propofol for 2 h, iv ; Fresenius Kabi, China) , wortmannin group (0.6 mg/kg wortmannin, iv ; Sigma Aldrich, USA), LPS group (5 mg/kgEscherichia coli B55:5, iv ; Sigma Aldrich), propofol+LPS group (20 mg·kg−1 ·h−1 propofol was administered 1 h before LPS, and followed by infusion of 20 mg·kg−1 ·h−1 propofol for 1 h), and wortmannin+propofol+LPS group (0.6 mg/kg wortmannin was administered 30 min before infusion of propofol, and followed by the same protocol as the propofol+LPS group).

    Techniques: Concentration Assay

    Histopathology of representative lung sections. Sections were stained with hematoxylin and eosin. A , Control group. B , Lipopolysaccharide (LPS) group: edematous changes of alveolar walls, swelling of alveolar epithelial cells, and massive polymorphonuclear infiltration were observed. C , Propofol+LPS group: less damage was observed compared to the LPS group. D , Wortmannin+propofol+LPS group: edematous changes of alveolar walls, swelling of alveolar epithelial cells, and massive polymorphonuclear infiltration were observed. E , Propofol group: no differences were observed compared to the control group. F , Wortmannin group: no differences were observed compared to the control group. Magnification bar: 50 μm (400×).

    Journal: Brazilian Journal of Medical and Biological Research

    Article Title: Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway

    doi: 10.1590/1414-431X20143949

    Figure Lengend Snippet: Histopathology of representative lung sections. Sections were stained with hematoxylin and eosin. A , Control group. B , Lipopolysaccharide (LPS) group: edematous changes of alveolar walls, swelling of alveolar epithelial cells, and massive polymorphonuclear infiltration were observed. C , Propofol+LPS group: less damage was observed compared to the LPS group. D , Wortmannin+propofol+LPS group: edematous changes of alveolar walls, swelling of alveolar epithelial cells, and massive polymorphonuclear infiltration were observed. E , Propofol group: no differences were observed compared to the control group. F , Wortmannin group: no differences were observed compared to the control group. Magnification bar: 50 μm (400×).

    Article Snippet: Thirty-six rats were randomly divided into 6 groups as follows: control group (saline only), propofol group (20 mg·kg−1 ·h−1 propofol for 2 h, iv ; Fresenius Kabi, China) , wortmannin group (0.6 mg/kg wortmannin, iv ; Sigma Aldrich, USA), LPS group (5 mg/kgEscherichia coli B55:5, iv ; Sigma Aldrich), propofol+LPS group (20 mg·kg−1 ·h−1 propofol was administered 1 h before LPS, and followed by infusion of 20 mg·kg−1 ·h−1 propofol for 1 h), and wortmannin+propofol+LPS group (0.6 mg/kg wortmannin was administered 30 min before infusion of propofol, and followed by the same protocol as the propofol+LPS group).

    Techniques: Histopathology, Staining

    Survival curve of LPS-challenged rats combined with saline or propofol pretreatment. Data are reported as means±SD for n=12 animals per experimental group. LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. *P

    Journal: Brazilian Journal of Medical and Biological Research

    Article Title: Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway

    doi: 10.1590/1414-431X20143949

    Figure Lengend Snippet: Survival curve of LPS-challenged rats combined with saline or propofol pretreatment. Data are reported as means±SD for n=12 animals per experimental group. LPS: lipopolysaccharide; Prop: propofol; Wort: wortmannin. *P

    Article Snippet: Thirty-six rats were randomly divided into 6 groups as follows: control group (saline only), propofol group (20 mg·kg−1 ·h−1 propofol for 2 h, iv ; Fresenius Kabi, China) , wortmannin group (0.6 mg/kg wortmannin, iv ; Sigma Aldrich, USA), LPS group (5 mg/kgEscherichia coli B55:5, iv ; Sigma Aldrich), propofol+LPS group (20 mg·kg−1 ·h−1 propofol was administered 1 h before LPS, and followed by infusion of 20 mg·kg−1 ·h−1 propofol for 1 h), and wortmannin+propofol+LPS group (0.6 mg/kg wortmannin was administered 30 min before infusion of propofol, and followed by the same protocol as the propofol+LPS group).

