SAT-LB127 Regulation of ENaC by Exosomal Lipids in the Diabetic Kidney

Effective treatment of hypertension (HTN) in patients with diabetes may help to significantly reduce the risk of those patients developing additional complications including vascular disease and diabetic nephropathy. Blockers of the renin-angiotensin system including angiotensin converting enzyme inhibitors and angiotensin receptor blockers are not always effective in treating HTN in diabetic patients. Therefore, the aim of this study was to use an animal model of type 2 diabetes to investigate a novel mechanism of diabetes associated HTN involving exosomal lipids in the upregulation of epithelial sodium channel (ENaC) activity in the kidney. We performed metabolic cages studies using male and female hypertensive (salt-loaded induced) diabetic db/db mice and healthy age-matched wild-type control mice in order to isolate and characterize urinary exosomes from each group by nanoparticle tracking analysis, Western blotting, and transmission electron microscopy. Our mass spectrometry based lipidomic studies identified key lipids that were differentially expressed in the kidney derived exosomes from the hypertensive diabetic mice compared to control mice. Sphingomyelin quantification assays showed total sphingomyelin content was elevated in the exosomes from the hypertensive diabetic mice compared to the control group. Single channel patch clamp studies showed urinary exosomes enriched in sphingomyelins from hypertensive diabetic mice compared to controls increase ENaC activity (at the level of channel density and open probability) in cultured distal tubule renal epithelial cells. Moreover, exogenous application of sphinomyelin-6 to cultured mouse cortical collecting duct (mpkCCD) cells resulted in an increase in amiloride-sensitive transepithelial current. Taken together, our data show various lipids are enriched in exosomes from hypertensive diabetic mice compared to controls and these exosomes positively regulate ENaC activity in distal tubule and collecting duct cells.