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缺氧复氧时肥大心肌细胞凋亡及其与能量代谢途径转换的关系

冯兵, 刘伟, 徐静, 何作云, 杨惠标

第三军医大学新桥医院肾内科.重庆 400037

摘要

心肌细胞凋亡是心肌肥大向心力衰竭转化的重要机制，因此，抑制肥大心肌细胞凋亡可能是防治心力衰竭的有效药物靶点之一。该研究以0.1 #mu#mol/L 血管紧张素II和1 #mu#mol/L去甲肾上腺素刺激培养心肌细胞，复制心肌细胞肥大模型，用三气孵箱培养。缺氧条件是95% N_(2)和5% CO_(2)，控制氧分压低于5 mmHg以下，8 h后常氧培养，液闪计数法测定丙酮酸脱氢酶(pyruvate dehydrogenase, PDH)和肉碱脂酰转移酶--1(carnitine palmitoyltransferase 1, CPT--1)活性，糖氧化、糖酵解、脂肪酸氧化率，以及细胞凋亡百分率，分析肥大心肌细胞能量代谢变化与细胞凋亡间的关系。结果如下：(1)与常氧培养比较，缺氧8 h后，活化型丙酮酸脱氢酶(PDHa)和CPT--1活性均有显著下降，但复氧早期肥大心肌细胞PDHa活性有轻度进一步降低({sl P}>0.05)，而CPT-1活性却较快恢复。 (2)缺氧时，正常和肥大心肌细胞葡萄糖氧化代谢率均有降低[分别下降(16±0.9)%、(48±1.1)%] ；复氧时，正常心肌细胞糖氧化代谢较快恢复，而肥大心肌细胞在复氧早期，糖氧化率进一步降低，此后才逐渐恢复。 (3)在缺氧时，肥大心肌细胞糖酵解率仅轻度下降，但在复氧后糖酵解率迅速升高，呈爆发样达峰值后又逐渐恢复到缺氧前水平。 (4)肥大心肌细胞在缺氧后脂肪酸代谢明显降低，但复氧后脂肪酸代谢呈爆发式上升，并大大高于缺血前的代谢水平。 (5)缺氧时肥大心肌细胞凋亡率即显著增加，在复氧早期细胞凋亡率继续大幅度上升，此后逐渐减少。 (6)预先用二氯乙酸处理肥大心肌细胞，可显著逆转缺氧复氧导致的细胞糖氧化受抑、糖酵解和脂肪酸代谢活化，同时，抑制细胞凋亡的发生。上述结果提示，缺氧复氧后的肥大心肌细胞能量代谢途径转换是导致细胞凋亡的重要原因。

关键词： 心肌肥大; 细胞凋亡; 缺氧复氧; 能量代谢

Relationship between apoptosis and alteration of the energetic metabolism pathways of hypertrophic cardiomyocytes induced by hypoxia--reoxygenation

Feng Bing, Liu Wei, Xu Jing, He Zuoyun, Yang Huibiao

Department of Kidney Disease, Xinqiao Hospital, the Third Military Medical University.Chongqing 400037

Abstract

The apoptosis of cardiomyocytes plays a pivotal role in the pathogenesis of cardiac failure transformed from cardiac hypertrophy, so that suppression of cardiomyocytes apoptosis is an effective pharmacotherapeutic target to prevent cardiac failure. This study focused on the relationship between apoptosis and alteration of the energetic metabolism pathways of hypertrophic cardiomyocytes induced by hypoxia-reoxygenation. Cardiomyocyte hypertrophy was induced by angiotensin II (0.1 #mu#mol/L ) and norepinephrine (1 #mu#mol/L), and the cells were cultured under the condition of hypoxia ( 95% N_(2) and 5% CO_(2), the O_(2) partial pressure was regulated at least lower than 5 mmHg ) for 8 h, then were recovered to normal culture environment. Apoptosis was detected with TUNEL. The activity of pyruvate dehydrogenase (PDH) and carnitine palmitoyltransferase 1 (CPT-1), the rate of glycose oxidation and glycolysis, and fatty acid metabolism were detected by liquid scintillation counting. The results are as follows: (1) The activity of active PDH (PDHa) was slightly higher in hypertrophic cardiomyocytes than that in normal cardiomyocytes, but the activity of CPT-1 was significantly lower in hypertrophic cardiomyoctes than that in normal cardiomyocytes.Compared with the hypertrophic cardiomyocytes cultured with normal oxygen concentration, the activities of PDHa and CPT-1 were decreased significantly after hypoxia for 8 h, and the activity of PDHa were decreased further after reoxygenation for 4 h, but the activity of CPT-1 recovered quickly after reoxygenation. (2) The rate of glucose oxidation in hypertrophic cardiomyocytes increased slightly when cultured under normal O_(2) partial pressure than that in normal cardiac cells. The rate of glucose oxidation reduced (16±0.9)% and (48±1.1)% in normal and hypertrophic cardiomyocytes, respectively, after hypoxia. It reduced further in hypertrophic cardiac cells at 4 h of reoxygenation, then recovered gradually. In normal cardiocytes, it recovered quickly after reoxygenation. (3) The rate of glycolysis of hypertrophic cardiocytes increased slightly than that of the normal cardiocytes when cultured in the general O_(2) environment. Compared with the normal cardiomyocytes, the rate of glycolysis of hypertrophic cardiac cells was the same during hypoxia-reoxygenation culture, i.e., the rate of glycolysis decreased slightly after hypoxia for 8 h, but increased rapidly and significantly after reoxygenation. (4) The rate of fatty acid oxidation was slightly lower in hypertrophic cardiocytes than that in normal cardiomyocytes. After hypoxia for 8 h, the rate of fatty acid oxidation decreased significantly in normal and hypertrophic cardiomyocytes, there was no difference between normal and hypertrophic cardiomyocytes. But the alterations of fatty acid oxidation after reoxygenation were different between normal and hypertrophic cardiac cells, namely, the fatty acid oxidation of normal cardiomyocytes were activated slowly and slightly, while the rate of fatty acid oxidation of hypertrophic cardiomyocytes increased markedly at the early stage of reoxygenation, and increased further at 8 h of reoxygenation. (5) The rate of apoptosis in hypertrophic cardiocytes increased obviously after hypoxia for 8 h, and increased further and markedly at the early stage of reoxygenation, then gradually decreased to normal level. (6) Dicholoroacetate could inhibit apoptosis of hypertrophic cardiocytes through increasing glucose oxidation and inhibiting the activation of glycolysis and fatty acid oxidation of hypertrophic cardiomyocytes induced by hypoxia-reoxygenation. These data demonstrate that apoptosis in hypertrophic cardiomyocytes after hypoxia-reoxygenation is mainly due to the inhibition of glucose oxidation and the activation of glucolysis and fatty acid oxidation. Furthermore, increasing glucose oxidation may be a new pharmacotherapeutic target to inhibit apoptosis of hypertrophic cardiac cells.

Key words: cardiac hypertrophy;Apoptosis;hypoxia-reoxygenation;Energy metabolism

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引用本文：

冯兵, 刘伟, 徐静, 何作云, 杨惠标. 缺氧复氧时肥大心肌细胞凋亡及其与能量代谢途径转换的关系[J]. 生理学报 2005; 57 (5): .

Feng Bing, Liu Wei, Xu Jing, He Zuoyun, Yang Huibiao. Relationship between apoptosis and alteration of the energetic metabolism pathways of hypertrophic cardiomyocytes induced by hypoxia--reoxygenation . Acta Physiol Sin 2005; 57 (5): (in Chinese with English abstract).