    Techniques:

    Roles of nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) and Pi3 kinase in the induction of heme oxygenase 1 ( HO-1 ) by certolizumab pegol ( CZP ). Monocytes were incubated, or not, with CZP 5 μg/ml in the presence or not of diphenyleneiodonium chloride ( DPI ) (10 μM) and wortmannin (0.4 μM) over 16 h. HO-1 was assessed by western blot. One representative blot is shown ( a ). Quantification of three western blot experiments was performed. *Paired t test

    Journal: Arthritis Research & Therapy

    Article Title: Anti-TNF certolizumab pegol induces antioxidant response in human monocytes via reverse signaling

    doi: 10.1186/s13075-016-0955-8

    Figure Lengend Snippet: Roles of nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) and Pi3 kinase in the induction of heme oxygenase 1 ( HO-1 ) by certolizumab pegol ( CZP ). Monocytes were incubated, or not, with CZP 5 μg/ml in the presence or not of diphenyleneiodonium chloride ( DPI ) (10 μM) and wortmannin (0.4 μM) over 16 h. HO-1 was assessed by western blot. One representative blot is shown ( a ). Quantification of three western blot experiments was performed. *Paired t test

    Article Snippet: The diphenyleneiodonium chloride (DPI) and wortmannin inhibitors were purchased from Sigma.

    Techniques: Incubation, Western Blot

    COX-2 and EP4 mediated regulation of miR655. ( A ) COX-2 protein expression in MCF7, MCF7-Mock, MCF7-COX-2 and MCF7-miR655 cells. MiR655 expression in ( B ) MCF7-COX-2 and ( C ) SKBR3-COX-2 cells treated with COX-2 inhibitor (NS-398, 10 µM) and EP4 antagonist (ONO-AE3-208, 10 µM). ( D ) Comparison of fold changes in miR655 expression relative to controls (vehicle treatment) in a panel of breast cancer cell lines treated with an EP ligand (PGE2; 10 µM), EP4 receptor agonist (PGE1OH; 10 µM), or vehicle (DMSO). Comparison of miR655 expression after stimulation with PGE2 and PGE1OH followed by treatment with two PI3K inhibitors, Wortmannin (WT), LY-240-002 (LY) both at (10 µM) and ERK inhibitor U0126 (10 µM) in ( E ) MCF7 and in ( F ) T47D cells. To test intermediary role of NF-κB in EP4 mediated regulation of miR655 in ( G ) MCF7 and ( H ) T47D cells, the cells were treated with NF-κB inhibitor BAY-11-7082 (10 µM) or vehicle (DMSO) along with PGE2 and PGE1OH. Quantitative data are presented as means of triplicate experiments ± SEM. *Indicates p

    Journal: Scientific Reports

    Article Title: COX-2 induces oncogenic micro RNA miR655 in human breast cancer

    doi: 10.1038/s41598-017-18612-3

    Figure Lengend Snippet: COX-2 and EP4 mediated regulation of miR655. ( A ) COX-2 protein expression in MCF7, MCF7-Mock, MCF7-COX-2 and MCF7-miR655 cells. MiR655 expression in ( B ) MCF7-COX-2 and ( C ) SKBR3-COX-2 cells treated with COX-2 inhibitor (NS-398, 10 µM) and EP4 antagonist (ONO-AE3-208, 10 µM). ( D ) Comparison of fold changes in miR655 expression relative to controls (vehicle treatment) in a panel of breast cancer cell lines treated with an EP ligand (PGE2; 10 µM), EP4 receptor agonist (PGE1OH; 10 µM), or vehicle (DMSO). Comparison of miR655 expression after stimulation with PGE2 and PGE1OH followed by treatment with two PI3K inhibitors, Wortmannin (WT), LY-240-002 (LY) both at (10 µM) and ERK inhibitor U0126 (10 µM) in ( E ) MCF7 and in ( F ) T47D cells. To test intermediary role of NF-κB in EP4 mediated regulation of miR655 in ( G ) MCF7 and ( H ) T47D cells, the cells were treated with NF-κB inhibitor BAY-11-7082 (10 µM) or vehicle (DMSO) along with PGE2 and PGE1OH. Quantitative data are presented as means of triplicate experiments ± SEM. *Indicates p

    Article Snippet: Wortmannin (WM), an irreversible PI3K inhibitor, and LY-204002 (LY), a reversible PI3K inhibitor all purchased from Sigma-Aldrich, ERK1/2 inhibitor U0126 from Cell Signalling, MA.

    Techniques: Expressing

    Protocol of various drugs intervention on rat myocardial ischemia/reperfusion model in vitro. C I/R: control ischemia and reperfusion; DM I/R: diabetes rats subjected to myocardial ischemia and reperfusion; DM + EtOH I/R: diabetes + ethanol subjected to myocardial ischemia and reperfusion; DM + EtOH + CYA I/R: diabetes + ethanol + cyanamide subjected to myocardial ischemia and reperfusion; DM + EtOH + Atr I/R: diabetes + ethanol + atractyloside subjected to myocardial ischemia and reperfusion; DM + EtOH + Wor I/R: diabetes + ethanol + wortmannin subjected to myocardial ischemia and reperfusion.

