Therapeutic Targeting of Rpl13a-Small Nucleolar RNAs Mitigates Atherosclerosis in Apoe/- Mice

Specific non-coding, small nucleolar (sno) RNAs encoded within introns of the Rpl13a gene elevate cellular levels of reactive oxygen species (ROS), and ROS are known to aggravate atherosclerosis (athero). We found that genetic deficiency of Rpl13a -snoRNAs reduces brachiocephalic artery athero in Apoe -/- mice by 50%. We therefore tested the hypothesis that athero could be reduced by acute pharmacologic intervention targeting Rpl13a -snoRNAs in Apoe -/- mice. To that end, we injected male and female Apoe -/- mice subcutaneously with 48 mg/kg antisense oligonucleotides (ASOs) targeting (a) the four Rpl13a -snoRNAs (ASO-snoRNA), or (b) green fluorescent protein (ASO-control). We tested two distinct strategies: ( i ) preventive, in which ASO injections and Western diet were started in 10-wk-old mice, with 4 weekly and 5 bi-weekly ASO treatments over 14 wk, then sacrifice; and ( ii ) “therapeutic,” in which ASO injections were started in 24-wk-old mice that had been fed Western diet for 14 wk and then were continued on Western diet with 6 weekly ASO treatments and then sacrificed. ASO injections were administered as indicated until 1 wk prior to sacrifice. Compared with saline-injected mice, ASO-control and ASO-snoRNA showed no hepatotoxicity (as assessed by serum ALT and AST). Compared with ASO-control, ASO-snoRNA administration for 6 wk reduced Rpl13a -snoRNA levels in the aorta and spleen by 54-80% for snoRNAs U32a , U33 , and U34 (RT-qPCR). In the “therapeutic” cohort, brachiocephalic artery cross-sectional athero was reduced by 38% in ASO-snoRNA-treated as compared with ASO-control-treated mice (n=7/group, p <0.03). Additionally, male ASO-snoRNA-treated mice exhibited a 98.1% decrease in circulating IL1-β transcripts (n=6-8/group, p<0.003). In the “preventive” cohort, aortic athero was reduced by 40% (13±5 vs 7±3 % aortic lesion area, n= 11-13/group, p <0.02) in ASO-snoRNA-treated as compared with ASO-control-treated mice. We conclude that therapeutic targeting of Rpl13a -snoRNAs with ASOs mitigates athero, both during the early and later stages of atherogenesis.