不同培养体系中施万细胞对BMSCs分化及分泌功能的影响

Effects of Schwann Cells on Differentiation and Secretion of BMSCs in Different Culture Systems

目的观察不同培养体系中施万细胞(Schwann cells, SCs)对骨髓基质干细胞(bone marrow stromal cells, BMSCs)分化以及分泌功能的影响。方法从预变性SD大鼠坐骨神经中分离纯化SCs,采用全骨髓贴壁法分离培养SD大鼠BMSCs,均行免疫荧光鉴定。根据培养体系不同将实验分为3组:A组,SCs与PKH26标记的BMSCs在Transwell中接触共培养组;B组,SCs与PKH26标记的BMSCs在Transwell中非接触培养组;C组,PKH26标记的BMSCs单独培养组。采用Western blot检测各组S-100蛋白表达,应用ELISA法检测各组上清液中神经营养因子3(neurotrophins-3, NT-3)的含量。结果成功分离并培养SCs和BMSCs,免疫荧光鉴定SCs呈S-100阳性表达,BMSCs呈CD44和CD90阳性表达;Western blot检测显示,A组S-100蛋白表达水平明显高于B、C组(P<0.05),B组S-100蛋白表达水平高于C组(P<0.05);ELISA检测显示,A组上清液中NT-3含量显著高于B、C组,且呈现时间依赖性增高(P<0.05)。结论在接触共培养体系中,SCs能够有效诱导BMSCs向外周胶质细胞分化,并增强BMSCs的分泌功能。

Objective To explore the effects of Schwann cells(SCs) on differentiation and secretion of bone marrow stromal cells(BMSCs) in different culture systems. Methods SCs were extracted and puri?ed from the injured distal sciatic nerves of Sprague-Dawiey(SD) rats. BMSCs were isolated from bone marrow of SD rats and cultured in vitro. The cells were identi?ed by immuno?uorescent staining. According to the different culture systems, the experiments were divided into three groups: group A, PKH26-labeled BMSCs were co-cultured with SCs on the lower layer of transwell;group B, PKH26-labeled BMSCs and SCs were cultured on the upper and lower layers of transwell;group C, PKH26-labeled BMSCs were cultured on the lower layers of transwell. The protein expression of S-100 in each group was detected by Western blot. The content of neurotrophins-3(NT-3) in the supernatant of each group was detected by ELISA. Results SCs and BMSCs were isolated and cultured successfully. The identi?cation of SCs showed positive expression of S-100 and BMSCs showed positive expressions of CD44 and CD90. Western blot analysis showed that the expression level of S-100 in group A was significantly higher than that in the group B and C(P<0.05),and the level of S-100 in group B was higher than that in group C(P<0.05). ELISA showed that the content of NT-3 in supernatant of group A was significantly higher than that in the group B and C, and showed time-dependent increase(P<0.05). Conclusion In the contact co-culture system, SCs can effectively induce BMSCs differentiation into peripheral glial cells and enhance BMSCs secretory function.