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: Activation of ALDH2 with Low Concentration of Ethanol Attenuates Myocardial Ischemia/Reperfusion Injury in Diabetes Rat Model

    doi: 10.1155/2016/6190504

    Figure Lengend Snippet: Protocol of various drugs intervention on rat myocardial ischemia/reperfusion model in vitro. C I/R: control ischemia and reperfusion; DM I/R: diabetes rats subjected to myocardial ischemia and reperfusion; DM + EtOH I/R: diabetes + ethanol subjected to myocardial ischemia and reperfusion; DM + EtOH + CYA I/R: diabetes + ethanol + cyanamide subjected to myocardial ischemia and reperfusion; DM + EtOH + Atr I/R: diabetes + ethanol + atractyloside subjected to myocardial ischemia and reperfusion; DM + EtOH + Wor I/R: diabetes + ethanol + wortmannin subjected to myocardial ischemia and reperfusion.

    Article Snippet: Chemicals and Reagents Streptozotocin (STZ), cyanamide (CYA), wortmannin (Wor), atractyloside (Atr), and Alda-1 were purchased from Sigma (St. Louis, MO, USA).

    Techniques: In Vitro

    Effects of peptide b11 and PLC-β3-CT on MAPKs and PI3K/Akt pathways. (A) Ba/F3 cells transduced with peptide b11 or PLC-β3-CT were analyzed by SDS-PAGE. Phosphorylation levels of MAPKs and PI3K/Akt were evaluated by immunoblotting. (B) Ba/F3 transduced cells were treated with wortmannin (3 nM), LY294062 (2 µM), SB203580 (0.1 µM), U0126 (1 µM) and SP600125 (0.1 µM). The live cells were counted on day 3 and day 7.

    Journal: PLoS ONE

    Article Title: Short Stat5-Interacting Peptide Derived from Phospholipase C-?3 Inhibits Hematopoietic Cell Proliferation and Myeloid Differentiation

    doi: 10.1371/journal.pone.0024995

    Figure Lengend Snippet: Effects of peptide b11 and PLC-β3-CT on MAPKs and PI3K/Akt pathways. (A) Ba/F3 cells transduced with peptide b11 or PLC-β3-CT were analyzed by SDS-PAGE. Phosphorylation levels of MAPKs and PI3K/Akt were evaluated by immunoblotting. (B) Ba/F3 transduced cells were treated with wortmannin (3 nM), LY294062 (2 µM), SB203580 (0.1 µM), U0126 (1 µM) and SP600125 (0.1 µM). The live cells were counted on day 3 and day 7.

    Article Snippet: Compounds Wortmannin, Ly294062, SB203580 and SP600125 were purchased from Calbiochem.

    Techniques: Planar Chromatography, Transduction, SDS Page

    Regulatory effect of RON160 and RON E5/6in on EMT-like activities in MDCK cells : A) Effect of RON, RON160, and RON E5/6in on epithelial/mesenchymal protein expression. Proteins (50 μg per sample) from cell lysates prepared after 72 h incubation were subjected to Western blot analysis using antibodies specific to vimentin and E-cadherin, respectively. Actin was used as the loading control. B ) Effect of RON, RON160 and RON E5/6in on cell morphological changes. MDCK, M-RON, M-RON160 and M-RON E5/6in cells were cultured for 24 h and then stimulated with 2 nM of MSP for 48 h. Cell morphological changes were observed under Olympus Inverted microscope and photographed. C ) Effect of RON, RON160 and RON E5/6in on spontaneous or MSP-induced MDCK cell migration. Cell monolayer was wounded as previously described [ 29 ] and stimulated with or without 2 nM of MSP for 16 h. Chemical inhibitors such as PD98059 (10 nM, specific to MAP kinase) and wortmannin (50 μg/ml, specific to PI-3 kinase) were added simultaneously. The wounded area covered by migrated cells was measured and shown as % of the covered space. Data shown here are from one of three experiments with similar results.

    Journal: Molecular Cancer

    Article Title: Deletion or insertion in the first immunoglobulin-plexin-transcription (IPT) domain differentially regulates expression and tumorigenic activities of RON receptor Tyrosine Kinase

