IL-17A在大鼠房颤模型中的作用和机制研究

发布时间：2018-09-17 18:19

【摘要】：背景房颤是临床上最常见的心律失常之一。近年来,一类新的辅助性T细胞亚群-Th17细胞(T help cell 17,Th17)及其特异性效应因子白介素-17(interleukin-17,IL17)备受关注,IL-17被证明是一种重要的炎症反应介质。研究发现,IL-17在多种心血管疾病的发生发展中发挥了重要的作用,包括动脉粥样硬化、慢性心力衰竭、急性冠状动脉综合征和病毒性心肌炎。但是目前关于IL-17与房颤发生相关的研究证据尚较少。为探索IL-17是否是房颤的独立危险因素和重要的炎症标记物,本课题组前期通过配对的病例对照研究发现,外周血浆中IL-17A与房颤发生相关联,且随着因子水平增高,关联强度越强,且可能通过影响心脏结构重构而影响房颤发生。但是IL-17A在房颤发生和维持中的具体作用和机制如何,外周血和心房组织中IL-17A水平是否存在一致性,仍有待进一步研究。目的本课题利用SD大鼠,通过食道快速心房起搏的方法来构建房颤动物模型,进一步探讨IL-17与房颤发生的关联,分析外周血中IL-17A水平与心房组织中是否存在一致性及二者与心房纤维化的相关性。同时,分析IL-17A与基质金属蛋白酶(matrix metalloproteinase,MMP)、基质金属蛋白酶抑制剂(tissue inhibitor metalloproteinase,TIMP)之间的相关性,进一步验证IL-17A通过引起MMPs/TIMPs失衡诱导心房纤维化的机制。研究结果将为IL-17作为房颤早期诊断和监测疾病进展的重要生物标记物提供依据,也将为全面认识房颤发生和维持的机制以及寻找新的治疗靶点提供参考依据。方法1.采用经食道快速心房起搏的方法构建大鼠房颤模型,比较分析房颤组与对照组心电数据的变化,包括QRS间期、QT间期等,以及两组心房纤维化程度的改变。2.通过酶联免疫吸附法测定房颤组与对照组血浆中IL-17A与C反应蛋白(c-reactive protein,CRP)水平的变化,以及两组血浆中MMP2、MMP9、TIMP2的水平。3.采用免疫组化法测定两组心房组织中IL-17A、MMP2、MMP9及TIMP2的水平。4.采用Pearson相关分析的方法,分析血浆和心房组织中IL-17A的一致性及二者与心房纤维化的相关性,同时分析血浆和心房组织中MMP2、MMP9、TIMP2的一致性及三者与心房纤维化的相关性,血浆和心房组织中MMP2、MMP9、TIMP2与IL-17A的相关性。结果第一章大鼠房颤模型的构建1.房颤组大鼠的心率显著加快,正常时为(159.63±18.25)次/min,房颤发作时达(203.24±33.56)次/min,同时,QRS间期和QT间期明显延长,分别由正常时的50.00±4.28 ms,103.00±1.85 ms增加至68.85±14.73 ms和134.80±22.05 ms,差异均有统计学意义(P0.05)。2.房颤组大鼠心房纤维化程度较对照组严重,胶原容积分数较对照组明显升高[(18.15±9.45)%vs(6.51±4.49)%,P0.01]。第二章大鼠房颤模型中IL-17A的作用及其与心房纤维化的关联研究1.大鼠外周血血浆中IL-17A和CRP的水平与房颤的发生密切相关,且随房颤发生次数的增多有上升趋势。2.房颤组大鼠心房组织中IL-17A的蛋白表达水平较对照组明显升高(0.17±0.05vs 0.04±0.01,P=0.003)。3.房颤组大鼠外周血血浆中IL-17A水平与心房组织中IL-17A水平存在高度相关性(r=0.721,P=0.019)。血浆中IL-17A水平与心房纤维化呈正相关(r=0.830,P=0.003),心房组织中IL-17A水平与心房纤维化也呈正相关(r=0.685,P=0.029)。第三章大鼠房颤模型中IL-17A与MMPs/TIMPs的关联研究1.尚未发现房颤组与对照组大鼠外周血血浆中MMP2、MMP9、TIMP2水平存在统计学差异。2.与对照组相比,房颤组大鼠心房组织中MMP2、MMP9水平明显升高(0.12±0.05vs 0.04±0.02,P=0.021;0.08±0.01 vs 0.03±0.01,P0.001),而TIMP2水平在房颤组明显降低(0.02±0.00 vs 0.06±0.02,P=0.004)。3.房颤组大鼠外周血血浆和心房组织中MMP2、MMP9、TIMP2的水平存在高度相关性,心房组织中MMP9水平与心房纤维化呈正相关(r=0.809,P=0.005),TIMP2水平与心房纤维化呈负相关(r=-0.884,P=0.001),未发现心房组织中MMP2水平与心房纤维化之间的相关性(r=0.599,P=0.067)。4.房颤组大鼠外周血血浆中MMP9水平与IL-17A呈正相关(r=0.648,P=0.043),TIMP2水平与IL-17A呈负相关(r=-0.695,P=0.037),未发现血浆中MMP2与IL-17A水平之间的相关性(r=0.576,P=0.082)。5.房颤组大鼠心房组织中MMP2、MMP9与IL-17A水平均呈正相关(r=0.801,P=0.005;r=0.778,P=0.008),TIMP2与IL-17A水平呈负相关(r=-0.841,P=0.002)。结论通过大鼠房颤动物模型研究,发现IL-17A与房颤的发生发展密切相关,IL-17A可能是通过影响MMPs/TIMPs平衡,引起心房结构重构,从而导致房颤的发生与发展。
[Abstract]:Background Atrial fibrillation (AF) is one of the most common arrhythmias in clinical practice. In recent years, a new helper T cell subset-Th17 (Th17) and its specific effector interleukin-17 (IL-17) have attracted much attention. IL-17 has been proved to be an important inflammatory mediator in many cardiovascular diseases. Interleukin-17 (IL-17) plays an important role in the development of atrial fibrillation, including atherosclerosis, chronic heart failure, acute coronary syndrome and viral myocarditis. Phase I matched case-control study showed that IL-17A in peripheral plasma was associated with AF, and with the increase of factor levels, the stronger the association was, and it may affect AF by affecting cardiac structural remodeling. However, the specific role and mechanism of IL-17A in the occurrence and maintenance of AF, and the role of IL-17A in peripheral blood and atrial tissue were also discussed. Objective To explore the relationship between IL-17 and atrial fibrillation in SD rats by rapid esophageal atrial pacing, and to analyze the consistency of IL-17A level in peripheral blood with atrial tissue and the relationship between them and atrial