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目的 观察纤维粘连蛋白 (FN)、层粘连蛋白 (LN)及胶原蛋白I (ColI)等主要细胞外基质成分 ,对气道平滑肌细胞 (ASMCs)增殖活性的影响 ,并探讨其作用机制。方法将原代培养的大鼠ASMCs,接种于不同细胞外基质蛋白包被的培养板 ,观察各时间点细胞的贴壁数量。在培养的ASMCs中分别加入两种浓度 (2 0和 6 0mg/L)的不同细胞外基质蛋白溶液 ,即FN组、LN组和ColI组 ,采用3 H TdR掺入法 ,检测作用 8、12和 2 4h后各组ASMCs增殖活性的变化。间接免疫荧光染色法检测RAS蛋白在各组ASMCs中的表达。结果 与无包被组相比较 ,FN、ColI包被组及LN包被组ASMCs的贴壁数量明显高 (P <0 .0 1或P <0 .0 5 )。与正常对照组相比较 ,FN和ColI各浓度组ASMCs的增殖活性增高显著 (P <0 .0 1) ,且与浓度和作用时间成正比 ;而LN组ASMCs的活性无明显差异。同时 ,FN和ColI组ASMCs的RAS染色呈强阳性 ;对照组及LN组ASMCs的染色则为弱阳性。结论 ASMCs对FN、LN和ColI均有良好的粘附性 ,但只有FN和ColI能明显提高ASMCs的增殖活性 ,其作用是通过RAS信号转导途径而实现的。表明细胞外基质是哮喘气道重塑的重要刺激因子之一
Objective To observe the effects of major extracellular matrix components such as fibronectin (FN), laminin (LN) and collagen I on the proliferation of airway smooth muscle cells (ASMCs) and to explore its mechanism. Methods Primary cultured rat ASMCs were inoculated into different kinds of extracellular matrix protein-coated culture plates and the number of adherent cells was observed at each time point. Two different concentrations of extracellular matrix proteins (20 and 60 mg / L) were added to the cultured ASMCs, namely FN group, LN group and ColI group, respectively, using 3 H TdR incorporation assay. And after 24 h the proliferation of ASMCs in each group activity changes. Indirect immunofluorescence staining was used to detect the expression of RAS protein in each group of ASMCs. Results Compared with non-coated group, the number of adherent ASMCs in FN, ColI-coated group and LN-coated group was significantly higher (P <0.01 or P <0.05). Compared with the normal control group, the proliferative activity of ASMCs in FN and ColI groups increased significantly (P <0.01), and was proportional to the concentration and duration of action; however, the activity of ASMCs in LN group showed no significant difference. At the same time, RAS staining of ASMCs in FN and ColI groups was strongly positive; staining of ASMCs in control group and LN group was weakly positive. Conclusions ASMCs have good adhesion to FN, LN and ColI, but only FN and ColI can significantly increase the proliferation activity of ASMCs through the RAS signal transduction pathway. The extracellular matrix is one of the important stimulators of airway remodeling in asthma