Role of nicotinamide riboside in lipotoxic cardiomyopathy: a mechanistic study
谢赛阳 1方文熙 1刘士强 1邓伟 1唐其柱 1刘新艳
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作者信息
1. 武汉大学人民医院心血管内科 代谢与相关慢病湖北省重点实验室，武汉 430060
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摘要
目的 探讨烟酰胺核糖对肥胖相关脂毒性心肌病的影响及其具体机制。 方法 将C57BL/6N雄性小鼠（8周龄）分为标准饮食组（对照组）、高脂饮食组和高脂+烟酰胺核糖饮食组（n=10），高脂饮食组通过高脂饲料喂养24周构建肥胖小鼠脂毒性心肌病模型，高脂+烟酰胺核糖饮食组小鼠在高脂饲料的基础上在饮用水中加入40 mmol/L烟酰胺核糖喂养12周。造模结束后检测小鼠尾动脉血压，收集小鼠外周血分析血糖、血脂等指标，应用超声心动图评估小鼠心功能，随后进行小鼠心脏取材并用于病理学和分子生物学分析。分别通过心脏大体拍照检测小鼠心重与胫骨长、麦胚凝集素（WGA）定量小鼠心肌细胞面积和苦味酸-天狼猩红（PSR）染色显示小鼠心肌纤维化水平评估烟酰胺核糖对小鼠心肌重构的影响。采用透射电镜检测小鼠心肌脂滴浸润情况；采用免疫印迹检测小鼠心肌细胞Sirt1及Serca2a蛋白表达水平。 结果 与对照组比较，高脂饮食小鼠的体质量、收缩压、舒张压、总胆固醇、三酰甘油和LDL-c水平均明显增高（均P<0.05），应用烟酰胺核糖后可缓解高脂所致的上述指标的增高（均P<0.05）；高脂饮食和应用烟酰胺核糖均不影响小鼠心率和血糖。与对照组比较，高脂饮食组小鼠左心室射血分数和心脏超声E峰、A峰比值（E/A）降低（均P<0.05），烟酰胺核糖则增高左心室射血分数和E/A比值（均P<0.05）。与对照组比较，高脂饮食组小鼠心重与胫骨长比值、左心室心肌细胞横截面积、胶原含量均增加（均P<0.05），烟酰胺核糖则可抑制小鼠心肌的上述肥大和纤维化表现（均P<0.05）。与对照组比较，高脂饮食导致小鼠左心室心肌脂滴沉积明显增多[（13.8±0.86）/100 μm2比（4.7±0.51）/100 μm2，P<0.05]，而烟酰胺核糖可减轻高脂诱导的心肌纤维化[（6.2±0.77）/100 μm2比（13.8±0.86）/100 μm2，P=0.021]。免疫印迹显示脂毒性心肌组织中Sirt1及心肌肌浆网Ca2+-ATP酶（Serca2a）蛋白表达水平均较对照组降低（均P<0.05），Serca2a的乙酰化水平增高（均P<0.05），但烟酰胺核糖可增加Sirt1及Serca2a的表达并促进Serca2a的去乙酰化（均P<0.05）。 结论 烟酰胺核糖通过刺激Sirt1介导的Serca2a去乙酰化缓解高脂所致的脂毒性心肌病。 Objective To investigate the role and mechanism of nicotinamide riboside in obesity-related lipotoxic cardiomyopathy. Methods Male C57BL/6N mice at 8 weeks of age were randomly divided into the normal diet group (control group), high-fat diet group, and high-fat diet with nicotinamide ribose group (n=10 each). The mice on high-fat diet were modeled for obese mice lipotoxic cardiomyopathy over a24-week feeding. The mice in the high-fat diet with nicotinamide ribose group were administered with 40 mmol/L nicotinamide ribose in drinking water for 12 weeks on top of the high-fat diet. After modeling, all mice were measured for blood pressure of the caudal artery, collected of peripheral blood samples for testing of blood sugar, and lipids, and evaluated for heart function by echocardiography. Then the mice hearts were harvested and subjected to pathological and molecular biological studies. The heart weight and tibia length of the mice were measured by gross photography. The area of mice cardiomyocytes was quantified by staining with wheat germ agglutinin (WGA). Picrosirius red (PSR) staining was used to show the myocardial fibrosis in mice such that the effect of nicotinamide riboside on mice myocardial remodeling was evaluated. The lipid droplets infiltration in mice cardiomyocytes was examined by transmission electron microscopy. The protein expression levels of Sirt1 and Serca2a in mice cardiomyocytes were detected by Western blotting. Results Compared with the control group, the mice on high-fat diet presented significant increases in body weight, systolic blood pressure, diastolic blood pressure, total cholesterol, triglyceride and LDL-c levels (all P<0.05), and the elevation in these indicators indiced by high fat diet was alleviated by the addition of nicotinamide riboside (allP<0.05). The heart rate and blood sugar level in mice were not affected by high-fat diet with or without nicotinamide riboside. Compared with the control group, the mice on high-fat diet had higher or greater left ventricular ejection fraction and lower ratio of E to A peaks (E/A) on echocardiograpy (bothP<0.05), and addition of nicotinamide riboside increased the left ventricular ejection fraction and E/A ratio (bothP<0.05). Compared with the control group, the mice on high-fat diet had higher or greater ratio of heart weight to tibia length, cross-sectional area and collagen contents in cardiomyocytes of the left ventricle (allP<0.05), whereas addition of nicotinamide riboside was found to inhibit the myocardial hypertrophy and fibrosis as described above (allP<0.05). Compared with the control group, high-fat diet led to a significant increase in deposition of lipid droplets in the left ventricular myocardium[ (13.8±0.86) /100 μm2 vs (4.7±0.51) /100 μm 2, P<0.05], while nicotinamide ribose could alleviate such lipid-induced myocardial fibrosis[ (6.2±0.77) /100 μm2 vs (13.8±0.86) /100 μm 2, P=0.021]. Western blotting showed lower protein expression levels of Sirt1 in the lipotoxic myocardial tissues and sarco (endo) plasmic reticulum Ca2+-ATPase (Serca2a), along with increased acetylation of Serca2a in mice on high-fat diet compared with the control group (all P<0.05), while nicotinamide riboside was shown to increase Sirt1 and Serca2a expression and promote Serca2a deacetylation (allP<0.05) . Conclusion Nicotinamide riboside may alleviate high fat-induced lipotoxic cardiomyopathy by promoting Sirt1-mediated deacetylation of Serca2a.
