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TRPC channels mediated calcium entry is required for proliferation of human airway smooth muscle cells induced by nicotine-nAChR
Biochimie ( IF 3.9 ) Pub Date : 2018-12-11 , DOI: 10.1016/j.biochi.2018.12.004
Yongliang Jiang , Yumin Zhou , Gongyong Peng , Heshen Tian , Dan Pan , Lei Liu , Xing Yang , Chao Li , Wen Li , Ling Chen , Pixin Ran , Aiguo Dai

The present study was designed to explore the role of transient receptor potential canonical 3 (TRPC3) in nicotine-induced chronic obstructive pulmonary disease (COPD) and its underlying mechanism. In this study, the expression and localization of α5 nicotinic acetylcholine receptor (α5-nAchR) in lung tissues were determined by western blotting and immunohistochemistry. The quantitative real-time PCR (qRT-PCR) analysis was performed to examine the mRNA expression levels of α5-nAchR and TRPC3 in human airway smooth muscle cells (HASMCs). Cell viability was assessed by CCK-8 assay. Proliferation was detected by cell counting and EdU immunofluorescent staining. Fluorescence calcium imaging was carried out to measure cytosolic Ca2+ ([Ca2+]cyt) concentration. The results showed that the α5-nAchR and TRPC3 expressions were significantly up-regulated in lung tissues of COPD smokers. Nicotine promoted HASMC proliferation, which was accompanied by elevated α5-nAchR and TRPC3 expressions, basal [Ca2+]cyt, store-operated calcium entry (SOCE) and the rate of Mn2+ quenching in HASMCs. Further investigation indicated that nicotine-induced Ca2+ response and TRPC3 up-regulation was reversibly blocked by small interfering RNA (siRNA) suppression of α5-nAChR. The knockdown of TRPC3 blunted Ca2+ response and HASMC proliferation induced by nicotine. In conclusion, nicotine-induced HASMC proliferation was mediated by TRPC3-dependent calcium entry via α5-nAchR, which provided a potential target for treatment of COPD.



中文翻译:

尼古丁-nAChR诱导人气道平滑肌细胞增殖需要TRPC通道介导的钙进入

本研究旨在探讨瞬态受体电位经典3(TRPC3)在尼古丁诱发的慢性阻塞性肺疾病(COPD)中的作用及其潜在机制。在这项研究中,通过蛋白质印迹和免疫组织化学方法确定了α5烟碱乙酰胆碱受体(α5-nAchR)在肺组织中的表达和定位。进行定量实时PCR(qRT-PCR)分析以检查人气道平滑肌细胞(HASMC)中α5-nAchR和TRPC3的mRNA表达水平。通过CCK-8测定法评估细胞生存力。通过细胞计数和EdU免疫荧光染色检测增殖。进行荧光钙成像以测量胞浆中的Ca 2 +([Ca 2+] cyt)浓度。结果表明,COPD吸烟者肺组织中α5-nAchR和TRPC3的表达明显上调。尼古丁促进了HASMC的增殖,并伴随着α5-nAchR和TRPC3表达的升高,基础[Ca 2+ ] cyt,贮藏钙输入(SOCE)和Mn 2+在HASMC中的淬灭速率。进一步的研究表明,尼古丁诱导的Ca 2+反应和TRPC3上调被α5-nAChR的小干扰RNA(siRNA)抑制可逆地阻断。TRPC3的敲低钝化了Ca 2+尼古丁诱导的肝癌反应和HASMC增殖。总之,尼古丁诱导的HASMC增殖是通过α5-nAchR介导的TRPC3依赖性钙进入介导的,这为治疗COPD提供了潜在的靶点。

更新日期:2018-12-11
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