Abstract 257: miR-217 Mediates the Protective Effects of the Dopamine D2 Receptor on Renal Inflammation and Fibrosis.

Abstract

Some common single nucleotide polymorphisms (SNPs; rs 6276, 6277, and 1800497) in the human dopamine D2 receptor (DRD2) gene are associated with decreased receptor expression and function, as well as high blood pressure. We have reported that human renal proximal tubule cells (hRPTCs) from subjects carrying D2R SNPs had decreased D2R expression and function and increased expression of the pro-fibrotic factor TGFβ1 and its signaling targets Smad3 and Snail1. These cells also showed induction of epithelial mesenchymal transition and production of extracellular matrix proteins fibronectin-1 (FN-1), Collagen 1a (Col 1a), and vimentin. To determine the mechanisms underlying the effects of the D2R SNPs, we studied the expression of several microRNAs (miRs) that are associated with the regulation of chemokines, anti-fibrosis, extracellular matrix remodeling, and cell adhesion through post-transcriptional repression of their target mRNAs. The expression of 4 miRs (miR-217, miR-224, miR-335, miR-1265) was downregulated (-10.3 to -6.0 fold) and that of one miRNA (miR-1290; 5.3 fold) was upregulated in hRPTCs with D2R SNPs. The expression of miR-217 and miR-224 is regulated by the D2R; the expression is decreased by D2R downregulation in cells not carrying D2R SNPs (WT) and normalized by treating cells carrying D2R SNPs with a Drd2 plasmid. Treatment of WT cells with a miR-217 mimic significantly decreased the mRNA expression of TGFβ1 (43±2%), MMP3 (65±4%), and FN-1 (30±4%) while treatment with a miR-217 inhibitor increased TGFβ1 (94± 9%), MMP3 (96±5%), and FN-1 (212±30%). miR-217 mimic treatment of D2R SNPs cells also significantly decreased the expression of TGFβ1, MMP3, and FN-1. However, treatment of these cells with a miR-217 inhibitor had non-significant effects probably because miR-217 level was already reduced. In contrast treatment of either WT or D2R SNPs cells with miR-224 mimic or inhibitor had no significant effects on the expression of TGFβ1, MMP3, or FN-1. These results indicate that the protective effects of D2R on inflammation and fibrosis are at least, in part, mediated by regulation of miR-217 expression. Dysregulation of miR-217 expression may be responsible for the profibrotic phenotype of hRPTCs in individuals with these D2R SNPs.