二十碳五烯酸对糖尿病大鼠冠状动脉的舒张作用及其机制

The relaxant effects and mechanisms of eicosapentaenoic acid on diabetic rat coronary artery

目的探讨不同浓度二十碳五烯酸(EPA)对糖尿病大鼠冠状动脉的舒张作用及其机制。方法 40只SD大鼠随机分为正常组和糖尿病组,分离冠状动脉。采用微血管张力测定技术评估不同浓度(0.1、1、3、10、30、100μmol/L)EPA对正常和糖尿病大鼠冠状动脉的舒张作用并观察孵育(100nmol/L)大电导钙激活钾通道(BK通道)特异性阻滞剂伊比蝎毒素(IBTX)后血管张力的变化;酶消化法分离正常和糖尿病大鼠冠状动脉平滑肌细胞并采用单通道膜片钳技术记录灌流不同浓度(0、10、30、100、300、1 000pmol/L)EPA时BK通道开放概率的变化;采用蛋白印迹技术测定正常和糖尿病大鼠冠状动脉BK通道亚单位蛋白表达。结果 EPA对正常组和糖尿病组大鼠冠状动脉舒张的比例随浓度升高而增加,孵育IBTX后舒张作用明显减弱;EPA对正常组和糖尿病组大鼠冠状动脉平滑肌细胞BK通道的激活随浓度升高而增强,且与正常组比较,EPA对糖尿病组大鼠冠状动脉的舒张作用及BK通道的激活作用较弱。两组大鼠冠状动脉BK通道蛋白α亚单位表达无差异,但糖尿病大鼠冠状动脉BK通道蛋白β1亚单位表达明显低于正常组(0.424 7±0.036 2vs 0.892 3±0.053 5,n=6,P<0.05)。结论 EPA对糖尿病冠状动脉具有舒张作用,其机制与激活BK通道有关,糖尿病时BK通道β1亚单位表达减少,EPA对BK通道的激活作用减弱。

Objective To investigate the relaxant effects and mechanisms of eicosapentaenoic （EPA） with different concentrations on diabetic rat coronary arterial rings. Methods All 40 male Sprague Dawley rats were randomly assigned to two groups： blank control group and diabetic group, coronary arteries were dissected.Coronary artery di- astolic function of normal and diabetic rats was evaluated by vascular tension measurement at different concentra- tions of EPA（0,10,30,100,300,1 000 pmol/L） with and without pre-incubation of large-conductance calcium-acti- vated potassium （BK） channel inhibitor IBTX; Coronary smooth muscle cells were isolated by enzyme digestion, and changes of I±K channel open probability at different concentrations of EPA（0,10,30,100,300,1 000 pmol/L） in coronary smooth muscle cells were recorded by patch clamp in single channel configuration. The protein expression of BK channels was determined by western blot analysis. Results EPA relaxed coronary arterial rings of both con- trol and diabetic groups in a concentration-dependent manner, which was significantly inhibited by pre-incubation with IBTX. EPA activated BK channels in coronary artery smooth muscle cells of both control and diabetic groups in a concentration-dependent manner.Compared with con- trol group, the relaxation and activation effects of EPA on coronary arteries and BK channels in diabetic group were weaker. There was no statistical difference in the protein expressions of BK-a subunit in the control and diabetic groups（0.424 7±0.036 2 vs 0.892 3±0.053 5,n = 6, P〈 0.05 ）. Compared with control group, BK-β1 subunit in diabetic group was significantly reduced. Conclusion EPA possesses a relaxant effect on isolated rat coronary arteries in a concentration-dependent man- ner. The mechanism involves activation of BK channels. The relaxant effects of EPA on diabetic rat coronary ar- teries are decreased due to down-regulation of BK- β1 subunit.