Abstract
Objective To investigate the role and mechanism of nicotinamide riboside in obesity-related lipotoxic cardiomyopathy. Methods Male C57BL/6N mice at 8 weeks of age were randomly divided into the normal diet group (control group), high-fat diet group, and high-fat diet with nicotinamide ribose group (n=10 each). The mice on high-fat diet were modeled for obese mice lipotoxic cardiomyopathy over a24-week feeding. The mice in the high-fat diet with nicotinamide ribose group were administered with 40 mmol/L nicotinamide ribose in drinking water for 12 weeks on top of the high-fat diet. After modeling, all mice were measured for blood pressure of the caudal artery, collected of peripheral blood samples for testing of blood sugar, and lipids, and evaluated for heart function by echocardiography. Then the mice hearts were harvested and subjected to pathological and molecular biological studies. The heart weight and tibia length of the mice were measured by gross photography. The area of mice cardiomyocytes was quantified by staining with wheat germ agglutinin (WGA). Picrosirius red (PSR) staining was used to show the myocardial fibrosis in mice such that the effect of nicotinamide riboside on mice myocardial remodeling was evaluated. The lipid droplets infiltration in mice cardiomyocytes was examined by transmission electron microscopy. The protein expression levels of Sirt1 and Serca2a in mice cardiomyocytes were detected by Western blotting. Results Compared with the control group, the mice on high-fat diet presented significant increases in body weight, systolic blood pressure, diastolic blood pressure, total cholesterol, triglyceride and LDL-c levels (all P<0.05), and the elevation in these indicators indiced by high fat diet was alleviated by the addition of nicotinamide riboside (allP<0.05). The heart rate and blood sugar level in mice were not affected by high-fat diet with or without nicotinamide riboside. Compared with the control group, the mice on high-fat diet had higher or greater left ventricular ejection fraction and lower ratio of E to A peaks (E/A) on echocardiograpy (bothP<0.05), and addition of nicotinamide riboside increased the left ventricular ejection fraction and E/A ratio (bothP<0.05). Compared with the control group, the mice on high-fat diet had higher or greater ratio of heart weight to tibia length, cross-sectional area and collagen contents in cardiomyocytes of the left ventricle (allP<0.05), whereas addition of nicotinamide riboside was found to inhibit the myocardial hypertrophy and fibrosis as described above (allP<0.05). Compared with the control group, high-fat diet led to a significant increase in deposition of lipid droplets in the left ventricular myocardium[ (13.8±0.86) /100 μm2 vs (4.7±0.51) /100 μm 2, P<0.05], while nicotinamide ribose could alleviate such lipid-induced myocardial fibrosis[ (6.2±0.77) /100 μm2 vs (13.8±0.86) /100 μm 2, P=0.021]. Western blotting showed lower protein expression levels of Sirt1 in the lipotoxic myocardial tissues and sarco (endo) plasmic reticulum Ca2+-ATPase (Serca2a), along with increased acetylation of Serca2a in mice on high-fat diet compared with the control group (all P<0.05), while nicotinamide riboside was shown to increase Sirt1 and Serca2a expression and promote Serca2a deacetylation (allP<0.05) . Conclusion Nicotinamide riboside may alleviate high fat-induced lipotoxic cardiomyopathy by promoting Sirt1-mediated deacetylation of Serca2a.
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