IRAK-1表达水平与COPD大鼠PASMCs分泌PDGF-AB和IL-6的相关性研究

Correlation between expression level of IRAK-1 and secretion of PDGF-AB and IL-6 by pulmonary artery smooth muscle cells treated with LPS in COPD rats

目的:探讨白细胞介素1受体相关激酶1(IRAK-1)与慢性阻塞性肺疾病(COPD)大鼠肺动脉平滑肌细胞(α-平滑肌肌动蛋白(α-SMA),PASMCs)分泌血小板源性生长因子AB(PDGF-AB)和白细胞介素6(IL-6)的相关性。方法:构建COPD大鼠模型,HE染色观察肺组织病理学变化,图像分析法测定远端肺动脉管壁厚度占动脉外径的百分比(WT%)和管壁面积占血管总面积的百分比(WA%)。消化、分离和纯化COPD大鼠远端PASMCs,并采用特异性抗α-平滑肌肌动蛋白(α-SMA)抗体进行细胞免疫荧光鉴定大鼠PASMCs。将COPD大鼠PASMCs随机分为对照组(不进行干预)、脂多糖(LPS)组(终浓度为1 mg/L)、IRAK-1/4抑制剂组(终浓度为10μmol/L)和LPS+IRAK-1/4抑制剂组(IRAK-1/4抑制剂终浓度为10μmol/L,预处理30 min后加入LPS,终浓度为1 mg/L),采用Western blot检测各组PASMCs中p-IRAK-1和IRAK-1的蛋白水平;ELISA方法检测各组PASMCs上清液中PDGF-AB和IL-6的浓度。结果:COPD模型组WT%和WA%较空白对照组升高(P<0.01)。光学显微镜下COPD大鼠PASMCs呈梭形,荧光镜下可见胞质α-SMA蛋白染成红色。与对照组相比,LPS组p-IRAK-1蛋白表达水平及PDGF-AB和IL-6的含量升高(P<0.05);与LPS组相比,LPS+IRAK-1/4抑制剂组p-IRAK-1的蛋白水平及PDGF-AB和IL-6的含量明显降低(P<0.05)。IRAK-1磷酸化水平与细胞上清液中PDGF-AB和IL-6的浓度呈正相关。结论:IRAK-1参与COPD大鼠PASMCs分泌PDGF-AB和IL-6的调控。这为COPD的早期干预提供了新依据。

AIM: To explore the expression level of interleukin-1 receptor-associated kinase-1(IRAK-1) and the secretion function of pulmonary artery smooth muscle cells(PASMCs) in chronic obstructive pulmonary disease(COPD) rats.METHODS: A rat model of COPD was established. Image analysis was used to calculate the ratio of thickness to outside diameter of pulmonary arterioles(WT%) and the ratio of vessel wall area to total vessel area(WA%). The PASMCs from COPD rats were digested, isolated and purified, and the cells were randomly divided into control group, lipopolysaccharide(LPS) group, IRAK-1/4 inhibitor group and LPS + IRAK-1/4 inhibitor group. The protein levels of p-IRAK-1 and IRAK-1 were determined by Western blot. The levels of platelet-derived growth factor-AB(PDGF-AB) and interleukin-6(IL-6) in the culture supernatant of PASMCs in each group were measured by ELISA. The correlation between p-IRAK-1 level and the concentrations of the secretion factors in the PASMCs was analyzed. RESULTS: The WT% and WA% in COPD model group were higher than those in control group(P<0.01). Under the optical microscope, the PASMCs showed in spindle in shape, and their α-smooth muscle actin was stained red under fluorescence microscope. Compared with control group, LPS up-regulated the protein level of phosphorylated IRAK-1(p-IRAK-1) and the levels of PDGF-AB and IL-6(P<0.05). IRAK-1/4 inhibitor was capable of suppressing the effects of LPS(P<0.05). The protein level of p-IRAK-1 was positively correlated with the concentrations of PDGF-AB and IL-6 in the PASMCs. CONCLUSION: IRAK-1 participates in the regulation of PASMCs to secrete PDGF-AB and IL-6, which provided a new target for the early intervention of COPD.