Inducible nitric oxide synthase (iNOS) expression in early diabetic nephropathy: Effect of ACE inhibitor and angiotensin II receptor antagonists

Öz

Amaç: Diabetes mellitusun en önemli komplikasyonlarından biri diabetik nefropatidir. Kaptopril bir ACE inhibitörüdür ve diabetik nefropati tedavisinde kullanılır. Son yıllarda anjiotensin reseptör antagonistlerinin proteinüriyi önlemek için ACE inhibitörleri gibi etki ettikleri gösterilmiştir. Anjiotensin II-AT1 reseptör antagonisti Losartan, bu çalışma alanında çalışılan yeni bir antihipertansif materyaldir. Nitrik oksit (NO), akut streptozosin verilerek oluşturulan diabetteki, akut meydana gelen hemodinamik Materials and methods: 32 male Wistar rats weighting 140-220 g were used for this study. The rats were made diabetic using a single intraperitoneal injection of 65 mg/kg streptozotocin (Sigma Chemical Co., St. Louis, MO, USA). Rats were divided in to four groups: Group A (n= 8): Control, Group B (n=8): STZ- diabetic control, Group C (n=8): STZ + Losartan (DM+ L) (10 mg/kg/day), Group D (n=8): STZ + Captopril (DM+C) (50 mg/kg/day). The experiment lasted 6 weeks. The levels of blood glucose, albuminuria, creatinine clearence was determined. Renal tissue samples were extracted under anesthesia for histopathologic study. Monoclonal antibodie against the following antigen was used: iNOS . Result: At the diabetic control group, the level of albuminuria was increased significantly and was decreased significantly in the Captopril and Losartan given groups (p<0.001). There was no statistically significant differences in albuminuria between the DM+L and DM+C groups (p>0.05). The values of creatinine clearence were increased in diabetic control group as compared to control group (p<0.001). The reducing creatinine clearence in DM+L and DM+C groups was statistically not significant (p>0.05). By immunohistochemistry, iNOS staining was only observed in glomeruli of diabetic control group. Conclusion: Glomerular iNOS expression was enhanced in diabetic nephropathy and the activation of angiotensin II may play a role in this enhancement. Materyal ve metod: Bu çalışmada 140-220 gr ağırlığında 32 adet Wistat ratları kullanılmıştır. Ratlarda tek doz intraperitoneal 65 mg/kg streptozotocin (Sigma Chemical Co., St. Loui, MO, USA) kullanılarak diabet oluşturulmuştur. Ratlar 4 gruba bölünmüştür: grup A (n=8): Kontrol, grup B (n=8): STZ-diabetik kontrol, grup C (n=8): STZ+Losartan (DM+L) (10 mg/kg/gün), grup D (n=8): STZ+Kaptopril (DM+C) (50 mg/kg/gün). Deney 6 hafta sürdürüldü. Kan şekeri düzeyi, albüminüri, kreatinin kleransı saptandı. Böbrek dokusu örnekleri histopatolojik çalışma için anestezi altında alındı. Monoklonal antikora karşı antijen kullanıldı: iNOS . Sonuçlar: Diabetik kontrol grubunda albiminüri seviyesi anlamlı derecede artmış ve Kaptopril ve Losartan verilen gruplarda önemli derecede azalmıştı (p<0.001). DM+L ve DM+C grupları arasında anlamlı bir istatiksel fark görülmemişti (p>0.05). Kreatinin kleransinin değeri kontrol grubuna benzer şekilde diabetik kontrol grubunda da artmıştı (p<0.001). DM+L ve DM+C gruplarında kreatinin kleransinindeki azalma istatiksel olarak anlamlı değildi (p>0.05). İmmün dokukimyasal olarak iNOS boyanması diabetik kontrol grubunda sadece glomerüllerde görüldü. Tartışma: Glomerüler iNOS ekspresyonu diabetik nefropatiyi arttırır ve anjiotensin II aktivasyonu bu artmada rol oynayabilir

Anahtar Kelimeler

• iNOS, diabetik nefropati, ACE inhibitörleri, Anjiotensin II

Erken diabetik nefropatide indüklenebilir nitrik oksid sentetaz (iNOS) ekspresyonu: ACE inhibitörlerinin ve anjiotensin II reseptör antagonistlerinin etkileri

