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  • 95
    Thermo Fisher ly 294002 hydrochloride
    Ly 294002 Hydrochloride, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Abcam ly294002
    <t>LY294002</t> down-regulated MMPs expression and blocked the effect of LV-AQP3 and aqp3shRNA in SGC7901 cells . SGC7901 cells were exposed to LY294002 for 48h and then were harvested to perform Western blot analysis. We found a significant decrease in MT1-MMP, MMP-2, and MMP-9 expression. However, with the addition of LY294002, the expression of MMPs could not be obviously reversed in LV-AQP3 or aqp3shRNA groups. * p
    Ly294002, supplied by Abcam, used in various techniques. Bioz Stars score: 95/100, based on 137 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ly294002/product/Abcam
    Average 95 stars, based on 137 article reviews
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    ly294002 - by Bioz Stars, 2020-05
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    92
    LKT Laboratories ly294002
    Activation of eIF4E induced by everolimus is independent of the growth factors but can be inhibited by a PI3K inhibitor. a Jurkat cells were serum starved for 24 h and then were treated with 40 nM of everolimus in the absence or presence of FBS for the indicated times. b The cells were cultured in serum-deficient medium for 24 h and then were treated with 40 nM of everolimus, 20 ng/mL of IGF-1, or their combination for 60, 120, 180 and 360 min, respectively. c Jurkat cells were pretreated with 10 μM of PQ401 (IGF-1R inhibitor) for 60 min and then were co-treated with everolimus (40 nM) for the indicated times. d Jurkat cells were incubated with different concentrations of everolimus and/or <t>LY294002</t> for 24 h. Whole-cell lysates were prepared and subjected to SDS-PAGE followed by immunoblotting with antibody that recognizes the corresponding antigen. The results are representative of three independent experiments
    Ly294002, supplied by LKT Laboratories, used in various techniques. Bioz Stars score: 92/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ly294002/product/LKT Laboratories
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    95
    Tocris ly294002
    Effects of UO126 and <t>LY294002</t> on the hCG-, EGF-, or IGF-I-induced phosphorylation of Akt or ERK1/2 in primary cultures of immature rat Leydig cells. Primary cultures of immature rat Leydig cells were preincubated for 30 min with DMSO (control), LY294003,
    Ly294002, supplied by Tocris, used in various techniques. Bioz Stars score: 95/100, based on 230 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ly294002/product/Tocris
    Average 95 stars, based on 230 article reviews
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    99
    Cell Signaling Technology Inc inhibitor ly 294002
    Functional phosphatidylinositol 3-kinase [PI3-kinase (PI3K)] and CFTR are required for protein kinase B (Akt) 2-stimulated paracellular barrier tightening in human airway epithelial cells. A : human airway epithelial cells express all three isoforms of PKB/Akt as shown by immunoblot. Treatment with bilateral (Bi), apical, or basolateral (Baso) insulin did not change total amounts of Akt protein. B : immunoblot of phospho-Akt1 and phospho-Akt2 after 30 min stimulation with bilateral 125 nM insulin. Calyculin A-treated phosphorylation-positive control Jurkat cells (+ve) were used as a control for phospho-Akt. C : fold-increase of phospho-Akt in n = 3 filters as in B . Akt stimulation was significantly higher for NuLi-1 cells than for CuFi-5 cells. D : insulin treatment significantly increased TER in NuLi-1 cells (red bar). Pretreatment with CFTRInh172 before and during insulin stimulation significantly diminished the insulin-stimulated increase in TER (blue bars). The insulin-stimulated increase in TER of NuLi-1 cells also was prevented by <t>LY-294002</t> treatment (green bars). E : pretreatment with CFTRInh172 before insulin stimulation also prevented the decrease in the rates of paracellular flux of both calcein (blue bar, left ) and dextran (blue bar, right ) that is normally seen with insulin alone (red bars). NuLi-1 cells pretreated with LY-294002 responded to subsequent exposure to insulin with an increase of both calcein (green bar, left ) and dextran (green bar, right ) flux, whereas insulin alone (red bar) selectively decreased paracellular flux of dextran ( right ). All data are shown as means ± SE and n = 3–4 experiments for each data point for each graph where P ≤ 0.05 (*), 0.01 (**), 0.001 (***), 0.0001 (****), and 0.06 (#) by unprotected two-way ANOVA Fisher’s LSD test.
    Inhibitor Ly 294002, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/inhibitor ly 294002/product/Cell Signaling Technology Inc
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    93
    BioVision ly294002
    TNF activates the PI3K/AKT pathway in HF stem cells. ( a ) Using TMT labelling and affinity enrichment followed by high-resolution LC–MS/MS and quantitative phosphor-proteomics, functional enrichment-based cluster analysis identified up-regulated signals in cultured epidermal stem cells after TNF-α treatment. ( b ) Western blot analysis of p-AKT (at Serine 473) and total AKT in cultured murine epidermal stem cells in the presence of different concentrations of TNF-α for 0.5 h. For all western blot analysis, data are representative of 3–5 independent experiments. ( c ) TNF-α treatment (8 h) resulted in no obvious changes in p-GSK-3β(ser9) and p-β-catenin (Ser33/37/Thr45, Ser675) but significantly increased the level of p-β-catenin (Ser552), which also showed in a TNF-dose-dependent manner, quite similar to p-AKT. ( d ) Perifosine diminished p-β-catenin (Ser552) levels that were elevated by TNF-α in cultured epidermal stem cells. ( e ) Western blot analysis indicated that TNF-α increased the level of β-catenin, and the effect was attenuated by Perifosine or <t>LY294002.</t> ( f ) TCF/LEF Dual-luciferase reporter analysis showed the relative transcription activity in differently treated groups. ( g ) At PWD-3, both p-AKT (Ser473) and p-β-catenin (Ser552) were detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the p-AKT (Ser473) and p-β-catenin (Ser552) were highly co-localized. ( h ) At PWD-3, accumulated β-catenin was detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the accumulated β-catenin was highly co-localized with p-AKT (Ser473). Scale bars, 30 μm.
    Ly294002, supplied by BioVision, used in various techniques. Bioz Stars score: 93/100, based on 37 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Selleck Chemicals pi3k inhibitor ly 294002
    <t>PI3K</t> inhibitors <t>(LY-294002)</t> increase nuclear translocation of BACH2 and enhance MCL cytotoxicity. ( A ) Effect of PI3K inhibitors on nuclear localization of BACH2 was analyzed by immunoblottings in Jeko cells. Cells were pretreated with PI3K inhibitors (LY294002, 50 µM) or Bruton's tyrosine kinase inhibitors (PCI-32765, 1 µM/1 h) followed by bortezomib treatment (20 nM/24h). Bruton's tyrosine kinase inhibitor (PCI-32765) was used as a control, which did not show increased nuclear translocation of BACH2. Cytoplasmic and nuclear fractions for each sample were separated by hypotonic method using a kit. TBP was shown as loading control for nuclear proteins. Controls are Jeko cells without any treatments. 293T cells were used as positive control for BACH2. Data represent four independent experiments. ( B ) BACH2 levels in each sample were plotted in the graph. ( C ) Inhibition of nuclear export of BACH2 by leptomycin-B (LTB) enhanced the bortezomib-induced apoptosis in Jeko cells. Apoptosis was determined by Annexin-V/7-AAD dual staining method using FACS. Error bars indicate the standard error of the means (SEM) from two independent experiments. Results are expressed as an increase in fold of percentage apoptosis compared to control. LTB treatment was given 6 h prior to BTZ treatment for 24 h.
    Pi3k Inhibitor Ly 294002, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    LY294002 down-regulated MMPs expression and blocked the effect of LV-AQP3 and aqp3shRNA in SGC7901 cells . SGC7901 cells were exposed to LY294002 for 48h and then were harvested to perform Western blot analysis. We found a significant decrease in MT1-MMP, MMP-2, and MMP-9 expression. However, with the addition of LY294002, the expression of MMPs could not be obviously reversed in LV-AQP3 or aqp3shRNA groups. * p

