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【目的】评价5-Aza联合20mL/LHS方案诱导人和大鼠BMMSC向肌细胞分化的可行性,揭示BMMSC肌分化的机制,尝试建立一种简便、有效、稳定的诱导BMMSC肌分化方法。【方法】首先分离纯化SD大鼠BMMSC,显微镜下观察人和大鼠BMMSC生长和形态。接着,应用5-Aza联合20mL/LHS诱导BMMSC向肌细胞分化,通过免疫细胞化学和RT-PCR方法分别检测肌标志性分子基因表达情况。【结果】①5-Aza联合20mL/LHS诱导人和大鼠BMMSC向肌系分化效能截然不同:对hBMMSC,无论形态学还是免疫细胞化学及RT-PCR均无明显改变;对SD BMMSC,显示了促肌分化的作用(刺激后desmin,MyoD,myogenin mRMA表达水平上调,Pax7表达水平下调),但形态学无改变。②高代数的MSC不利诱导分化。③Pax3和MRF4分子在诱导前后均无表达。【结论】①5-Aza联合20mL/LHS诱导BMMSC向肌细胞分化结果因品系不同而异。②5-Aza作为MSCs肌分化诱导剂,有早期启蒙作用,但其作用和机制还需进一步明确。③一些肌相关分子的表达受细胞品系、细胞来源、细胞状态等因素影响。
【Objective】 To evaluate the feasibility of 5-Aza combined with 20mL / LHS regimen in inducing BMMSCs to differentiate into muscle cells in human and rat, and to reveal the mechanism of myogenic differentiation in BMMSC. It is an attempt to establish a simple, effective and stable method to induce myogenic differentiation in BMMSCs. 【Method】 BMMSCs were isolated and purified from SD rats. The growth and morphology of BMMSCs in human and rat were observed under microscope. Next, 5-Aza and 20 mL / LHS were used to induce BMMSCs to differentiate into muscle cells. The expression of muscle-specific genes was detected by immunocytochemistry and RT-PCR respectively. 【Results】 ①The differentiation of myoblasts from BMMSCs induced by 5-Aza and 20 mL / LHS was completely different: there was no significant difference in hBMMSC between morphological and immunocytochemistry and RT-PCR; for SD BMMSCs, MyoD, myogenin, mRMA and Pax7 were down-regulated, but the morphology was unchanged. ② High algebraic MSC detrimental differentiation. ③Pax3 and MRF4 molecules were not expressed before and after induction. 【Conclusion】 (1) Differentiation of BMMSCs into muscle cells by 5-Aza combined with 20mL / LHS results in different strains. ② As an inducer of MSCs differentiation, 5-Aza has early enlightenment, but its role and mechanism need to be further clarified. The expression of some muscle-related molecules is affected by cell lines, cell sources, cell status and other factors.