Fig 1: Inhibition of BCL-xL protein function increases apoptosis from D7 onwards.a Schematic showing the usage of WEHI-539 or shBCL2L1 to inhibit BCL-xL at D7 during pancreatic specification. b Western blot showing the expression of BCL-2 proteins upon treatment with WEHI-539 on D7 cells. c Immunofluorescence staining for cleaved CASP3 protein on cells treated with DMSO or WEHI-539. Scale bar represents 100 µm. d LIVE/DEAD Viability/Cytotoxicity to quantify the percentage of live and dead cells after treatment with DMSO or WEHI-539 on all eight timepoints during pancreatic differentiation. e Expression of anti-apoptotic and pro-apoptotic gene transcripts upon treatment with WEHI-539 on D7 cells. f Expression of BCL-xL gene transcripts upon treatment with WEHI-539 and QVD-OPh on D7 cells. Error bars indicate standard deviation of three biological replicates undergoing independent differentiations. Asterisk (*) indicates P < 0.05 compared to DMSO control by one-way ANOVA. A representative of at least two independent experiments is shown. “See also Fig. S2”.
Fig 2: Perturbation of BCL-xL early on during pancreas specification has detrimental long-term impact on pancreatic beta cell formation.a Schematic showing 35D differentiation protocol used to differentiate hPSCs into pancreatic beta-like cells. Respective growth factors used are depicted at each time point. b Expression of PDX1, MAFA, INS, and BCL-xL transcripts over the course of 35D differentiation in H9 hESCs. c Expression of PDX1, NKX6.1, MAFA, and INS transcripts upon treatment with WEHI-539 on D8 cells followed by the completion of 35D differentiation in H9 hESCs. Error bars indicate standard deviation of three biological replicates undergoing independent differentiations. Asterisk (*) indicates P < 0.05 compared to DMSO control by one-way ANOVA. A representative of at least two independent experiments is shown.
Fig 3: LINC00173 knockdown restricts the proliferation, migration, and invasiveness but induces the apoptosis of A375 and HT144 cells. (A) LINC00173 expression was evaluated in A375 and HT144 cells through RT-qPCR analysis following transfection of either si-LINC00173 or si-NC. (B) CCK-8 assay was performed to analyze the proliferation of A375 and HT144 cells after LINC00173 silencing. (C) The percentage of apoptotic A375 and HT144 cells transfected with either si-LINC00173 or si-NC was determined by flow cytometry. (D) The cell cycle status of LINC00173-deficient A375 and HT144 cells was determined by flow cytometry. (E) Western blotting was conducted to detect the expression levels of apoptotic-associated proteins (Bcl-2, BAX and Bcl-XL) and cell cycle-associated proteins (CDK-2, CDK-4 and cyclin D1) in A375 and HT144 cells after LINC00173 knockdown. (F and G) Migratory and invasive abilities were determined in LINC00173-deficient A375 and HT144 cells by migration and invasion assays. *P < 0.05 and **P < 0.01.
Fig 4: Inhibition of BCL-xL function perturbs Wnt signaling that may play a role in pancreatic specification.a Hierarchical clustering heatmap analysis of Wnt signaling-associated genes in D7 cells treated with DMSO or WEHI-539. Colors in the heat map depict gene expression in units of SD from the mean across all samples (upregulation in red, downregulation in blue). Expression of b WNT or c SFRP family of transcripts upon treatment with WEHI-539 on D7 cells. d Expression of SFRP5 transcripts over the course of 17D differentiation in H9 hESCs. Error bars indicate standard deviation of three biological replicates undergoing independent differentiations. Asterisk (*) indicates P < 0.05 compared to DMSO control or D0 by one-way ANOVA. A representative of at least two independent experiments is shown. e Immunofluorescence staining for SFRP5 protein on cells treated with DMSO or WEHI-539. Scale bar represents 100 µm. “See also Fig. S3”.
Fig 5: Effect of Res on the expression of apoptotic proteins in H446 cells are assessed by western blotting. (A-D) Res at 40 µg/ml decreased the expression of (B) Bcl-2 and (D) Bcl-xL while it increased (C) Bax expression compared with that of the control (*P<0.05). By contrast, pretreatment of NAC at 2 mmol/l alleviated the effects of Res on the expression of Bcl-2, Bax and Bcl-xL, respectively (#P<0.05 vs. the Res group). Res, resveratrol; NAC, N-acetyl-L-cysteine.
Supplier Page from Abcam for Anti-Bcl-XL antibody [EPR16642]