Journal: Theranostics

Article Title: Cardiomyocyte S1PR1 promotes cardiac regeneration via AKT/mTORC1 signaling pathway

doi: 10.7150/thno.103797

Figure Lengend Snippet: S1PR1 inhibits cardiomyocyte apoptosis via AKT/BCL2 signaling pathways. A-C . Western blot analysis was conducted to assess the levels of total and phosphorylated AKT (Ser473) and BCL2 protein in neonatal mouse cardiomyocytes (NMCMs) treated with or without SEW2871 under the normoxia or 24h-hypoxia/12h-reoxygenation condition ( A ). Quantification of the ratio of p-AKT(Ser473)/t-AKT ( B ), the ratio of BCL2/GAPDH ( C ) (n = 4). D-E . Flow cytometry analysis using Annexin V/PI staining was utilized to evaluate apoptosis in NMCMs treated with or without SEW2871, LY294002 or Bcl2-siRNA under 24h-hypoxic/12h-reoxygenation condition ( D ) and quantification of the percentage of apoptotic NMCMs (Annexin V FITC + /PI - ) ( E ) (n = 5). F-G . Representative immunostaining images of TUNEL positive and α-SA positive NMCMs treated with or without SEW2871, LY294002 or Bcl2-siRNA under 24h-hypoxia/12h-reoxygenation condition (n = 5) ( G ). The arrows indicate α-SA (green) cardiomyocytes positive for TUNEL (magenta). DAPI, nuclear staining (blue). Quantification of the percentage of TUNEL + α-SA + cardiomyocytes ( F ). H/R, 24h-hypoxia/12h-reoxygenation condition. BCL2, B-cell lymphoma 2. SEW2871, S1PR1 agonist. LY294002 (LY), AKT inhibitor. siBcl2 , Bcl2-siRNA . Data are represented as means ± S.E.M. P < 0.05 indicates significant statistical differences. n.s., no statistical significance. Scale bar: G , 50 µm. One-way ANOVA ( B - C and E - F ).

Article Snippet: Cells were then stained propidium iodide (PI) and FITC Annexin V according to the manufacturer's instructions using FITC Annexin V Apoptosis Detection Kit (#C1062M, Beyotime).

Techniques: Protein-Protein interactions, Western Blot, Flow Cytometry, Staining, Immunostaining, TUNEL Assay

Journal: Theranostics

Article Title: Cardiomyocyte S1PR1 promotes cardiac regeneration via AKT/mTORC1 signaling pathway

doi: 10.7150/thno.103797

Figure Lengend Snippet: A work model for cardiomyocyte S1PR1 promotes cardiomyocyte proliferation and cardiac regeneration. S1PR1 plays a key regulator to regulate cardiomyocyte proliferation during the process of cardiac regeneration. Mechanistically, S1P-S1PR1 signaling promoted cardiomyocyte proliferation and inhibits CM apoptosis via AKT/mTORC1/CYCLIN D1 and BCL2 pathway, respectively. CM-S1PR1 overexpression promotes cardiac regeneration and improves cardiac functions of injured hearts, providing a future promising CM-target therapy for myocardial infarction and heart failure through the S1PR1 signal pathway.

Article Snippet: Cells were then stained propidium iodide (PI) and FITC Annexin V according to the manufacturer's instructions using FITC Annexin V Apoptosis Detection Kit (#C1062M, Beyotime).

Techniques: Over Expression

Journal: Nature Communications

Article Title: Targeted delivery of celastrol to mesangial cells is effective against mesangioproliferative glomerulonephritis

doi: 10.1038/s41467-017-00834-8

Figure Lengend Snippet: Anti-inflammatory effects of CLT and CLT-AN in anti-Thy1.1 nephritic rats. a – d Real-time PCR analysis of renal mRNA levels of MCP-1 ( a ), ICAM-1 ( b ), IL-6 ( c ), IL-1β ( d ) in anti-Thy1.1 nephritic rats on day 1 after treatment with CLT or CLT-AN. Data are shown as normalized fold expressions relative to normal group using β-actin mRNA as internal control. e Representative photomicrographs of immunostaining for MCP-1, ICAM-1, IL-6, and IL-1β in kidney tissue sections taken from anti-Thy1.1 nephritic rats on day 1 after treatment with CLT or CLT-AN. Scale bars , 20 μm. f The levels of MCP-1, ICAM-1, IL-6, and IL-1β were semiquantitatively scored as described in Methods on the basis of immunohistochemical results. In panels a – d and f data are mean ± s.d. ( n = 5), results are representative of two independent experiments. * P < 0.05 vs. PBS group; # P < 0.05 vs. CLT group. Statistical significance was determined by one-way ANOVA with Tukey post hoc test

Article Snippet: After addition of 100 μL Agent A (horseradish peroxidase-conjugated ChemMate Envision reagent) from the anti-rabbit/rat general immunohistochemical test kit (Envision Detection Kit, GK500705), the color reaction was performed using 3,3-diaminobenzidine (Sigma-Aldrich, USA), then each section was counterstained with hematoxylin.

