COMPREHENSIVE METABOLOMIC ANALYSIS OF DIABETIC ATHEROSCLEROSIS

Abstract

Cardiovascular disease (CVD) is responsible for the majority of deaths in people with diabetes. Our lack of understanding of the biochemical mechanisms linking diabetes and CVD complicates the development of effective prevention/treatment strategies. Comprehensive metabolomics, which is the study of small molecules, offers a novel approach to this problem as it is capable of providing a global snapshot of the dynamic intracellular changes associated with a particular pathological state. This allows the identification of novel pathways and biomarkers indicative of metabolic alterations related to diabetic atherosclerosis. Diabetic atherosclerosis was studied using two different murine models of hyperglycemia. Multiple low-dose intraperitoneal streptozotocin (STZ) injections were used to induce severe hyperglycemia/insulinopenia (fasting blood glucose (FBG)=21.0±5.2mM) in ApoE-/- mice. Moderate hyperglycemia/insulinopenia (FBG=11.8±3.9mM) was achieved in ApoE-/- mice carrying a point mutation in one copy of insulin 2 gene (Ins2+/Akita). Normoglycemic ApoE-/- controls had FBG of 9.1±1.6mM. Subsets (n=10) of mice were harvested at 5, 10 and 15 weeks of age to study the progression of atherosclerosis. Atherosclerotic lesions of hyperglycemic ApoE-/- mice were significantly (p<0.05) larger compared to the controls at 15 weeks of age. Liquid chromatography coupled to mass spectrometry was used to analyze plasma, artery wall and liver tissue. Multivariate analysis showed that plasma metabolomic profiles of 5034 detected metabolite features of hyperglycemic mice were readily distinguished from each other (severe versus moderate hyperglycemia) and from the controls. Univariate analysis including p-value and correlation coefficient were used to determine metabolites that are significantly different (p<0.05) between hyperglycemic mice and controls. Diverse compound classes including amino acids and derivatives, dipeptides and several classes of bioactive lipids such as fatty acids, glycerolipids and sphingolipids were identified. The amino acid ergothioneine (antioxidant and anti-inflammatory) and ceramide phosphate (inflammation mediator) were elevated, whereas dimethylarginine (nitric oxide inhibitor) and diacylglycerol (precursor of bioactive lipids) were diminished, in the hyperglycemic models. Disturbed amino acid and lipid metabolism suggest a metabolic complexity in diabetic atherosclerosis. These clues are being used to better understand the specific mechanisms and pathways involved in this multifactorial disease. These findings will facilitate the identification of potential targets for the development of novel therapeutic strategies to slow or block atherogenesis in diabetic individuals.