Research Paper Volume 13, Issue 2 pp 2822—2850

Figure 7. shVEGFA and shMMP-2 attenuate the shRac1-mediated effects in BMMSCs. (A) qRT-PCR analysis confirmed shRNA knockdown of each hub gene. (N.S. no significance, *P < 0.05, **P < 0.01, ***P < 0.001, statistically analyzed by one-way ANOVA followed by the Bonferroni correction, n = 3). (B, C) VEGFA and MMP2 protein expression in shLuci, shRac1, shRac1-shVEGFA and/or shRac1-shMMP2 BMMSCs post-OGD 12 h, β-actin was used as the housekeeping protein. (*P < 0.05, ***P < 0.001, statistically analyzed by one-way ANOVA followed by the Bonferroni correction, n = 3). (D–G) Images showing trypan blue-stained shLuci, shRac1, shRac1+shVEGFA, and shRac1+shMMP-2 cells captured under a bright field microscope with scale bar of 100 μm. (H) Quantitative analyses of trypan blue staining showing increased apoptotic rate of BMMSCs-shRac1 co-transfected with shVEGFA or shMMP-2 lentiviruses in comparison with the BMMSCs transfected with shRac1 alone. (N.S. no significance, *P < 0.05, **P < 0.01, statistically analyzed by one-way ANOVA followed by the Bonferroni correction, n = 3). (I) BMMSCs-shRac1 survival was attenuated by the co-transfection of shVEGFA and shMMP-2 lentiviruses, but not by other shRNA, as measured by the CCK-8 analysis (N.S. no significance, *P < 0.05, **P < 0.01, ***P < 0.001, statistically analyzed by one-way ANOVA followed by the Bonferroni correction, n = 3). (J–N) Flow cytometry measured apoptosis of different BMMSCs cell lines. Data further show anti-apoptosis effect of BMMSCs-shRac1 can be weakened by shVEGFA or shMMP-2 co-transfection. (*P < 0.05, ***P < 0.001, statistically analyzed by one-way ANOVA followed by the Bonferroni correction, n = 3). Data are presented as mean ± SD.