血管活性肠肽在气道上皮损伤修复中的作用及其分子机制研究

发布时间：2018-11-13 13:54

【摘要】：血管活性肠肽(vasoactive intestinal peptides，VIP)是肺内重要的感觉神经肽，主要由非肾上腺素能非胆碱能神经纤维释放。VIP阳性神经纤维广泛分布于气道上皮细胞、支气管平滑肌层、肺血管和支气管血管的管壁以及粘膜下的腺体周围。VIP的受体则主要分布于肺血管平滑肌、气道平滑肌、气道上皮层、粘膜下的腺体、巨噬细胞、肺泡上皮和淋巴细胞等。VIP除扩张血管外，还具有舒张气道平滑肌、减少肺炎性介质释放、清除氧自由基和抗细胞凋亡等生物学作用，其在机体内的多种效应日益受到重视。支气管上皮细胞是炎症和物理刺激因子的靶细胞，在哮喘、支气管炎等气道高反应性疾病中，常伴有不同程度气道上皮结构破坏和功能受损。气道上皮的修复包括损伤边缘细胞变平、铺展、向损伤部位迁移和基细胞增殖分化成纤毛上皮细胞以及功能的恢复等环节。作为肺内重要的神经肽，VIP对气道上皮损伤后的结构及功能的恢复有何影响尚未见报道。因此，探讨VIP在气道上皮损伤修复中的作用及其分子机制，有助于揭示VIP在维持气道稳态中的作用，寻找促进肺损伤修复的可能途径，具有重要的学术价值和临床应用前景。目的：研究血管活性肠肽(VIP)对气道上皮损伤后修复过程的影响，并探讨其可能的信号转导途径。方法：(1) VIP对人气道上皮损伤修复过程的影响：以人支气管上皮细胞(human bronchial epithelial cells，HBECs)株为研究对象，观察VIP对HBECs损伤修复指数的影响；采用blind-well Boyden chamber法测定VIP对HBECs趋化性的影响；采用免疫细胞化学方法检测HBECs增殖相关抗原Ki67的表达；流式细胞仪检测VIP对HBECs S期细胞百分率(SPF)和细胞增殖指数(PI)的影响；采用免疫细胞化学、RT-PCR法检测人支气管上皮细胞钙粘素(E-cd)蛋白及其mRNA的表达。(2)信号转导机制的研究：以HBECs为研究对象，，双察PKC抑制剂H-7、CaM抑制剂W-7、PKA抑制剂H-89和ERK抑制剂PD98059对VIP促损伤修复作用的影响。(3)CREB在VIP促气道损伤
[Abstract]:Vasoactive intestinal peptide (vasoactive intestinal peptides,VIP) is an important sensory neuropeptide in the lung, which is mainly released by non-adrenergic non-cholinergic nerve fibers. VIP positive nerve fibers are widely distributed in airway epithelial cells and bronchial smooth muscle cells. VIP receptors are mainly distributed in pulmonary vascular smooth muscle, airway epithelium, submucosal glands, macrophages, and so on. Alveolar epithelium and lymphocytes. VIP not only dilates blood vessels, but also relaxes airway smooth muscle, reduces the release of pneumonic mediators, scavenges oxygen free radicals and inhibits apoptosis. Bronchial epithelial cells (BECs) are the target cells of inflammatory and physical stimulating factors. In airway hyperresponsiveness diseases such as asthma and bronchitis, bronchial epithelial cells are often accompanied with various degree of destruction of airway epithelial structure and impairment of function. The repair of airway epithelium includes flattening and spreading of injured edge cells, migration to injury site, proliferation and differentiation of basal cells into ciliated epithelial cells, and functional recovery. As an important neuropeptide in lung, the effect of VIP on the recovery of structure and function after airway epithelial injury has not been reported. Therefore, to explore the role of VIP in the repair of airway epithelial injury and its molecular mechanism is helpful to reveal the role of VIP in maintaining airway homeostasis and to find possible ways to promote the repair of lung injury, which has important academic value and clinical application prospect. Aim: to investigate the effect of vasoactive intestinal peptide (VIP) on the repair of airway epithelium after injury and to explore its possible signal transduction pathway. Methods: (1) the effect of VIP on the repair process of human airway epithelial injury: the effect of VIP on the repair index of HBECs injury was observed in human bronchial epithelial cell line (human bronchial epithelial cells,HBECs. The effect of VIP on chemotaxis of HBECs was determined by blind-well Boyden chamber assay, and the expression of HBECs proliferation-associated antigen Ki67 was detected by immunocytochemistry. The effect of VIP on the percentage of HBECs S phase (SPF) and cell proliferation index (PI) was detected by flow cytometry. The expression of cadherin (E-cd) protein and its mRNA in human bronchial epithelial cells were detected by immunocytochemistry and RT-PCR method. (2) the mechanism of signal transduction. The effects of CaM inhibitor W-7 PKA inhibitor H-89 and ERK inhibitor PD98059 on VIP induced injury repair. (3) the effect of CREB on VIP induced airway injury
【学位授予单位】：中南大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R363

【参考文献】