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: Aquaporin-3 positively regulates matrix metalloproteinases via PI3K/AKT signal pathway in human gastric carcinoma SGC7901 cells

    doi: 10.1186/1756-9966-30-86

    Figure Lengend Snippet: LY294002 down-regulated MMPs expression and blocked the effect of LV-AQP3 and aqp3shRNA in SGC7901 cells . SGC7901 cells were exposed to LY294002 for 48h and then were harvested to perform Western blot analysis. We found a significant decrease in MT1-MMP, MMP-2, and MMP-9 expression. However, with the addition of LY294002, the expression of MMPs could not be obviously reversed in LV-AQP3 or aqp3shRNA groups. * p

    Article Snippet: LY294002, MT1-MMP, MMP-2, and MMP-9 antibodies were purchased from Abcam (Hong Kong, China).

    Techniques: Expressing, Western Blot

    Effect of LY294002 on Akt/GSK-3β/PDK-1 signaling and the inhibition of GSK-3β activity on PDK-1 expression. The cells were exposed to PDGF, or with increasing LY294002 concentrations of 1, 2, 5 and 20 µ M for 24 h, prior to detection of the expression of (A and B) p-Akt, (C and D) GSK-3β or (E and F) PDK-1. (G and H) The cells were treated with PDGF, or with 5 µ M LY294002 with/without 30 min pre-exposure to 5 µ M SB216763 for 24 h, prior to the detection of PDK-1 expression. # P

    Journal: International Journal of Molecular Medicine

    Article Title: Reversal of the Warburg effect with DCA in PDGF-treated human PASMC is potentiated by pyruvate dehydrogenase kinase-1 inhibition mediated through blocking Akt/GSK-3β signalling

    doi: 10.3892/ijmm.2018.3745

    Figure Lengend Snippet: Effect of LY294002 on Akt/GSK-3β/PDK-1 signaling and the inhibition of GSK-3β activity on PDK-1 expression. The cells were exposed to PDGF, or with increasing LY294002 concentrations of 1, 2, 5 and 20 µ M for 24 h, prior to detection of the expression of (A and B) p-Akt, (C and D) GSK-3β or (E and F) PDK-1. (G and H) The cells were treated with PDGF, or with 5 µ M LY294002 with/without 30 min pre-exposure to 5 µ M SB216763 for 24 h, prior to the detection of PDK-1 expression. # P

    Article Snippet: Lactate measurement The PASMCs were seeded into 25 cm2 tissue culture flasks at a cell density of 5×105 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation with 0.5% FBS for 24 h. The cells were then incubated at 37°C with 20 ng/ml PDGF with 5 µ M LY294002, DCA at 10 or 20 mM, a combination of LY294002 and 10 mM DCA, or alone for 48 h, subsequent to which the medium was removed from cells and lactate levels in the extracellular medium were measured at room temperature using the Lactate Colorimetric Assay kit (Abcam, Cambridge, MA, USA) according to the manufacturer's protocol.

    Techniques: Inhibition, Activity Assay, Expressing

    Effect of DCA, LY294002 or combination of DCA and LY294002 on extracellular lactate concentration, key glycolysis-associated enzymes expression and HK-2 activation. The PASMCs were seeded into 25 cm 2 tissue culture flask at a density of 5×10 5 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation for 24 h. (A) The cells were then exposed to PDGF or 5 µ M LY294002, DCA at 10 or 20 mM, and a combination of LY294002 and 10 mM DCA for 48 h prior to measurement of lactate concentration. * P

    Journal: International Journal of Molecular Medicine

    Article Title: Reversal of the Warburg effect with DCA in PDGF-treated human PASMC is potentiated by pyruvate dehydrogenase kinase-1 inhibition mediated through blocking Akt/GSK-3β signalling

    doi: 10.3892/ijmm.2018.3745

    Figure Lengend Snippet: Effect of DCA, LY294002 or combination of DCA and LY294002 on extracellular lactate concentration, key glycolysis-associated enzymes expression and HK-2 activation. The PASMCs were seeded into 25 cm 2 tissue culture flask at a density of 5×10 5 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation for 24 h. (A) The cells were then exposed to PDGF or 5 µ M LY294002, DCA at 10 or 20 mM, and a combination of LY294002 and 10 mM DCA for 48 h prior to measurement of lactate concentration. * P

    Article Snippet: Lactate measurement The PASMCs were seeded into 25 cm2 tissue culture flasks at a cell density of 5×105 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation with 0.5% FBS for 24 h. The cells were then incubated at 37°C with 20 ng/ml PDGF with 5 µ M LY294002, DCA at 10 or 20 mM, a combination of LY294002 and 10 mM DCA, or alone for 48 h, subsequent to which the medium was removed from cells and lactate levels in the extracellular medium were measured at room temperature using the Lactate Colorimetric Assay kit (Abcam, Cambridge, MA, USA) according to the manufacturer's protocol.