    doi: 10.1186/1476-4598-9-307

    Figure Lengend Snippet: Regulatory effect of RON160 and RON E5/6in on EMT-like activities in MDCK cells : A) Effect of RON, RON160, and RON E5/6in on epithelial/mesenchymal protein expression. Proteins (50 μg per sample) from cell lysates prepared after 72 h incubation were subjected to Western blot analysis using antibodies specific to vimentin and E-cadherin, respectively. Actin was used as the loading control. B ) Effect of RON, RON160 and RON E5/6in on cell morphological changes. MDCK, M-RON, M-RON160 and M-RON E5/6in cells were cultured for 24 h and then stimulated with 2 nM of MSP for 48 h. Cell morphological changes were observed under Olympus Inverted microscope and photographed. C ) Effect of RON, RON160 and RON E5/6in on spontaneous or MSP-induced MDCK cell migration. Cell monolayer was wounded as previously described [ 29 ] and stimulated with or without 2 nM of MSP for 16 h. Chemical inhibitors such as PD98059 (10 nM, specific to MAP kinase) and wortmannin (50 μg/ml, specific to PI-3 kinase) were added simultaneously. The wounded area covered by migrated cells was measured and shown as % of the covered space. Data shown here are from one of three experiments with similar results.

    Article Snippet: PD98059 (PD), SB203580 (SB) and wortmannin (WT) were from Calbiochem (San Diego, CA).

    Techniques: Expressing, Incubation, Western Blot, Cell Culture, Inverted Microscopy, Migration

    IGF-1 induced F-actin assembly. The F-actin content of the MM cells was measured by FACS analysis. The mean fluorescence intensity is shown as the relative value compared to unstimulated cells. The cells were stimulated with or without 100ng ml −1 IGF-1, after a 30 min incubation with Wortmannin (100 n M ) or PD98059 (20 μ M ) where indicated. The cells were then labelled with phalloidin FITC. Mean values±s.d. for three independent experiments are shown ( * : P

    Journal: British Journal of Cancer

    Article Title: Specific roles for the PI3K and the MEK-ERK pathway in IGF-1-stimulated chemotaxis, VEGF secretion and proliferation of multiple myeloma cells: study in the 5T33MM model

    doi: 10.1038/sj.bjc.6601613

    Figure Lengend Snippet: IGF-1 induced F-actin assembly. The F-actin content of the MM cells was measured by FACS analysis. The mean fluorescence intensity is shown as the relative value compared to unstimulated cells. The cells were stimulated with or without 100ng ml −1 IGF-1, after a 30 min incubation with Wortmannin (100 n M ) or PD98059 (20 μ M ) where indicated. The cells were then labelled with phalloidin FITC. Mean values±s.d. for three independent experiments are shown ( * : P

    Article Snippet: The inhibitors Wortmannin (Sigma, Irvine, UK), Ly294002 (Sigma), PD98059 (Alexis, CA) and UO126 (Alexis) were added 30 min prior to IGF-1 stimulation at a concentration of 100 nM , 10 μ M , 20 μ M and 25 μ M, respectively.

    Techniques: FACS, Fluorescence, Incubation

    Stimulation by IGF-1 of VEGF secretion. The 5T33MM cells were stimulated with or without 100 ng ml −1 IGF-1 for 24 h, after a 30 min incubation with Wortmannin (100 n M ) or PD98059 (20 μ M ) where indicated. Concentrations of VEGF are shown relative to unstimulated cells. The maximum IGF-1-stimulated VEGF secretion reaches 250pg ml −1 . Mean values±s.d. for three independent experiments are shown ( * : P

    Journal: British Journal of Cancer

    Article Title: Specific roles for the PI3K and the MEK-ERK pathway in IGF-1-stimulated chemotaxis, VEGF secretion and proliferation of multiple myeloma cells: study in the 5T33MM model

    doi: 10.1038/sj.bjc.6601613

    Figure Lengend Snippet: Stimulation by IGF-1 of VEGF secretion. The 5T33MM cells were stimulated with or without 100 ng ml −1 IGF-1 for 24 h, after a 30 min incubation with Wortmannin (100 n M ) or PD98059 (20 μ M ) where indicated. Concentrations of VEGF are shown relative to unstimulated cells. The maximum IGF-1-stimulated VEGF secretion reaches 250pg ml −1 . Mean values±s.d. for three independent experiments are shown ( * : P

    Article Snippet: The inhibitors Wortmannin (Sigma, Irvine, UK), Ly294002 (Sigma), PD98059 (Alexis, CA) and UO126 (Alexis) were added 30 min prior to IGF-1 stimulation at a concentration of 100 nM , 10 μ M , 20 μ M and 25 μ M, respectively.

    Techniques: Incubation

    Crosstalk between the PI3K pathway and the MEK–ERK pathway. The MEK inhibitors PD98059 and UO126 abolish stimulation by IGF-1 of ERK phosphoryation but have no influence on the phosphorylation of Akt (first and third panel). The inhibitors of the PI3K pathway Wortmannin and Ly294002, on the other hand, inhibit the phosphorylation of Akt (PKB), confirming that Akt becomes phosphorylated through activation of the PI3K pathway (third panel), but also reduces the phosphorylation of ERK1 and 2 (first panel). The cells were stimulated with 100 ng ml −1 IGF-1 for 10 min and lysates were treated as in the previous figure. One experiment representative of four is shown.

