Fu Dan-Ting, Tu Jue, Cai Yue-Qin, Cai Zhao-Wei, Zhang Li-Zong, Liu Jing-Yan, Xu Shan-Chun, Wang De-Jun
Laboratory Animal Research Center/Institute of Comparative Medicine Zhejiang Chinese Medical University, Hangzhou 310053.
Department of Internal Medicine, the First Affiliated Hospitial of Zhejiang Chinese Medicine University, Hangzhou 310006, China.
Zhongguo Ying Yong Sheng Li Xue Za Zhi. 2020 Jan 28;36(1):33-38. doi: 10.12047/j.cjap.5834.2020.007.
To investigate the mRNA, protein expression levels and the phosphorylation levels of key factors in rapidly accelerated fibrosarcoma/mitogen-activated protein kinase kinase/extracellular regulated protein kinases (Raf/MEK/ERK) pathway, and to clarify the regulatory function of Raf/MEK/ERK pathway in myocardial hypertrophy.
Twenty SD rats were divided into sham-operated group and model group. The myocardial hypertrophy model was established by transverse aortic constriction (TAC). At 12 weeks after TAC, blood samples were collected from the submandibular vein, and the serum was separated to detect the content of N terminal pro B type natriuretic peptide (NT-proBNP). After that, the rats were subjected to echocardiography and hemodynamic measurement. Then the pathological changes of myocardial tissue were observed. And the levels of mRNA, protein expression and the phosphorylation of key factors in Raf/MEK/ERK pathway were detected in myocardial tissue.
Compared with sham-operated group, left ventricular end-diastolic interventricular septal thickness (IVSd), left ventricular end-systolic interventricular septal thickness (IVSs), left ventricular end-diastolic posterior wall thickness (LVPWd) and left vebtricular end-systolic posterior wall thickness (LVPWs) in TAC model group were increased significantly (P<0.05,P<0.01), left ventricular end-systolic diameter (LVIDs) was decreased significantly (P<0.01), LV Mass and LW(LV Mass/Weight)were increased significantly (P<0.05, P<0.01). The levels of heart rate (HR), left ventricular pressure maximal rate of rise (+dp/dtmax), left ventricular pressure maximal rate of fall (-dp/dtmax) were decreased significantly (P<0.01). The serum level of NT-proBNP in TAC rat was increased significantly (P<0.01). The myocardial cells in TAC model group were arranged disorderly, myocardial cell hypertrophy, cytoplasm were increased significantly, and inflammatory cells infiltrated. A large amount of collagen fibers were deposited and large area of myocardial cells were stained blue in TAC rat. The expression levels of phospho-c-Raf (Ser259) and phospho-c-Raf (Ser338) in myocardial tissue were significantly increased (P<0.01), meanwhile the expression levels of phospho- MEK1/2(Ser217/Ser221) and phospho-ERK1/2 (Thr202/Tyr204) were also significantly increased (P<0.01).
The regulatory role of Raf / MEK / ERK pathway in cardiac hypertrophy may be through the activation of phosphorylation of c-raf, MEK1, Mek2, ERK1 and ERK2 at specific sites.
研究快速进展性纤维肉瘤/丝裂原活化蛋白激酶激酶/细胞外调节蛋白激酶(Raf/MEK/ERK)通路关键因子的mRNA、蛋白表达水平及磷酸化水平,阐明Raf/MEK/ERK通路在心肌肥大中的调控作用。
将20只SD大鼠分为假手术组和模型组。采用主动脉弓缩窄术(TAC)建立心肌肥大模型。TAC术后12周,从下颌下静脉采集血样,分离血清检测N末端B型利钠肽原(NT-proBNP)含量。之后,对大鼠进行超声心动图和血流动力学测量。然后观察心肌组织的病理变化。并检测心肌组织中Raf/MEK/ERK通路关键因子的mRNA、蛋白表达及磷酸化水平。
与假手术组相比,TAC模型组左心室舒张末期室间隔厚度(IVSd)、左心室收缩末期室间隔厚度(IVSs)、左心室舒张末期后壁厚度(LVPWd)和左心室收缩末期后壁厚度(LVPWs)显著增加(P<0.05,P<0.01),左心室收缩末期内径(LVIDs)显著降低(P<0.01),左心室质量和左心室重量比(LV Mass/Weight)显著增加(P<0.05,P<0.01)。心率(HR)、左心室压力最大上升速率(+dp/dtmax)、左心室压力最大下降速率(-dp/dtmax)显著降低(P<0.01)。TAC大鼠血清NT-proBNP水平显著升高(P<0.01)。TAC模型组心肌细胞排列紊乱,心肌细胞肥大,细胞质显著增加,有炎性细胞浸润。TAC大鼠大量胶原纤维沉积,大面积心肌细胞呈蓝色染色。心肌组织中磷酸化c-Raf(Ser259)和磷酸化c-Raf(Ser338)表达水平显著升高(P<0.01),同时磷酸化MEK1/2(Ser217/Ser221)和磷酸化ERK1/2(Thr202/Tyr204)表达水平也显著升高(P<0.01)。
Raf/MEK/ERK通路在心肌肥大中的调控作用可能是通过特异性位点激活c-raf、MEK1、Mek2、ERK1和ERK2的磷酸化实现的。