Abstract 190: Serum HDL-C Decreases in MicroRNA-33b Knock-in Mice for an Intron of Sterol Regulatory Element-binding Factor 1 (Srebf1)
Background: MicroRNAs (miRs) are small non-protein-coding RNAs that bind to specific mRNAs and inhibit translation or promote mRNA degradation. Recent reports, including ours, indicated that miR-33 (miR-33a) located within the intron of sterol regulatory element-binding factor (SREBF) 2 controls cholesterol homeostasis. Primates, but not rodents, express a second miR-33 gene (miR-33b) from an intron of SREBF1. To address miR-33b function in vivo, we developed humanized mice, in which a miR-33b transgene is inserted within a Srebf1 intron.
Methods: The human miR-33b sequence was introduced into intron 16 of mouse Srebf1 because miR-33b is located in intron 16 of human SREBF1 and there are high homologies in exons 16 and 17 between human and mouse. The expression of serum miRNA was normalized with cel-miR-39 as a spike-in control.
Results: We successfully established miR-33b knock-in (KI) mice with C57BL/6 background and this miR-33b KI strategy did not alter Srebf1 intron 16 splicing, which was confirmed by RT-PCR and sequencing. An LXR agonist T0901317, which induces Srebf1 expression, enhanced miR-33b expression in primary hepatocytes and the liver of miR-33 KI mice. The protein levels of known miR-33a target genes, such as ABCA1, ABCG1, and SREBP-1, were reduced compared with those in wild-type mice. Peritoneal macrophages from the miR-33b KI mice had a reduced cholesterol efflux capacity via apoA-I and HDL-C. Serum HDL-C levels were reduced by almost 35% in miR-33b KI mice. HPLC elution analysis showed that the decreased HDL-C levels were mainly composed of very large-, large-, medium sized HDL-C, which was compatible with the previous results of miR-33a deficient mice. Next, we measured miR-33b levels in serum of human. The expressions of miR-33b-3p were inversely correlated with serum HDL-C levels (P=0.025, R=0.336).
Conclusions: miR-33b KI mice for an intron of Srebf1 showed reduced HDL-C level and serum miR-33b-3p levels were inversely correlated with HDL-C levels in human. These results indicate that miR-33b can be a potential target for raising HDL-C and may account for lower HDL-C levels in humans than those in mice. These mice will aid in elucidating the roles of miR-33s and screening of the drugs that can alter miR-33s levels and activities.
Author Disclosures: T. Horie: None. T. Nishino: None. Y. Kuwabara: None. O. Baba: None. T. Kimura: None. K. Ono: None.
- © 2015 by American Heart Association, Inc.