Techniques: Real-time Polymerase Chain Reaction, Control, Immunostaining, Immunohistochemical staining

Journal: Nature Communications

Article Title: Targeted delivery of celastrol to mesangial cells is effective against mesangioproliferative glomerulonephritis

doi: 10.1038/s41467-017-00834-8

Figure Lengend Snippet: Anti-proliferative effects of CLT and CLT-AN in vitro and in vivo. a Effects of CLT and CLT-AN on PDGF-BB-induced proliferation of HBZY-1 cells. b Flow cytometry analysis of the effects of CLT and CLT-AN on the cell cycle distribution of HBZY-1 cells in the presence of PDGF-BB ( C CLT = 0.25 μg mL −1 ). c Representative quadrant plot obtained by flow cytometry analysis showing the ability of CLT and CLT-AN to induce apoptosis in HBZY-1 cells in the presence of PDGF-BB. d Flow cytometry analysis of the proportions of apoptotic HBZY-1 cells after 24-h exposure to CLT or CLT-AN in the presence of PDGF-BB ( C CLT = 0.5 μg mL −1 ). In panels a , b , and d data are mean ± s.d. ( n = 3), results are representative of three independent experiments. * P < 0.05. Statistical significance was determined by one-way ANOVA with Tukey post hoc test. e Real-time PCR analysis of renal mRNA levels of PDGF-BB in anti-Thy1.1 nephritic rats on day 5 after treatment with CLT or CLT-AN. Data are shown as normalized fold expressions relative to normal group using β-actin mRNA as internal control. f Representative photomicrographs of immunostaining for PDGF-BB in kidney tissue sections from anti-Thy1.1 nephritic rats on day 5 after treatment with CLT or CLT-AN, and the levels of PDGF-BB were semiquantitatively scored as described in Methods on the basis of immunochemical results. Scale bars , 20 μm. In panels e , f data are mean ± s.d. ( n = 5), results are representative of two independent experiments. * P < 0.05 vs. PBS group; # P < 0.05 vs. CLT group. Statistical significance was determined by one-way ANOVA with Tukey post hoc test

Article Snippet: After addition of 100 μL Agent A (horseradish peroxidase-conjugated ChemMate Envision reagent) from the anti-rabbit/rat general immunohistochemical test kit (Envision Detection Kit, GK500705), the color reaction was performed using 3,3-diaminobenzidine (Sigma-Aldrich, USA), then each section was counterstained with hematoxylin.

Techniques: In Vitro, In Vivo, Flow Cytometry, Real-time Polymerase Chain Reaction, Control, Immunostaining

Journal: Nature Communications

Article Title: Targeted delivery of celastrol to mesangial cells is effective against mesangioproliferative glomerulonephritis

doi: 10.1038/s41467-017-00834-8

Figure Lengend Snippet: Anti-fibrotic effects of CLT and CLT-AN in anti-Thy1.1 nephritic rats. a – d Real-time PCR analysis of renal mRNA levels of Col I ( a ), Col IV ( b ), FN-1 ( c ), TGF-β 1 ( d ) in anti-Thy1.1 nephritic rats on day 5 after treatment with CLT or CLT-AN. Data are shown as normalized fold expressions relative to normal group using β-actin mRNA as internal control. e Representative photomicrographs of immunostaining for TGF-β 1 in kidney tissue sections taken from anti-Thy1.1 nephritic rats on day 5 after treatment with CLT or CLT-AN. Scale bars , 20 μm. f The levels of TGF-β 1 were semiquantitatively scored as described in Methods on the basis of immunochemical results. In panels a – d , f data are mean ± s.d. ( n = 5), results are representative of two independent experiments. * P < 0.05 vs. PBS group; # P < 0.05 vs. CLT group. Statistical significance was determined by one-way ANOVA with Tukey post hoc test

Article Snippet: After addition of 100 μL Agent A (horseradish peroxidase-conjugated ChemMate Envision reagent) from the anti-rabbit/rat general immunohistochemical test kit (Envision Detection Kit, GK500705), the color reaction was performed using 3,3-diaminobenzidine (Sigma-Aldrich, USA), then each section was counterstained with hematoxylin.

Techniques: Real-time Polymerase Chain Reaction, Control, Immunostaining