壮骨强筋片含药血清对老年大鼠骨髓基质干细胞成脂分化的影响

Effects of Serum Containing Zhuanggu Qiangjin Tablets on Adipogenic Differentiation of Bone Marrow Stromal Stem Cells of Aged Rats

【目的】观察壮骨强筋片含药血清对老年大鼠骨髓基质干细胞(bone marrow stromal stem cells,BMSCs)成脂分化的影响。【方法】取18月龄雄性SD大鼠,采用全骨髓贴壁法培养BMSCs,传代3次后用于实验。采用流式细胞仪检测细胞表面蛋白标志CD34、CD31、CD29。将细胞分为成脂诱导组、成脂诱导中药组、中药组、空白组。14 d后进行油红O染色,计算脂肪细胞转化率,并进行油红O染色定量分析;采用逆转录—聚合酶链反应(RT-PCR)法检测脂蛋白脂酶(lipoprotein lipase,LPL)和过氧化物酶增殖活化受体(peroxisome proliferator activated receptor,PPARγ2)mRNA表达。【结果】CD29表达为阳性,CD31和CD34表达均为阴性。油红O染色结果显示:成脂诱导组、成脂诱导中药组镜下观察可见有红色脂滴,成脂诱导中药组脂肪细胞转化率显著低于成脂诱导组(P<0.05)。油红O染色定量分析吸光度(D)值检测结果显示:成脂诱导中药组油红O含量显著低于成脂诱导组(P<0.01)。成脂诱导组LPL基因相对D值显著高于空白组(P<0.01)。成脂诱导中药组LPL基因相对D值较成脂诱导组显著降低(P<0.05),成脂诱导组PPARγ2基因相对D值显著高于空白组(P<0.01),中药组PPARγ2基因相对D值显著低于空白组(P<0.05),成脂诱导中药组PPARγ2基因相对D值显著低于成脂诱导组(P<0.01)。【结论】壮骨强筋片含药血清可抑制老年大鼠骨髓基质干细胞的成脂分化,可能是其防治老年性骨质疏松症的机制之一。

Objective To study the the influence of serum containing Zhuanggu Qiangjin Tablets （ZQT） on adipogenic differentiation of bone marrow stromal stem cells （BMSCs） of aged rats. Methods The whole bone marrow of male SD rats aged 18 months was got out for the culture of BMSCs by adherence method. After subcuhuring for 3 generations, the cultured BMSCs were used for the experiment. Flow cytometry was used to detect the cell surface protein markers of CD34, CD31 and CD29. BMSCs were divided into adipogenic induction group, Chinese medicine ZQT plus adipogenic induction group, ZQT group, and the blank group. After 14 days, oil red 0 dyeing was performed for the calculation of fat cell differentiation ratio, and oil red O staining quantitative analysis was carried out. Lipoprotein lipase （LPL） and peroxidase proliferation activated receptor （PPAR＇y 2） mRNA contents were detected with reverse transcription -polymerase chain reaction （PCR）. Results CD29 expression was positive, but CD31 expression and CD34 expression were negative. The results of oil red O staining showed that red lipid drops were visible in adipogenic induction group and ZQT plus adipogenic induction group, and ZQT plus adipogenic induction group had lower fat cell differentiation ratio than adipogenic induction group （P〈0.05） The results of oil red O staining quantitative analysis showed ZQT plus adipogenic induction group had lower oil red content than adipogenic induction group （P 〈 0.01） . LPL gene relative OD value was markedly higher in adipogenic induction group than that in the blank group （P〈0.01）, while was lower in ZQT plus adipogenic induction group than that in adipogenic induction group （P〈0.05）. PPARγ2 gene relative OD value was higher in adipogenic induction group （P〈0.01） while was lower in ZQT group （P〈0.05） than that in the blank group, and was lower in ZQT plus adipogenic induction group than that in adipogenic induction group （P 〈 0.01） . Conclusion Serum containing ZQT can inhibit the adipogenic differentiation of BMSCs of aged rats, which may contribute to its preventive and therapeutic mechanism for senile osteoporosis.