左归丸含药血清对间充质干细胞向软骨分化过程中骨粘连蛋白基因表达的影响

Effects of Zuogui Pill Medicated Serum on the Expressions SPARC during the Differentiation of Rat Marrow Mesenchymal Stem Cells towards Chondrocytes

目的研究左归丸含药血清对间充质干细胞向软骨分化过程中骨粘连蛋白基因(SPARC)表达的影响。方法诱导间充质干细胞向软骨分化,诱导同时以左归丸含药血清刺激细胞。分别在在0、7、14、21d通过Real-time PCR检测左归丸含药血清对诱导前后SPARC mRNA表达的影响。构建SPARC启动子报告载体并转染骨髓间质细胞系,双荧光素酶试剂盒检测左归丸含药血清SPARC启动子转录活性的影响。结果与空白对照组和单纯诱导组相比,在诱导第21天时,左归丸能明显促进诱导细胞中SPARC mRNA的表达(P<0.05)。与空白组比,当予以左归丸含药血清刺激后,SPARC基因启动子转录活性能增强近1倍(P<0.05)。结论左归丸含药血清能促进间充质干细胞向软骨分化过程中SPARC基因表达,可能主要是通过增强SPARC基因转录活性。

OBJECTIVE To investigate the effects of Zuogui Pill （ZP） medicated serum on the expressions SPARC during the differentiation of rat marrow mesenchymal stem cells towards chondrocytes. METHODS Mesenchymal stem cells （MSCs） isolated from rat bone-marrow were induced differentiation towards ehondrocytes and stimulated with ZP in the meantime. The impact of Zuogui Pill medicated serum on the expression of SPARC mRNA around the induced 0, 7, 14 and 21d respectively was assayed by Real-time PCR. SPARC promoter reporter vector was constructed and transfeeted into bone marrow mesenehy- mal cell line. After cells treated with or without ZP for 48h, cell lysates were prepared and assayed for promoter activity using the Dual-Luciferase reporter assay. RESULTS The expression of the SPARC mRNA was significant increased in ZP groups than those in blank or non-ZP group on the induced 21 day （P〈0.05）. The promoter activity of SPARC displayed a one-fold increasing after ZP stimulation compared to blank group （P〈0.05）. CONCLUSION Zuogui Pill medicated serum can promote SPARC mRNA expression during the differentiation of rat MSCs to chondrocytes mainly through enhancing SPARC gene tran- scription activity.