    Techniques: Concentration Assay, Expressing, Activation Assay, Cell Culture

    Effect of DCA, LY294002 or combination of DCA and LY294002 on the apoptosis and mitochondria membrane potential of human PASMCs. The PASMCs were seeded into 25 cm 2 tissue culture flask at a density of 5×10 5 cell/flask and cultured in RPMI-1640 complete culture medium for 16 h followed by serum starvation for 24 h. The cells were then exposed to PDGF alone or 5 µ M LY294002, DCA at 10 mM or a combination of 5 µ M LY294002 and 10 mM DCA for 48 h prior to (A) apoptosis or (B) JC-1 assay. (C) The expression levels of caspase-3 and cleaved caspase-3 were analyzed with western blot analysis. The representative change of one of the three experiments is presented, as all assays exhibited identical results. (D) Results were pooled from three separate experiments and are presented as mean ± standard deviation. * P

    Journal: International Journal of Molecular Medicine

    Article Title: Reversal of the Warburg effect with DCA in PDGF-treated human PASMC is potentiated by pyruvate dehydrogenase kinase-1 inhibition mediated through blocking Akt/GSK-3β signalling

    doi: 10.3892/ijmm.2018.3745

    Figure Lengend Snippet: Effect of DCA, LY294002 or combination of DCA and LY294002 on the apoptosis and mitochondria membrane potential of human PASMCs. The PASMCs were seeded into 25 cm 2 tissue culture flask at a density of 5×10 5 cell/flask and cultured in RPMI-1640 complete culture medium for 16 h followed by serum starvation for 24 h. The cells were then exposed to PDGF alone or 5 µ M LY294002, DCA at 10 mM or a combination of 5 µ M LY294002 and 10 mM DCA for 48 h prior to (A) apoptosis or (B) JC-1 assay. (C) The expression levels of caspase-3 and cleaved caspase-3 were analyzed with western blot analysis. The representative change of one of the three experiments is presented, as all assays exhibited identical results. (D) Results were pooled from three separate experiments and are presented as mean ± standard deviation. * P

    Article Snippet: Lactate measurement The PASMCs were seeded into 25 cm2 tissue culture flasks at a cell density of 5×105 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation with 0.5% FBS for 24 h. The cells were then incubated at 37°C with 20 ng/ml PDGF with 5 µ M LY294002, DCA at 10 or 20 mM, a combination of LY294002 and 10 mM DCA, or alone for 48 h, subsequent to which the medium was removed from cells and lactate levels in the extracellular medium were measured at room temperature using the Lactate Colorimetric Assay kit (Abcam, Cambridge, MA, USA) according to the manufacturer's protocol.

    Techniques: Cell Culture, Expressing, Western Blot, Standard Deviation

    Effect of DCA, LY294002 or combination of DCA and LY294002 on the growth of human PASMCs. The human PASMCs were seeded in 96-well plates in RPMI-1640 medium supplemented with 10% FBS followed by 48 h serum starving prior to exposure to 20 ng/ml PDGF or with increased concentrations of (A) DCA or (B) LY294002 in fresh culture medium with 0.5% foetal bovine serum for 72 h. * P

    Journal: International Journal of Molecular Medicine

    Article Title: Reversal of the Warburg effect with DCA in PDGF-treated human PASMC is potentiated by pyruvate dehydrogenase kinase-1 inhibition mediated through blocking Akt/GSK-3β signalling

    doi: 10.3892/ijmm.2018.3745

    Figure Lengend Snippet: Effect of DCA, LY294002 or combination of DCA and LY294002 on the growth of human PASMCs. The human PASMCs were seeded in 96-well plates in RPMI-1640 medium supplemented with 10% FBS followed by 48 h serum starving prior to exposure to 20 ng/ml PDGF or with increased concentrations of (A) DCA or (B) LY294002 in fresh culture medium with 0.5% foetal bovine serum for 72 h. * P

    Article Snippet: Lactate measurement The PASMCs were seeded into 25 cm2 tissue culture flasks at a cell density of 5×105 cell/flask and cultured with RPMI-1640 complete culture medium for 16 h followed by serum starvation with 0.5% FBS for 24 h. The cells were then incubated at 37°C with 20 ng/ml PDGF with 5 µ M LY294002, DCA at 10 or 20 mM, a combination of LY294002 and 10 mM DCA, or alone for 48 h, subsequent to which the medium was removed from cells and lactate levels in the extracellular medium were measured at room temperature using the Lactate Colorimetric Assay kit (Abcam, Cambridge, MA, USA) according to the manufacturer's protocol.

    Techniques:

    The E-cadherin expression of EpH4 cells 7 days after incubation of EpH4 cells with MM, GM, and matrigel-coated GM or without microspheres and EpH4 cells were cultured by the conventional 3D and 2D monolayer methods in the absence (□) or presence of PI3K inhibitor LY294002 (■). The number of EpH4 cells added initially was 1 × 10 4 /well while that of microspheres was 1 × 10 2 or 1 × 10 3 /well. *, p

    Journal: Regenerative Therapy

    Article Title: Preparation of epithelial cell aggregates incorporating matrigel microspheres to enhance proliferation and differentiation of epithelial cells

    doi: 10.1016/j.reth.2017.07.001

    Figure Lengend Snippet: The E-cadherin expression of EpH4 cells 7 days after incubation of EpH4 cells with MM, GM, and matrigel-coated GM or without microspheres and EpH4 cells were cultured by the conventional 3D and 2D monolayer methods in the absence (□) or presence of PI3K inhibitor LY294002 (■). The number of EpH4 cells added initially was 1 × 10 4 /well while that of microspheres was 1 × 10 2 or 1 × 10 3 /well. *, p

    Article Snippet: In the experiment of phosphotidylinositol 3-kinase (PI3K, Sigma–Aldrich Inc., St. Louis, America) inhibition experiment of EpH4 cells, the standard medium containing a PI3K inhibitor LY294002 (50 μM, Abcam Inc., Cambridge, British) was used and the cells were cultured for 72 h.

