低氧训练诱导miR-27/PPARγ调控肥胖大鼠肝脏脂肪酸代谢变化的研究

Study on Hypoxia Exercise Inducing miR-27/PPARγ to Regulate Fatty Acids Metabolism in Obese Rat's Liver

目的:探讨低氧训练对肥胖大鼠肝脏中miR-27/PPARγ及其下游脂肪酸代谢相关基因、蛋白表达水平的时序性影响。方法:13周龄雄性SD肥胖大鼠50只随机平均分成5组(n=10×5):低氧对照组(C组)、低氧训练1周组(E1组)、低氧训练2周组(E2组)、低氧训练3周组(E3组)和低氧训练4周组(E4组),所有训练组用水平跑台进行耐力训练,训练强度常氧25 m/min,低氧20 m/min(低氧浓度13.6%),持续运动1 h/天、5天/周,共4周。检测血清TC、TG、LDL-C和HDL-C水平;实时荧光定量PCR检测miR-27、PPARγ基因表达,Western Blot和免疫组化检测PPARγ、CD36、ATGL、LPL、L-FABP、SREBP1蛋白表达水平。结果:4周低氧训练过程中,肥胖大鼠脂体比、TC、TG和LDL-C浓度逐步降低,而HDL-C逐步升高;伴随着低氧训练时间的延长,肥胖大鼠肝脏中miR-27表达逐步降低,其中,E3和E4组极显著高于其余各组(P≤0.01);与之相反,PPARγ、CD36、ATGL、LPL的表达逐步升高,其中E4组CD36表达量显著高于C组(P≤0.05),C组AGTL表达量显著低于E2组(P≤0.05),且极显著低于E3和E4组(P≤0.01),C组LPL表达量显著低于E2和E3组(P≤0.05),而极显著低于E4组(P≤0.01);L-FABP表达量在E1组最高,显著高于C组(P≤0.05);SREBP1表达量无显著变化(P≥0.05)。结论:肥胖大鼠肝脏miR-27表达与低氧训练时间呈现负相关,低氧训练通过miR-27影响PPARγ及其下游脂肪酸代谢相关靶基因和靶蛋白的表达,改善血脂水平,最终机体脂含量伴随低氧训练时间的延长而逐渐下降。

Objective： To explore the sequential effects of hypoxic exercising on miR-27/PPARγ and lipid metabolism target gene and protein expression levels in the obesity rat’s liver. Methods： Fifty 13-week-old male diet-induced obesity rats were randomly divided into five groups（n=10 each）： Control group（C）, 1-week hypoxic exercise group（E1）, 2-week hypoxic exercise group（E2）, 3-week hypoxic exercise group（E3） and 4-week hypoxic exercise group（E4）. All the rats were trained with a treadmill for 1 h/day, 5 d/week for a total of 4 weeks, and the control group was trained in the normal oxygen environment with the speed of 25 m/min, but the hypoxic groups were trained in the 13.6% hypoxic environment. To detect the concentrations of serum total cholesterol（TC）, triglyceride（TG）, low density lipoprotein（LDL-C）, high-density lipoprotein（HDL-C） levels. Micro RNA-27（miR-27） expression levels in the liver were determined by Real-time PCR. Protein and m RNA expression levels of PPARγ 、 CD36 、 ATGL 、 LPL 、 L-FABP 、 SREBP1 were tested by western blot and immunohistochemistry in the obesity rat liver. Results： The fat/body weight ratio and the concentrations of TC, TG, LDL-C were gradually decreased in obese rats during four weeks hypoxic exercise, but the concentrations of HDL-C were gradually increased. With the prolonged hypoxic exercise time, the expression of miR-27 was gradually decreased in the liver of obese rat, E3 and E4 were significantly higher than other groups（P≤0.01）. However, the expression levels of PPARγ、CD36、ATGL、LPL were gradually increased, and the levels of CD36 in E4 group were significantly higher than C group（P≤0.05）, the levels of AGTL were significantly lower than E2 group（P≤0.05）, E3 and E4 group（P≤0.01）, the levels of LPL were significantly lower than E2 and E3 group（P≤0.05）, especially lower than E4 group（P≤0.01）. Meanwhile, the levels of L-FABP were the highest in E1 group, which were significantly higher than C group（P≤0.05）, but the expression levels of SREBP1 were no significant change. Conclusion： The expression of miR-27 in the obese rat liver was negatively correlated with the hypoxic exercise time, and hypoxic exercise could affect PPARγ expression and its downstream fatty acid metabolism related target genes and target proteins expression, improve the blood lipid level through miR-27. Finally, the lipid content was gradually decreased with the prolonged hypoxic exercise time.