Penehyclidine hydrochloride alleviates lipopolysaccharide-induced acute lung injury in rats: Potential role of caveolin-1 expression upregulation
Penehyclidine hydrochloride alleviates lipopolysaccharide-induced acute lung injury in rats: Potential role of caveolin-1 expression upregulation
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DOI：
10.3892/ijmm.2019.4117
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发表时间：
2019-05-01
影响因子：
5.4
通讯作者：
Song, Xuemin
中科院分区：
文献类型：
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作者：
Wu, Xiaojing;Kong, Qian;Song, Xuemin
本研究的目的是在体内和体外检查caveolin-1（Cav-1）在基于盐酸戊乙奎醚（PHC）的脂多糖（LPS）诱导的急性肺损伤（ALI）抑制中的保护作用，以及潜在的潜在机制。在体内，通过气管内给予LPS（5 mg/kg）建立ALI大鼠模型，并在LPS治疗30分钟后给予PHC（2 mg/kg）。在体外，J774A.1 细胞中的 Cav-1 基因被小干扰 (si)RNA 敲低。将细胞与LPS (1μg/ml)一起孵育2小时，随后与PHC(2μg/ml)一起孵育另外2小时。通过肺组织学评估肺损伤，并评估支气管肺泡灌洗液（BALF）中多形核白细胞（PMN）与总细胞的比率、髓过氧化物酶（MPO）活性、BALF蛋白含量和肺湿/干（W/D）比。采用ELISA法测定大鼠血清和细胞培养上清液中促炎因子的水平，包括肿瘤坏死因子-α（TNF-α）、白细胞介素（IL）-6和IL-1β。通过蛋白质印迹法分析肺组织和 J774A.1 细胞中 Cav-1、Toll 样受体 4 (TLR4)、磷酸化 (p)-p38 丝裂原激活蛋白激酶 (p38 MAPK) 和活化 B 细胞转录因子 p65 亚基的核因子 kappa 轻链增强子 (NF-kappa B p65) 的蛋白表达水平。结果表明，PHC通过减少BALF中的中性粒细胞浸润和蛋白质浓度，以及LPS诱导的肺W/D比值、MPO活性和促炎细胞因子的产生，有效减轻肺损伤。此外，PHC 显着降低了组织病理学变化和肺功能障碍的程度。在体外，PHC 治疗可抑制 LPS 刺激的 J774A.1 细胞中的促炎细胞因子水平和 MPO 活性。然而，Cav-1基因敲低的J774A.1细胞中的结果却相反。此外，在体内和体外，PHC 降低了 TLR4、p-p38 MAPK 和核 NF-kappa B p65 的表达水平，并上调了 Cav-1 的表达水平。这些数据表明，PHC 对大鼠和 LPS 刺激的 J774A.1 细胞中 LPS 诱导的 ALI 表现出保护作用，这可能是由于 Cav-1 上调抑制 p38 MAPKs 磷酸化和 TLR4/NF-kappa B 信号通路所致。
The aim of the present study was to examine the protective effect of caveolin-1 (Cav-1) in the penehyclidine hydrochloride (PHC)-based inhibition of lipopolysaccharide (LPS)-induced acute lung injury (ALI) in vivo and in vitro, in addition to the potential underlying mechanisms. In vivo, an ALI rat model was established via intratracheal administration of LPS (5 mg/kg), and PHC (2 mg/kg) was administered 30 min following LPS treatment. In vitro, the Cav-1 gene was knocked down by small interfering (si)RNA in J774A.1 cells. Cells were incubated with LPS (1 mu g/ml) for 2 h, and subsequently incubated with PHC (2 mu g/ml) for an additional 2 h. Lung injury was assessed by lung histology and the ratio of polymorphonuclear leukocytes (PMNs) to total cells was assessed in bronchoalveolar lavage fluid (BALF), myeloperoxidase (MPO) activity, BALF protein content and lung wet/dry (W/D) ratio. The levels of pro-inflammatory factors, including tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-6 and IL-1 beta, in the sera of rats and cell culture supernatant were determined by ELISA. The protein expression levels of Cav-1, toll-like receptor 4 (TLR4), phosphorylated (p)-p38 mitogen activated protein kinases (p38 MAPKs) and nuclear factor kappa-light-chain-enhancer of activated B cells transcription factor p65 subunit (NF-kappa B p65) in lung tissues and J774A.1 cells were analyzed by western blot analysis. The results indicated that PHC effectively alleviated lung injury by decreasing neutrophil infiltration and protein concentration in BALF, and the lung W/D ratio and MPO activity and pro-inflammatory cytokine production induced by LPS. Furthermore, PHC significantly decreased the degrees of histopathological changes and pulmonary dysfunction. In vitro, treatment with PHC inhibited pro-inflammatory cytokine levels and MPO activity in LPS-stimulated J774A.1 cells. However, the results in the J774A.1 cells with Cav-1 gene knockdown were contrary. In addition, PHC decreased TLR4, p-p38 MAPKs and nuclear NF-kappa B p65 expression levels and upregulated the expression level of Cav-1, in vivo and in vitro. These data demonstrated that PHC exhibited a protective effect against LPS-induced ALI in rats and LPS-stimulated J774A.1 cells, which may be due to the inhibition of p38 MAPKs phosphorylation and TLR4/NF-kappa B signaling pathway by Cav-1 upregulation.