2019 年第 1 期 第 14 卷

Smad泛素化调节因子和Arkadia在大鼠肺纤维化中的作用研究

Effect of Smad ubiquitination regulatory factor and Arkadia on pulmonary fibrosis in rats

作者：黄洁玉张敏

英文作者：

单位：510220广州市红十字会医院暨南大学医学院附属广州红十字会医院呼吸内科

英文单位：

关键词：肺纤维化；Smad泛素化调节因子；Arkadia；Smads信号传导

英文关键词：

• 摘要：

• 【摘要】目的    探讨Smad泛素化调节因子（Smurf）和Arkadia在博莱霉素诱导的大鼠肺纤维化形成中的作用及可能的分子机制。方法    将24只无特定病原体级健康雄性SD大鼠随机分为对照组、模型7 d组、模型14 d组、模型 28 d组，每组6只。模型组大鼠气管内滴入博莱霉素（10 mg/kg）建立肺纤维化动物模型，对照组气管内滴入等体积0.9%氯化钠注射液。于造模后第7、14和28天分批处死大鼠。应用苏木精-伊红（HE）染色和Masson染色观察肺组织炎症及纤维化情况；应用逆转录聚合酶链反应和蛋白质印迹法检测Ⅰ型胶原α1（COL1A1）、Arkadia、Smurf1、Smurf2、Smad7、Ski和转化生长因子βⅠ型受体（TβRⅠ）的表达。结果    HE染色和Masson染色显示肺纤维化模型建立成功。模型7、14、28 d组大鼠肺组织COL1A1蛋白表达水平均明显高于对照组，差异均有统计学意义（均P＜0.05）。模型7、14、28 d组大鼠肺组织Arkadia mRNA和蛋白表达水平均明显高于对照组，Smurf2 mRNA和蛋白表达水平均明显低于对照组（均P＜0.05）。模型7 d组大鼠肺组织Smad7和Ski蛋白表达水平明显高于对照组，随后Smad7和Ski蛋白表达水平随时间延长而逐渐降低，至第28天均低于对照组水平（均P＜0.05），但各组间Smad7和Ski mRNA表达水平差异均无统计学意义（均P＞0.05）。模型7、14、28 d组大鼠肺组织TβRⅠ mRNA和蛋白表达水平均明显高于对照组［(0.340±0.056)、(0.520±0.021)、(1.014±0.190)比(0.244±0.033)；(0.453±0.033)、(0.617±0.060)、(0.720±0.038)比(0.309±0.031)］，差异均有统计学意义（均P＜0.05）。结论    Smurf2和Arkadia均参与了博莱霉素诱导的大鼠肺纤维化形成；Arkadia可能通过泛素化降解蛋白途径实现对Smad7和Ski蛋白表达的下调，促进肺纤维化形成。

• 【Abstract】Objective    To investigate the effect and possible molecular mechanisms of Smad ubiquitination regulatory factor(Smurf) and Arkadia on bleomycin-induced pulmonary fibrosis in rats. Methods    Twenty-four specific pathogen free healthy male rates were randomly divided into control group and pulmonary fibrosis model groups（7 d model group, 14 d model group, 28 d model group）, with 6 rats in each group. Pulmonary fibrosis model was made by intratracheal instillment of bleomycin(10 mg/kg); the control group was intratracheally instilled 0.9% sodium chloride solution. Pulmonary fibrosis model rats were separately sacrificed on the 7th, 14th and 28th day after modeling. Pulmonary inflammation and fibrosis were observed by hematoxylin-eosin（HE） and Masson staining. Expressions of collagen type Ⅰ α1(COL1A1), Arkadia, Smurf1, Smurf2, Smad7, Ski and transforming growth factor β receptor type Ⅰ（TβRⅠ） were detected by reverse transcriptase polymerase chain reaction and western blotting. Results    Pulmonary fibrosis model in rats was confirmed by HE and Masson staining. COL1A1 protein expressions in the 7, 14, 28 d model groups were significantly higher than that in the control group(P＜0.05). Expressions of Arkadia mRNA and protein in the 7, 14, 28 d model groups were significantly higher than those in the control group(P＜0.05). Expressions of Smurf2 mRNA and protein in the 7, 14, 28 d model groups were significantly lower than those in the control group(P＜0.05). Expressions of Smad7 and Ski protein gradually decreased in the 7, 14, 28 d model groups; the expression levels in the 7 d model group were higher than those in the control group; the expression levels in the 28 d group were lower than those in the control group（P＜0.05）; Smad7 and Ski mRNA expression showed no significant changes(P＞0.05). Expressions of TβRⅠ mRNA and protein in the 7, 14, 28 d model groups were significantly higher than those in the control group［(0.340±0.056),(0.520±0.021),(1.014±0.190) vs (0.244±0.033)； (0.453±0.033),(0.617±0.060),(0.720±0.038) vs (0.309±0.031)］(P＜0.05). Conclusions    Smurf2 and Arkadia are associated with bleomycin-induced pulmonary fibrosis in rats. Arkadia can contribute to pulmonary fibrosis through inducing ubiquitin-dependent degradation of Smad7 and Ski protein.