白藜芦醇对脓毒症大鼠心肌能量代谢的影响

Effect of resveratrol on myocardial energy metabolism in sepsis rats

目的探讨白藜芦醇对脓毒症大鼠心肌能量代谢的影响及相关机制。方法将20只雄性SD大鼠按随机数字表法分为4组，每组5只。以腹腔注射脂多糖（LPS）10mg／kg$1]备脓毒症模型；假手术组（Sham组）腹腔注射等量生理盐水。白藜芦醇治疗组（RES组）于注射LPS前10min静脉注射白藜芦醇8mg／kg；白藜芦醇联合沉默信息调节因子1（sirt1）抑制剂sirtinol组（RES＋sirtinol组）于注射LPS前先静脉注射sirtinol溶液1mg／kg，5min后再静脉注射白藜芦醇8mg／kg。6h后处死大鼠取心脏，透射电镜下观察各组大鼠心肌超微结构；蛋白质免疫印迹试验（WesternBlot）检测心肌sirt1蛋白表达；紫外分光光度法检测心肌细胞色素C氧化酶（CCO）活性；高效液相色谱法检测心肌三磷酸腺苷（ATP）、二磷酸腺苷（ADP）、一磷酸腺苷（AMP）含量。结果透射电镜下显示，Sham组心肌线粒体结构正常；LPS组及RES＋sirtinol组心肌线粒体肿胀、排列紊乱，线粒体嵴减少；RES组上述症状不明显。与Sham组比较，LPS组sirt1蛋白表达下降（灰度值：0．602±0．038比0．902±0．037，p〈0．05），CCO活性降低（U／mg：0．344±0．019比0．600±0．023，P〈0．05），ATP、ADP、AMP含量下降[ATP（μg／g）：89．958±7．570比157．460±6．737，ADP（μg／g）：164．658±6．742比251．608±5．191，AMP（μg／g）：179．926±7．303比276．262±5．546，均P〈0．05]。与LPS组比较，RES组sirt1蛋白表达增加（灰度值：0．874±0．017比0．602±0．038，P〈0．05），CCO活性提高（U／mg：0．574±0．013比0．344±0．019，P〈0．05），ATP、ADP、AMP的含量升高[ATP（μg／g）：147．686±8．853比89．958±7．570，ADP（μg／g）：245．860±7．111比164．658±6．742，AMP（μg）：271．260±6．658比179．926±7．303，均P〈0．05]。与RES组比较，RES＋sirtinol组sirt1蛋白表达下降（灰度值：0．636±0．030比0．874±0．017，P〈0．05）、CCO活性减弱（U／mg：0．369±0．040比0．574±0．013，P〈0．05），ATP、ADP、AMP含量下降[ATP（μg／g）：97．942±11．257比147．686±8．853，ADP（μg）：168．888±8．965比245．860±7．111，AMP（μg／g）：175．498±6．993比271．260±6．658，均P〈0．05]。结论白藜芦醇能明显改善脓毒症大鼠心肌能量代谢，sirt1是白藜芦醇影响心肌能量代谢的重要调节因子。

Objective To investigate the influence of resveratrol （RES） on myocardial energy metabolism in sepsis rats and its mechanism. Methods Twenty Sprague-Dawley （SD） rats were divided into four groups according to the random number table, 5 rats in each group. The sepsis model was reproduced by intraperitoneal injection of lipopolysaccharide （LPS） 10 mg/kg, and the rats in sham group were injected intraperitoneally with the same amount of saline. The rats in RES treatment group （RES group ） were injected RES 8 mg/kg 10 minutes before LPS injection, and those in RES combined with the silent information regulator 1 （sirtl） inhibitor group （RES ＋ sirtinol group ） were injected sirtinol 1 mg/kg followed by RES 8 mg/kg 10 minutes before LPS injection. The rats were sacrificed 6 hours later, and the heart was harvested. The myocardial uhrastructure was observed by transmission electron microscope. The expression of sirtl protein was detected by Western Blot. The cytochrome C oxidase （CCO） activity was determined by ultraviolet spectrophotometry. The content of adenosine triphosphate （ATP）, adenosine diphosphate （ADP）, and adenosine monophosphate （AMP） were detected by high-performance liquid chromatography （HPLC）. Results Under transmission electron microscopy, the mitoehrondria in myocardium in sham group was normal, and mitochrondria was swollen and derangement in LPS group and RES ＋ sirtinol group, but it was not significant in RES group. Compared with sham group, the sirtl protein expression in LPS group was decreased （gray value： 0.602± 0.038 vs. 0.902±0.037, P 〈 0.05 ）, the CCO activity was declined （U/mg： 0.344± 0.019 vs. 0.600±0.023, P 〈 0.05 ）, the contents of ATP, ADP, and AMP were decreased [ ATP （μg/g）： 89.958 ± 7.570 vs. 157.460± 6.737,ADP （μg/g）： 164.658± 6.742 vs. 251.608 ± 5.191, AMP （μg/g）： 179.926 ± 7.303 vs. 276.262 ± 5.546, all P 〈 0.05 ]. Compared with LPS group, the sirtl protein expression in RES group was increased （gray value： 0.874±0.017 vs. 0.602±0.038, P 〈 0.05）, the CCO activity was increased （U/nag： 0.574±0.013 vs. 0.344±0.019, P 〈 0.05）, the contents of ATP, ADP, and AMP were increased [ ATP （μ/g）： 147.686±8.853 vs. 89.958 ±7.570, ADP （μg/g）： 245.860±7.111 vs. 164.658±6.742, AMP （μg/g： 271.260±6.658 vs. 179.926±7.303, all P 〈 0.05 ]. Compared with RES group, the sirtl protein expression in RES ＋ sirtinol group was decreased （gray value： 0.636±0.030 vs. 0.874±0.017, P 〈 0.05）, the CCO activity was declined （U/mg： 0.369±0.040 vs. 0.574±0.013, P 〈 0.05）, the contents of ATP, ADP, and AMP were decreased [ ATP （μg/g）： 97.942± 11.257 vs. 147.686±8.853, ADP （μg/g）： 168.888±8.965 vs. 245.860±7.111, AMP （μg/g）： 175.498±6.993 vs. 271.260±6.658, all P 〈 0.05]. Conclusion RES could improve myocardial energy metabolism in sepsis rats, in which sirtl is an important factor.