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Abstract: SA-PO383

Effect of Pioglitazone on the Shedding of Urinary Angiotensin Converting Enzyme (ACE) 2 and Neprilysin (NEP) in db/db Mice and Their Role as Urinary Biomarkers for Diabetic Kidney Disease

Session Information

Category: Chronic Kidney Disease (Non-Dialysis)

  • 301 CKD: Risk Factors for Incidence and Progression


  • Kumbaji, Meenasri, Wright State University, Dayton, Ohio, United States
  • Fadnavis, Rucha, Wright State University, Dayton, Ohio, United States
  • El-Amouri, Salim, Wright State University, Dayton, Ohio, United States
  • Saklayen, Mohammad G., VA Medical Center, Dayton, Ohio, United States
  • Elased, Khalid M., Wright State University, Dayton, Ohio, United States
  • Grobe, Nadja, Wright State University, Dayton, Ohio, United States

Diabetic kidney disease (DKD) is one of the major causes of end-stage renal disease. Angiotensin (Ang) II is the major biological active peptide of the renin angiotensin system. Elevated levels of Ang II contribute to initiation and progression of DKD. The actions of Ang II could be antagonized by its conversion to the vasodilator Ang (1-7), partly generated by the action of ACE2 and NEP. Although there is an emergence of some urinary biomarkers such as angiotensinogen, kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL), there is still a need for new early biomarkers for DKD. The aim is to investigate the effect of hyperglycemia on urinary albuminuria, ACE2, NEP and KIM-1 and to test the hypothesis that ACE2 and NEP can be used as early biomarkers for DKD.


8 weeks old control and db/db diabetic mice were subjected to pioglitazone treatment (20 mg/kg/day) for 10 weeks. Metabolic and renal parameters were measured. Urine was collected for the evaluation of ACE2, NEP, KIM-1 protein expression and RAS activity.


At 7 weeks old, there was no significant difference in urinary albumin, NEP and KIM-1 between db/db and control mice. However, western blot showed a significant increased shedding of urinary ACE2 fragment (60KDa and 70kDa immunoreactive bands) in db/db mice compared to controls. Although there was a prominent immunoreactive band for NEP in control mice at 9 weeks, there was no detectable immunoreactive band for ACE2. After the development of albuminuria in older db/db mice (9-17week), there was a significant increase of full length (95KDa) and fragment of ACE2, NEP (90kDa) and KIM-1 (75KDa) compared to control mice. Urinary NEP and ACE2 activities significantly increased in 17 weeks db/db mice compared to controls. Pioglitazone normalized hyperglycemia and attenuated urinary albumin, glucose and ACE2 shedding. However, it has no effect on urinary NEP and KIM-1.


In db/db diabetic mice, increased shedding of enzymatically active ACE2 precedes albuminuria. Depletion of tubular renal ACE2, NEP, could lead to accumulation of Ang II, with concomitant development of microalbuminuria. Urinary ACE2 and NEP could be used as biomarkers for DKD.