金黄色葡萄球菌肠毒素C_2对兔骨髓间充质干细胞增殖和成骨分化的实验研究

Staphylococcus aureus enterotoxin C_2 induced differentiation of rabbit bone marrow mesenchymal stem cells in vitro experimental research

目的观察不同浓度金黄色葡萄球菌肠毒素C2(SEC2)对体外兔骨髓间充质干细胞(rBMSCs)的诱导分化作用。方法以梯度密度离心法获得rBMSCs并通过激光共聚焦对细胞进行形态学鉴定后传代培养,分别应用1、10、100、500μM SEC2处理细胞,检测各组细胞的碱性磷酸酶(ALP)活性;四甲基偶氮唑蓝(MTT)比色法检测各组细胞的增殖情况;逆转录-聚合酶链反应(RT-PCR)法检测骨桥蛋白和胶原蛋白的基因表达。结果 rBMSCs在应用SEC2诱导时,随着浓度的增加,rBMSCs的增殖逐渐被抑制,其中100μM SEC2共培养细胞增殖情况明显高于对照组(P<0.01);应用100μM SEC2诱导rBMSCs 16d后,成骨诱导检测茜素红染色可见矿化和钙沉积;同时进行RT-PCR检测,随着培养时间的延长,骨桥蛋白(Osteopontin)和胶原蛋白-1(Collagen I)有明显上调趋势,差异均有统计学意义(P<0.05)。结论 SEC2能够使rBMSCs成骨分化,分化的效果与肠毒素剂量及诱导时间相关。

Objective To observe the induction and differentiation of Staphylococcus aureus enterotoxin C 2 （SEC2） on the growth of cultured rabbit bone marrow mesenchymal stem cells （rBMSCs） in vitro co-cultivation. Methods rBMSCs were obtained through the density gradient centrifugation and were subcultured after morphologi -cal identification.Cells were processed by applying 1,10,100,500 μM SEC2,the expression of alkaline phospha-tase （ALP） activity was detected,the cell proliferation was detected by using tetrazolium blue （MTT） assay,and os-teopontin and collagen gene expressions were detected by RT-PCR assay .Results When applying SEC 2 for induc-tion of rBMSCs,rBMSCs proliferation was inhibited with the increase of concentration （P〈0.01）;The co-cultivation cell proliferation with 100μM SEC2 was obviously higher than that of control group （P〈0.01）;16 days after induc-tion of rBMSCs with 100μM SEC2 ,mineralization and Calcium deposition were observed with osteogenesis induction detection;at the same time,on RT-PCR test,osteopontin and Collagen I were obviously increased along with the ex-tension of incubation time,indicating statistically significant differences （P〈0.05）.Conclusion Staphylococcus aureus enterotoxin C 2 can induce RBMSCs osteogenic differentiation ,the effect of differentiation is related to the en-terotoxin dose and the induction time .