    Techniques: Expressing, Incubation, Cell Culture

    The β-casein expression of EpH4 cells 7 days after incubation of EpH4 cells with MM, GM, and matrigel-coated GM or without microspheres and EpH4 cells were cultured by the conventional 3D and 2D monolayer methods in the absence (□) or presence of PI3K inhibitor LY294002 (■). The number of EpH4 cells added initially was 1 × 10 4 /well while that of microspheres was 1 × 10 2 or 1 × 10 3 /well. *, p

    Journal: Regenerative Therapy

    Article Title: Preparation of epithelial cell aggregates incorporating matrigel microspheres to enhance proliferation and differentiation of epithelial cells

    doi: 10.1016/j.reth.2017.07.001

    Figure Lengend Snippet: The β-casein expression of EpH4 cells 7 days after incubation of EpH4 cells with MM, GM, and matrigel-coated GM or without microspheres and EpH4 cells were cultured by the conventional 3D and 2D monolayer methods in the absence (□) or presence of PI3K inhibitor LY294002 (■). The number of EpH4 cells added initially was 1 × 10 4 /well while that of microspheres was 1 × 10 2 or 1 × 10 3 /well. *, p

    Article Snippet: In the experiment of phosphotidylinositol 3-kinase (PI3K, Sigma–Aldrich Inc., St. Louis, America) inhibition experiment of EpH4 cells, the standard medium containing a PI3K inhibitor LY294002 (50 μM, Abcam Inc., Cambridge, British) was used and the cells were cultured for 72 h.

    Techniques: Expressing, Incubation, Cell Culture

    Activation of eIF4E induced by everolimus is independent of the growth factors but can be inhibited by a PI3K inhibitor. a Jurkat cells were serum starved for 24 h and then were treated with 40 nM of everolimus in the absence or presence of FBS for the indicated times. b The cells were cultured in serum-deficient medium for 24 h and then were treated with 40 nM of everolimus, 20 ng/mL of IGF-1, or their combination for 60, 120, 180 and 360 min, respectively. c Jurkat cells were pretreated with 10 μM of PQ401 (IGF-1R inhibitor) for 60 min and then were co-treated with everolimus (40 nM) for the indicated times. d Jurkat cells were incubated with different concentrations of everolimus and/or LY294002 for 24 h. Whole-cell lysates were prepared and subjected to SDS-PAGE followed by immunoblotting with antibody that recognizes the corresponding antigen. The results are representative of three independent experiments

    Journal: Acta Pharmacologica Sinica

    Article Title: MNK1 inhibitor CGP57380 overcomes mTOR inhibitor-induced activation of eIF4E: the mechanism of synergic killing of human T-ALL cells

    doi: 10.1038/s41401-018-0161-0

    Figure Lengend Snippet: Activation of eIF4E induced by everolimus is independent of the growth factors but can be inhibited by a PI3K inhibitor. a Jurkat cells were serum starved for 24 h and then were treated with 40 nM of everolimus in the absence or presence of FBS for the indicated times. b The cells were cultured in serum-deficient medium for 24 h and then were treated with 40 nM of everolimus, 20 ng/mL of IGF-1, or their combination for 60, 120, 180 and 360 min, respectively. c Jurkat cells were pretreated with 10 μM of PQ401 (IGF-1R inhibitor) for 60 min and then were co-treated with everolimus (40 nM) for the indicated times. d Jurkat cells were incubated with different concentrations of everolimus and/or LY294002 for 24 h. Whole-cell lysates were prepared and subjected to SDS-PAGE followed by immunoblotting with antibody that recognizes the corresponding antigen. The results are representative of three independent experiments

    Article Snippet: LY294002 was purchased from LKT Laboratories, Inc. (St. Paul, MN, USA).

    Techniques: Activation Assay, Cell Culture, Incubation, SDS Page

    Effects of UO126 and LY294002 on the hCG-, EGF-, or IGF-I-induced phosphorylation of Akt or ERK1/2 in primary cultures of immature rat Leydig cells. Primary cultures of immature rat Leydig cells were preincubated for 30 min with DMSO (control), LY294003,

    Journal: Endocrinology

    Article Title: Activation of the Lutropin/Choriogonadotropin Receptor Inhibits Apoptosis of Immature Leydig Cells in Primary Culture

    doi: 10.1210/en.2009-0207

    Figure Lengend Snippet: Effects of UO126 and LY294002 on the hCG-, EGF-, or IGF-I-induced phosphorylation of Akt or ERK1/2 in primary cultures of immature rat Leydig cells. Primary cultures of immature rat Leydig cells were preincubated for 30 min with DMSO (control), LY294003,

    Article Snippet: UO126 and LY294002 were from Tocris (Ellisville, MO) or Biomol (Plymouth Meeting, PA).

    Techniques:

    Effects of UO126 and LY294002 on the proliferative effects of hCG, EGF, or IGF-I on primary cultures of rat Leydig cells. A, Primary cultures of immature rat Leydig cells were incubated with buffer only, hCG, EGF, or IGF-I (each at 100 ng/ml) for 24 h

    Journal: Endocrinology

    Article Title: Activation of the Lutropin/Choriogonadotropin Receptor Inhibits Apoptosis of Immature Leydig Cells in Primary Culture

    doi: 10.1210/en.2009-0207

    Figure Lengend Snippet: Effects of UO126 and LY294002 on the proliferative effects of hCG, EGF, or IGF-I on primary cultures of rat Leydig cells. A, Primary cultures of immature rat Leydig cells were incubated with buffer only, hCG, EGF, or IGF-I (each at 100 ng/ml) for 24 h

    Article Snippet: UO126 and LY294002 were from Tocris (Ellisville, MO) or Biomol (Plymouth Meeting, PA).

    Techniques: Incubation

    Effects of UO126 and LY294002 on the antiapoptotic effects of hCG, EGF, or IGF-I on primary cultures of rat Leydig cells. A, Primary cultures of immature rat Leydig cells were preincubated with buffer only, hCG, EGF, or IGF-I (each at 100 ng/ml) for 2

    Journal: Endocrinology

    Article Title: Activation of the Lutropin/Choriogonadotropin Receptor Inhibits Apoptosis of Immature Leydig Cells in Primary Culture

    doi: 10.1210/en.2009-0207

    Figure Lengend Snippet: Effects of UO126 and LY294002 on the antiapoptotic effects of hCG, EGF, or IGF-I on primary cultures of rat Leydig cells. A, Primary cultures of immature rat Leydig cells were preincubated with buffer only, hCG, EGF, or IGF-I (each at 100 ng/ml) for 2

    Article Snippet: UO126 and LY294002 were from Tocris (Ellisville, MO) or Biomol (Plymouth Meeting, PA).

