补肾健脾方对尾部悬吊大鼠骨髓间充质干细胞成脂分化的影响

Effects of Bushen Jianpi Decoction on Adipogenic Differentiation of Bone Mesenchymal Stem Cells in Tail Suspension Model Rats

目的:观察补肾健脾方对尾部悬吊大鼠骨髓间充质干细胞(BMSCs)成脂分化能力的影响。方法:大鼠随机分为正常组、悬吊组、补肾健脾方高、中、低剂量组共5组。除正常组外,其余各组大鼠头低位-30度尾部悬吊连续21 d模拟失重,后3组大鼠从实验第1天开始分别按剂量11.4,5.7,2.8 g·kg-1·d-1给予补肾健脾方ig 21 d,其余各组大鼠ig等容积的生理盐水。全骨髓贴壁法体外分离培养大鼠骨髓间充质干细胞,并进行成脂分化诱导,油红O染色法检测脂滴形成情况,Real Time PCR法和Western bolt法分别检测过氧化物酶体增殖物激活受体γ(PPARγ)mRNA和蛋白表达,在成脂诱导分化的第7,14,21天分别检测细胞内甘油三酯(TG)含量。结果:较正常组,悬吊组大鼠BMSCs经成脂诱导后7,14,21 d TG含量均明显升高(P<0.01),成脂诱导21 d成脂能力,PPARγmRNA和蛋白表达明显增强(P<0.01);较悬吊组,补肾健脾高、中、低剂量组大鼠BMSCs经成脂诱导7,14,21 d后TG含量降低(P<0.01,P<0.05),经成脂诱导21 d后成脂能力均明显减弱(P<0.01),PPARγmRNA和蛋白表达减少(P<0.01,P<0.05);较补肾健脾中剂量组,补肾健脾高剂量组大鼠BMSCs经成脂诱导14,21 d后TG含量降低(P<0.05),经成脂诱导21 d后补肾健脾高剂量组成脂能力均减弱(P<0.05),PPARγmRNA和蛋白表达均减少(P<0.05,P<0.01),补肾健脾低剂量组PPARγ蛋白表达增多(P<0.05)。结论:补肾健脾方具有抑制尾部悬吊大鼠骨髓间充质干细胞成脂分化的作用,以补肾健脾方高剂量作用为优,其机制可能与下调PPARγ表达有关。

Objective： To observe the action of Bushen Jianpi decoction （BSJP） on the adipogenic differentiation potential of bone marrow mesenchymal stem cells （BMSCs） in tail suspension model rats. Method： Rats were assigned to five groups randomly： control （C）, tail suspended （S）, high dose of BSJP （H）, middle dose of BSJP （M） and low dose of BSJP （L） groups. Rats in S, H, M and L groups were suspended by head down tilt - 30° for 21 days to simulate weightlessness. Rats of H, M and L groups were orally administrated with BSJP and other groups with equivalent saline from the first day. BMSCs were isolated and cultured by adherence method, and the third generation cells were induced by adipogenic differentiation solution. The lipid droplet formation was detected by oil red O staining and the expression of peroxisome proliferators-activated receptor γ （PPARγ） was analyzed by Real time PCR and Western blot. The content of triglycerides （TG） in cells was examined on day 7, 14, 21 of adipogenic differentiation. Result： Compared with the C group, the content of TG was increased significantly on day 7, 14, 21 of adipogenic differentiation, the adipogenic differentiation potential and the mRNA and protein expression of PPARγ were also increased on day 21 of adipogenic differentiation in S group. Compared with the S group, the content of TG was declined on day 7, 14, 21 of adipogenic differentiation in BSJP H, M and L dose groups, the adipogenic differentiation potential and the mRNA and protein expression of PPARγ were also decreased on day 21 of adipogenic differentiation in the three groups. Compared with the M group, the content of TG on day 14, 21 of adipogenic differentiation were reduced in H group, the adipogenic differentiation potential and the expression of PPARγ were decreased in H group. Conclusion： The BSJP can inhibit the adipogenic differentiation of BMSCs by down-regulating the expression of PPARγ and the inhibition effect of high dose in the BSJP is best.