    Journal: British Journal of Cancer

    Article Title: Specific roles for the PI3K and the MEK-ERK pathway in IGF-1-stimulated chemotaxis, VEGF secretion and proliferation of multiple myeloma cells: study in the 5T33MM model

    doi: 10.1038/sj.bjc.6601613

    Figure Lengend Snippet: Crosstalk between the PI3K pathway and the MEK–ERK pathway. The MEK inhibitors PD98059 and UO126 abolish stimulation by IGF-1 of ERK phosphoryation but have no influence on the phosphorylation of Akt (first and third panel). The inhibitors of the PI3K pathway Wortmannin and Ly294002, on the other hand, inhibit the phosphorylation of Akt (PKB), confirming that Akt becomes phosphorylated through activation of the PI3K pathway (third panel), but also reduces the phosphorylation of ERK1 and 2 (first panel). The cells were stimulated with 100 ng ml −1 IGF-1 for 10 min and lysates were treated as in the previous figure. One experiment representative of four is shown.

    Article Snippet: The inhibitors Wortmannin (Sigma, Irvine, UK), Ly294002 (Sigma), PD98059 (Alexis, CA) and UO126 (Alexis) were added 30 min prior to IGF-1 stimulation at a concentration of 100 nM , 10 μ M , 20 μ M and 25 μ M, respectively.

    Techniques: Activation Assay

    IGF-1 induced DNA synthesis. For the thymidine incorporation assays, the MM cells were incubated with RPMI in the absence or the presence of 10 ng ml −1 IGF-1. Before stimulation with or without IGF-1, the cells were preincubated for 30 min with Wortmannin (100 n M ), PD98059 (20 μ M ) or both where indicated. Mean values±s.d. for four independent experiments are shown. ( * : P

    Journal: British Journal of Cancer

    Article Title: Specific roles for the PI3K and the MEK-ERK pathway in IGF-1-stimulated chemotaxis, VEGF secretion and proliferation of multiple myeloma cells: study in the 5T33MM model

    doi: 10.1038/sj.bjc.6601613

    Figure Lengend Snippet: IGF-1 induced DNA synthesis. For the thymidine incorporation assays, the MM cells were incubated with RPMI in the absence or the presence of 10 ng ml −1 IGF-1. Before stimulation with or without IGF-1, the cells were preincubated for 30 min with Wortmannin (100 n M ), PD98059 (20 μ M ) or both where indicated. Mean values±s.d. for four independent experiments are shown. ( * : P

    Article Snippet: The inhibitors Wortmannin (Sigma, Irvine, UK), Ly294002 (Sigma), PD98059 (Alexis, CA) and UO126 (Alexis) were added 30 min prior to IGF-1 stimulation at a concentration of 100 nM , 10 μ M , 20 μ M and 25 μ M, respectively.

    Techniques: DNA Synthesis, Incubation

    Arrest of double strand break repair by inhibitors of DNA-PKcs phosphorylation. (A) Phosphorylation of DNA-PKcs on threonine-2609 (green) in cells irradiated and incubated without or with wortmannin (100 µM) or (C) without or with NU7441 (10 µM) assayed by immunofluorescence; DNA was stained by DRAQ (red). Below, quantitation of the signal from DNA-PKcs2609Thr-P (green pixel intensity/nuclear area). (B) Repair in cells incubated with wortmannin (100 µM) or (C) NU7441 (10 µM). (D) Quantitation of linear and supercoiled DNA during repair. Error bars show SEM from three independent experiments, or two independent experiments for NU7441.

    Journal: PLoS ONE

    Article Title: Repair of DNA Strand Breaks in a Minichromosome In Vivo: Kinetics, Modeling, and Effects of Inhibitors

    doi: 10.1371/journal.pone.0052966

    Figure Lengend Snippet: Arrest of double strand break repair by inhibitors of DNA-PKcs phosphorylation. (A) Phosphorylation of DNA-PKcs on threonine-2609 (green) in cells irradiated and incubated without or with wortmannin (100 µM) or (C) without or with NU7441 (10 µM) assayed by immunofluorescence; DNA was stained by DRAQ (red). Below, quantitation of the signal from DNA-PKcs2609Thr-P (green pixel intensity/nuclear area). (B) Repair in cells incubated with wortmannin (100 µM) or (C) NU7441 (10 µM). (D) Quantitation of linear and supercoiled DNA during repair. Error bars show SEM from three independent experiments, or two independent experiments for NU7441.

