抑制HIF-1α表达对糖尿病小鼠RhoA/ROCK信号转导通路的影响

Effect of inhibiting HIF-1αexpression on RhoA/ROCK signal transduction pathway in diabetic mice

目的探讨抑制缺氧诱导因子1(HIF-1)α表达对糖尿病小鼠RhoA/Rho相关卷曲螺旋形成的蛋白激酶(ROCK)信号转导通路的影响。方法糖尿病小鼠33只随机数字表法分为3组-模型组、NC组与抑制组,每组各11只小鼠。NC组与抑制组在玻璃体腔分别注射含siRNA NC序列、HIF-1α序列和转染试剂的混合液,模型组注射等剂量的磷酸盐缓冲液。建模后12周,测定3组小鼠的血糖水平、体重、心脏重量、心脏重量/体重比;采用黄嘌呤氧化法测超氧化物歧化酶(SOD)含量,采用硫代巴比妥酸法测定丙二醛(MDA)含量;用Image-Pro图像分析软件计算RhoA、ROCK蛋白的相对表达量。结果建模后12周,抑制组小鼠的血糖水平、体重、心脏重量、心脏重量/体重比、SOD含量、MDA含量、RhoA、ROCK蛋白相对表达量分别为(16.83±1.22)mmol/L、(30.11±2.13)g、(0.09±0.02)g、(2.99±0.12)mg/g、(294.29±12.40)U/mgprot、(1.30±0.22)nmol/mgprot、(0.77±0.09)、(0.87±0.10)。NC组分别为(16.29±1.44)mmol/L、(23.98±1.11)g、(0.12±0.01)g、(5.00±0.14)mg/g、(176.88±14.85)U/mgprot、(2.70±0.42)nmol/mgprot、(2.39±0.15)、(2.48±0.17)。模型组分别为(8.22±0.33)mmol/L、(23.10±0.32)g、(0.12±0.02)g、(5.19±0.26)mg/g、(178.09±16.02)U/mgprot、(2.68±0.87)nmol/mgprot、(2.37±0.16)、(2.45±0.22)。抑制组小鼠的血糖水平、心脏重量、心脏重量/体重比、MDA含量、RhoA、ROCK蛋白相对表达水平低于NC组与模型组,差异均有统计学意义(P<0.05)。抑制组小鼠体重、血清SOD含量高于NC组与模型组,差异均有统计学意义(P<0.05)。NC组与模型组各指标比较,差异无统计学意义(P>0.05)。结论抑制HIF-1α表达在糖尿病小鼠的应用能抑制RhoA/ROCK信号转导通路的激活,降低血清MDA表达,促进SOD的释放,从而降低糖尿病小鼠的血糖水平与心脏重量、心脏重量/体重比,促进恢复正常体重。

Objective To investigate the inhibition of hypoxia inducible factor 1(HIF-1)alpha expression on RhoA/Rho-associated coiled-coil forming protein serine/thmonine kinase(ROCK)signal transduction pathway in diabetic mice.Methods Thirty-three cases of diabetic mice were divided into three groups-model group,NC group and inhibition group withe 11 mice in each group by random number table.The NC group and the inhibition group were injected with mixture of siRNA NC sequence,HIF-1α sequence and transfection reagent in the vitreous cavity,and the model group were injected with equal dose of phosphate buffer.Twelve weeks after modeling,the blood glucose level,body weight,heart weight,and heart weight/body weight ratio of the three groups of mice were measured and compared;the content of superoxide dismutase(SOD)was measured using the xanthine oxidation method,malondialdehyde(MDA)content was determined using the thiobarbituric acid method;the relative expression of RhoA and ROCK protein was calculated using Image-Pro image analysis software.Results After 12 weeks of modeling,the blood glucose level,body weight,heart weight,heart weight/body weight ratio,SOD content,MDA content,the relative expression levels of RhoA in the inhibition group mice were(16.83±1.22)mmol/L,(30.11±2.13)g,(0.09±0.02)g,(2.99±0.12)mg/g,(294.29±12.40)U/mgprot,(1.30±0.22)nmol/mgprot,(0.77±0.09),(0.87±0.10)respectively.The NC groups were(16.29±1.44)mmol/L,(23.98±1.11)g,(0.12±0.01)g,(5.00±0.14)mg/g,(176.88±14.85)U/mgprot,(2.70±0.42)nmol/mgprot,(2.39±0.15),(2.48±0.17)respectively.The model groups were(8.22±0.33)mmol/L,(23.10±0.32)g,(0.12±0.02)g,(5.19±0.26)mg/g,(178.09±16.02)U/mgprot,(2.68±0.87)nmol/mgprot,(2.37±0.16),(2.45±0.22)respectively.The level of blood glucose,heart weight,heart weight/body weight ratio,the MDA content,the relative expression levels of RhoA and ROCK protein the suppression group were lower than those in the NC group and the model group,the differences were statistically significant(P<0.05).After 12 weeks of modeling,the body weight and serum SOD content of the suppression group were higher than those of the NC group and the model group,the differences were statistically significant(P<0.05).There were no statistically significant differences in the indexes between the NC group and the model group(P>0.05).Conclusion The application of inhibiting the expression of HIF-1α in diabetic mice can inhibit the activation of RhoA/ROCK signal transduction pathway,reduce the expression of serum MDA,and promote the release of SOD,thereby reduce the blood glucose level,heart weight,and heart weight/body weight ratio of diabetic mice,promote the return to normal weight.