Publication Date

2009

Document Type

Thesis

Committee Members

Khalid Elased (Advisor), James Lucot (Committee Member), Mariana Morris (Committee Member)

Degree Name

Master of Science (MS)

Abstract

Alterations in the renin-angiotensin system (RAS) are considered to be crucial for the development of diabetic complications like hypertension and nephropathy. Our previous work demonstrated role of AT1 receptors (AT1R) in the development of hypertension in db/db diabetic mice. The aim of this study was to test the hypothesis that there is upregulation of renal AT1R and imbalance in renal ACE/ACE2 homeostasis in db/db mice. In addition, we hypothesize that treatment with an anti-hyperglycemic or an AT1R blocker will correct this imbalance. Five week old control and db/db mice were housed in metabolic cages for 24 hour collection of urine. At early age of 5 weeks, db/db mice were obese and hyperglycemic. Urinary albumin excretion was also significantly high in db/db mice. Changes in RAS were evaluated using enzyme activities, western blots and immunohistochemistry. There was a significant increase in urinary ACE2 activity and ACE2 content in db/db mice at 5 weeks. There was a significant increase in plasma ACE activity and Ang II content in db/db mice compared to controls at 8 weeks. Western blot analysis showed significant increase in AT1R protein expression in 8, 18 and 31 week db/db mice compared to controls. There was upregulation of ACE2 and down-regulation of ACE in kidney to compensate the effects of high plasma Ang II. To study the effect of reduction in blood glucose and AT1R blockade, mice were treated with metformin and losartan for 12 weeks. Chronic treatment with metformin (150 mg/kg/day) and losartan (10 mg/kg/day) significantly decreased urinary albumin and protein excretion. Metformin improved blood glucose and glucose tolerance db/db mice, but did not affect renal expression of ACE, ACE2 and AT1R. Although chronic losartan treatment did not alter blood glucose levels, it improved the morphology of pancreatic islets. There was a significant increase in renal AT1R protein expression and decrease in renal ACE2 protein expression following losartan treatment. Losartan treatment significantly increased urinary ACE2 activity. Western blot of concentrated urine from 8 week db/db mice revealed immunoreactive bands of ACE, ACE2 and AT1R protein. Conclusion: 1) There is upregulation in renal AT1R protein expression in db/db mice. 2) Chronic metformin treatment significantly reduces blood glucose and microalbuminuria in db/db mice without affecting ACE/ACE2 balance. 3) Chronic losartan treatment had no effect on blood glucose, but it up-regulates renal AT1R and down-regulates renal ACE2. 4) Enzyme activity and western blot shows increased excretion of ACE2 in the urine of db/db mice. These data show that urinary ACE and ACE2 provide good index of intra-renal RAS status and could be used in early diagnosis and prognosis of diabetic renal disease.

Page Count

101

Department or Program

Department of Pharmacology and Toxicology

Year Degree Awarded

2009

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