    Article Snippet: Inhibition of Enzymes Involved in Repair Wortmannin and caffeine (Sigma-Aldrich), NU1025 and 1,5-IQD (Calbiochem), and NU7441, KU55933, and Mirin (Tocris) were dissolved in DMSO.

    Techniques: Irradiation, Incubation, Immunofluorescence, Staining, Quantitation Assay

    Analysis of SOD activities (A), MDA contents (B) in the renal tissue, as well as plasma IL-10 (C) and TNF-α levels (D) in each groups. Electroacupuncture at bilaterally Zusanli and Neiguan acupoints significantly increased SOD activities, decreased MDA contents accompanied with increased levels of IL-10 and decreased TNF-α levels. Pretreatment with wortmannin significantly restrained the above effects of electroacupuncture in group ELW. Values were presented as mean ± SD (n = 10). The data were analyzed using one-way ANOVA and the Bonferroni test for multiple comparisons. Significant differences were indicated with an asterisk: *P

    Journal: PLoS ONE

    Article Title: Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways

    doi: 10.1371/journal.pone.0141622

    Figure Lengend Snippet: Analysis of SOD activities (A), MDA contents (B) in the renal tissue, as well as plasma IL-10 (C) and TNF-α levels (D) in each groups. Electroacupuncture at bilaterally Zusanli and Neiguan acupoints significantly increased SOD activities, decreased MDA contents accompanied with increased levels of IL-10 and decreased TNF-α levels. Pretreatment with wortmannin significantly restrained the above effects of electroacupuncture in group ELW. Values were presented as mean ± SD (n = 10). The data were analyzed using one-way ANOVA and the Bonferroni test for multiple comparisons. Significant differences were indicated with an asterisk: *P

    Article Snippet: To block PI3K/Akt signaling pathway, wortmannin (the selective PI3K inhibitor, 0.6mg/kg; Selleck, USA) was administered in group ELW intravenously 30min before LPS injection [ ].

    Techniques: Multiple Displacement Amplification

    Immunofluorescence assays of Nrf2 nucleoprotein by fluorescence microscope. (original magnification×400): (A) The pictures of immunofluorescence staining. Green stands for Nrf2-FITC stained sections, while blue stands for images of DAPI stained nuclei. The overlay color of blue staining in nucleus, accompanied with green staining both in cytoplasm and nucleus was considered to be positive. (B) Quantification of nuclear localization of Nrf2 protein among six groups. Electroacupuncture protocols dramatically increased translocation of Nrf2 from cytoplasm into nucleus compared with group L or SEL. To some degree, pretreatment with wortmannin counteracted nuclear accumulation of Nrf2 protein, while wortmannin alone had no effects. Data were representative of three independent experiments. Values were expressed as mean ± SD (n = 10; **P

    Journal: PLoS ONE

    Article Title: Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways

    doi: 10.1371/journal.pone.0141622

    Figure Lengend Snippet: Immunofluorescence assays of Nrf2 nucleoprotein by fluorescence microscope. (original magnification×400): (A) The pictures of immunofluorescence staining. Green stands for Nrf2-FITC stained sections, while blue stands for images of DAPI stained nuclei. The overlay color of blue staining in nucleus, accompanied with green staining both in cytoplasm and nucleus was considered to be positive. (B) Quantification of nuclear localization of Nrf2 protein among six groups. Electroacupuncture protocols dramatically increased translocation of Nrf2 from cytoplasm into nucleus compared with group L or SEL. To some degree, pretreatment with wortmannin counteracted nuclear accumulation of Nrf2 protein, while wortmannin alone had no effects. Data were representative of three independent experiments. Values were expressed as mean ± SD (n = 10; **P

    Article Snippet: To block PI3K/Akt signaling pathway, wortmannin (the selective PI3K inhibitor, 0.6mg/kg; Selleck, USA) was administered in group ELW intravenously 30min before LPS injection [ ].

    Techniques: Immunofluorescence, Fluorescence, Microscopy, Staining, Translocation Assay

    Western blot analysis of phospho-Akt protein (A), HO-1 protein (B), Nrf2 total (C) and nucleoprotein (D) expressions in the renal tissue of six groups. β-actin was monitored as the internal standard to ensure similar gel loading of the starting materials in each sample. Blot images were cropped for comparison (E). Electroacupuncture treatment significantly increased the levels of phospho-Akt protein, HO-1 protein, Nrf2 total and nucleoprotein compared with group L or group SEL. In wortmannin-treated rabbits, phosphorylation of Akt was significantly decreased while HO-1 protein, Nrf2 total and nucleoprotein was slightly decreased compared with group EL. All values were expressed as mean ±SD (n = 10; *P

