Janus激酶／信号转导和转录激活因子3对重症急性胰腺炎大鼠肺泡Ⅱ型上皮细胞损伤的体外作用研究

The role of Janus kinase/signal transducer and activator of transcription 3 in injury of alveolar type Ⅱepithelial cells of rat with severe acute pancreatitis in vitro

目的探讨Janus激酶／信号转导和转录激活因子3（JAK／STAT3）信号转导通路在重症急性胰腺炎（SAP）肺泡Ⅱ型上皮细胞损伤中的作用。方法用牛磺胆酸钠建立SAP大鼠模型，取血清备用。将经原代培养的肺泡Ⅱ型上皮细胞随机分组，对照组加入不含SAP大鼠血清的Dulbecco改良Eagle培养基（DMEM）培养液；SAP组加入含SAP大鼠血清的DMEM培养液；SAP＋AG490组用JAK激酶抑制剂AG490预处理细胞，加入含SAP大鼠血清的DMEM培养液。用电泳迁移率改变分析法（EMSA）检测STAT3活化状态；逆转录-聚合酶链反应（RT—PCR）检测mRNA表达；蛋白质免疫印迹法（Westernblotting）检测蛋白水平；流式细胞技术检测肺泡表面活性物质相关蛋白C（SP—C）表达和肺泡Ⅱ型上皮细胞凋亡。结果与对照组比较，SAP组STAT3活性增强，STAT3 mRNA和蛋白表达均增强，SP—C蛋白表达下降（2hSP—C荧光指数0．69±0．02比1．02±0．03，P〈0．01），肺泡Ⅱ型上皮细胞凋亡增加[（11．55±1．10）％比（5．30±0．36）％，P〈0．053；与SAP组比较，SAP＋AG490组STAT3活性减弱，STAT3mRNA和蛋白表达减弱，SP—C蛋白表达下降（2hSP—C荧光指数0．48±0．10比0．69±0．02，P〈0．01），肺泡Ⅱ型上皮细胞凋亡增加[（13．92±0．82）％比（11．55±1．10）％，P〈O．05]。结论提示JAK／STAT3信号转导通路参与了SAP肺泡Ⅱ型上皮细胞损伤的病理生理过程。

Objective To study the role of Janus kinase/signal transducer and activator of transcription 3 （JAK/STAT3） in injury of alveolar type Ⅱepithelial cells （AT Ⅱ ） in severe acute pancreatitis （SAP）. Methods The rat model of SAP was reproduced intraductal injection of sodium tauroeholate. Serum was collected for examinations. AT I cells in primary culture were randomized to be treated with serum from rats with SAP （SAP group）, or SAP serum＋AG490 （JAK inhibitor, SAP-bAG490 group）, while the normal cell control was cultured with Dulbecco improvemed Eagle medium （DMEM）, AT Ⅱ cells were collected after the treatment to determine the activation of STAT3 by electrophoretic mobility shift assay （EMSA）, STAT3 mRNA expression by reverse transcription-polymerase chain reaction （RT-PCR）, STAT3 protein expression by Western blotting, surfactant protein C （SP-C） level in AT Ⅱ and apoptosis of AT Ⅱ with flow cytometry. Results Compared with control group, activation of STAT3 was enhanced in SAP group, and expression of STAT3 mRNA and protein was also enhanced, level of SP-C protein declined （2 hours SP-C fluorescence index 0.69± 0.02 vs. 1.02 ±0.03, P 〈 0.01）, and apoptosis of AT Ⅱ increased [（11.55 ± 1.10）% vs. （5.30±0. 36）%, P〈0.05]. Compared with SAP group, activation of STAT3 was attenuated in SAP＋AG490 group, expression of STAT3 mRNA and protein was lowered, and level of SP-C protein declined （2 hours SP-C fluorescence index 0.48± 0. 10 vs. 0. 69 ±0.02, P〉0.01）, and the apoptosis of AT Ⅱincreased [（13.92±0.82）% vs. （11.55±1.10）%, P〈0.05]. Conclusion The results suggest that JAK/STAT3 pathway might be involved in the pathogenesis of injury to AT Ⅱ in SAP.