P物质受体在大鼠骨髓间充质干细胞成骨分化过程中的表达

Expression of substance P receptor in bone marrow mesenchymal stem cells during osteogenic differentiation

目的 比较大鼠骨髓间充质干细胞（BMSCs）成骨分化过程中P物质（SP）受体的表达变化.方法 采用全骨髓法体外分离、培养SD大鼠BMSCs,分为诱导成骨组与未诱导组,在传代培养的不同时期（1、2、3周）,采用免疫细胞化学、Western blot对BMSCs的成骨细胞诱导进行鉴定,采用逆转录-聚合酶链反应（RT-PCR）、Western blot检测各组细胞SP受体mRNA、蛋白的表达,观察时间-效应关系.结果 （1）rBMSCs及其诱导的成骨细胞有SP受体表达.（2）SP受体mRNA在诱导1周时下降（P〈0.05）,在诱导至3周时显著增高（P〈0.05）.（3）同一时间点诱导组SP受体的蛋白水平高于未诱导组（P〈0.05）,诱导组SP受体蛋白水平在第3周时显著增高（P〈0.05）,呈明显的时间依赖性.结论 大鼠BMSCs及其诱导的成骨细胞表达SP受体,随着成骨诱导的不断进行,SP受体在成骨分化的末期大量表达.

Objective To compare the expression of substance P （SP） receptor in rat bone marrow mesenchymal stem cells （BMSCs） according to the stage of osteoblast differentiation. Methods The rBMSCs were isolated in vitro using whole bone marrow adherence method, and classified as osteoblast-in-duced group and non-induced group. In the different periods of culture （1,2, and 3 weeks） , identification of osteoblasts was performed by immunocytochemistry and reverse transcriptase polymerase chain reaction （ RT-PCR）. The expression of the SP receptor was detected by using Western blotting and RT-PCR. Results There was SP receptor expression in the cultured BMSCs and osteoblasts. RT-PCR demonstrated that the expression of SP receptor in osteoblast-induced group was lower than in non-induced group at the week one （P〈0.05 ） , but significantly increased at the week three（P〈0.05 ）. In osteoblast-induced group the expression of SP receptor was higher than in non-induced group at the same time point （P 〈0.05）, and that was significantly increased time-dependently at the week three （P〈0.05）. Conclusion SP receptor exists in the BMSCs and osteoblasts. SP receptor expression was significantly increased at late-stage of osteogenic differentiation.