    Journal: PLoS ONE

    Article Title: Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways

    doi: 10.1371/journal.pone.0141622

    Figure Lengend Snippet: Western blot analysis of phospho-Akt protein (A), HO-1 protein (B), Nrf2 total (C) and nucleoprotein (D) expressions in the renal tissue of six groups. β-actin was monitored as the internal standard to ensure similar gel loading of the starting materials in each sample. Blot images were cropped for comparison (E). Electroacupuncture treatment significantly increased the levels of phospho-Akt protein, HO-1 protein, Nrf2 total and nucleoprotein compared with group L or group SEL. In wortmannin-treated rabbits, phosphorylation of Akt was significantly decreased while HO-1 protein, Nrf2 total and nucleoprotein was slightly decreased compared with group EL. All values were expressed as mean ±SD (n = 10; *P

    Article Snippet: To block PI3K/Akt signaling pathway, wortmannin (the selective PI3K inhibitor, 0.6mg/kg; Selleck, USA) was administered in group ELW intravenously 30min before LPS injection [ ].

    Techniques: Western Blot

    Microphotographs of histopathologic changes of kidney sections stained with hematoxylin and eosin. (original magnification×400): (W, C) Normal morphology of kidneys; (L, SEL) Rabbits treated with LPS or sham electroacupuncture showing infiltration of inflammatory cells, tubular epithelial cell vacuolization, swelling and desquamation. (EL) Marked attenuation of tubular damages were displayed in treatment with electroacupuncture at ST36 and PC6 acupoints. (ELW) Preprocessing with wortmannin suppressed the protective efficacy of electroacupuncture to some extent. Black arrows: the destruction of renal capsule and capsular space in group ELW while infiltration of inflammatory cells in group EL or group SEL.

    Journal: PLoS ONE

    Article Title: Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways

    doi: 10.1371/journal.pone.0141622

    Figure Lengend Snippet: Microphotographs of histopathologic changes of kidney sections stained with hematoxylin and eosin. (original magnification×400): (W, C) Normal morphology of kidneys; (L, SEL) Rabbits treated with LPS or sham electroacupuncture showing infiltration of inflammatory cells, tubular epithelial cell vacuolization, swelling and desquamation. (EL) Marked attenuation of tubular damages were displayed in treatment with electroacupuncture at ST36 and PC6 acupoints. (ELW) Preprocessing with wortmannin suppressed the protective efficacy of electroacupuncture to some extent. Black arrows: the destruction of renal capsule and capsular space in group ELW while infiltration of inflammatory cells in group EL or group SEL.

    Article Snippet: To block PI3K/Akt signaling pathway, wortmannin (the selective PI3K inhibitor, 0.6mg/kg; Selleck, USA) was administered in group ELW intravenously 30min before LPS injection [ ].

    Techniques: Staining

    Effects of the inhibitors on class I PI3K/Akt/mTOR axis signaling in canine C2 cells. Cells were treated with pan-class I PI3K inhibitor Wortmannin (W) at 1 μM and ZSTK474 (Z) at 1 μM, mTOR inhibitor Rapamycin (R) at 100 nM ( A ) and Akt inhibitor KP372-1 at 0, 150, 200 and 400 nM ( B ) for the indicated period of time. Whole cell lysates (comprising 50 μg total protein) were subjected to western blot with the indicated antibodies. β-actin was used as a loading control. N/A indicates data unavailable due to induction of apoptosis in all cells.

    Journal: BMC Veterinary Research

    Article Title: The class I PI3K/Akt pathway is critical for cancer cell survival in dogs and offers an opportunity for therapeutic intervention

    doi: 10.1186/1746-6148-8-73

    Figure Lengend Snippet: Effects of the inhibitors on class I PI3K/Akt/mTOR axis signaling in canine C2 cells. Cells were treated with pan-class I PI3K inhibitor Wortmannin (W) at 1 μM and ZSTK474 (Z) at 1 μM, mTOR inhibitor Rapamycin (R) at 100 nM ( A ) and Akt inhibitor KP372-1 at 0, 150, 200 and 400 nM ( B ) for the indicated period of time. Whole cell lysates (comprising 50 μg total protein) were subjected to western blot with the indicated antibodies. β-actin was used as a loading control. N/A indicates data unavailable due to induction of apoptosis in all cells.

    Article Snippet: Drug compounds and pathway inhibitors ZSTK474 (pan-PI3K inhibitor, Z-1066, LC Laboratories, USA), Wortmannin (pan-PI3K inhibitor, 1232, Tocris bioscience, USA), KP372-1 (Akt inhibitor, B-0102, Echelon, USA) and Rapamycin (mTOR inhibitor, R0395, Sigma-Aldrich, USA) were dissolved in dimethyl sulfoxide (DMSO) as concentrated stocks that were stored at -70 °C and diluted freshly in cell medium before use.