    Techniques:

    Reactivation stimuli increase the number of VZV genomes and transcripts in quiescently-infected human neurons. Wells of quiescently infected neurons were induced to reactivate VZV using growth factor withdrawal (GF n = 2 for each time point) or treatment with PI3K inhibitor LY294002 (LY) 2, 4, or 7 weeks (n = 5 for each time point) after infection. DNA and RNA were extracted from the wells and VZV genomes (A) and transcripts (B) of ORF63 and ORF31 were quantified. Both treatments increased the levels of both viral genomes and transcripts to varying degrees at all time points tested, indicating at least a partial reactivation of VZV. In all experiments reactivating VZV with LY, changes in nucleic acid levels measured were statistically significant.

    Journal: PLoS Pathogens

    Article Title: An In Vitro Model of Latency and Reactivation of Varicella Zoster Virus in Human Stem Cell-Derived Neurons

    doi: 10.1371/journal.ppat.1004885

    Figure Lengend Snippet: Reactivation stimuli increase the number of VZV genomes and transcripts in quiescently-infected human neurons. Wells of quiescently infected neurons were induced to reactivate VZV using growth factor withdrawal (GF n = 2 for each time point) or treatment with PI3K inhibitor LY294002 (LY) 2, 4, or 7 weeks (n = 5 for each time point) after infection. DNA and RNA were extracted from the wells and VZV genomes (A) and transcripts (B) of ORF63 and ORF31 were quantified. Both treatments increased the levels of both viral genomes and transcripts to varying degrees at all time points tested, indicating at least a partial reactivation of VZV. In all experiments reactivating VZV with LY, changes in nucleic acid levels measured were statistically significant.

    Article Snippet: Reactivation using PI3 kinase inhibition was performed using LY294002 hydrochloride (LY, 10 μM, Tocris, cat.# 1130).

    Techniques: Infection

    LY-294002 prevents the increase of CFTR activity induced by dexamethasone in Calu-3 cells. Graphs represent the mean + SEM of I SC in response to 100 nM dexamethasone and LY-294002 for 24 h measured in Ussing chambers. A: Forskolin-induced I SC (n = 12–18, ** p

    Journal: PLoS ONE

    Article Title: Glucocorticoids Distinctively Modulate the CFTR Channel with Possible Implications in Lung Development and Transition into Extrauterine Life

    doi: 10.1371/journal.pone.0124833

    Figure Lengend Snippet: LY-294002 prevents the increase of CFTR activity induced by dexamethasone in Calu-3 cells. Graphs represent the mean + SEM of I SC in response to 100 nM dexamethasone and LY-294002 for 24 h measured in Ussing chambers. A: Forskolin-induced I SC (n = 12–18, ** p

    Article Snippet: To determine the involvement of the phosphoinositide 3-kinase (PI3K), the inhibitor LY-294002 (10 μM, # 1130 TOCRIS Bioscience, Bristol, UK) was used.

    Techniques: Activity Assay

    Functional phosphatidylinositol 3-kinase [PI3-kinase (PI3K)] and CFTR are required for protein kinase B (Akt) 2-stimulated paracellular barrier tightening in human airway epithelial cells. A : human airway epithelial cells express all three isoforms of PKB/Akt as shown by immunoblot. Treatment with bilateral (Bi), apical, or basolateral (Baso) insulin did not change total amounts of Akt protein. B : immunoblot of phospho-Akt1 and phospho-Akt2 after 30 min stimulation with bilateral 125 nM insulin. Calyculin A-treated phosphorylation-positive control Jurkat cells (+ve) were used as a control for phospho-Akt. C : fold-increase of phospho-Akt in n = 3 filters as in B . Akt stimulation was significantly higher for NuLi-1 cells than for CuFi-5 cells. D : insulin treatment significantly increased TER in NuLi-1 cells (red bar). Pretreatment with CFTRInh172 before and during insulin stimulation significantly diminished the insulin-stimulated increase in TER (blue bars). The insulin-stimulated increase in TER of NuLi-1 cells also was prevented by LY-294002 treatment (green bars). E : pretreatment with CFTRInh172 before insulin stimulation also prevented the decrease in the rates of paracellular flux of both calcein (blue bar, left ) and dextran (blue bar, right ) that is normally seen with insulin alone (red bars). NuLi-1 cells pretreated with LY-294002 responded to subsequent exposure to insulin with an increase of both calcein (green bar, left ) and dextran (green bar, right ) flux, whereas insulin alone (red bar) selectively decreased paracellular flux of dextran ( right ). All data are shown as means ± SE and n = 3–4 experiments for each data point for each graph where P ≤ 0.05 (*), 0.01 (**), 0.001 (***), 0.0001 (****), and 0.06 (#) by unprotected two-way ANOVA Fisher’s LSD test.

    Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

    Article Title: Insulin signaling via the PI3-kinase/Akt pathway regulates airway glucose uptake and barrier function in a CFTR-dependent manner

    doi: 10.1152/ajplung.00364.2016

    Figure Lengend Snippet: Functional phosphatidylinositol 3-kinase [PI3-kinase (PI3K)] and CFTR are required for protein kinase B (Akt) 2-stimulated paracellular barrier tightening in human airway epithelial cells. A : human airway epithelial cells express all three isoforms of PKB/Akt as shown by immunoblot. Treatment with bilateral (Bi), apical, or basolateral (Baso) insulin did not change total amounts of Akt protein. B : immunoblot of phospho-Akt1 and phospho-Akt2 after 30 min stimulation with bilateral 125 nM insulin. Calyculin A-treated phosphorylation-positive control Jurkat cells (+ve) were used as a control for phospho-Akt. C : fold-increase of phospho-Akt in n = 3 filters as in B . Akt stimulation was significantly higher for NuLi-1 cells than for CuFi-5 cells. D : insulin treatment significantly increased TER in NuLi-1 cells (red bar). Pretreatment with CFTRInh172 before and during insulin stimulation significantly diminished the insulin-stimulated increase in TER (blue bars). The insulin-stimulated increase in TER of NuLi-1 cells also was prevented by LY-294002 treatment (green bars). E : pretreatment with CFTRInh172 before insulin stimulation also prevented the decrease in the rates of paracellular flux of both calcein (blue bar, left ) and dextran (blue bar, right ) that is normally seen with insulin alone (red bars). NuLi-1 cells pretreated with LY-294002 responded to subsequent exposure to insulin with an increase of both calcein (green bar, left ) and dextran (green bar, right ) flux, whereas insulin alone (red bar) selectively decreased paracellular flux of dextran ( right ). All data are shown as means ± SE and n = 3–4 experiments for each data point for each graph where P ≤ 0.05 (*), 0.01 (**), 0.001 (***), 0.0001 (****), and 0.06 (#) by unprotected two-way ANOVA Fisher’s LSD test.

