小鼠心肌转化生长因子β1表达与依普利酮的影响

Effect of eplerenone on myocardial transforming growth factor-beta1 expression in mice

目的:观察高选择性醛固酮受体拮抗剂依普利酮对鸟苷酸环化酶偶联受体-A基因敲除小鼠心肌转化生长因子β1表达的影响,并对其逆转心室重构的分子机制进行探讨。方法:实验于2005-06/2006-04在河北省人民医院心内科、日本奈良县立医科大学第一内科完成。①实验材料:鸟苷酸环化酶偶联受体-A基因敲除小鼠20只由日本国立循环器病中心岸本一郎教授馈赠,雄性,12周龄,基因背景为清洁级C57BL/小鼠,随机数字表法分为空白对照组6只、依普利酮组7只、肼苯哒嗪组7只。另取同龄野生型小鼠7只作为野生组。实验过程中对动物的处置符合动物伦理学标准。②实验方法:依普利酮组每天通过饲料给予100mg/kg依普利酮,肼苯哒嗪组每天通过饮用水给予10mg/kg肼苯哒嗪,均给药4周。空白对照组与野生组不进行任何干预。③实验评估:每周检测小鼠尾动脉收缩压及体质量。各组小鼠于16周龄时采用腹主动脉抽血法处死,称取心脏重量,计算心脏重量与体质量的比值。心脏切片行Masson三色染色,并利用图像分析系统测量心肌胶原容积分数、心肌血管周围胶原面积和管腔面积之比。实时定量PCR法检测心室胶原纤维Ⅰ、胶原纤维Ⅲ、心肌转化生长因子β1 mRNA在心肌组织的表达。结果:①心室重构指标检测:与野生组比较,空白对照组收缩压、心脏重量/体质量、心肌胶原容积分数、心肌血管周围胶原面积/管腔面积均显著升高(P<0.01)。与空白对照组比较,依普利酮组上述4项指标均显著下降(P<0.05或0.01);肼苯哒嗪组收缩压明显下降(P<0.01),心脏重量/体质量、心肌胶原容积分数、心肌血管周围胶原面积/管腔面积则明显升高(P<0.05)。②心肌纤维化情况:与空白对照组比较,依普利酮治疗4周后小鼠心肌纤维化得到改善,心脏重量/体质量、心肌胶原容积分数、心肌血管周围胶原面积/管腔面积3项量化指标均降低;肼苯哒嗪治疗4周后表现为心肌纤维化加重,3项量化指标相应增加。③心肌胶原纤维Ⅰ、胶原纤维Ⅲ及心肌转化生长因子β1 mRNA的表达:与空白对照组比较依普利酮治疗4周后小鼠胶原纤维Ⅰ、胶原纤维Ⅲ及心肌转化生长因子β1的mRNA表达均下降;肼苯哒嗪治疗4周后三者mRNA的表达均升高。结论:依普利酮可独立于血压改善小鼠心肌纤维化和心室重构,其作用机制可能与抑制心室肌高表达的心肌转化生长因子β1有关。

AIM： To investigate the effects of high selective mineralocorticoid receptor （MR） antagonist eplerenone on the expression of transforming growth factor-beta1 （TGFβ1） in myocardium of guanyl cyclase-A （GC-A） knockout mice and to explore its molecular mechanism for reversing left ventricular remodeling. METHODS： The experiment was conducted in the Department of Cardiology, Hebei Provincial People＇s Hospital and the First Department of Internal Medicine, Nara Medical University, Japan between June 2005 and April 2006. ①Twenty 12-week-old male GC-A knockout （KO） mice （clean C57BL/6） were gifts from Kishimoto I in National Cardiovascular Center of Japan. They were randomly divided into blank control group （n =6）, eplerenone group （n =7） and hydralazine group （n =7）. Meanwhile, 7 wild mice with the same genetic background were selected as wild group. The disposition of all animals was accorded with the animal ethics standards during the experiments. ②Eplerenone was given in food to the mice in eplerenone group （100 mg/kg/day） and hydralazine was given in drinking water （10 mg/kg/day） for four weeks. No intervention was given to the control group and wild group. ③The tail artery systolic blood pressure （SBP） and body mass of mice were measured every week. The mice were executed by exsanguinated through abdominal aorta at 16 weeks. The ratios of the heart weight to the body mass （HW/BM） were calculated. Myocardial tissues were stained with Masson＇s Trichrome; The collagen volume fraction （CVF） and the ratio of perivascular collagen area （PVCA） to lumen area were measured with image analysis system. The mRNA expression of collagen Ⅰ, collagenⅢ and TGF- β1 in myocardium were evaluated by real-time quantitative PCR technique. RESULTS： ①Indexes of ventricular remodeling： Compared with wild group, the SBP, HW/BM, CVF and PVCA were significantly higher in control group （P 〈 0.01）. Compared with control group, the 4 indicators were all decreased significantly in eplerenone group （P 〈 0.05 or 0.01）; The SBP was significantly decreased （P 〈 0.01）, but the HW/BM, CVF and PVCA were obviously higher in hydralazine group （P 〈 0.05）. ②Compared with control group, the cardiac fibrosis was attenuated after 4 weeks of eplerenone treatment, and the three indexes of HW/BM, CVF and PVCA were all lower; But cardiac fibrosis was aggravated after 4 weeks of hydralazine treatment, and those three indexes were all increased. ③Compared with control group, the mRNA expression of collagen Ⅰ, collagen Ⅲ and TGF-β1 were all decreased after 4 weeks of eplerenone treatment; But they were all increased after 4 weeks of hydralazine treatment. CONCLUSION： Eplerenone improves cardiac fibrosis and ventricular remodeling in GC-A-KO mice by suppressing the expression of TGFβ1 in myocardial tissue.