    Techniques: Western Blot

    Schematic diagram of the class I PI3K/Akt/mTOR axis pathway. The targets of the inhibitors (ZSTK474, Wortmannin, KP372-1 and Rapamycin) used in this study are indicated.

    Journal: BMC Veterinary Research

    Article Title: The class I PI3K/Akt pathway is critical for cancer cell survival in dogs and offers an opportunity for therapeutic intervention

    doi: 10.1186/1746-6148-8-73

    Figure Lengend Snippet: Schematic diagram of the class I PI3K/Akt/mTOR axis pathway. The targets of the inhibitors (ZSTK474, Wortmannin, KP372-1 and Rapamycin) used in this study are indicated.

    Article Snippet: Drug compounds and pathway inhibitors ZSTK474 (pan-PI3K inhibitor, Z-1066, LC Laboratories, USA), Wortmannin (pan-PI3K inhibitor, 1232, Tocris bioscience, USA), KP372-1 (Akt inhibitor, B-0102, Echelon, USA) and Rapamycin (mTOR inhibitor, R0395, Sigma-Aldrich, USA) were dissolved in dimethyl sulfoxide (DMSO) as concentrated stocks that were stored at -70 °C and diluted freshly in cell medium before use.

    Techniques:

    Inhibition of Nrf2 activity by PI3K inhibitors. ARPE cells were treated with 10 μ M LY294002 for 16 hours ( A ) or wortmannin for 6 hours ( B ) at the indicated concentrations. The Nrf2 activity was measured by transient transfection with an ARE reporter

    Journal:

    Article Title: Essential Roles of the PI3 Kinase/Akt Pathway in Regulating Nrf2-Dependent Antioxidant Functions in the RPE

    doi: 10.1167/iovs.07-1099

    Figure Lengend Snippet: Inhibition of Nrf2 activity by PI3K inhibitors. ARPE cells were treated with 10 μ M LY294002 for 16 hours ( A ) or wortmannin for 6 hours ( B ) at the indicated concentrations. The Nrf2 activity was measured by transient transfection with an ARE reporter

    Article Snippet: Sulforaphane, γ -glutamyl glutamate ( γ GG), wortmannin, and LY294002 were purchased from LKT Laboratories (St. Paul, MN), MP Biomedicals (Irvine, CA), Upstate (Lake Placid, NY), and Promega (Madison, WI), respectively.

    Techniques: Inhibition, Activity Assay, Transfection

    Decreased expression of the antioxidant genes after PI3K inhibition. RPE cells were treated with either 10 μ M LY294002 for 16 hours ( A, C ) or wortmannin for 6 hours ( B ) at indicated concentrations. mRNA levels of the catalytic (GCLC) and modulatory

    Journal:

    Article Title: Essential Roles of the PI3 Kinase/Akt Pathway in Regulating Nrf2-Dependent Antioxidant Functions in the RPE

    doi: 10.1167/iovs.07-1099

    Figure Lengend Snippet: Decreased expression of the antioxidant genes after PI3K inhibition. RPE cells were treated with either 10 μ M LY294002 for 16 hours ( A, C ) or wortmannin for 6 hours ( B ) at indicated concentrations. mRNA levels of the catalytic (GCLC) and modulatory

    Article Snippet: Sulforaphane, γ -glutamyl glutamate ( γ GG), wortmannin, and LY294002 were purchased from LKT Laboratories (St. Paul, MN), MP Biomedicals (Irvine, CA), Upstate (Lake Placid, NY), and Promega (Madison, WI), respectively.

    Techniques: Expressing, Inhibition

    Decreased intracellular GSH in ARPE-19 cells exposed to PI3K inhibitors. Cells were treated with either wortmannin for 6 hours or LY294002 for 16 hours at indicated concentrations. Akt phosphorylation was measured by Western blot analyses ( A, C ) and was

    Journal:

    Article Title: Essential Roles of the PI3 Kinase/Akt Pathway in Regulating Nrf2-Dependent Antioxidant Functions in the RPE

    doi: 10.1167/iovs.07-1099

    Figure Lengend Snippet: Decreased intracellular GSH in ARPE-19 cells exposed to PI3K inhibitors. Cells were treated with either wortmannin for 6 hours or LY294002 for 16 hours at indicated concentrations. Akt phosphorylation was measured by Western blot analyses ( A, C ) and was

    Article Snippet: Sulforaphane, γ -glutamyl glutamate ( γ GG), wortmannin, and LY294002 were purchased from LKT Laboratories (St. Paul, MN), MP Biomedicals (Irvine, CA), Upstate (Lake Placid, NY), and Promega (Madison, WI), respectively.

    Techniques: Western Blot