The Role of miR-486-5p in Renal Vascular Endothelium in Ischemic Acute Kidney Injury

Abstract

Acute kidney injury (AKI) refers to a rapid decline in kidney function and increases the risk for chronic kidney disease (CKD). Yet, no effective treatments exist. Ischemia-reperfusion (I/R) injury is a major cause of AKI in clinical settings such as cardiac surgery and kidney transplant. In I/R AKI, endothelial injury contributes to capillary rarefaction, tubulointerstitial fibrosis, and the transition to CKD. microRNAs are short non-coding RNAs that potently regulate gene expression. We previously showed that miR-486-5p protects against kidney I/R injury in mice. However, the effect of miR-486-5p on long-term vascular outcomes after kidney I/R injury were unexplored, and its targets in endothelial cells undefined. The aims of this project were to (1) evaluate the effect of miR-486-5p on kidney I/R injury in rats, focusing on long-term vascular function and the AKI to CKD transition, (2) determine the effect of miR-486-5p in cultured endothelial cells, and (3) conduct a systematic review of dysregulated miRNAs in human AKI.

We demonstrated that only early delivery of miR-486-5p to rats with kidney I/R prevents kidney injury and protects against CKD development and endothelial dysfunction. Unexpectedly, early and delayed delivery of miR-486-5p attenuated I/R-induced kidney endothelial nitric oxide synthase (eNOS) expression. In cultured endothelial cells, miR-486-5p decreases eNOS protein levels without targeting the eNOS 3' untranslated region. miR-486-5p also inhibits angiogenesis in vitro in an eNOS-dependent manner. Biotinylated miR-486-5p pull-down identified multiple predicted and novel transcripts, including MAML3 - a transcriptional coactivator of Notch signaling - as targeted by miR-486-5p. Selective MAML3 knockdown inhibits eNOS protein levels, suggesting that miR-486-5p inhibits angiogenesis via targeting of MAML3 and resultant eNOS downregulation. For the third aim, our systematic review of miRNAs in human AKI identified potential AKI biomarkers and highlighted important clinical miRNA study methodological limitations.

This thesis advances our earlier work on the therapeutic potential of miR-486-5p in pre-clinical AKI. The prevention of capillary rarefaction and systemic vascular dysfunction only by early miR-486-5p administration suggest that early gene responses to I/R injury are critical for the transition from acute to chronic kidney disease. Although no adverse vascular effects were observed in vivo, miR-486-5p conferred an eNOS-dependent anti-angiogenic effect in cultured endothelial cells, possibly by targeting MAML3. These findings support a novel pathway regulating endothelial function.