脑源性神经营养因子前体在脊髓损伤后对少突胶质细胞存活及生长的影响

Effects of precursor of brain-derived neurotrophic factor on the survival and growth of oligodendrocytes after spinal cord injury

目的 观察脑源性神经生长因子前体（proBDNF）在少突胶质细胞前体细胞（OPCs）的表达及其对OPCs存活及生长的调控作用.方法 取新生C57BL/6小鼠大脑皮层分离少突胶质细胞前体细胞,体外原代培养.应用Western blot法观察proBDNF及其受体p75NTR和sortilin在OPCs内的表达水平;设置100 μg/L胎牛血清白蛋白（BSA）对照组和proBDNF梯度浓度组（1、10、100 μg/L）,每组6孔细胞.应用结晶紫及β-tublin染色,观察OPCs细胞形态,评估proBDNF对OPCs生长的影响.结果 Western blot结果显示OPCs细胞内源性表达proBDNF及其受体p75 NTR和sortilin;与BSA对照组比较,proBDNF治疗组OPCs的数量显著减少[（537.67±34.93）个比（440.67±9.29）、（390.67±9.45）、（337.25±7.27）个],差异有统计学意义（P＜0.01）;同时proBDNF可以抑制OPCs的生长,与BSA对照组比较,proBDNF治疗组OPCs的长突起减少,照片显示细胞平均面积减小显著[（2.44±0.71） μm2比（1.65±0.12）、（1.30±0.11）、（0.95±0.08）μm2],差异均有统计学意义（1 μg/L proBDNF组P＜0.05,其他组P＜0.01）.结论 OPCs内源性表达proBDNF及其受体.proBDNF对OPCs的存活及生长具有抑制作用,该作用可能通过p75 NTR-sortilin通路介导.

Objective To investigate whether the precursor of Brain-drived neurotrophic factor （proBDNF） can be expressed on oligodenerocyte precursor cells （OPCs） and the effects of proBDNF on the survival and growth of OPCs in the acute period of spinal cord injury （SCI）.Methods Primary cuhure of OPCs from neonatal C57BL/6 mice was applied for the studies.Western blotting was used to investigate the expression levels of proBDNF and its receptors p75NTR and sortilin in OPCs.100 μg/L bovine serum albumin （BSA） was used as a control group and gradient concentrations of proBDNF （1,10,100 μg/L） were used to treat the cells.Each group had 6 wells of cultured cells.After the treatment,crystal violet staining and β-tublin immunofluorescence staining were used to observe the size and growth of cells.Results Western blotting showed that proBDNF,p75NTR and sortilin were endogeneously expressed on OPCs.As compared with the BSA control,proBDNF inhibited survival of OPCs （537.67 ± 34.93 vs.440.67 ± 9.29,390.67 ± 9.45 and 337.25 ± 7.27） and the difference was statistically significant （P ＜ 0.001）.Meanwhile,proBDNF also inhibited the OPCs growth as compared with BSA group.The OPCs in proBDNF treated groups had less long projections and smaller size [（2.44 ± 0.71） vs.（1.65 ± 0.10）,（1.30 ± 0.11） and （0.95 ± 0.08） μm2].The difference was statistically significant （P ＜ 0.05 in 1 μg/L proBDNF group,and P ＜ 0.01 in other groups）.Conclusion OPCs express proBDNF and its receptors endogenously.proBDNF can inhibit the survival and growth of OPCs and the inhibitory effects may be induced by p75NTR-sortilin pathway.