Abstract 18260: Apolipoprotein A-I Mimetic Peptide, 4F Targets Lipoxygenases via miR193-3p Induction to Rescue Severe Pulmonary Hypertension

Abstract

Background: Pulmonary hypertension (PH) causes increase in pulmonary arterial pressure causing right ventricular (RV) hypertrophy and RV failure (RVF). We have shown that apolipoprotein A-I mimetic peptide 4F rescues advanced PH and decreases circulating levels of oxidized fatty acids [hydroxyoctadecadienoic acids (HODEs) and hydroxyeicosatetraenoic acids(HETEs)] in the MCT rat-model of PH. Here, we show that 4F rescues PH in rats and mice by inducing miR-193-3p (miR193), which regulates the enzymes responsible for the production of HETEs and HODEs. Methods and results: PH was induced in rats by monocrotaline (MCT, 60mg/kg,s.c. single injection) or in mice by hypoxia (O 2 ≤10%) for 21 days. 4F daily therapy or intratracheal instillation of miR193 were started after the establishment of PH in both models (4F: 50mg/kg/day, s.c. from day 21-30 in MCT model and day 14-21 in hypoxia model; miR193 (20nM) at days 16, 21 and 26 in MCT model or at days 14 and 18 in hypoxia model). RV pressure (RVP) was measured by direct catheterization. Rats developed PH 21 days after MCT (RVP=67.12±1 in PH vs. 29.8±1mmHg in the ctrl group) which progressed to RVF by day 30 (RVP=74±1, p<0.05 vs ctrl). 4F therapy rescued PH in both models (MCT model: RVP=46±3 mmHg, p<0.05 vs PH and RVF; Hypoxia model: RVP=22±3.8 mmHg vs 36.91±5.74 in PH and 20.93±2.52 in ctrl normoxic mice). MicroRNA microarray and qPCR showed a ~3 fold downregulation of miR193 in PH that was normalized to ctrl levels by 4F treatment in both models. OE of miR193 (20nM) resulted in ~2.5 fold gain of miR193 in the lung tissue and rescued preexisting PH as the RVPs were significantly lower than PH group in both models (38.2±5.5 mmHg in MCT-rat and ~25.3±0.97mmHg in hypoxic-mice). MiR193 therapy reversed increased arteriolar muscularization in PH. In vivo gain of miR193 suppressed transcription of lipooxygenases ALOX5, ALOX12 and ALOX15. In vitro treatment of human Pulmonary Arterial SMCs (hPASMC) with HETEs and HODEs suppressed miR193 levels in the absence of 4F. Lastly, miR193 OE decreased hPASMCs proliferation in the presence of serum or 12-HETE (100ng/ml) by >2 folds (p<0.05 vs ctrl) whereas miR193 KD increased proliferation. Conclusion: 4F rescues preexisting severe PH in rats and mice by inducing miR193 and targeting lipoxygenases.