Diabetes Metab J.  2011 Apr;35(2):130-137. 10.4093/dmj.2011.35.2.130.

Effects of Spironolactone and Losartan on Diabetic Nephropathy in a Type 2 Diabetic Rat Model

• Lee MY
• Shim MS
• Kim BH
• Hong SW
• Choi R
• Lee EY
• Nam SM
• Kim GW
• Shin JY
• Shin YG
• Chung CH

• KMID: 2281711
• DOI: http://doi.org/10.4093/dmj.2011.35.2.130

Abstract

BACKGROUND
While there is an evidence that the anti-inflammatory properties of spironolactone can attenuate proteinuria in type 2 diabetes, its effects on vascular endothelial growth factor (VEGF) expression in diabetic nephropathy have not been clearly defined. In this study, we examined the effects of spironolactone, losartan, and a combination of these two drugs on albuminuria, renal VEGF expression, and inflammatory and oxidative stress markers in a type 2 diabetic rat model.
METHODS
Thirty-three Otsuka-Long-Evans-Tokushima-Fatty (OLETF) rats were divided into four groups and treated with different medication regimens from weeks 25 to 50; OLETF diabetic controls (n=5), spironolactone-treated (n=10), losartan-treated (n=9), and combination of spironolactone- and losartan-treated (n=9).
RESULTS
At week 50, the albumin-to-creatinine ratio was significantly decreased in the losartan and combination groups compared to the control OLETF group. No decrease was detected in the spironolactone group. There was a significant reduction in renal VEGF, transforming growth factor (TGF)-beta, and type IV collagen mRNA levels in the spironolactone- and combination regimen-treated groups. Twenty-four hour urine monocyte chemotactic protein-1 levels were comparable in all four groups but did show a decreasing trend in the losartan and combination regimen groups. Twenty-four hour urine malondialdehyde levels were significantly decreased in the spironolactone- and combination regimen-treated groups.
CONCLUSION
These results suggest that losartan alone and a combined regimen of spironolactone and losartan could ameliorate albuninuria by reducing renal VEGF expression. Also, simultaneous treatment with spironolactone and losartan may have protective effects against diabetic nephropathy by decreasing TGF-beta and type IV collagen expression and by reducing oxidative stress in a type 2 diabetic rat model.

Keyword

Diabetic nephropathy; Losartan; Spironolactone; Vascular endothelial growth factor

MeSH Terms

Albuminuria
Animals
Chemokine CCL2
Collagen Type IV
Diabetic Nephropathies
Losartan
Malondialdehyde
Oxidative Stress
Proteinuria
Rats
RNA, Messenger
Spironolactone
Transforming Growth Factor beta
Transforming Growth Factors
Vascular Endothelial Growth Factor A
Chemokine CCL2
Collagen Type IV
Losartan
Malondialdehyde
RNA, Messenger
Spironolactone
Transforming Growth Factor beta
Transforming Growth Factors
Vascular Endothelial Growth Factor A

Figure

• 

  Fig. 1 PAS staining of glomeruli (A) and glomerular matrix indices (GMI) (B). PAS staining of glomeruli showed marked mesangial expansion and sclerosis in the CO group compared to other groups. Compared with the CO group, GMI scores were significantly decreased in the SPR, LO, and COM groups. CO, control Otsuka-Long-Evans-Tokushima-Fatty (OLETF) group; SPR, spironolactone-treated OLETF group; LO, losartan-treated OLETF group; COM, spironolactone- and losartan-treated OLETF group. aP<0.05 compared with CO group, bP<0.05 compared with SPR group, cP<0.05 compared with LO group. Scale bar, 100 µm, ×400.

• 

  Fig. 2 Analysis of renal vascular endothelial growth factor (VEGF) expression. (A) Optical densities of immunohistochemical staining for VEGF. (B) VEGF mRNA expression by real-time RT-PCR, and (C) Western blot for renal VEGF expression. Optical densities of glomerular VEGF in the LO and COM groups were significantly lower than those of the CO and SPR groups. Quantitative analysis revealed that expression of VEGF mRNA was 0.35-fold in the SPR group, 0.67-fold in the LO group, and 0.22-fold in the COM group when compared with the CO group. Western blot did not show any differences among groups. CO, control Otsuka-Long-Evans-Tokushima-Fatty (OLETF) group; SPR, spironolactone-treated OLETF group; LO, losartan-treated OLETF group; COM, spironolactone- and losartan-treated OLETF group. aP<0.05 compared with CO group.

• 

  Fig. 3 Transforming growth factor (TGF)-β and collagen type IV mRNA expression by real time RT-PCR. Compared with the CO and LO groups, TGF-β and collagen type IV mRNA expression of the SPR and COM groups were significantly decreased. CO, control Otsuka-Long-Evans-Tokushima-Fatty (OLETF) group; SPR, spironolactone-treated OLETF group; LO, losartan-treated OLETF group; COM, spironolactone- and losartan-treated OLETF group. aP<0.05 compared with CO group, bP<0.05 compared with LO group.

• 

  Fig. 4 Twenty-four hour urinary monocyte chemotactic protein- 1 (MCP-1) (A) and malondialdehyde (MDA) (B) levels. MCP-1 levels were not different among the four groups, but showed a decreasing trend in the LO and COM groups. MDA levels were significantly decreased in the COM group compared to the CO group. CO, control Otsuka-Long-Evans-Tokushima-Fatty (OLETF) group; SPR, spironolactone-treated OLETF group; LO, losartan-treated OLETF group; COM, spironolactone- and losartan-treated OLETF group. aP<0.05 compared with CO group.

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