α-硫辛酸对高盐诱导的大鼠高血压反应过程的抑制作用

The inhibitory effect ofα-lipoic acid on the high salt-induced hypertension rats

目的研究α硫辛酸能否抑制头端延髓腹外侧区(RVLM)线粒体中活性氧(ROS)的过量产生来缓解高盐诱导的高血压反应。方法将雄性大鼠32只分配到两组:分别给予正常盐饮食(0.3%NaCl)8周(n=16)和高盐饮食(8%NaCl)8周(n=16,诱导高血压模型)。之后将正常盐饮食大鼠及高盐饮食大鼠各自再分别分为两组,每两组分别给予溶于0.9%生理盐水的α硫辛酸(60mg/kg)或单独给予生理盐水灌胃饲养9周。由此共4组,即:实验组(n=8,高血压模型样本喂养α硫辛酸)、模型组(n=8,高血压模型样本喂养生理盐水)、对照组(n=8,正常盐饮食样本喂养α硫辛酸)和空白对照组(n=8,正常盐饮食样本喂养生理盐水)。监测各组样本动脉压的变化并在实验结束时通过免疫荧光检测超氧化物含量表达情况,Western blot检测头端延髓腹外侧区内还原型烟酰胺腺嘌呤二核苷酸磷酸[NAD(P)H]氧化酶2(NOX2)、NOX4及铜/锌超氧化物歧化酶(Cu/Zn-SOD)的表达情况;用ELISA测定RVLM中线粒体丙二醛(MDA)等氧化应激相关物质的表达差异。结果实验组平均动脉压(MAP)低于模型组差异有统计学意义(P<0.05);实验组与模型组二氢乙啶(DHE)荧光强度分别为60.2±3.1、99.1±3.8;Cu/Zn-SOD阳性神经元的数量分别为(20.8±1.1)、(6.9±1.2)个;NOX2阳性神经元数量分别为(12.3±3.5)、(25.1±5.4)个;NOX4阳性神经元数量分别为(10.1±2.2)、(13.3±4.1)个,差异均有统计学意义(P<0.05)。Western blot显示实验组与模型组NOX2水平分别为78.9±2.0、112.7±3.8;NOX4水平分别为63.2±2.1、99.4±1.7;实验组与模型组RVLM中Cu/Zn-SOD水平分别为19.7±1.6、10.3±1.2,差异均有统计学意义(P<0.05);线粒体超氧化物歧化酶(SOD)水平分别为(33.1±3.8)、(15.2±1.7)U/mg,差异均有统计学意义(P<0.05)。实验组与模型组线粒体谷胱甘肽(GSH)水平分别为(5.2±0.9)、(2.3±0.5)μmol/g;血清去甲肾上腺素(NE)水平分别为(325.8±7.3)、(467.9±6.1)pg/mL,差异均有统计学意义(P<0.05)。结论α硫辛酸可通过降低NOX2和NOX4的表达、线粒体酶生物能量及提高高血压发展中的RVLM在细胞内的抗氧化能力来抑制高血压发展过程的氧化应激反应。

Objective To investigate whether the excessive production of reactive oxygen species （ROS） in the mitochondria of the rostral ventrolateral medulla （RVLM） can inhibit the high salt-induced hypertension response. Methods A total of 32 male rats were divided into two groups： two groups were given normal salt diet （0.3% NaC1） for 8 weeks （n=16） and high salt diet （8% NaC1） for 8 weeks （n=16 , induced hypertension model） respectively. The two groups were divided into four groups,two groups were given α-li- poic dissolving in 0. 9% normal saline （60 mg/kg）, two groups were fed with saline for 9 weeks. There were four groups：the experimental group （n= 8,the hypertension model sample fed α-lipoic acid） ,the model group （n= 8, the hypertension model sample fed saline）, the control group （n= 8, normal salt diet sample fed α-lipoic acid） and the blank control group （n=8,normal salt diet sample fed saline）. Monitored the change of the arterial pressure and detected the expression of superoxide by immunofluorescence at the end of the experiment, measured the expression of NAD（P）H NOX2 ,NOX4 and Cu/Zn-SOD in RVLM by Western blot; determined the expression differences of oxidative stress related substances such as mitochondrial malondialdehyde（MDA） in RVLM by ELISA. Results The mean arterial pressure （MAP） in the experimental group was lower than that in the model group, the difference was statistically significant （P〈0.05）; in the experimental group and the model group the intensities of fluorescent-labled dihydroethidium（DHE） were 60. 2 ± 3.1,99.1± 3.8 ; the numbers of positive neurons in Cu/Zn-SOD were 20.8±1.1,6.9±1. 2;the numbers of NOX2 positive neurons were 12.3 ±3.5,25.1 ±5.4 ; the numbers of NOX4 positive neurons were 10.1±2.2,13.3 ± 4.1, the difference was statistically significant （P〈0.05）. Western blot showed that the NOX2 levels of the experimental group and the model group were 78.9±2.0,112.7±3.8,the levels of NOX4 were 63.2±2.1,99.4±1.7. The levels of Cu/Zn-SOD in RVLM of the experimental group and the model group were 19.7 4±1.6,10.3± 1.2, the difference was statistically significant （P〈0.05） ; the levels of mitochondrial superoxide dismutase （SOD） were （33.1 ± 3.8）, （15.2 ±1.7） U/mg, the difference was statistically significant （P〈0.05）. The levels of mitochondrial glutathione （GSH） in the experimental group and the model group were （5.2±0.9）, （2.3±0.5） μmot/g;the levels of norepinephrine （NE） were （325.8±7.3）, （467.9±6.1） pg/mL,the difference was sta- tistically significant （P〈0. 05）. Conclusion α-lipoic acid could decrease the expression of NOX2,NOX4 and the bioenergy of mitochondria enzyme, and increase the intracellular antioxidant ability in the RVLM during the development of hypertension to inhibit the oxidative stress response in the development of hypertension.