2型糖尿病BTBR ob/ob小鼠肝损伤的脂质代谢组学变化

Lipid metabolomic changes in liver injury in BTBR ob/ob mice with type 2 diabetes mellitus

目的采用生物化学结合代谢组学的方法,探讨2型糖尿病BTBR ob/ob小鼠肝损伤相关脂类生物标志物及脂质代谢通路变化。方法选择7周龄BTBR野生型(WT)小鼠16只(WT组),BTBR ob/ob肥胖小鼠14只(ob/ob组),在SPF环境下喂养至20周龄。比较两组小鼠肝损伤的情况:高分辨率呼吸仪检测肝组织线粒体呼吸酶复合体活性,并使用超高效液相-四极杆飞行时间质谱对两组小鼠肝脏组织样本进行脂质代谢组学分析,主要检测内源性代谢产物,采用主成分分析(PCA)、正交偏最小二乘法判别分析(OPLS-DA)等模型筛选潜在的生物标志物,并通过MetaboAnalyst 5.0对已鉴定的代谢产物进行代谢通路分析。结果与WT组小鼠相比,ob/ob组小鼠体质量和空腹血糖均明显升高,血清天冬氨酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT)、低密度脂蛋白(LDL-C)和胆固醇(CHO)水平也明显升高(P<0.01);肝脏苏木精-伊红(HE)染色显示,ob/ob组小鼠出现肝小叶结构紊乱,肝脏细胞肿胀,细胞内可见大量脂肪空泡,且呈弥散性分布,胞质疏松化;油红O染色显示ob/ob组小鼠肝细胞中有大量脂质沉积;高分辨率呼吸仪检测结果显示ob/ob组小鼠出现线粒体氧化磷酸化障碍,复合体Ⅳ活性下降;脂质代谢组学分析结果显示ob/ob组小鼠脂质代谢谱发生改变,涉及的代谢通路主要有甘油磷脂代谢、糖基磷脂酰肌醇(GPI)锚定生物合成、甘油酯代谢、亚油酸代谢、α-亚麻酸代谢、花生四烯酸代谢。结论ob/ob组小鼠出现肝损伤表现,其机制可能与脂质代谢紊乱相关,其中甘油磷脂代谢紊乱是最为关键的一条代谢通路。

Objective To investigate the changes of lipid biomarkers and lipid metabolic pathways related to liver injury in BTBR ob/ob mice with type 2 diabetes mellitus by biochemical and metabolomics methods.Methods 16 BTBR wild-type(WT)mice(WT group)and 14 BTBR ob/ob obese mice(ob/ob group)at 7 weeks of age were selected and fed in SPF environment until 20 weeks of age.Liver injury was compared between the two groups:The activities of mitochondrial respiratory enzyme complex in liver tissue were detected by high-resolution respirators,and the lipid metabolomic analysis of liver tissue samples in the two groups of mice was performed by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry,mainly detecting endogenous metabolites.Principal component analysis(PCA),orthogonal partial least squares discriminant analysis(OPLS-DA)and other models were used to screen potential biomarkers,and the metabolic pathway analysis of the identified metabolites was performed by MetaboAnalyst 5.0.Results Compared with the WT group,the ob/ob group had significantly increased body weight,fasting blood glucose,serum levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT),low-density lipoprotein(LDL-C)and cholesterol(CHO)(P<0.01).Liver hematoxylin-eosin staining(HE)staining showed that the mice in ob/ob group had structural disorder of liver lobules,swelling of liver cells,a large number of fat vacuoles in cells,diffuse distribution and loose cytoplasm.Oil red O staining showed that there was a large amount of lipid deposition in the hepatocytes of ob/ob mice.The high resolution spirometer showed that the ob/ob mice had mitochondrial oxidative phosphorylation disorder and the activity of complexⅣdecreased.Lipid metabolomic analysis showed that the lipid metabolic profile of ob/ob mice changed,and the metabolic pathways involved mainly included glycerophospholipid metabolism,glycosylphosphatidylinositol(GPI)anchor biosynthesis,triglyceride metabolism,linoleic acid metabolism,α-linolenic acid metabolism and arachidonic acid metabolism.Conclusion The liver injury of ob/ob group mice may be related to the disorder of lipid metabolism,in which the disorder of glycerophospholipid metabolism is the most critical metabolic pathway.