【摘要】 成肌细胞分化机理至今尚不明确,为此主要研究细胞外基质蛋白模式对成肌细胞分化的影响。采用微接触印刷技术制备4种不同的纤维粘连蛋白微图案(平行组、随机组、垂直组、对照组)培养成肌细胞,用免疫荧光染色法分析7 d后各组成肌细胞的分化情况,并分析2、6、18 h后各组中的黏附分子、Vinculin以及微丝蛋白F-actin的分布情况,每组实验重复3次,每次取3个样本。肌管形成个数平行排列组与对照组相比,无显著性差异(17.33±0.58 vs 16.00±1.73),与垂直组(7.00±1.00)和无极性组(10.89±0.19)相比则显著高(P<0.05)。在2 h时,微丝在细胞中排列与局部微图案一致;在6 h时,微丝开始沿整体图案长轴方向排列;在12 h时,微丝分布与条带整体图案长轴方向一致;18 h时,所形成的黏着斑会沿着微图案区域分布。细胞外基质蛋白模式会对成肌细胞的分化有影响,不同FN极性排列微图案对成肌细胞的骨架和黏附影响不同,这也可能是后期产生不同分化结果的一个原因。更多还原

【Abstract】 The differentiation mechanism of skeletal myoblast differentiation is still unclear. The main purpose of this research is to explore the effect of the extracellular matrix(ECM) protein pattern on myogenic differentiation. In this study, four different fibronectin micropatterns(parallel, random, vertical, and control group) were prepared by microcontact printing technology, and the differentiation of C2 C12 cells after 7 day incubation was analyzed by immunofluorescence staining and analyzes the adhesion molecule, Vinculin and F-actin distribution after 2 h, 6 h, 18 h between groups by immunofluorescence staining method, each group experiment repeated 3 times, each time point had 3 parallel samples. The myotubes number formed in the control group had no significant difference compared with that in the parallel alignment group(17.33±0.58 vs 16±1.73), but was significantly higher than the vertical group(7.00±1.00) and the random group significantly(10.89±0.19)(P<0.05). At 2 h, the arrangement of microfilaments in cells was consistent with the local micropatterns. At 6 h, the microfilament began to align along the long axis direction of the whole strips pattern. At 12 h, the distribution of microfilaments was consistent with that of the long axis direction of the whole strips pattern. At 18 h, the adhesion plaques formed was distributed along the micropattern area. In conclusion, the extracellular matrix protein patterns are influential on the differentiation of myoblasts, different fibronectin polarity micropatterns have distinct effects on myoblasts cytoskeleton and adhesion, and it might be one of the reasons that later differentiation of C2 C12 results in different outcomes.更多还原