mTORC2介导的信号通路影响肺泡上皮钠通道对急性肺损伤肺组织的保护作用

发布时间：2018-07-05 09:03

本文选题：肺泡上皮钠通道 + mTORC/SGK　；参考：《第三军医大学学报》2015年24期

【摘要】：目的通过调控肺泡上皮细胞α钠离子通道(epithelial sodium channelα,ENa C-α)上游信号通路,探讨m TORC2/SGK1途径在急性肺损伤(acute lung injury,ALI)情况下对肺水清除方面的作用机制。方法将50只BALB/c雄性小鼠按随机数表法分成对照组、模型组、胰岛素组、雷帕霉素组及PP242抑制剂组,每组10只。滴鼻给予5 mg/kg LPS构建ALI动物模型。建模2 h后,分别腹腔注射0.8 U/kg胰岛素、4 mg/kg雷帕霉素、60 mg/kg PP242干预,8 h后处死小鼠。A549细胞分别以培养基、10 mmol/m L胰岛素、5 mmol/m L雷帕霉素、5 mmol/m L PP242处理。采用ELISA检测各组肺泡灌洗液(BALF)中IL-6及TNF-α浓度,并测量各组右上肺组织的湿/干质量比(W/D)评估肺水清除(alveolar fluid clearance,AFC),HE染色观察各组肺组织病理改变。采用Real-time PCR、Western blot、免疫荧光法检测A549细胞rictor、糖皮质激素调节蛋白激酶1(serum and glucocorticoid-regulated protein kinase1,SGK1)、磷酸泛素化连接酶(E3 ubiquitin protein ligase,Nedd4-2)及ENa C-α的表达。结果体内实验发现,与模型组比较,胰岛素组小鼠BALF中IL-6[(77.48±6.04)vs(49.17±8.49)pg/m L]和TNF-α[(116.2±13.5)vs(92.4±8.37)pg/m L]水平显著降低,并伴有湿/干质量比减低,病检示肺损伤程度减轻(P0.05)。PP242抑制剂组BALF中IL-6[(77.48±6.04)vs(100.6±14.69)pg/m L]和TNF-α[(116.2±13.5)vs(192.5±16.01)pg/m L]水平上升,湿/干质量比显著升高,肺组织损伤明显,程度较模型组更重(P0.05)。而雷帕霉素组与模型组中各项指标及病理学改变差异无统计学意义(P0.05)。体外实验中,A549细胞中rictor、SGK1、Nedd4-2及ENa C-αmRNA和蛋白表达在胰岛素组明显增高,在PP242抑制剂组中明显降低(P0.05),而在雷帕霉素组中无明显改变(P0.05)。结论调控m TORC2/SGK1信号通路可影响ENa C-α的表达,从而发挥其在ALI情况下对肺组织的保护作用。
[Abstract]:Objective to investigate the role of m TORC2 / SGK1 pathway in the clearance of lung water in acute lung injury (acute lung injury-induced Ali) by regulating the upstream signaling pathway of 伪 -sodium channel (epithelial sodium channel 伪 -ENa C- 伪 in alveolar epithelial cells. Methods 50 BALB / c male mice were randomly divided into control group, insulin group, rapamycin group and PP242 inhibitor group with 10 rats in each group. Ali animal model was established by nasal administration of 5 mg/kg LPS. After modeling for 2 h, the mice were treated with 10 mmol/m / L insulin / 5 mmol/m / L rapamycin 5 mmol/m / L PP242 for 8 h respectively by intraperitoneal injection of 0. 8 U / kg insulin with 4 mg/kg rapamycin for 60 mg/kg PP242. The levels of IL-6 and TNF- 伪 in alveolar lavage fluid (BALF) were detected by Elisa, and the wet / dry mass ratio (WR / D) of right upper lung tissue was measured to evaluate the lung water clearance and (alveolar fluid clearance. The pathological changes of lung tissue in each group were observed by HE staining. The expression of rictor-1, glucocorticoid regulated protein kinase 1 (serum and glucocorticoid-regulated protein kinase1 (SGK1), E3 ubiquitin protein ligase Nedd4-2 and ENAC- 伪 in A549 cells were detected by Real-time PCR Western blot. Results compared with the model group, the levels of IL-6 [(77.48 卤6.04) vs (49.17 卤8.49) pg/m L] and TNF- 伪 [(116.2 卤13.5) vs (92.4 卤8.37) pg/m L] in insulin group were significantly lower than those in the model group, and the wet / dry weight ratio was decreased. The lung injury degree was alleviated (P0.05) .PP242 inhibitor group increased IL-6 [(77.48 卤6.04) vs (100.6 卤14.69) pg/m L] and TNF- 伪 [(116.2 卤13.5) vs (192.5 卤16.01) pg/m L]. The wet / dry weight ratio increased significantly and the lung injury was more severe than the model group (P0.05). However, there was no significant difference between rapamycin group and model group in all indexes and pathological changes (P0.05). In vitro, the mRNA and protein expressions of rictorin SGK1Nedd4-2 and ENAC- 伪 were significantly increased in insulin group and decreased significantly in PP242 inhibitor group (P0.05), but not in rapamycin group (P0.05). Conclusion regulating the signaling pathway of m TORC 2 / SGK 1 may affect the expression of ENAC-伪, thus exerting its protective effect on lung tissue in Ali.
【作者单位】：重庆医科大学附属第二医院呼吸与危重症医学科;
【基金】：国家自然科学基金面上项目(81270141)~~
【分类号】：R563.8

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