A novel non-genetic murine model of hyperglycemia and hyperlipidemia-associated accelerated atherosclerosis

Objective Atherosclerosis, the main pathology underlying cardiovascular diseases is accelerated in diabetic patients. Genetic mouse models require breeding efforts which are time-consuming and costly. To establish a new non-genetic model of inducible metabolic risk factors mimicking hyperlipidemia, hyperglycemia, or both and allowing to detect phenotypic differences dependent on the metabolic stressor(s). Methods and Results Wild type mice were injected with gain-of-function PCSK9D377Y (proprotein convertase subtilisin/kexin type 9) mutant adeno-associated viral particles (AAV) and streptozotocin and fed either a high-fat diet (HFD) or high-cholesterol/high fat-diet (Paigen diet, PD). Combined hyperlipidemic and hyperglycemic (HGHCi) mice, but not hyperlipidemia (HCi) alone, display characteristic features of accelerated atherosclerosis. Atherosclerotic plaques of HGHCi animals were larger, showed a less stable phenotype, contained more macrophages and less smooth muscle cells. These findings were observed both at early (12 weeks) and late (20 weeks) time points on both HFD or PD diet. Differences between the HGHCi and HCi model were confirmed using RNAseq analysis revealing that significantly more genes are dysregulated in mice with combined hyperlipidemia and hyperglycemia as compared to the hyperlipidemia only group. The HGHCi-associated genes were related to pathways regulating inflammation, cellular metabolism and collagen degradation. PD accelerates atherosclerosis in mice and shows plaque formation already after 8 weeks, therefore, representing a fast direct inducible hyperglycemic atherosclerosis model. Conclusion We established a non-genetic inducible mouse model allowing comparative analyses of atherosclerosis in HCi and HGHCi conditions and its modification by diet, allowing analyses of multiple metabolic hits in mice. ### Competing Interest Statement The authors have declared no competing interest. * ApoE : apolipoprotein E BSA : bovine serum albumin BCA : bicinchoninic acid DM : diabetes mellitus DAPI : 4’,6-Diamidino-2-Phenylindole, Dihydrochloride HFD : high fat diet HRP : horseradish peroxidase LDLR : low-density lipoprotein receptor NOD : non-obese diabetic OCT : optimal cutting temperature PD : paigen diet PCSK9 : proprotein convertase subtilisin/kexin type 9 PBS : phosphate buffer saline RIPA : radioimmunoprecipitation assay RNAseq : RNA sequencing rAAV : recombinant adeno-associated virus STZ : streptozotocin α-SMA : α smooth muscle actin