Abstract P120: Mir-338-3p is Down-regulated in Subcutaneous Small Arteries of Hypertensive Patients With Chronic Kidney Disease

Abstract

Background: Hypertension (HTN) and chronic kidney disease (CKD) are among the most prevalent global health conditions that cause millions of deaths per year. They are associated with vascular damage characterized by vascular remodeling, stiffening and endothelial dysfunction. miRNAs (miRs) are a class of small non-coding RNA that regulate gene expression by binding to their target messenger RNAs (mRNAs), thereby leading to mRNA degradation or translational repression. Their implication in vascular injury remains unclear. We aimed to identify differentially expressed (DE) miRs in small arteries of HTN and CKD human subjects to gather insight into pathophysiological molecular mechanisms in these conditions. Methods and Results: Normotensive, HTN [systolic blood pressure (BP) >135 mmHg or diastolic BP of 85-115 mmHg with BpTRU] and CKD subjects (eGFR<60mL/min/m 2 ) (n=15-16) were studied. Small arteries were isolated from subcutaneous gluteal biopsies and RNA extracted for small and total RNA sequencing using Illumina HiSeq-2500. EdgeR identified DE miRs ( P <0.05) uniquely associated with HTN (3 up and 6 down) or CKD (42 up and 39 down) or with both groups (2 down). Reverse transcription-quantitative PCR (RT-qPCR) was used to confirm miRNA differential expression. Correlation between RNA sequencing and RT-qPCR data was demonstrated for 3 miRs from 14 tested: mir-146a-5p (r=0.56, P <10 -4 ), miR-338-3p (r=0.91, P <10 -16 ) and mir-374a-3p (r=0.57, P <10 -4 ). The best correlated DE miRNA, miR-338-3p was down-regulated by 76% ( P <10 -4 ) and uniquely associated with CKD. TargetScan predicted that the up-regulated mRNA encoding glutathione peroxidase 3 ( GPX3 , 1.57 fold, P <0.001) to be a miR-338-3p target. Both miR-338-3p and GPX3 were found to be highly expressed in human aortic endothelial cells (HAECs) by RT-qPCR. Target prediction was validated by showing using RT-qPCR that GPX3 expression was up-regulated 1.73±0.28 fold in HAECs transfected with anti-miR-338-3p ( P <0.05). Conclusion: miR-338-3p down-regulation was found in small arteries, uniquely associated with CKD. GPX3 , which may be important for preservation of endothelial function, is a miR-338-3p potential target. miR-338-3p and associated target may play a role in vascular remodeling in CKD.