    Article Snippet: The CFTR-specific inhibitor CFTRInh172 (20 µM, no. 219674; EMD/Millipore) and the phosphatidylinositol 3-kinase (PI3-kinase)-specific inhibitor LY-294002 (20 µM, no. 9901; CST) were used to pretreat ALI cultures bilaterally for 30 min before insulin stimulation and flux/TER measurements and were included with the insulin stimulation solutions during the experiments.

    Techniques: Functional Assay, Positive Control

    Reelin promotes the activation of Syk and STAT3 via integrin β1 A. Reelin induces the activation of Syk and STAT3. H929 cells transfected with pCrl or control plasmid were cultured in 5% BSA- or FN-coated plates for 1 hour. The cells were then subjected to western blotting with phospho-STAT3 (Tyr705), total STAT3, phospho-Syk (Tyr525/526), and total Syk-specific antibodies. B. Densitometric quantification of phospho-STAT3 over total STAT3 from the blot shown in (A). C. Reelin promotes activation of Syk and STAT3 in H929 cells treated with Doxorubicin. H929 cells were transfected with pCrl or reelin-specific siRNAs for 40 hours and were then treated with Dox (2 μM) in the presence or absence of FN. Twenty-four hours later, the cells were subjected to western blotting for the activation of Jak2 (phospho-Jak2, Tyr1007/1008), Syk, and STAT3. D. Densitometric quantification of phospho-STAT3 over total STAT3 from the blot shown in (C). E. The reduced activation of FAK, Src, Syk, and STAT3 caused by reelin-specific siRNAs is ameliorated by the addition of recombinant reelin. Reelin-specific siRNA-transfected H929 cells were treated with Dox in FN-coated wells in the presence of BSA or recombinant reelin. Cell harvesting and western blotting was performed 24 hours later. F. The effects of integrin β1 inhibitory antibody, Src inhibitor, Syk inhibitor and PI3K inhibitor on the activation of β1 signaling pathway. H929 cells were transfected with pCrl or control plasmid. Forty hours later, the cells were cultured in FN-coated plates and were treated with integrin β1 inhibitory antibody P4C10, Src inhibitor PP2, Syk inhibitor BAY 61-3606, or PI3K inhibitor LY 294002 for 24 hours. The cells were then lysed and subjected to western blotting. Data are representative of three independent experiments.

    Journal: Oncotarget

    Article Title: Reelin promotes the adhesion and drug resistance of multiple myeloma cells via integrin β1 signaling and STAT3

    doi: 10.18632/oncotarget.7151

    Figure Lengend Snippet: Reelin promotes the activation of Syk and STAT3 via integrin β1 A. Reelin induces the activation of Syk and STAT3. H929 cells transfected with pCrl or control plasmid were cultured in 5% BSA- or FN-coated plates for 1 hour. The cells were then subjected to western blotting with phospho-STAT3 (Tyr705), total STAT3, phospho-Syk (Tyr525/526), and total Syk-specific antibodies. B. Densitometric quantification of phospho-STAT3 over total STAT3 from the blot shown in (A). C. Reelin promotes activation of Syk and STAT3 in H929 cells treated with Doxorubicin. H929 cells were transfected with pCrl or reelin-specific siRNAs for 40 hours and were then treated with Dox (2 μM) in the presence or absence of FN. Twenty-four hours later, the cells were subjected to western blotting for the activation of Jak2 (phospho-Jak2, Tyr1007/1008), Syk, and STAT3. D. Densitometric quantification of phospho-STAT3 over total STAT3 from the blot shown in (C). E. The reduced activation of FAK, Src, Syk, and STAT3 caused by reelin-specific siRNAs is ameliorated by the addition of recombinant reelin. Reelin-specific siRNA-transfected H929 cells were treated with Dox in FN-coated wells in the presence of BSA or recombinant reelin. Cell harvesting and western blotting was performed 24 hours later. F. The effects of integrin β1 inhibitory antibody, Src inhibitor, Syk inhibitor and PI3K inhibitor on the activation of β1 signaling pathway. H929 cells were transfected with pCrl or control plasmid. Forty hours later, the cells were cultured in FN-coated plates and were treated with integrin β1 inhibitory antibody P4C10, Src inhibitor PP2, Syk inhibitor BAY 61-3606, or PI3K inhibitor LY 294002 for 24 hours. The cells were then lysed and subjected to western blotting. Data are representative of three independent experiments.

    Article Snippet: Immunoblotting and confocal microscopy Following incubation under the indicated conditions, including the addition of integrin β1-blocking antibodies (20 μg/ml), control mouse IgG antibodies (20 μg/ml), Src inhibitor PP2 (1 μmol/L, Merck Millipore), Syk inhibitor IV, BAY 61-3606 (1 μmol/L, Merck Millipore), PI3K inhibitor LY 294002 (50 μmol/L, Cell Signaling Technology), MM cells were collected and washed twice with ice-cold PBS, and incubated for 10 minutes at 4°C in Triton X-100 lysis buffer (30 mM Tris-HCl pH7.5, 150 mM NaCl, 25 mM NaF, 1% Triton X-100, 10% glycerol, 2 mM Naorthovanadate).

    Techniques: Activation Assay, Transfection, Plasmid Preparation, Cell Culture, Western Blot, Recombinant, Cell Harvesting

    Analysis of signal pathways mediating the A2AR-inhibitory effect on LPS-induced neutrophis’ autophagy. Investigation of relative protein, ROS and GSSG during A2AR activation. ( A) WB for p-p38 and the quantification. ( B) WB for p-ERK and the quantification. ( C) Western blot for p-AKT and the quantification. ( D) WB for p-JNK and the quantification. (E) WB for p110γ and the quantification. ( F) The fluorescence of DCF in neutrophils were analyzed by flow cytometry, reflecting the intracellular ROS levels. The cells were treated as indicated for 2 hours and incubated with DCFH-DA (100 μg/ml) for 30 min. The ROS inhibitor NAC (10 mM) were used as a negtive control. ( G) Quantification of DCF fluorescence intensity in ( F ) from each group. (H) The antioxidents, glutathione (GSH) were detected in each neutrophils’ group. The primary neutrophils were treated as indicated for 2 hours. Then cell lysis were processed with GSH detection kit and detected through microplate spectrophotometer. Total glutathione except doubled oxidized glutathione (GSSG) were the concentrations of GSH. ( I) Isolated neutrophil were treated as indicated for 2 hours. The ROS inhibitor NAC (10 mM) were add 15 min before other drugs were used. Then alterations in LC3 were detected by WB, in which GAPDH was used as the internal controls. ( J) Isolated neutrophil were treated as indicated for 2 hours, during which βγ-PI3K inhibitor Gallein (10 μM) and PI3K-AKT inhibitor LY294002 (10 μM) were pretreated for 15 min. Subsequently the total proteins were extracted and probed by WB to detect the LC3 levels. The data are represented as mean ± SEM for three independent experiments. (*p

