麝香含药血清促进骨髓间充质干细胞迁移及其机制

Mechanism of musk-containing serum in promoting the migration of bone marrow mesenchymal stem cells

目的探讨麝香含药血清对体外骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)迁移的作用及机制。方法将60只SD大鼠随机分为麝香高、中、低剂量组,制备含药血清,另设空白对照组。15只SD大鼠,分离培养BMSCs,将第3代BMSCs通过形态学、表型鉴定、成骨及成脂诱导后鉴定。采用高、中、低组(16.8、8.4、4.2μL/100 g)麝香含药血清干预BMSCs,培养不同时间,MTT法检测细胞增殖情况;Transwell实验检测麝香含药血清及蛋白激酶C(PKC)信号通路阻断剂(GF109203X)干预对BMSCs体外迁移的影响。结果大鼠BMSCs贴壁生长,排列整齐规则,细胞活力好;表型鉴定显示CD44、CD90阳性表达,CD45、CD34阴性表达;可分化为成骨、成脂细胞。不同浓度麝香组在不同时间点的BMSCs增殖率均高于空白对照组(P<0.05);低浓度麝香组(4.2μL/100 g)在24、48、72 h的BMSCs迁移数量较对照组明显增加(P<0.05),低浓度麝香组+阻断剂(GF109203X)组的不同时间点的迁移数量明显减少,但各时间点的组间比较无统计学意义(P>0.05)。结论麝香含药血清可能通过活化PKC信号通路促进BMSCs迁移。

Objective To investigate the effect and mechanism of musk-containing serum on the migration of bone marrow mesenchymal stem cells(BMSCs).Methods Sixty SD rats were randomly divided into four groups:musk-high-,medium-and low-dose groups and blank control group;medicated serum was prepared.Fifteen SD rats were isolated and cultured with BMSCs,and the third generation of BMSCs were identified by morphology,phenotype,osteogenic and adipogenic induction.BMSCs received medicinal healing intervention with high-,medium-and low-(16.8,8.4,and 4.2μL/100 g)musk,and the cell proliferation rate was detected by MTT assay.Under the intervention of the protein kinase C(PKC)signaling pathway(GF109203X),the effect of musk with pharmacition on the migration of BMSCSs was detected with the Transwell test.Results The rat BMSCs were attached to the wall,with orderly arrangement and good cell viability.Phenotypic identification revealed that the expressions of CD44 and CD90 were positive,while the expressions of CD45 and CD34 were negative,and the cells could differentiate into osteoblasts and adipocytes.The proliferation rates of BMSCSs with different concentrations at different time periods were higher than those in the blank control group(P<0.05).The number of BMSCs in the low-concentration musk group(4.2μL/100 g)was significantly increased at 24 h,48 h and 72 h after the addition of the blocking agent GF109203X(P<0.05).The migration quantity of the low-concentration musk group+blocker group(GF109203X)significantly decreased at different time periods,and there was no significant difference between different time groups(P>0.05).Conclusion The mechanism of musk-containing serum in promoting BMSCs migration may be related to the activation of PKC signaling pathway.