Nephropathy secondary to Diabetes Mellitus (DM)-induced hyperglycemia is a leading cause of morbidity and mortality. Prolonged hyperglycemia activates the sorbitol pathway, resulting in osmotic damage, oxidative stress, impairment of renal function, and eventual kidney failure. The enzyme aldose reductase (AR) is believed to be the primary regulator of entry into this pathway by converting excess glucose to sorbitol and is a prime target for pharmacological therapy to preserve kidney function in diabetics. However, recent findings suggest that other aldo-keto reductases (AKRs) may also be important in synthesizing sorbitol. Many AR inhibitors have failed clinical trials due to lack of potency in human kidney and inability to target other AKR family enzymes, which serve redundant function in vivo. New strategies for developing more potent inhibitors with broader activity are needed. During our interrogation of the metabolic targets of Nasonia wasp venom in human cells, we identified non-canonical Aldo-keto reductase (AKR) enzymes (AKR1C* and 1D1) that may synthesize sorbitol more effectively and at lower substrate concentrations than the canonical enzyme AR. We hypothesize that non-canonical AKRs increase sorbitol accumulation, resulting in glomerular dysregulation in response to hyperglycemia. This proposal outlines an approach to study the role of non-canonical AKRs in sorbitol metabolism in human glomerular cells by investigating three primary aims: 1) Determine which AKR1C* sub-family (and AKR1D1) enzymes catalyze sorbitol synthesis in human glomerular cells by candidate AKR overexpression and knockdown. 2) Determine whether AKR1C* and 1D1 isoforms contribute to oxidative stress during sorbitol pathway activation by measuring site-specific ROS, and compensation by ROS scavengers (i.e. TEMPOL/Mito-TEMPOL). 3) Determine if AKR1C* sub-family (and AKR1D1) enzymes alter osmotic sensitivity in human glomerular cells by overexpressing candidate AKRs and measuring response to altered osmotic environment. The end result of this investigation will be identification of novel, non-canonical AKR regulators of te glucose to sorbitol conversion in human kidney cells. Identification of the enzymes affected will provide new insight into strategies for stratifying risk, blocking AR/AKR function in vivo, and preventing sorbitol accumulation and development of glomerular dysfunction. The formal mentoring program combines and integrates training and research opportunities in biomedical and basic biology highly tailored to the PD/PI's career goals, greatly enhancing the training potential of the applicant's dual MD/PhD program. A highly accomplished mentoring team with extensive mentoring experience and complementary skills in genetics, cell biology, molecular biology and biochemistry is in place.
Diabetic nephropathy secondary to prolonged periods of hyperglycemia is a common source of morbidity and mortality in the United States, and the leading cause of diabetes mellitus (DM)-related End Stage Renal Disease (ESRD). With the growing number of Americans developing DM earlier in life and the significant cost of treatment, new strategies are needed to understand the development of ESRD and to devise new treatments to protect kidney function from chronic hyperglycemia-induced damage. Sorbitol pathway activation in mesangial cells and podocytes may contribute to the pathophysiological process of glomerular dysfunction observed in long-standing DM. This study will elucidate the role of non-canonical enzymes that may mediate this process, and which could be used for risk stratification and serve as future therapeutic targets.