    Journal: Scientific Reports

    Article Title: Activation of Adenosine 2A receptor inhibits neutrophil apoptosis in an autophagy-dependent manner in mice with systemic inflammatory response syndrome

    doi: 10.1038/srep33614

    Figure Lengend Snippet: Analysis of signal pathways mediating the A2AR-inhibitory effect on LPS-induced neutrophis’ autophagy. Investigation of relative protein, ROS and GSSG during A2AR activation. ( A) WB for p-p38 and the quantification. ( B) WB for p-ERK and the quantification. ( C) Western blot for p-AKT and the quantification. ( D) WB for p-JNK and the quantification. (E) WB for p110γ and the quantification. ( F) The fluorescence of DCF in neutrophils were analyzed by flow cytometry, reflecting the intracellular ROS levels. The cells were treated as indicated for 2 hours and incubated with DCFH-DA (100 μg/ml) for 30 min. The ROS inhibitor NAC (10 mM) were used as a negtive control. ( G) Quantification of DCF fluorescence intensity in ( F ) from each group. (H) The antioxidents, glutathione (GSH) were detected in each neutrophils’ group. The primary neutrophils were treated as indicated for 2 hours. Then cell lysis were processed with GSH detection kit and detected through microplate spectrophotometer. Total glutathione except doubled oxidized glutathione (GSSG) were the concentrations of GSH. ( I) Isolated neutrophil were treated as indicated for 2 hours. The ROS inhibitor NAC (10 mM) were add 15 min before other drugs were used. Then alterations in LC3 were detected by WB, in which GAPDH was used as the internal controls. ( J) Isolated neutrophil were treated as indicated for 2 hours, during which βγ-PI3K inhibitor Gallein (10 μM) and PI3K-AKT inhibitor LY294002 (10 μM) were pretreated for 15 min. Subsequently the total proteins were extracted and probed by WB to detect the LC3 levels. The data are represented as mean ± SEM for three independent experiments. (*p

    Article Snippet: And all the drugs were purchased as follows: Specific A2AR agonist CGS21680 (CGS), A2AR antagonist ZM24138 (ZM) and βγ- phosphatidylinositol kinase (PI3K) inhibitor Gallein (10 μM) were all bought from TOCRIS (Bristol, UK); highly selective PI3K-Akt inhibitor LY 294002 was bought from Cell Signaling Technology (Boston, MA); H-89 from Beyotime (Tianjin,China) or GF109203X (GFX) from SelleckChem (San Diego, CA) were used to inhibit protein kinase A (PKA) or protein kinase C (PKC) substrate phosphorylation and related cellular functions as previous (Both the working dose of the drugs are listed in the ).

    Techniques: Activation Assay, Western Blot, Fluorescence, Flow Cytometry, Cytometry, Incubation, Lysis, Spectrophotometry, Isolation

    TNF activates the PI3K/AKT pathway in HF stem cells. ( a ) Using TMT labelling and affinity enrichment followed by high-resolution LC–MS/MS and quantitative phosphor-proteomics, functional enrichment-based cluster analysis identified up-regulated signals in cultured epidermal stem cells after TNF-α treatment. ( b ) Western blot analysis of p-AKT (at Serine 473) and total AKT in cultured murine epidermal stem cells in the presence of different concentrations of TNF-α for 0.5 h. For all western blot analysis, data are representative of 3–5 independent experiments. ( c ) TNF-α treatment (8 h) resulted in no obvious changes in p-GSK-3β(ser9) and p-β-catenin (Ser33/37/Thr45, Ser675) but significantly increased the level of p-β-catenin (Ser552), which also showed in a TNF-dose-dependent manner, quite similar to p-AKT. ( d ) Perifosine diminished p-β-catenin (Ser552) levels that were elevated by TNF-α in cultured epidermal stem cells. ( e ) Western blot analysis indicated that TNF-α increased the level of β-catenin, and the effect was attenuated by Perifosine or LY294002. ( f ) TCF/LEF Dual-luciferase reporter analysis showed the relative transcription activity in differently treated groups. ( g ) At PWD-3, both p-AKT (Ser473) and p-β-catenin (Ser552) were detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the p-AKT (Ser473) and p-β-catenin (Ser552) were highly co-localized. ( h ) At PWD-3, accumulated β-catenin was detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the accumulated β-catenin was highly co-localized with p-AKT (Ser473). Scale bars, 30 μm.

    Journal: Nature Communications

    Article Title: Macrophages induce AKT/β-catenin-dependent Lgr5+ stem cell activation and hair follicle regeneration through TNF

    doi: 10.1038/ncomms14091

    Figure Lengend Snippet: TNF activates the PI3K/AKT pathway in HF stem cells. ( a ) Using TMT labelling and affinity enrichment followed by high-resolution LC–MS/MS and quantitative phosphor-proteomics, functional enrichment-based cluster analysis identified up-regulated signals in cultured epidermal stem cells after TNF-α treatment. ( b ) Western blot analysis of p-AKT (at Serine 473) and total AKT in cultured murine epidermal stem cells in the presence of different concentrations of TNF-α for 0.5 h. For all western blot analysis, data are representative of 3–5 independent experiments. ( c ) TNF-α treatment (8 h) resulted in no obvious changes in p-GSK-3β(ser9) and p-β-catenin (Ser33/37/Thr45, Ser675) but significantly increased the level of p-β-catenin (Ser552), which also showed in a TNF-dose-dependent manner, quite similar to p-AKT. ( d ) Perifosine diminished p-β-catenin (Ser552) levels that were elevated by TNF-α in cultured epidermal stem cells. ( e ) Western blot analysis indicated that TNF-α increased the level of β-catenin, and the effect was attenuated by Perifosine or LY294002. ( f ) TCF/LEF Dual-luciferase reporter analysis showed the relative transcription activity in differently treated groups. ( g ) At PWD-3, both p-AKT (Ser473) and p-β-catenin (Ser552) were detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the p-AKT (Ser473) and p-β-catenin (Ser552) were highly co-localized. ( h ) At PWD-3, accumulated β-catenin was detected in the wound adjacent to Lgr5 + hair follicle stem cells, and the accumulated β-catenin was highly co-localized with p-AKT (Ser473). Scale bars, 30 μm.

