雷帕霉素对再生障碍性贫血患者骨髓间充质干细胞生物学功能的影响

本研究探讨雷帕霉素对再生障碍性贫血（aplasticanemia，AA）患者的骨髓间充质干细胞的生物学功能和自噬现象的影响，为研究雷帕霉素应用于临床再生障碍性贫血的治疗提供资料。不同浓度雷帕霉素（0、10、50、100nmol／L）处理AA患者BM—MSC48h，然后用Westernblot检测自噬相关蛋白LC3B的表达，应用流式细胞术检测细胞凋亡和细胞周期的变化，CCK-8法测定细胞增殖情况，以BM—MSC成脂分化诱导2周后油红染色方法检测成脂分化能力，并且用real—timePCR检测成脂相关基因（LPL、CFD和PPART）的表达。结果显示：雷帕霉素引起AA患者的BM—MSC自噬增加，促进其凋亡（P〈0．05），且呈剂量依赖性地将细胞阻滞于Go／G1期，阻止其进入S期（P〈0．05）；BM-MSC增殖率随雷帕霉素浓度增加而降低，并且雷帕霉素抑制AA患者BM—MSC的成脂分化，且呈剂量依赖性。结论：雷帕霉素诱导AA患者BM—MSC发生自噬和凋亡，明显促进G，期细胞周期阻滞，抑制细胞增殖和降低成脂分化，以上现象的机制可能与雷帕霉素通过抑制mTOR信号通路而激活自噬有关。

Effects of Rapamycin on Biological Characteristics of Bone Marrow Mesenchymal Stem Cells from Patients with Aplastic Anemia

This study was aimed to investigate the effects of rapamycin on biological function and autophagy of bone marrow mesenchymal stem cells（BM-MSC） from patients with aplastic anemia so as to provide experimental basis for the clinical treatment of aplastic anemia（AA） with rapamycin. BM-MSC were treated with different concentrations of rapamycin （0,10,50,100 μmol/L） for 48 h, the expression of LC3B protein was detected by Western blot to observe the effect of rapamycin on cell autophagy; cell apoptosis and cell cycles were detected by flow cytometry; the proliferation of BM-MSC of AA patients was measured by cell counting kit-8 ; the adipogenic differentiation of BM-MSC were tested by oil red O staining after adipogenic induction for 2 weeks; the adipogenic related genes （ LPL, CFD, PPART） were detected by real-time PCR. The results showed that the proliferation and adipogenesis of BM-MSC of AA patients were inhibited by rapamycin. Moreover, the autophagy and apoptosis of BM-MSC were increased by rapamycin in a dose-dependent way. Rapamycin arrested the BM-MSC in G0/G1 phase and prevented them into S phase （ P 〈 0. 05）. It is concluded that rapamycin plays an critical role in inhibiting cell proliferation, cell cycles, and adipogenesis, these effects may be related with the autophagy activation and mTOR inhibition resulting from rapamycin.