The effect of homocysteine on Human Aortic Endothelial Cells [RNA]

Hyperhomocysteinemia (HHcy) is an established and potent independent risk factor for degenerative diseases, including cardiovascular disease (CVD), Alzheimer disease, type II diabetes mellitus, and chronic kidney disease. HHcy has been shown to inhibit proliferation and promote inflammatory responses in endothelial cells (EC), and impair endothelial function, a hallmark for vascular injury. However, metabolic processes and molecular mechanisms mediating HHcy-induced endothelial injury remains to be elucidated. This study examined the effects of HHcy on the expression of mRNA and microRNA (miRNA) in human aortic endothelial cells treated with a pathophysiologically relevant concentration of homocysteine (Hcy 500 μM). This is the first study to consider the effects of HHcy on both global mRNA and miRNA expression changes for mechanism identification. Molecular axes and biochemical processes identified in this study are useful not only for the understanding of mechanisms underlying HHcy-induced endothelial injury, but also for discovering therapeutic targets for CVD in general. RNA was extracted from three biological replicates for each treatment group using the miRNeasy Mini Kit (Qiagen, Valencia, CA). RNA quality was assessed using the Nanodrop 2000 Spectrophotometer (Thermo Scientific). mRNA microarrays were performed using Affymetrix GeneChip® Human Gene 1.0 ST Arrays and miRNA microarrays were performed using Affymetrix GeneChip® miRNA 1.0 Arrays (Affymetrix, Santa Clara, CA).