Fig 2: Anti-apoptotic effects of the SVF cell secretome on TNF-α-induced apoptosis. a Western blot showing the effects of SVF on the activation of caspase-3 and caspase-7 as well as the cleavage of PARP in myoblasts treated with 10 ng/ml TNF-α for 24 h. b, c Caspase 3/7 activity (b) and caspase 8 activity (c) in TNF-α treated myoblasts (10 ng/ml) after 24 h of indirect co-culture with SVF. d The percentage of Annexin-V positive myoblasts in SVF co-cultures after 24 h of TNF-α treatment (10 ng/ml). e The percentage of TUNEL-positive myoblasts in SVF co-cultures after 48 h of TNF-α treatment (10 ng/ml). n = 6. f TUNEL labeling in myoblasts indirectly co-cultured with SVF cells. n = 3. DAPI staining shown in the upper left corner. × 20 magnification. Statistical significance was calculated using one-way ANOVA with post hoc test (Bonferroni correction). **P < 0.01, ***P < 0.001. ns = not significant

Fig 3: Effect of transfection with a PARP1-knockdown lentivirus on the apoptosis of chondrocytes. (A) eGFP fluorescence intensity under different treatment conditions and transfection with the PARP1-knockdown lentivirus. Magnification, ×100. PARP1 (B) mRNA and (C) protein expression levels after transfection with three different PARP1-knockdown lentiviruses. *P<0.05 vs. si-NC group. (D) Quantitative analysis of the relative protein expression levels of PARP1. *P<0.05 vs. si-NC group. (E) Apoptosis of chondrocytes in groups following PARP1 knockdown. (F) Analysis of chondrocyte apoptosis rate in the different groups. *P<0.05 vs. si-NC group; #P<0.05 vs. si-NC+ H2O2 group. si, small interfering RNA; NC, negative control; MOI, multiplicity of infection; PARP1, poly [ADP-ribose] polymerase-1; H2O2, hydrogen peroxide.

Fig 4: Mithramycin’s effect on immunofluorescent markers of caspase activation in neurons is cell death model-specific.RCN were treated with 200 nM doxorubicin, 50 µM etoposide, 50 μM C2-ceramide, or 0.5 μM staurosporine +/−200 nM mithramycin. After 6 h, cells were fixed and stained for neuronal markers (MAB2300, Neuro-Chrom™ Pan-Neuronal Marker), DAPI and cleaved Casp3 or cleaved PARP, imaged, and quantified. A representative field for each image at 63x magnification is shown. The Milli-MarkTM Pan-Neuronal Marker is not able to successfully stain those neurons who are close to the end stages of cell death (caspase activation); however, white arrows indicate cleaved Casp3/cleaved PARP cells that show a clear neuronal morphology. Quantification was performed on at least seven fields for each treatment. Data all represent mean ± SD. Significance assigned based on one-way ANOVA and Tukey post hoc test; *p < 0.05, ***p < 0.001, ****p < 0.0001 vs control; ^^p < 0.01, ^^^p < 0.001, ^^^^p < 0.0001 vs cell death inducer alone.

Fig 5: The ROCK1 activation inhibitor Y-27632 attenuates MPP+-induced Drp1-dependent aberrant mitochondrial fission and apoptosis through the inhibition of Drp1 dephosphorylation/activation.a PC12 cells were treated with MPP+ (1 mM) alone or pretreated with ROCK1 activation inhibitor Y-27632 (50 µM) for 2 h and then treated with MPP+ (1 mM) for 24 h. The expression of ROCK1 and CF of ROCK1 in whole-cell lysates was determined by western blot analysis. b The expression of p-Drp1 (Ser 656) was determined by western blot analysis. c The expression of Drp1 in mitochondrial lysates (Mito) and in cytosolic fractions (Cyto) was determined by western blot analysis. d Cells were transfected with a DsRed-Mito plasmid, and the mitochondria were viewed by confocal microscopy. Scale bars: 5 μm. e Mitochondrial length was quantified using the Imaris software. f The expression of C-Cas 3 and C-PARP in whole-cell lysates was determined by western blot analysis. g, h The apoptosis rate was measured by flow cytometry using annexin V-FITC/PI staining. The data are expressed as the means ± S.D. (n = 3). ***P < 0.001