Peer Review #2 of "Vitamin C supplementation reduces expression of circulating miR-451a in subjects with poorly controlled type 2 diabetes mellitus and high oxidative stress (v0.2)"

Abstract

Background: Vitamin C is an essential element required for normal metabolic function. We investigated the effect of vitamin C supplementation on circulating miRNA (miR) expression in subjects with poorly controlled type 2 diabetes mellitus (T2DM). Changes in miR expression were also correlated with clinical measures of disease. Methods: Pre-and post-vitamin C supplementation samples from five participants who had increased vitamin C levels, improved oxidative status and polymorphonuclear (PMN) function after receiving 1,000 mg of vitamin C daily for six weeks were screened for miRNA expression using the NanoString miRNA assay. Differences in miRNA expression identified from the miRNA screen were validated by qRT-PCR. Results: Four miRNAs showed significantly different expression post-vitamin C supplementation relative to baseline, including the downregulation of miR-451a (-1.72 fold change (FC), p=0.036) and up-regulation of miR-1253 (0.62 FC, p=0.027), miR-1290 (0.53 FC, p=0.036) and miR-644a (0.5 FC, p=0.042). The validation study showed only miR-451a expression was significantly different from baseline with vitamin C supplementation. MiR-451a expression was negatively correlated with vitamin C levels (r=-0.497, p=0.049) but positively correlated with levels of malondialdehyde (MDA) (r=0.584, p=0.017), cholesterol (r=0.564, p=0.022) and lowdensity lipoproteins (LDL) (r=0.522, p=0.037). Bioinformatics analysis of the putative target genes of miR-451a indicated gene functions related to signaling pathways involved in cellular processes, such as the mammalian target of rapamycin (mTOR) signaling