骨碎补总黄酮对大鼠骨髓间充质干细胞倍增反应和分化的作用机制实验

王军松; 李佳; 刘桂奇

doi:10.3969/j.issn.2095-5227.2021.02.015

骨碎补总黄酮对大鼠骨髓间充质干细胞倍增反应和分化的作用机制实验

Mechanism of total flavonoids of rhizoma drynariae in multiplication response and differentiation of BMSCs in rats

摘要

摘要:
背景骨碎补总黄酮(total flavonoids of rhizome drynariae，TFRD)是骨碎补的主要活性成分，近年来许多研究表明TFRD可有效提高骨质疏松症患者骨密度，进而促进骨愈合，但目前其作用的具体分子机制尚不完全明确。
目的探讨TFRD对大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells，BMSCs)倍增反应和分化作用的分子机制。
方法获取SD大鼠的骨髓间充质干细胞。细胞培养基中加入TFRD，根据TFRD浓度的不同分为低浓度组(TFRD 0.1 mg/L)、中浓度组(TFRD 1.0 mg/L)、高浓度组(TFRD 10.0 mg/L)，细胞培养基中未加入TFRD的作为对照组。采用CCK8 法检测各组BMSCs细胞增殖情况，茜素红染色法检测各组成骨分化情况，RT-PCR法和Western blot法检测中浓度组与对照组β-catenin、RunX2、PPARG的mRNA和蛋白表达情况。
结果不同时间点低浓度、中浓度、高浓度TFRD组吸光度值与对照组比较均显著升高(P＜0.05)；4组矿化结节数统计，中浓度组高于其他三组，差异有统计学意义(P＜0.05)；中浓度组与对照组比较，β-catenin和RunX2 mRNA表达明显升高，PPARG mRNA表达明显降低(P＜0.05)；中浓度组与对照组比较，β-catenin和RunX2 蛋白表达明显升高，PPARG 蛋白表达明显降低(P＜0.05)。
结论 TFRD可促进BMSCs细胞增殖和成骨分化，其作用机制可能与上调β-catenin、RunX2表达，抑制PPARG表达有关。

Abstract:
Background The total flavonoids of rhizoma drynariae (TFRD) are the main active components of drynaria. In recent years, many studies have shown that TFRD can effectively increase bone mineral density in patients with osteoporosis, thereby promoting bone healing. However, the specific molecular mechanism of its effect is not yet fully understood.
Objective To explore the molecular mechanism of TFRD in multiplication response and differentiation of bone marrow mesenchymal stem cells (BMSCs) in rats.
Methods BMSCs from SD rats were obtained, cultured and identified. TFRD was added to the cell culture medium for intervention. According to the concentration of TFRD, BMSCs were divided into low concentration group (TFRD 0.1 mg/L), medium concentration group (TFRD 1.0 mg/L), high concentration group (TFRD 10.0 mg/L)and control group (without TFRD in the cell culture medium). The proliferation of BMSCs in each group was detected by CCK8 assay, the osteogenic differentiation of each group was detected by alizarin red staining, and the mRNA and protein expressions of β-catenin, RunX2 and PPARG in medium concentration group and control group were detected by RT-PCR and Western blot.
Results Compared with the control group, the absorbance level in the low concentration group, the medium concentration group and the high concentration group were significantly higher at different time points (all P＜0.05). The number of mineralized nodules in the medium concentration group was significantly higher than that in the other three groups (P＜0.05). The mRNA expressions of β-catenin and RunX2 and the protein expressions of β-catenin and RunX2 in the medium concentration group were also significantly higher than those in the control group, while the PPARG mRNA expression and the protein expression of PPARG were significantly lower than that in the control group (all P＜0.05).
Conclusion TFRD can promote cell proliferation and osteogenic differentiation of BMSCs, and the mechanism may be related to up-regulation of β-catenin and RunX2 expressions, and inhibition of PPARG expression.

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