Effect of spironolactone on diabetic nephropathy in albino rats: ultrastructural and immunohistochemical study
Keywords:Spironolactone, WT-1 protein, Podocytes, Glomerulofibrosis, Diabetic nephropathy
Background: Diabetic nephropathy (DN) has become one of the most common causes of end stage renal disease (ESRD). Hyperglycemia induces oxidative stress in renal tubular epithelial cells that initiate tubulointerstitial fibrosis, which is a characteristic feature of diabetic nephropathy that becomes progressively complicated by renal failure. Aim: To assess the effect of spironolactone (SPL) on WT-1 protein expression and ultrastructural changes associated with the progression of experimental diabetic nephropathy (DN).
Methods: Forty female albino rats were divided into five groups. Group I (control group), Group II (untreated diabetic rats), Group III (insulin-treated diabetic rats), Group IV (spironolactone-treated diabetic rats) and Group V (insulin and spironolactone-treated diabetic rats). At the 4th and 8th weeks, 4 rats from each group were sacrificed and renal tissue and blood samples were obtained. The rats were anaesthetized using ether inhalation. Each kidney was longitudinally divided and processed for immunohistochemical analysis with rabbit polyclonal anti-WT-1 Antibody and electron microscopic examination.
Results: Treatment of STZ-induced diabetic rats with insulin and spironolactone (Group V) showed improvement in renal corpuscles as well as their capsular space, the basement membrane became normal with preserved minor and major processes and subpodocytic space, most of the proximal convoluted tubules retained their brush border, and their cells showed normal euchromatic nuclei and scattered mitochondria with apical microvilli, which is similar to the findings of the control group. Quantitative analyses showed significant increase in area of fibrosis and focal thickening of the glomerular basement membrane in non-SPL treated groups. There was a marked decrease in proteinuria compared to other treated groups. The results were better after 8 weeks compared to those after 4 weeks.Conclusions: The administration of SPL significantly prevented the extent of interstitial fibrosis in the diabetic kidney.
Barton M, Sorokin A. Endothelin and the Glomerulus in Chronic Kidney Disease. Semin Nephrol. 2015;35:156–67.
Weil J, Lemley K, Mason C, Yee B, Jones L, Blouch K, et al. Podocyte detachment and reduced glomerular capillary endothelial fenestration promote kidney disease in type 2 diabetic nephropathy. Kidney Int. 2012;82:1010–7.
Nishikawa T, Matsuzawa Y, Suematsu S, Saito J, Omura M, Kino T. Effect of Atorvasatin on Aldosterone Production, induced by Glucose, LDL or Angiotensin II in Human Renal Mesangial Cells. Arzneimittelforschung. 2010;60:445–51.
Pagtalunan E, Miller L, Jumping-Eagle S, Nelson G, Myers D, Rennke G, et al. Podocyte loss and progressive glomerular injury in type II diabetes. J Clin Invest. 1997;99:342-8.
Reidy K, Susztak K. Epithelial mesenchymal transition and podocyte loss in diabetic kidney disease. Am J kidney Dis. 2009;54:590-3.
Shibata S1, Nagase M, Yoshida S, Kawachi H, Fujita T. Podocyte as the target for aldosterone: roles of oxidative stress and Sgk1. Hypertension. 2007;49(2):355-64.
Waldstorm M, Grove A. Immunohistochemical expression of Wilms tumor gene protein in different histologic subtypes of ovarian carcinomas. Arch Pathol Lab Med. 2005;129:85-8.
Su J, Li J, Chen H, Zeng H, Zhou H, Li S, et al. Evaluation of podocyte lesion in patients with diabetic nephropathy: Wilms' tumor-1 protein used as a podocyte marker. Diabetes Res Clin Pract. 2010;87:167-75.
Hsueh A, Wyne K. Renin-Angiotensin-Aldosterone System in Diabetis and Hypertension. J clini hypertension. 2011;13:224-37.
Athyros G, Mikhailidis P, Kakafika I, Tziomalos K, Karagiannis A. Angiotensin II reactivation and aldosterone escape phenomena in renin-angiotensin-aldosterone system blockade: is oral renin inhibition the solution? Expert Opin Pharmacother. 2007;8:529-35.
Mehdi F, Adams-Huet B, Raskin P, Vega L, Toto D. Addition of Angiotensin Receptor blockade or mineralocorticoid antagonism to maximal angiotensin-converting enzyme inhibition in diabetic nephropathy. J Am Soc Nephrol. 2009;20:2641-50.
Ceron CS, Castro MM, Tanus-Santos JE. Spirlonolactone And Hydrochlorothiazide Exert Antioxidant Effects And Reduce Vascular Matrix Metalloproteinase-2 Activity And Expression In A Model Of Renovascular Hypertension. British J Pharmacol. 2010;160:77-87.