fibrosis. The correlation between IL-17A and matrix metalloproteinase (MMP) and tissue inhibitor metalloproteinase (TIMP) was analyzed to further validate the mechanism of IL-17A inducing atrial fibrosis by inducing imbalance of MMPs/TIMPs. Methods 1. A rat model of atrial fibrillation was established by transesophageal rapid atrial pacing, and the changes of electrocardiogram data, including QRS, were compared and analyzed between atrial fibrillation group and control group. The changes of plasma levels of IL-17A and C-reactive protein (CRP) and plasma levels of MMP2, MMP9 and TIMP 2 in AF group and control group were measured by enzyme-linked immunosorbent assay. 3. The levels of IL-17A, MMP2 and MMP in atrial tissues were determined by immunohistochemistry. Pearson correlation analysis was used to analyze the consistency of IL-17A in plasma and atrial tissues and their correlation with atrial fibrosis. The consistency of MMP2, MMP9, TIMP2 in plasma and atrial tissues and their correlation with atrial fibrosis were also analyzed. Result The first chapter was the establishment of rat atrial fibrillation model. 1. The heart rate of rats in AF group increased significantly from (159.63 (+ 18.25) beats per minute in normal time to (203.24 (+ 33.56) beats per minute in AF attack. Meanwhile, the QRS interval and QT interval prolonged significantly from 50.00 (+ 4.28 ms), 103.00 (+ 1.85 ms) to 68.85 (+ 14.73 ms) and 134.80 (+ 22.05) beats in normal time, respectively. The degree of atrial fibrosis in AF group was more serious than that in control group. The volume fraction of collagen in AF group was significantly higher than that in control group [(18.15+9.45)% vs (6.51+4.49)%, P 0.01]. The level of IL-17A in atrial tissue of AF group was significantly higher than that of control group (0.17.05 vs 0.04.01, P = 0.003). 3. The level of IL-17A in plasma of AF group was significantly higher than that of control group (0.17.05 vs 0.04.01, P = 0.003). The level of IL-17A in plasma was positively correlated with atrial fibrosis (r = 0.830, P = 0.003), and the level of IL-17A in atrial tissue was positively correlated with atrial fibrosis (r = 0.685, P = 0.029). Compared with the control group, the levels of MMP 2 and MMP 9 in atrial tissue of AF group were significantly higher (0.12.05 vs 0.04.02, P = 0.021; 0.08.01 vs 0.03.01, P 0.001), while the levels of TIMP 2 were significantly lower in AF group (0.02.00 vs 0.06.02, P = 0.004). The levels of MMP 2, MMP 9 and TIMP 2 in atrial tissues were positively correlated with atrial fibrosis (r = 0.809, P = 0.005). The levels of TIMP 2 were negatively correlated with atrial fibrosis (r = - 0.884, P = 0.001). No correlation was found between the levels of MMP 2 in atrial tissues and atrial fibrosis (r = 0.599, P = 0.067). 4. The levels of MMP-9 in plasma were positively correlated with IL-17A (r = 0.648, P = 0.043), and TIMP-2 was negatively correlated with IL-17A (r =-0.695, P = 0.037). No correlation was found between the levels of MMP-2 and IL-17A (r = 0.576, P = 0.082). MMP-2, MMP-9 and IL-17A were positively correlated in atrial tissues of AF rats (r = 0.801, P = 0.005; r = 0.778, P = 0.008). Conclusion IL-17A is closely related to the occurrence and development of atrial fibrillation in the rat model of atrial fibrillation. IL-17A may induce atrial remodeling by affecting the balance of MMPs/TIMPs, leading to the occurrence and development of atrial fibrillation.
【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】：R541.75;R-332

【参考文献】