Cholesterol homeostasis regulation by miR-223: basic science mechanisms and translational implications.

C holesterol homeostasis is a tightly regulated process. Previous work has been dominated by sterol-response element—related mechanisms, but the influence of micro-RNAs (miRs) is increasingly recognized, with miR-223 becoming a recent member of this group. Several micro-RNAs (miRNAs or miR-) have been reported to be involved in cholesterol homeostasis. 1–4 In a recent study, Vickers et al 5 presented miR-223 as an important new player in this arena, one that they propose to be a coordinator of cholesterol homeostasis. 5 This miRNA had been previously found to be rich in myeloid cells, where it regulates differentiation and inflammation, 6,7 but the authors found that its abundance in hepatic cells is at least comparable with that of many functionally validated miRNAs. A relationship between miR-223 and cholesterol metabolism was suggested by finding its expression in human hepatoma (Huh7) cells was positively associated with intracellular cholesterol levels. In a series of studies in Huh7 cells, it was then shown that miR-223 (1) inhibited high-density lipoprotein cholesterol (HDL-C) uptake, (2) promoted cholesterol efflux, and (3) suppressed cholesterol biosynthesis. The authors also showed increased cholesterol levels in liver and plasma of miR-223 knockout mice, extending its involvement in cholesterol homeostasis in vitro to the in vivo setting. The experimental approaches taken are well established in the miRNA field and were coupled to a productive use of bioinformatics. This included investigating whether the effects of miR-223 on a particular RNA were direct, by cloning into a luciferase reporter plasmid the putative target sequence in the 3ʹ-UTR of that mRNA (or this sequence deleted in nucleotides required for miR-223 binding). The effect of exogenously supplied miR-223 on luciferase activity in transfected HEK293 cells was then measured. Thus, they determined that the molecular mechanisms for the functional reductions in HDL-C uptake and cholesterol biosynthesis after miR-223 treatment included its direct effect on the mRNAs encoding SR-BI (SCARB1) for the former process and HMG CoA synthase 1 (HMGCS1) and methylsterol monooxygenase 1 (SC4MOL) for the latter. In contrast to HDL-C uptake and cholesterol synthesis, the promotion of cholesterol efflux from Huh7 cells to apoA1, which is mediated by ABCA1, was shown to be indirect. Transcription factor analyses led them to consider Sp1 and Sp3 as the effectors; Sp1 is a known transcriptional activator of ABCA1 and Sp3 is an antagonist of Sp1. Both mRNAs harbor putative miR-223 target sites, but only Sp3 mRNA levels were reduced by miR-223 …