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抑制G蛋白偶联受体35对小鼠缺血性心肌损伤的保护作用
作者:贺磊 杨怡 田玥 王挺 沈阳 裴海峰 杨大春 
单位:400038  重庆  陆军军医大学研究生管理大队(贺磊、杨大春) 610083  成都  成都军区总医院心血管内科(杨怡、田玥、王挺、沈阳、裴海峰、杨大春) 646000  四川泸州  西南医科大学临床医学院(杨怡、田玥、沈阳) 
关键词:G蛋白偶联受体35 缺氧/缺血 细胞凋亡 心脏功能 
分类号:R542.2
出版年,卷(期):页码:2018,43(2):101-106
摘要:

[摘要]  目的  探讨抑制G蛋白偶联受体35(GPR35)对缺氧/缺血条件下心肌细胞的保护作用。方法  细胞实验:为检测GPR35在细胞缺氧情况下的变化,将小鼠心肌细胞系分为常氧组、缺氧组,6h后采用q-PCRWestern blotting检测GPR35的表达情况;为进一步验证GPR35抑制剂的作用,将小鼠心肌细胞系分为常氧组、缺氧组、缺氧+溶剂组、缺氧+CID2745678(GPR35活性抑制剂3μmol/L)6h后通过流式细胞仪及TUNEL染色检测各组心肌细胞凋亡水平。动物实验:为检测GPR35在心肌缺血条件下的变化,将雄性C57小鼠分为假手术组(n=6),心梗组(n=8),术后3d心脏取材,采用q-PCRWestern blotting检测GPR35的表达情况;为进一步验证GPR35抑制剂的作用,将小鼠分为假手术组(n=6)、心梗组(n8)、心梗+溶剂组(n=8,腹腔注射等量溶剂)、心梗+CID2745678(n=8,腹腔注射12μg/kg CID2745678),术后4周通过小动物超声检测左室短轴缩短率(LVFS)、左室射血分数(LVEF),并通过Masson染色检测心肌纤维化水平。结果  细胞实验结果显示,小鼠心肌细胞缺氧6h后,缺氧组GPR35 mRNA及蛋白表达水平与常氧组比较明显增加(P<0.01P<0.05);与缺氧+溶剂组比较,缺氧+CID2745678组凋亡细胞明显减少(P<0.05)。动物实验结果显示,小鼠心梗后3d,心梗组心肌组织GPR35 mRNA及蛋白表达水平与假手术组比较明显增加(P<0.01P<0.05);与心梗+溶剂组比较,心梗+CID2745678组心脏LVFSLVEF明显升高(P<0.05),心肌纤维化水平明显降低(P<0.05)结论  抑制GPR35可以减轻缺氧/缺血心肌损伤。

[Abstract]  Objective  To investigate the protective role of GPR35 inhibition on hypoxic myocardial cell line and mouse myocardial infarction (MI) model. Methods  For investigating the changes of GPR35 expression in hypoxic environment, the murine myocardial cells (MCM) were divided into normoxia group and hypoxia group, the mRNA expression of GPR35 was determined by q-PCR and the protein level was measured by Western blotting 6h after incubation. For further studying the role of GPR35, MCM were divided into four groups: normoxia, hypoxia, hypoxia+vehicle, hypoxia+CID2745678 (GPR35 inhibitor, 3μmol/L) group. Accordingly, the apoptosis of cardiomyocytes was measured by flow cytometer and TUNEL. For investigating the changes of GPR35 expression in the state of myocardial ischemia, the C57 male mice were divided into sham group (n=6) and MI group (n=8), the mRNA expression of GPR35 was determined by q-PCR and the protein level was measured by Western blotting 3 days after MI. For further studying the role of GPR35, the C57 mice were divided into four groups: sham (n=6), MI (n=8), MI+vehicle (n=8) and MI+CID2745678 (n=8) group. Ultrasound echocardiography was performed 4 weeks after MI. Mice were then sacrificed and the hearts were removed and stained with Masson to measure the myocardial fibrosis area. Results Compared with normoxia group, the levels of GPR35 mRNA and protein increased obviously in hypoxia group (P<0.01, P<0.05); Compared with hypoxia+vehicle group, the myocardial cells apoptosis in hypoxia+CID2745678 group decreased markedly (P<0.05). Three days after MI, compared with the sham group, the levels of GPR35 mRNA and protein increased obviously (P<0.01, P<0.05) in MI group; Compared with MI+vehicle group, the left ventricular fraction shortening (LVFS) and left ventricular ejection fraction (LVEF) relieved obviously (P<0.05) and myocardial fibrosis level declined markedly in MI+CID2745678 group (P<0.05). Conclusion Inhibition of GPR35 could decrease the apoptosis of cardiomyocytes cultured in hypoxia and attenuated the injury of myocardial ischemia.

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