国际麻醉学与复苏杂志   2018, Issue (10): 3-3
    
脂多糖对肺成纤维细胞胸腺细胞分化抗原?蛳1 及整合素β3表达的调控并抑制自噬的研究
万晗曦, 谢婷婷, 徐侨翌, 胡晓婷, 邢顺鹏, 皋源, 何征宇1()
1.上海交通大学医学院附属仁济医院
Lipopolysaccharide regulates the expression of thymocyte differentiation antigen?蛳1 and integrin β3 of lung fibroblasts and inhibit autophagy
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摘要:

目的 明确脂多糖(lipopolysaccharide, LPS)对小鼠肺成纤维细胞胸腺细胞分化抗原?蛳1(thymocyte differentiation antigen-1, Thy-1)和整合素β3蛋白表达的调控作用,并观察其与肺成纤维细胞自噬抑制的关系。 方法 将原代培养的小鼠肺成纤维细胞采用随机数字法分为4组(每组3孔):PBS对照组(Con组)、0.25 mg/L LPS组(LPS0.25组)、0.50 mg/L LPS组(LPS0.50组)、1.00 mg/L LPS组(LPS1.00组),随后以不同浓度LPS(0.25、0.50、1.00 mg/L)刺激,于刺激后6 h通过Western blot检测不同浓度LPS刺激小鼠肺成纤维细胞后Thy-1和整合素β3蛋白的表达情况。于刺激后30 h比较肺成纤维细胞自噬相关蛋白LC3-Ⅱ/Ⅰ和Beclin-1的表达情况,同时通过透射电子显微镜观察细胞内自噬小体形成情况。 结果 随着LPS刺激浓度的增加,小鼠肺成纤维细胞的Thy-1蛋白表达量下降,其中LPS1.00组较Con组有明显下降(P<0.05);整合素β3蛋白表达量升高,其中LPS1.00组较Con组有明显升高(P<0.05)。LPS1.00组刺激小鼠肺成纤维细胞30 h后明显抑制其自噬相关蛋白LC3-Ⅱ/Ⅰ和Beclin-1的表达(P<0.05),并抑制自噬小体形成(P<0.05)。 结论 LPS可下调Thy?蛳1蛋白表达、上调整合素β3蛋白表达,同时抑制自噬相关蛋白LC3-Ⅱ/Ⅰ和Beclin-1的表达及自噬小体的生成,该过程可能与LPS诱导肺成纤维细胞自噬抑制以及肺纤维化进展有关。

关键词: 脂多糖; 肺成纤维细胞; 自噬; 胸腺细胞分化抗原?蛳1; 整合素β3
Abstract:

Objective To clarify the effect of lipopolysaccharide (LPS) on the expression of thymocyte differentiation antigen-1(Thy-1) and integrin β3 in mouse lung fibroblasts and its relationship with autophagy inhibition. Methods Primary cultured mouse lung fibroblasts were divided into four groups (n=3) based on different concentration of LPS stimulation: PBS control group (Con group),0.25 mg/L LPS group (LPS0.25 group), 0.50 mg/L LPS group (LPS0.50 group), 1.00 mg/L LPS group (LPS1.00 group). Then the total cellular proteins were extracted. The expression of Thy-1 and integrin β3 were detected by Western blot. The expression of autophagy-related proteins LC3-Ⅱ/Ⅰ and Beclin-1 in cells was compared at 30 h after LPS stimulatin. The formation of autophagic bodies was observed by using transmission electron microscopy. Results Thy-1 protein expression was decreased in lung fibroblasts with increasing concentration of LPS stimulation. Compared with the control group(P<0.05), the level of Thy-1 protein in lung fibroblasts was significantly decreased in 1.00 mg/L LPS group. Integrin β3 protein expression was increased with increasing concentration of LPS stimulation. As a result, the expression leve of integrin in 1.00 mg/L LPS group was significantly higher than the level in control group (P<0.05). At the same time, LPS significantly inhibited the expression of autophagy-related proteins LC3-Ⅱ/Ⅰ(P<0.05) and Beclin-1 (P<0.05) in mouse lung fibroblasts and the formation of autophagic bodies(P<0.05). Conclusions LPS can down-regulate the expression of Thy-1 but up-regulate the expression of integrin β3. It also inhibits the expression of autophagy-related proteins LC3-Ⅱ/Ⅰ, Beclin-1 and the formation of autophagic bodies. This process may be associated with LPS-induced autophagy inhibition of lung fibroblasts and the progress of lung fibrosis.

Key words: Lipopolysaccharide;Lung fibroblasts; Autophagy; Thy-1; Integrin β3