Figure 6 | Scientific Reports

Figure 6

From: hBcl2 overexpression in BMSCs enhances resistance to myelin debris-induced apoptosis and facilitates neuroprotection after spinal cord injury in rats

Figure 6

BMSCs overexpressing hBcl2 could effectively degrade myelin debris through the lysosomal pathway to promote cell survival. (A) Immunofluorescence staining was used to measure Dil-myelin (red, 500 μg/mL) phagocytosis by BMSCs in the hBcl2 group at 1, 3, and 5 days after treatment with Dil-myelin (red, 500 μg/mL) CM, and all cell nuclei were labeled with DAPI (blue). (B) Quantitative analysis of the proportion of cells in which Dil-myelin (red, 500 μg/mL) accounted for 1/3 of the total cell volume and more in the total cell number swallowed by BMSCs in the hBcl2 group at 1, 3, and 5 days after treatment with Dil-myelin (red, 500 μg/mL) CM (At least three independent replicates were performed for each experiment. Data are presented as mean ± SD, ****P < 0.0001, **P < 0.01, Day1 vs. Day3, Day1 vs. Day5 analyzed by one-way ANOVA, followed by Tukey’s post hoc test). (C) At 1, 3, and 5 days after treatment with Dil-myelin (red, 500 μg/mL) CM, immunofluorescence labeling was employed to determine the activation of BMSCs (GFP, green) and lysosomes (Lamp1, purple) in the hBcl2 group. (D) Quantitative analysis of BMSCs in the hBcl2 group's lysosomal (Lamp1, purple) fluorescence intensity after treatment with Dil-myelin CM at 1, 3, and 5 days. (At least three independent replicates were performed for each experiment. Data are presented as mean ± SD, *P < 0.05, **P < 0.01, vs. Day1, NS, no significant difference, Day3 vs. Day5 analyzed by one-way ANOVA, followed by Tukey’s post hoc test). (E) Apoptosis was measured by cleaved caspase3 immunofluorescence labeling of BMSCs in the hBcl2 group at 1, 3 and 5 days following treatment with Dil-myelin CM. (F) quantification of the proportion of cleaved caspase3 positive cells in the total number of cells (At least three independent replicates were performed for each experiment. Data are presented as mean ± SD, ****P < 0.0001, vs. Day1, NS, no significant difference, Day3 vs. Day5 analyzed by one-way ANOVA, followed by Tukey’s post hoc test). Scale bars = 20 μm (A,C), scale bar = 50 μm (E).

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