microRNA-483 ameliorates hypercholesterolemia by inhibiting PCSK9 production
Jianjie Dong, … , Sotirios Tsimikas, John Y.J. Shyy
Jianjie Dong, … , Sotirios Tsimikas, John Y.J. Shyy
Published October 29, 2020
Citation Information: JCI Insight. 2020;5(23):e143812. https://doi.org/10.1172/jci.insight.143812.
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Research Article Metabolism Vascular biology

microRNA-483 ameliorates hypercholesterolemia by inhibiting PCSK9 production

Abstract

Proprotein convertase subtilisin/kexin type 9 (PCSK9) affects cholesterol homeostasis by targeting hepatic LDL receptor (LDLR) for lysosomal degradation. Clinically, PCSK9 inhibitors effectively reduce LDL-cholesterol (LDL-C) levels and the incidence of cardiovascular events. Because microRNAs (miRs) are integral regulators of cholesterol homeostasis, we investigated the involvement of miR-483 in regulating LDL-C metabolism. Using in silico analysis, we predicted that miR-483-5p targets the 3′-UTR of PCSK9 mRNA. In HepG2 cells, miR-483-5p targeted the PCSK9 3′-UTR, leading to decreased PCSK9 protein and mRNA expression, increased LDLR expression, and enhanced LDL-C uptake. In hyperlipidemic mice and humans, serum levels of total cholesterol and LDL-C were inversely correlated with miR-483-5p levels. In mice, hepatic miR-483 overexpression increased LDLR levels by targeting Pcsk9, with a significant reduction in plasma total cholesterol and LDL-C levels. Mechanistically, the cholesterol-lowering effect of miR-483-5p was significant in mice receiving AAV8 PCSK9-3′-UTR but not Ldlr-knockout mice or mice receiving AAV8 PCSK9-3′-UTR (ΔBS) with the miR-483-5p targeting site deleted. Thus, exogenously administered miR-483 or similarly optimized compounds have potential to ameliorate hypercholesterolemia.

Authors

Jianjie Dong, Ming He, Jie Li, Ariane Pessentheiner, Chen Wang, Jin Zhang, Yameng Sun, Wei-Ting Wang, Yuqing Zhang, Junhui Liu, Shen-Chih Wang, Po-Hsun Huang, Philip L.S.M. Gordts, Zu-Yi Yuan, Sotirios Tsimikas, John Y.J. Shyy

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Figure 3

miR-483 targets PCSK9 and enhances LDLR expression in hepatocytes.

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miR-483 targets PCSK9 and enhances LDLR expression in hepatocytes. (A) B...
(A) Bioinformatics prediction of miR-483-5p binding sites in the 3′-UTR of human and mouse PCSK9 mRNA. In B–G, HepG2 cells were transfected with pre–miR-483 mimic (pre-483) or anti–miR-483 (anti-483) for 24 hours. (B and C) mRNA and protein levels of PCSK9 and LDLR. (D) HepG2 cells transfected with pre-483 or anti-483 were cotransfected with Luc-PCSK9-3′-UTR (WT) or Luc-mutated PCSK9-3′-UTR (MT). Luciferase activity was measured with pRL-TK activity as a transfection control. (E) Ago-1 or Ago-2 immunoprecipitation was performed, and miRISCs-associated miR-483, PCSK9, and CTGF mRNA levels were quantified by qPCR. (F and G) mRNA and protein levels of PCSK9 and LDLR in WT HepG2 and mPCSK9 HepG2 cells were transfected with pre-483 or anti-483. #LDLR in the same samples were detected in parallel in a separate gel (G). Data are mean ± SEM from 3–4 independent experiments. In B–G, non-normally distributed data were analyzed using Mann-Whitney U test between indicated group and control. *P < 0.05 vs. control. miR, microRNA; PCSK9, proprotein convertase subtilisin/kexin type 9; HepG2, human hepatocellular carcinoma; LDLR, LDL receptor; Luc-PCSK9-3′-UTR (WT), WT PCSK9 3′-UTR; Luc-mutated PCSK9-3′-UTR (MT), mutant PCSK9 3′-UTR; miRISCs, miRNA-induced silencing complexes; CTGF, connective tissue growth factor.