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背景:近年的研究表明,肿瘤坏死因子α对不同组织成纤维细胞的作用具有组织特异性及浓度依赖性。目的:观察肿瘤坏死因子α及其信号传导途径中特异性激酶抑制剂对小鼠胚胎成纤维细胞成熟化所起的作用。方法:体外培养小鼠胚胎成纤维细胞,将细胞分为3组:第1组用含体积分数2%血清的DMEM高糖培养基培养作为空白对照组;第2组用含100μg/L肿瘤坏死因子α的培养基培养;第3组先加入质量浓度为50μg/L的Anti-TNFRSF1B作用1h后,倒出培养基再加入含有肿瘤坏死因子α的培养基继续培养。采用RT-PCR法测定各组Ⅰ型胶原蛋白和基质金属蛋白酶3mRNA表达、Western Blot法测定各组Ⅰ型胶原蛋白和基质金属蛋白酶3蛋白表达。结果与结论:小鼠胚胎成纤维细胞在一定质量浓度肿瘤坏死因子α作用下,其信号传导途径特异性激酶发生磷酸化或蛋白被激活,信号通路被激活,促进基质金属蛋白酶3的活化,明显降低Ⅰ型胶原的表达。加入其信号传导途径的抑制剂Anti-TNFRSF1B后,肿瘤坏死因子α的效应得到了一定的抑制,但并未完全消除,这更进一步证明肿瘤坏死因子α对小鼠胚胎成纤维细胞活化的作用。
BACKGROUND: Recent studies have shown that tumor necrosis factor α has tissue-specific and concentration-dependent effects on fibroblasts in different tissues. OBJECTIVE: To observe the role of tumor necrosis factor-α (TNF-α) and specific kinase inhibitors in signal transduction pathway on the maturation of mouse embryonic fibroblasts. Methods: Mouse embryonic fibroblasts were cultured in vitro and the cells were divided into 3 groups: Group 1 was cultured in DMEM high glucose medium containing 2% serum by volume as control group; Group 2 was treated with 100 μg / L tumor necrosis Factor α medium. Group 3 was treated with Anti-TNFRSF1B at a concentration of 50 μg / L for 1 h, then the medium was decanted, then the medium containing tumor necrosis factor α was added to continue the culture. The expression of type Ⅰ collagen and matrix metalloproteinase-3 mRNA in each group were determined by RT-PCR. The expressions of type Ⅰ collagen and matrix metalloproteinase-3 in each group were determined by Western Blot. RESULTS AND CONCLUSION: The mouse embryonic fibroblasts were activated by phosphorylation or protein activation of signaling pathway-specific kinases by certain concentration of tumor necrosis factor-α and the activation of the signal pathway, which promoted the activation of matrix metalloproteinase-3 Decrease the expression of type I collagen. The anti-TNFRSF1B, an inhibitor of its signal transduction pathway, inhibited tumor necrosis factor-α (TNF-α) to a certain extent but did not completely eliminate the effect of tumor necrosis factor-α (TNF-α) on the activation of mouse embryonic fibroblasts.