Abstract

Objective: One of the most important complications of diabetes mellitus is diabetic nephropathy. Captopril is an ACE inhibitor and used in the diabetic nephropathy treatment. In the last years, it has been shown that angiotensin receptor antagonists have similar efficiency as ACE inhibitors for prevention of proteinuria. The angiotensin II-AT1 receptor antagonist Losartan is a new antihypertansive material that has been worked on this field. Nitric oxide (NO) is a multifunctional mediator that has been implicated in the short-term hemodynamic alterations that occur acute streptozocin induced değişikliklerde etkisi bulunan çok fonksiyonlu bir mediadiabetes. Nitric oxide is synthesized exclusively from its tördür. Nitrik oksit, nitrik oksit sentez (NOS) enziminin precursor L-arginine under the catalytic effect of nitric oxide synthase (NOS), which there are three isoforms. Diabetic nephropathy represents a complex metabolic arena characterized with patho-physiological events that both stimulate end depress intrarenal NO production. The aim of this study was to assess whether the iNOS pathway is pathologically altered in experimental diabetic nephropathy, and in therapy with ACE inhibitor (Captopril) or angiotensin II-AT1 receptor antagonist (Losartan). Materials and methods: 32 male Wistar rats weighting 140-220 g were used for this study. The rats were made diabetic using a single intraperitoneal injection of 65 mg/kg streptozotocin (Sigma Chemical Co., St. Louis, MO, USA). Rats were divided in to four groups: Group A (n= 8): Control, Group B (n=8): STZ- diabetic control, Group C (n=8): STZ + Losartan (DM+ L) (10 mg/kg/day), Group D (n=8): STZ + Captopril (DM+C) (50 mg/kg/day). The experiment lasted 6 weeks. The levels of blood glucose, albuminuria, creatinine clearence was determined. Renal tissue samples were extracted under anesthesia for histopathologic study. Monoclonal antibodie against the following antigen was used: iNOS . Result: At the diabetic control group, the level of albuminuria was increased significantly and was decreased significantly in the Captopril and Losartan given groups (p<0.001). There was no statistically significant differences in albuminuria between the DM+L and DM+C groups (p>0.05). The values of creatinine clearence were increased in diabetic control group as compared to control group (p<0.001). The reducing creatinine clearence in DM+L and DM+C groups was statistically not significant (p>0.05). By immunohistochemistry, iNOS staining was only observed in glomeruli of diabetic control group. Conclusion: Glomerular iNOS expression was enhanced in diabetic nephropathy and the activation of angiotensin II may play a role in this enhancement

Keywords

• iNOS, diabetic nephropathy, ACE inhibitor

References

1. Veelken R, Hilgers KF, Hartner A, Haas A, et al. Nitric oxide synthase isoforms and glomerular hyperfiltration in early diabetic nephropathy. J Am Soc Nephrol 2000; 11: 71-79.
2. Ibrahim HN, Hostetter TH. Diabetic nephropathy. J Am Soc Nephrol 1997; 8: 487-493.
3. Craven PA, DeRubertis FR, Melhem M. Nitric oxide in diabetic nephropathy. Kidney Int 1997; 52: 46-53.
4. Ignarro LJ, Buga GM, Woods KS, et al. Endothelium derived relaxing factor produced and realesed from artery and vein is nitric oxide. Proc Natl Acad Sci. 1987; 84: 9265- 9269.
5. Narita I, Border WA, Ketteler M, Noble NA. Nitric oxide mediates immunologic injury to kidney mesangium in experimental glomerulonephritis. Lab Invest 1995; 72: 17- 24.
6. Sharma K, Danoff TM, DePiero A, Ziyadeh FN. Enhanced expression of inducible nitric oxide synthase in murine macrophages and glomerular mesangial cells by elevated glucose levels: Possible mediation via protein kinase C. Biochem Biophys Res Commun 1995; 207: 80-88.
7. Lewis EJ, Hunsicker LG, Bain RP, Rohde RD. The effect of angiotensin converting enzyme inhibition on diabetic nephropathy. N Engl J Med 1993; 329: 1456-1462.
8. Fischer JW, Tschope C, Reinecke A, Giachelli CM, et al. Upregulation of osteopontin expression in renal cortex of streptozocin induced diabetic rats is mediated by bradykinin. Diabetes 1998; 47: 1512-1518.