Lozano-Maneiro L, Puente-Garcia A. Renin-Angiotensin-Aldosterone Blockade in Diabetic Nephropathy. Present Evidences. J Clin Med. 2015;4:1908-37.
Wen Y, Ouyang J, Yang R, Chen J, Liu Y, Zhou X. Reversal of new onset type 1 diabetes in mice by synergic bone marrow transplantation. Bioch Biophy Res Commu. 2008;374:282-7.
Balkis Budin S, Othman F, Louis SR, Abu Bakar M, Radzi M, Osman K, et al. Effect of alpha lipoic acid on oxidative stress and vascular wall of diabetic rats, Rom J Morphol Embryol. 2009;50:23–30.
Michael T, Ganesh N, Viswanathan P. Effect of long acting insulin supplementation on diabetic nephropathy in Wistar rats. Indian J Experim Biol. 2012;50:867-74.
Aguilar C, Rodríguez-Delfín L. Effects of spironolactone administration on the podocytes loss and progression of experimental diabetic nephropathy. Rev Peru Med Exp Salud Publica. 2012;29:490-7.
ILAR (Institute of Laboratory Animal Resources). Guide for the Care and Use of Laboratory Animals NIH Publication No. 86-23. National Academy Press, Washington, D.C., 1985.
Schreier C, Kremer W, Huber F, Neumann S, Pagel P, Lienemann K, et al. Reproducibility of NMR Analysis of Urine Samples: Impact of Sample Preparation, Storage Conditions, and Animal Health Status. BioMed. Res Int. 2013;2013(878374):19.
Mahmoodi K, Gansevoort T, Veeger J, Matthews G, Navis G, Hillege L, et al. Prevention of Renal Vascular End-stage Disease (PREVEND) Study Group: "Microalbuminuria and risk of venous thromboembolism". JAMA. 2009;301:1790–7.
Susztak K, Raff C, Schiffer M, Böttinger P. Glucose-induced reactive oxygen species cause apoptosis of podocytes and podocyte depletion at the onset of diabetic nephropathy. Diabetes. 2006;55:225–33.
Qian Y, Feldman E, Pennathur S, Kretzler M, Brosius F. Mechanisms of Glomerulosclerosis in Diabetic Nephropathy. Diabetes. 2008;57:1439–45.
Wharram L, Goyal M, Wiggins E, Sanden K, Hussain S, Filipiak E, et al. Podocyte depletion causes glomerulosclerosis: diphtheria toxin-induced podocyte depletion in rats expressing human diphtheria toxin receptor transgene. J Am Soc Nephrol. 2005;16:2941–52.
Pagtalunan E, Miller L, Jumping-Eagle S, Nelson G, Myers D, Rennke G, et al. Podocyte loss and progressive glomerular injury in type II diabetes. J Clin Invest. 1997;99:342–8.
Fornoni A. Proteinuria, the Podocyte, and Insulin Resistance. N Engl J Med. 2010;363:2068-9.
Makhlough A, Kashi Z, Akha O, Zaboli E, Yazdanicharati J. Effect of Spironolactone on Diabetic Nephropathy Compared to the Combination of Spironolactone and Losartan. Nephro Urol Mon. 2014;6:e12148.
Ng P, Jain P, Heer G, Redman V, Chagoury L, Dowswell G, et al. Spironolactone to prevent cardiovascular events in early-stage chronic kidney disease (STOP-CKD): study protocol for a randomized controlled pilot trial. Trials. 2014;15:158.
Lin M, Yiu H, Wu J, Chan Y, Leung C, Au S, et al. Toll-like receptor 4 promotes tubular inflammation in diabetic nephropathy. J Am Soc Nephrol. 2012;23:86–102.
Elsherbiny N, El-Sherbiny M, Said E. Amelioration of experimentally induced diabetic nephropathy and renal damage by nilotinib. J Physiol Biochem. 2015;71:635-48.
Alsaad O, Herzenberg M. Distinguishing diabetic nephropathy from other causes of glomerulosclerosis: An update. J Clin Pathol. 2007;60:18-26.
Kim K, Lee H, Kim C, Cha R, Kang S. A case of primary aldosteronism combined with acquired nephrogenic diabetes insipidus. Kidney Res Clin Pract. 2014;33:229–33.
Toyonaga J, Tsuruya K, Ikeda H, Noguchi H, Yotsueda H, Fujisaki K, et al. Spironolactone inhibits hyperglycemia-induced podocyte injury by attenuating ROS production. Nephrol Dial Transplant. 2011;26:2475-84.
Lin S, Li D, Jia J, Zheng Z, Jia Z, Shang W. Spironolactone ameliorates podocytic adhesive capacity via restoring integrin α3 expression in streptozotocin-induced diabetic rats. J Renin Angiotensin Aldosterone Syst. 2010;11(3):149-57.