Figure 7

Overexpression of hBcl2 enhances the survival of BMSCs in the adverse microenvironment and promotes the repair of the injured spinal cord. (A) Schematic representation. BMSCs in the hBcl2, cb, and NC groups were transplanted in situ 3 days after SCI, and samples were collected 11 days later. (B) A spinal cord sagittal fluorescence labelling was used to determine the survival and distribution of BMSCs (GFP, green) in the hBcl2, cb, and NC transplantation groups 14 days post injury (d.p.i.). The arrows show the survived transplanting BMSCs, the asterisks denote the damage sites, and DAPI denotes all viable cells, scale bar = 200 μm (left), scale bar = 20 μm (right); (C) The apoptosis of BMSCs transplanted rats at the injury sites at 14 d.p.i. was evaluated by using spinal cord sagittal immunofluorescence staining. Dead cells were designated by cleaved caspase3 positive cells (red), BMSCs overexpressing hBcl2 protein in the transplanted area were labeled by hBcl2 protein (green), and nuclei were labeled by DAPI, scale bar = 100 μm. (D) Quantification the intensity of cleaved caspase3 staining in the spinal cord segment spanning the injured core at 14 d.p.i. (At least three independent replicates were performed for each experiment. Data are presented as mean ± SD, **P < 0.01, ***P < 0.001, vs. hBcl2, analyzed by one-way ANOVA, followed by Tukey’s post hoc test). (E) Spinal cord sagittal immunofluorescence was used to evaluate the expression and distribution of GFAP in BMSCs transplanted rats in each transplantation group, scale bar = 200 μm. (F) Quantification the intensity of GFAP staining in the spinal cord segment spanning the injured core at 14 d.p.i. (At least three independent replicates were performed for each experiment. Data are presented as mean ± SD, ***P < 0.001, **P < 0.01, vs. hBcl2, analyzed by one-way ANOVA, followed by Tukey’s post hoc test). (G) In the injured core of rats in the hBcl2 and cb BMSCs transplantation groups, the formation of glial scar (GFAP, green) and the regeneration and preservation of neuronal axons (NF200, red) were measured using immunofluorescence staining. Scale bar = 200 μm (left), scale bar = 100 μm (right). (H) Quantification the intensity of NF200 staining in the spinal cord segment spanning the injured core at 14 d.p.i. (Three independent replicates were performed in the experiment. Data are presented as mean ± SD, **P < 0.01, analyzed by t-test).