Protective role of CH25h/25-HC in diabetic kidney disease.

James J Peters Va Medical Center, Bronx NY

Investigators

Abstract

Abstract Diabetic kidney disease (DKD) is the leading cause of chronic kidney disease (CKD) worldwide. Endothelial cell (EC) dysfunction is a key event leading to the progression of DKD. Loss of ECs occurs in both glomerular endothelial cells (GECs) and peritubular endothelial cells (PTECs). Through unbiased screening of GEC transcriptome in animal models with early DKD, we identified cholesterol 25-hydroxylase (CH25H) as one of the highly regulated genes in diabetic kidneys. CH25H is the main enzyme that hydroxylates cholesterol to generate 25-hydroxycholesterol (25-HC). Previous studies have shown that CH25H and 25-HC are involved in a wide range of biological processes, including regulation of inflammation, lipogenesis, and EC function. However, these studies reveal controversial findings, suggesting that the exact role of CH25H and its metabolite 25-HC have not been well-defined. CH25H is known to be highly expressed in the kidney cells but its role in renal cell biology and pathology has never been reported. Over the last several years, we have generated a large amount of preliminary data to support the critical role of CH25H/25-HC in the pathogenesis of DKD. Our key observations show that CH25H is expressed mostly in GECs and PTECs and that its expression is increased in human and mouse kidneys with early DKD. Global deletion of CH25H in db/db mice aggravated DKD and was associated with increased EC apoptosis, while treatment of db/db mice with 25-HC was able to alleviate the kidney injury. Mechanistically, we found that 25-HC increased ARF4 activity in ECs by disrupting the interaction between ARF4 and its inhibitor ASAP1. We also confirmed the key role of ARF4 in the Golgi function in ECs. We found that in diabetic conditions, ASAP1 expression was increased in ECs and resulted in ARF4 deficiency, Golgi dysfunction, and EC apoptosis, but these abnormalities could be rescued by overexpression of CH25H or treatment of 25-HC. An intronic single nucleotide polymorphism of ASAP1 and a missense mutation of ASAP1 were also identified to be associated with DKD in diabetic patients. Based on these preliminary data, we hypothesized that CH25H/25-HC has a major protective role in DKD via maintaining normal Golgi function in ECs by inhibiting the negative regulatory effect of ASAP1 on ARF4. To test this hypothesis, we propose the following aims: Aim 1: Determine how CH25H/25-HC are regulated in ECs and how CH25H/25-HC contributes to Golgi dysfunction and EC injury in diabetic conditions. Aim 2: Determine the cell-specific effects of CH25H on the development of DKD by using inducible EC-specific CH25H knockout or overexpression mice. Aim 3: Determine the role of ARF4 and Asap1 in the regulation of Golgi function and EC injury in DKD in mice.