Cellular and plasma lipid levels are tightly controlled by complex gene regulatory mechanisms. Elevated plasma lipid content, or hyperlipidemia, is a significant risk factor for cardiovascular morbidity and mortality. MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression and have emerged as important modulators of lipid homeostasis, but the extent of their role has not been systematically investigated. In this study, we performed high-throughput small RNA sequencing and detected approximately 150 miRNAs in mouse liver. We then employed an unbiased, in silico strategy to identify miRNA regulatory hubs in lipid metabolism, and miR-27b was identified as the strongest such hub in human and mouse liver. In addition, hepatic miR-27b levels were determined to be sensitive to plasma hyperlipidemia, as evidenced by its ~3-fold up-regulation in the liver of mice on a high-fat diet (42% calories from fat). Further, we showed in a human hepatocyte cell line (Huh7) that miR-27b regulates the expression (mRNA and protein) of several key lipid-metabolism genes, including Angptl3 and Gpam. Finally, we demonstrated that hepatic miR-27b and its target genes are inversely altered in a mouse model of dyslipidemia and atherosclerosis. Conclusion miR-27b is responsive to lipid levels, and controls multiple genes critical to dyslipidemia.

INTRODUCTION Red blood cell (RBC) hemolysis represents an intrinsic mechanism for human vascular disease. Intravascular hemolysis releases hemoglobin and other metabolites that inhibit nitric oxide signaling and drive oxidative and inflammatory stress. While these pathways are important in disease pathogenesis, genetic and population modifiers of hemolysis including sex have not been established. MATERIALS AND METHODS We studied sex differences in storage or stress-induced hemolysis in RBC units from the US and Canada, in 22 inbred mouse strains, and in sickle cell disease using measures of hemolysis in 315 homozygous SS sickle cell patients from the Walk-PHASST cohort. We used a mouse model to evaluate post-transfusion recovery of stored RBCs, and gonadectomy to determine mechanisms related to sex hormones. RESULTS An analysis of predisposition to hemolysis based on sex revealed that male RBCs consistently exhibit increased susceptibility to hemolysis than females in response to routine cold storage, under osmotic or oxidative stress, after transfusion in mice, and in sickle cell disease. The sex difference is intrinsic to the erythrocyte and not mediated by plasmatic factors or female sex hormones. Importantly, orchiectomy in mice improves RBC storage stability and post-transfusion recovery, whereas testosterone repletion therapy exacerbates hemolytic response to osmotic or oxidative stress. DISCUSSION Our findings suggest that testosterone increases susceptibility to hemolysis across human diseases, suggesting that male sex may modulate clinical outcomes in blood storage and sickle cell disease, and establishing a role for donor genetic variables in the viability of stored erythrocytes and in human hemolytic diseases.

Apolipoprotein mimetic peptides are short synthetic peptides that share structural, as well as biological features of native apolipoproteins. The early positive clinical trials of intravaenous preparations of apoA-I, the main protein component of high density lipoproteins (HDL), have stimulated great interest in the use of apolipoprotein mimetic peptides as possible therapeutic agents. Currently, there are a wide variety of apolipoprotein mimetic peptides at various stages of drug development. These peptides typically have been designed to either promote cholesterol efflux or act as anti-oxidants, but they usually exert other biological effects, such as anti-inflammatory and anti-thrombotic effects. Uncertainty about which of these biological properties is the most important for explaining their anti-atherogenic effect is a major unresolved question in the field. Structure-function studies relating the in vitro properties of these peptides to their ability to reduce atherosclerosis in animal models may uncover the best rationale for the design of these peptides and may lead to a better understanding of the mechanisms behind the atheroprotective effect of HDL.