    Article Snippet: A total of 50 μl of LY294002 (PI3K inhibitor, BioVision, USA) in Matrigel (growth factor reduced, BD Biosciences) at a concentration of 200 μM, or an equal amount of Matrigel alone (control), was injected intracutaneously into the backs of 7–8-week-old mice after hair depilation every 48 h for 15 days (7 injections in total).

    Techniques: Liquid Chromatography with Mass Spectroscopy, Mass Spectrometry, Functional Assay, Cell Culture, Western Blot, Luciferase, Activity Assay

    Changes in intracellular ROS levels and WSSV genome copy number after chemical inhibition of the PI3K-Akt-mTOR pathway with LY294002 and inhibition of G6PDH activity with DHEA. Hemocytes were collected from shrimp that had been pretreated with ( A , C ) LY294002/DMSO or PBS/DMSO (control) or ( B,D ) DHEA/DMSO or PBS/DMSO (control), and then injected with WSSV or PBS. ( A,B ) The ROS levels in shrimp hemocytes were quantified by flow cytometry scanning. Each bar indicates the mean ± SD for the WSSV-challenged shrimps and the PBS-injected controls at the indicated time points. ( C,D ) Each bar indicates the mean ± SD of the number of copies of WSSV genomic DNA in the WSSV-injected shrimp pretreated with ( C ) LY294002 or ( D ) DHEA. Single (P

    Journal: Scientific Reports

    Article Title: Six Hours after Infection, the Metabolic Changes Induced by WSSV Neutralize the Host’s Oxidative Stress Defenses

    doi: 10.1038/srep27732

    Figure Lengend Snippet: Changes in intracellular ROS levels and WSSV genome copy number after chemical inhibition of the PI3K-Akt-mTOR pathway with LY294002 and inhibition of G6PDH activity with DHEA. Hemocytes were collected from shrimp that had been pretreated with ( A , C ) LY294002/DMSO or PBS/DMSO (control) or ( B,D ) DHEA/DMSO or PBS/DMSO (control), and then injected with WSSV or PBS. ( A,B ) The ROS levels in shrimp hemocytes were quantified by flow cytometry scanning. Each bar indicates the mean ± SD for the WSSV-challenged shrimps and the PBS-injected controls at the indicated time points. ( C,D ) Each bar indicates the mean ± SD of the number of copies of WSSV genomic DNA in the WSSV-injected shrimp pretreated with ( C ) LY294002 or ( D ) DHEA. Single (P

    Article Snippet: The LY294002 solution (BioVision, Inc., USA) was preserved in 10% DMSO and diluted with PBS before use.

    Techniques: Inhibition, Activity Assay, Injection, Flow Cytometry, Cytometry

    PI3K inhibitors (LY-294002) increase nuclear translocation of BACH2 and enhance MCL cytotoxicity. ( A ) Effect of PI3K inhibitors on nuclear localization of BACH2 was analyzed by immunoblottings in Jeko cells. Cells were pretreated with PI3K inhibitors (LY294002, 50 µM) or Bruton's tyrosine kinase inhibitors (PCI-32765, 1 µM/1 h) followed by bortezomib treatment (20 nM/24h). Bruton's tyrosine kinase inhibitor (PCI-32765) was used as a control, which did not show increased nuclear translocation of BACH2. Cytoplasmic and nuclear fractions for each sample were separated by hypotonic method using a kit. TBP was shown as loading control for nuclear proteins. Controls are Jeko cells without any treatments. 293T cells were used as positive control for BACH2. Data represent four independent experiments. ( B ) BACH2 levels in each sample were plotted in the graph. ( C ) Inhibition of nuclear export of BACH2 by leptomycin-B (LTB) enhanced the bortezomib-induced apoptosis in Jeko cells. Apoptosis was determined by Annexin-V/7-AAD dual staining method using FACS. Error bars indicate the standard error of the means (SEM) from two independent experiments. Results are expressed as an increase in fold of percentage apoptosis compared to control. LTB treatment was given 6 h prior to BTZ treatment for 24 h.

    Journal: PLoS ONE

    Article Title: Nuclear Translocation of B-Cell-Specific Transcription Factor, BACH2, Modulates ROS Mediated Cytotoxic Responses in Mantle Cell Lymphoma

    doi: 10.1371/journal.pone.0069126

    Figure Lengend Snippet: PI3K inhibitors (LY-294002) increase nuclear translocation of BACH2 and enhance MCL cytotoxicity. ( A ) Effect of PI3K inhibitors on nuclear localization of BACH2 was analyzed by immunoblottings in Jeko cells. Cells were pretreated with PI3K inhibitors (LY294002, 50 µM) or Bruton's tyrosine kinase inhibitors (PCI-32765, 1 µM/1 h) followed by bortezomib treatment (20 nM/24h). Bruton's tyrosine kinase inhibitor (PCI-32765) was used as a control, which did not show increased nuclear translocation of BACH2. Cytoplasmic and nuclear fractions for each sample were separated by hypotonic method using a kit. TBP was shown as loading control for nuclear proteins. Controls are Jeko cells without any treatments. 293T cells were used as positive control for BACH2. Data represent four independent experiments. ( B ) BACH2 levels in each sample were plotted in the graph. ( C ) Inhibition of nuclear export of BACH2 by leptomycin-B (LTB) enhanced the bortezomib-induced apoptosis in Jeko cells. Apoptosis was determined by Annexin-V/7-AAD dual staining method using FACS. Error bars indicate the standard error of the means (SEM) from two independent experiments. Results are expressed as an increase in fold of percentage apoptosis compared to control. LTB treatment was given 6 h prior to BTZ treatment for 24 h.

    Article Snippet: PI3K inhibitor (LY-294002) (#L9908), bruton's tyrosine kinase inhibitor (PCI-32765, #S2680) and CAL-101 (#S2226) were purchased from Selleckchem.

    Techniques: Translocation Assay, Positive Control, Inhibition, Staining, FACS