ERK1/2信号转导通路在大鼠冠状动脉微栓塞致心肌炎症及心功能损伤的作用机制

目的 探讨ERK1/2信号通路对大鼠冠状动脉微栓塞(CME)致心肌炎症及心功能损伤的影响.方法 SD大鼠分为假手术组、微栓塞组和PD98059组(每组15只).采用夹闭升主动脉经左心室注射42 μm微球0.1 ml(3×10~4/ml,3000个)建立大鼠CME模型.PD98059组大鼠术前30 min静脉注射细胞外信号调节激酶(ERK1/2)抑制剂PD98059.应用Western印迹和免疫组化检测心肌组织ERK1/2磷酸化程度及分布;采用心脏超声评价心功能;HE染色观察心肌组织炎症细胞浸润情况;RT-PCR分析肿瘤坏死因子α(TNF-α)和巨噬细胞移动抑制因子(MIF)mRNA表达水平;应用电泳迁移率转变分析(EMSA)评价核转录因子(NF-κB)的活性.结果 微栓塞组比假手术组显著增加心肌组织ERK1/2蛋白磷酸化(0.92±0.10比0.61±0.04)、心肌炎症细胞数(455±16)个比(47±7)个]、TNF-αmRNA(0.94±0.04比0.60±0.09)和MIFmRNA表达(1.30±0.44比0.63±0.25)以及NF-κB活化(541±25比311±65)(均P<0.05);左室射血分数显著下降(73±3)%比(83±4)%,P<0.05].与微栓塞组相比,PD98059组阻断ERK1/2蛋白磷酸化(0.48±0.11比0.92±0.10,P<0.05),同时抑制了心肌炎症反应,TNF-α mRNA的表达减少(0.42±0.06比0.94±0.04,P<0.05),但MIFmRNA表达未见明显改变(1.17±0.37比1.30±0.44,P>0.05),NF-κB的活性降低(105±14比541±25,P<0.05),炎症细胞浸润减少(401±12)个比(455±16)个,P<0.05],左室射血分数提高(76±4)%比(73±3)%,P<0.05].结论 ERK1/2信号转导通路激活是CME致心肌炎症反应及心功能损伤的重要机制,提示抑制ERK1/2信号转导通路可能成为CME防治的潜在新靶点.

Myocardial inflammation and injury induced by coronary microembolization in rats: role of ERK1/2 signaling pathway

Objective To determine the role of extraceilular signal-regulated kinases1/2 ERK1/2) signaling pathway in regulating myocardial inflammation and cardiac function in a rat model of coronary microembolization (CME). Methods The Sprague-Dawley rats were randomly divided into three groups:sham-operated group( n = 15 ), coronary microcmbolization group ( n = 15 ) and PD98059 group ( n = 15 ).CME model was established by injection of 42 μm microspheres 0. 1 ml (3×10~4/ml, 3000 ) into left ventricle while occluding the ascending aorta. At 30 minutes pre-operation, rats of PD98059 group were injected with PD98059 IV, a specific ERK1/2 inhibitor. Western blot and immunochemical analysis were used to determine the activation and distribution of ERK1/2. Echocardiography was employed to evaluate cardiac functions. The hematoxyhn-eosin staining was used to assay myocardial inflammation. Expression of TNF-α and MIF mRNA was determined by RT-PCR analysis and activity of NF-κB assessed by electrophoretic mobility shift assay. ResultsIn comparison with sham-operated group, CME increased phosphorylatian of ERK1/2 (0. 92±0. 10 vs 0. 61±0. 04), local myocardial inflammatory cells(455±16 vs47± 7), expression of TNF-α mRNA (0. 94± 0. 04 vs 0. 60 ± 0. 09 ) and MIF mRNA ( 1.30 ± 0. 44 vs0. 63 ± 0. 25 ) and activity of NF-κB ( 541 ± 25 vs 311 ± 65 ) in myocardium ( all P<0. 05 ). All of these dramatically induced cardiac dysfunctionLVEF (73 ± 3 ) % vs (83 ± 4) %, P<0. 05]. To compare with CME group, treatment of specific ERK1/2 inhibitor PD98059 blocked the activation of ERK1/2 (0. 48 ±0. 11 vs 0.92 ±0. 10, P <0.05), decreased inflammatory cells (401 ± 12 vs 455 ± 16, P<0.05),decreased expression of TNF-α mRNA ( 0. 42 ± 0. 06 vs 0. 94 ± 0. 04, P<0. 05 ) and suppressed activity of NF-κB (105 ± 14 vs 541 ± 25, P<0. 05 ). Most importantly, PD98059 treatment ameliorated cardiac functions dramaticallyLVEF (76±4)% vs (73 ± 3)%, P <0. 05]. However there was no significant change in the expression of MIF mRNA ( 1.17 ± 0. 37 vs 1.30 ± 0. 44, P>0. 05 ). Conclusion The present study demonstrates a novel role of ERK1/2 signaling pathway in promoting myocardial inflammation in CME. And ERK1/2 may be a novel drug target for CME therapy.