白介素17在心房颤动模型大鼠中的作用机制研究

The Mechanism of Interleukin-17 in Atrial Fibrillation of Rats

目的观察白介素17(IL-17)在心房颤动模型大鼠中的作用及主要来源细胞,并探讨IL-17在心房颤动发病中的作用机制。方法构建SD大鼠心房颤动模型,采用酶联免疫吸附法测定(ELISA)和实时定量聚合酶链式反应(Real-Time PCR)测定心房颤动大鼠IL-17表达水平。流式细胞术分选心房组织中浸润的淋巴细胞,将分选后的T淋巴细胞培养并利用ELISA检测上清液IL-17水平,明确IL-17来源。经腹腔注射鼠IL-17单克隆抗体中和IL-17,探讨IL-17中和后SD大鼠(对照组、心房颤动组和心房颤动干预组)心房组织间质纤维化水平及纤维化相关蛋白表达水平。结果ELISA结果显示:心房颤动组SD大鼠血浆IL-17含量高于对照组(P<0.01);Real-Time PCR结果显示:IL-17基因表达高于对照组(P<0.001);流式细胞术分选结果显示:心房颤动辅助T淋巴细胞占成熟T淋巴细胞比例升高,差异有统计学意义(P=0.007)。ELISA检测结果发现心房颤动血清高水平IL-17主要来源于CD3+/CD4+辅助T淋巴细胞。给予IL-17单克隆中和抗体后,心房颤动SD大鼠房性期前收缩、房性二联律、房性三联律较心房颤动组降低,心房不应期和动作电位时程较心房颤动组增加。给予IL-17单克隆中和抗体后心房颤动SD大鼠心房组织间质纤维化水平较心房颤动组降低,Western Blotting结果显示:心房颤动组转化生长因子β1(TGF-β1)、平滑肌肌动蛋白(α-SMA)、基质金属蛋白酶-9(MMP-9)纤维化相关蛋白表达水平显著增加,而给予IL-17抗体干预后这些纤维化相关蛋白表达水平较心房颤动组显著降低。结论心房颤动时高水平IL-17主要来源于辅助T淋巴细胞17,其细胞分泌的IL-17可促进心房纤维化,导致心房颤动发生。

Objective To observe the role of interleukin-17(IL-17)in atrial fibrillation and its main source cells,and to explore the mechanism of IL-17 in the pathogenesis of atrial fibrillation.Methods The model of atrial fibrillation in SD rats was established,and the expression of IL-17 in atrial fibrillation rats was detected by enzyme-linked immunosorbent assay(ELISA)and real-time quantitative polymerase chain reaction.The infiltrating lymphocytes in atrial tissue were sorted by flow cytometry.The T lymphocytes after flow cytometry sorting were cultured and the expression of IL-17 in the supernatant was detected by ELISA to identify the source of IL-17.Intraperitoneal injection of monoclonal antibody against IL-17 was used to neutralize IL-17 in SD rats,and the levels of interstitial fibrosis and fibrosis-related proteins in atrial tissues of SD rats(control group,atrial fibrillation group and atrial fibrillation intervention group)were compared after the neutralization of IL-17.Results ELISA showed that the expression of IL-17 in SD rats with atrial fibrillation was significantly higher than that in control group(P<0.01).Real-time PCR results showed that the expression of IL-17 gene was significantly higher than that in control group(P<0.001).Flow cytometry results showed that the proportion of helper T lymphocytes in mature T lymphocytes significantly increased(P=0.007).The results of ELISA showed that the high level of IL-17 in atrial fibrillation was mainly derived from CD3+/CD4+helper T lymphocytes.After administration of IL-17 monoclonal neutralizing antibody,atrial premature beats,atrial diploid rhythm,and atrial triplet rhythm in SD rats were reduced more than those in the atrial fibrillation group,and the durations of atrial anechoic phase and action potential in SD rats increased more than those in the atrial fibrillation group.After administration of IL-17 monoclonal neutralizing antibody,the level of interstitial fibrosis in atrial tissue in SD rats reduced more than that in atrial fibrillation group.Western Blotting results showed that the expressions of transformed growth factor 1,smooth muscle actin,and matrix metalloproteinase-9 fibrosis related proteins significantly increased in the atrial fibrillation group,while the expressions of these fibrosis related proteins were significantly decreased after IL-17 neutralization.Conclusion The high level of IL-17 in atrial fibrillation was mainly derived from helper T lymphocytes 17.IL-17 secreted by Th17 cells could promote atrial fibrosis and promote the occurrence of atrial fibrillation.