硫化氢对LPS致小鼠急性肺损伤的保护及机制研究

发布时间：2018-05-05 20:35

本文选题：急性肺损伤 + 内皮祖细胞　；参考：《第三军医大学》2017年硕士论文

【摘要】：背景与目的:急性肺损伤(acute lung injury,ALI)是多种繁杂病因造成的毛细血管内皮细胞和肺泡上皮细胞的损伤,伴有不断加重的低氧血症和ARDS。ALI具有病死率高、治疗困难、预后差等特点,寻找有效的ALI治疗靶点仍然是目前危重病医学亟待解决的关键问题之一[1]。H_2S被视为和CO、NO一类的第三种气态递质[2]。前期研究发现,在肢体损伤导致的远隔肺损伤中,肺H_2S产生减少、循环H_2S浓度降低,给以外源性H_2S供体Na HS可减轻肢体损伤导致的肺损伤[3]。大量文献报道,外源性H_2S可减轻高压氧、LPS、缺血再灌注、机械通气引起的肺损伤[4]。但是,H_2S在LPS诱发小鼠ALI中发挥保护效用的机制尚未阐明。本研究通过经气管内注射LPS来构建小鼠ALI模型,检测炎症因子、H_2S、内皮祖细胞(endothelialprogenitorcel,EPCs)、肺巨噬细胞(alveolar macrophage,AM)等在ALI小鼠体内的含量变化,观察H_2S对于ALI损伤程度的影响,探讨:(1)H_2S对EPCs动员和再内皮化的作用,(2)H_2S与ALI肺内AM表型转化的关联,为ALI的治疗提供新的解决思路。方法:1.气管内注射LPS构建小鼠ALI模型。2.肉眼下比较各组小鼠肺大体标本,光学显微镜下比较肺病理学改变,测定肺系数和肺病理学评分。3.测定不同时相点各组小鼠血清炎症因子IL-6、IL-8和TNF-α浓度。4.测定各组小鼠血清H_2S在不同时相点的浓度。5.检测各组小鼠肺组织CD34和VEGFR2的表达,计数CD34+/VEGFR2+细胞数目。6.检测各组小鼠肺组织i NOS和Arginase的表达。结果:1.成功建立ALI模型:LPS组各时相点肺组织呈现不同程度的毛细血管上皮和肺泡壁破坏、肺泡壁和肺间质显著增厚,肺间质及肺泡腔内大量红细胞渗出和炎性细胞浸润;与Sham组相比,肺系数、IQA评分显著升高。2.LPS组第3天肺大体标本的出血面积显著大于其它时相点,肺组织肺间质增厚、红细胞渗出和炎性细胞浸润程度最为严重,肺系数和IQA病理学评分高于其他时相点。3.Na HS组小鼠肺组织IQA值及肺系数比LPS组显著降低,PAG组和AOAA组小鼠肺组织IQA值及肺系数比LPS组显著升高。4.ALI小鼠血清H_2S浓度显著减少,Na HS组血清H_2S浓度比LPS组显著增加。5.Na HS组小鼠肺组织CD34和VEGFR2表达较LPS组显著增多,CD34+/VEGFR2+细胞数目显著增加。6.Na HS组肺组织中Arginase/i NOS比值较LPS组显著升高。结论:1.通过气管内滴注LPS构建小鼠ALI模型的方法具有重复性好,稳定性强的优点。为进一步研究ALI相关生理病理机制提供稳定的动物模型基础。2.LPS气管内滴注后第3天是ALI小鼠肺损伤最严重的时间节点,因此作为后续研究H_2S对ALI的作用及机制的时相点。3.外源性H_2S可通过促进EPCs动员及在肺部的再内皮化、纠正ALI时AM的极化失衡,抑制炎症反应、促进损伤肺组织愈合,进而减轻ALI肺损伤。
[Abstract]:Background & objective: acute lung injury (Ali) is a kind of injury of capillary endothelial cells and alveolar epithelial cells caused by many complicated causes. It is accompanied by increasing hypoxemia and ARDS.ALI with the characteristics of high mortality, difficult treatment and poor prognosis. Finding an effective therapeutic target for ALI is still one of the key problems in critical medicine [1] .Hs / s _ 2S is regarded as the third gaseous transmitter such as CONO _ 2 [2]. Previous studies showed that in the distant lung injury caused by limb injury, H2S production and circulating H2S concentration decreased in the lung. The exogenous H2S donor NaHS could reduce the lung injury caused by limb injury [3]. It is reported that exogenous H2S can reduce lung injury induced by hyperbaric oxygen, ischemia reperfusion and mechanical ventilation. However, the mechanism of the protective effect of H2S in LPS induced ALI in mice has not been elucidated. In this study, the ALI model of mice was established by intratracheal injection of LPS, and the levels of inflammatory factor, endothelial-progenitor cells, endothelial progenitor cells, alveolar macrophage AM) were detected in ALI mice, and the effects of H2S on the degree of ALI damage were observed. To explore the role of H2S in mobilization and re-endothelialization of EPCs and its relationship with the phenotypic transformation of AM in the lung of ALI, and to provide a new solution for the treatment of ALI. Method 1: 1. Establishment of mouse ALI model by intratracheal injection of LPS. Gross lung samples of each group were compared under naked eye, lung pathological changes were compared under optical microscope, lung coefficient and lung pathological score were measured. The levels of IL-6, IL-8 and TNF- 伪 in serum of mice at different time points were determined. The concentration of H 2S in serum of each group of mice was determined at different time points. The expression of CD34 and VEGFR2 in lung tissue of each group was detected, and the number of CD34 / VEGFR2 cells. The expression of I NOS and Arginase in lung tissue of each group was detected. The result is 1: 1. The ALI model was successfully established at different time points in the ALI group. The pulmonary tissue showed various degrees of destruction of capillary epithelium and alveolar wall, significant thickening of alveolar wall and interstitial tissue, massive exudation of erythrocytes and infiltration of inflammatory cells in the interstitial and alveolar lumen of the lung, as compared with that in the Sham group. IQA score of lung coefficient was significantly increased in LPS group. 2. The hemorrhage area of lung gross specimens in LPS group was significantly larger than that in other time points on the 3rd day. The pulmonary interstitial thickening, erythrocyte exudation and inflammatory cell infiltration were the most serious in LPS group. Lung coefficient and IQA pathological score were higher than those of other time point. 3.The lung tissue IQA and lung coefficient of NaHS group were significantly lower than that of LPS group. 4. Serum H2S concentration of Ali mice was significantly higher than that of LPS group. Compared with LPS group, the expression of CD34 and VEGFR2 in lung tissue of Na-HS group was significantly higher than that of LPS group. The number of CD34 / VEGFR2 cells was significantly increased. 6. The ratio of Arginase/i NOS in lung tissue of NaHS group was significantly higher than that of LPS group. Conclusion 1. The method of establishing mouse ALI model by intratracheal instillation of LPS has the advantages of good repeatability and strong stability. In order to provide a stable animal model basis for further study on the physiological and pathological mechanism of ALI. 2.The third day after intratracheal instillation of ALI was the most severe time node of lung injury in ALI mice. Exogenous H2S can correct the imbalance of AM polarization in ALI by promoting EPCs mobilization and pulmonary re-endothelialization, inhibit inflammatory reaction, promote the healing of injured lung tissue, and then alleviate the lung injury of ALI.
【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R563.8

【参考文献】