9. Shin SJ, Lee YJ, Tan MS, Hsieh TJ, et al. Increased atrial natriuretic peptide mRNA expression in the kidney of diabetic rats. Kidney Int 1997; 51: 1100-1105.
10. Haynes WG, Hand MF, Dockrell ME, Eadington DW, et al. Physiological role of nitric oxide in regulation of renal function in humans. Am J Physiol 1997; 272: 364-F371.
11. Trachtman H, Futterweit S, Pine E, Mann J, et al. Chronic diabetic nephropathy: role of inducible nitric oxide synthase. Pediatr Nephrol 2002; 17: 20-29.
12. Komers R, Lindsley JN, Oyama TT, Allison KM, et al. Role of neuronal nitric oxide synthase in the pathogenesis of renal hemodynamic changes in diabetes. Am J Physiol Renal Physiol 2000; 279: 573-583.
13. Bank NB, Aynedjian HS. Role of EDRF (nitric oxide) in diabetic renal hyperfiltration. Kidney Int 1993; 43: 1306-1312.
14. Pieper GM. Review of alterations in endothelial nitric oxide production in diabetes: protective role of arginine suplementation. Hypertension 1998; 31: 1047-1060.
15. Kone BC, Baylis C. Biosynthesis and homeostatic roles of nitric oxide in the normal kidney. Am J Physiol 1997; 272: F561-F578.
16. Mattar AL, Fujihara CK, Ribeiro MO, de Nucci G, et al. Renal effects of acute and chronic nitric oxide inhibition in experimental diabetes. Nephron 1996; 74: 136-143.
17. Trachtman H, Futterweit S, Crimmins DL. High glucose inhibits nitric oxide production in cultered rat mesangial cells. J Am Soc Nephrol 1997; 8: 1276-1282.
18. Noh H, Ha H, Yu MR, Kang SW, et al. High glucose increases inducible NO production in cultered rat mesangial cells: Possible role in fibronectin production. Nephron 2002; 90: 78-85.
19. Lee HY, Noh H, Gang J, Xu ZG, et al. Inducible nitric oxide synthase expression is increased in lipopolysaccharide- stimulated diabetic rat glomeruli: effect of ACE inhibitor and angiotensin II receptor blocker. Yonsei Med J 2002; 43: 183-192.
20. Sugimoto H, Shikata K, Wada J, Horiuchi S, et al. Advanced glycation end products-cytokine-nitric oxide sequence pathway in the development of diabetic nephro- pathy: aminoguanidine ameliorates the overexpression of tumor necrosis factor-α and inducible nitric oxide synthase in diabetic rat glomeruli. Diabetologia 1999; 42: 878-886.
21. Yates M, Rendon A, Prabhakar SS. Inducible nitric oxide synthesis in normal human mesangial cells by augmenting cellular uptake of L-arginine. Neph Dial and Transplant 2003; 18: 49.
22. Mehta JL, Chen H, Li D, Phillips IM. Modulation of myocardial SOD and iNOS during ischemia-reperfusion by antisense directed at ACE mRNA. J Mol Cell Cardiol 2000; 32: 2259-2268.
23. Onozato ML, Tojo A, Goto A, Fujita T. Effects of combination therapy with dipyridamole and quinapril in diabetic nephropathy. Diab Res Clin Prac 2003; 59: 83-92.
24. Andersen S, Tarnow L, Rossing P, Hansen BV, et al. Renoprotective effects of angiotensin II receptor blockade in type I diabetic patients with diabetic nephropathy. Kidney Int 2000; 57: 601-606.
25. Zhang H, Schmeisser A, Garlichs CD, Plotze K, et al. Angiotensin II-induced superoxide anion generation in human vascular endothelial cells: role of membrane-bound NADH-/ NADPH oxidase. Cardiovasc Res 1999; 44: 215-222.
26. Guzik TJ, West NE, Black E, McDonald D, et al. Vascular superoxide production by NAD(P)H oxidase: association with endothelial dysfunction and clinical risk factors. Circ Res 2000; 86: 85-90.
27. Schwobel J, Fischer T, Lanz B, Mohaupt M. Angiotensin II receptor subtypes determine induced NO production in rat glomerular mesangial cells. Am J Physiol Renal Physiol 2000; 279: F1092-F1100.
28. Choi KC, Lee SC, Kim SW, Kim NH, et al. Role of nitric oxide in the pathogenesis of diabetic nephropathy in streptozotocin-induced diabetic rats. Korean J Intern Med 1999; 14: 32-41.
29. Addres for correspondence
30. Kısmet Bildirici, MD
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