J Korean Diabetes Assoc.  2007 May;31(3):220-229. 10.4093/jkda.2007.31.3.220.

Effects of Troglitazone on the Expression of VEGF and TGF-beta in Cultured Rat Mesangial Cells

• Kim DL
• Kim NH
• Choi DS

• KMID: 1523010
• DOI: http://doi.org/10.4093/jkda.2007.31.3.220

Abstract

BACKGROUND: Clinical study reported that troglitazone ameliorated microalbuminuria in diabetic nephropathy. However, the mechanism of action is not fully understood. Vascular endothelial growth factor (VEGF) is known as vascular permeability factor and it is considered the most likely cause of glomerular hyperfiltration and proteinuria in diabetic nephropathy. Transforming growth factor-beta (TGF-beta) is a potent inducer of extracellular matrix production and fibrosis in renal cells and one of the important cytokine in the pathogenesis of diabetic nephropathy. To determine whether troglitazone affects VEGF and TGF-beta production in diabetic nephropathy, we examined the effects of troglitazone on the VEGF and TGF-beta expression in cultured rat mesangial cells exposed to high glucose concentration.
METHODS
Rat mesangial cells were cultured in media with D-glucose 5.5 mM (NG) or D-glucose 30 mM (HG), or D-glucose 30 mM/troglitazone 20 micrometer(HTz) and for 6, 24, or 72 hours, respectively. VEGF and TGF-beta expression were assessed by semiquantitative RT-PCR and western blot analysis.
RESULTS
Troglitazone decreased the VEGF164 and VEGF120 mRNA expressions in cultured rat mesangial cells exposed to high glucose concentration with incubation for 24 and 72 hours, respectively. VEGF protein was also decreased in experimental group treated with troglitazone (HTz) than in those with HG for 24 and 72 hours. However troglitazone had no effect on the expression of TGF-beta mRNA in mesangial cells.
CONCLUSION
This study suggested that troglitazone may modulate the development and progression of diabetic nephropathy by reducing the expression of VEGF in mesangial cells

Keyword

Diabetic nephropathy; Mesangial cell; TGF-beta; VEGF

MeSH Terms

Animals
Blotting, Western
Diabetic Nephropathies
Extracellular Matrix
Fibrosis
Glucose
Mesangial Cells*
Proteinuria
Rats*
RNA, Messenger
Transforming Growth Factor beta*
Vascular Endothelial Growth Factor A*
Glucose
RNA, Messenger
Transforming Growth Factor beta
Vascular Endothelial Growth Factor A

Figure

• 

  Fig. 1 The effects of troglitazone on the VEGF mRNA expression under high glucose concentration. Three alternative splicing variants encoding peptides VEGF120, VEGF164, and VEGF188 were identified. VEGF120 and VEGF164 were expressed more highly than VEGF188, which was faintly expressed. VEGF mRNA expression was increased in HG 6, 24 and 72 hour groups than NG group. Troglitazone decreased the VEGF mRNA expression in high glucose exposed mesangial cells with incubation for 24 and 72 hours, respectively. NG, normal glucose concentration of media (5.5 mM); HG, high glucose concentration (30 mM); HTz, high glucose concentration treated with troglitazone (glucose 30 mM and troglitazone 20 µM); Mann, mannitol.

• 

  Fig. 2 The ratio of the expression of VEGF164/β-actin was increased in HG 6, 24 and 72 hour groups than NG group. VEGF mRNA expression was significantly reduced in troglitazone treated groups than HG groups for 24 and 72 hours, respectively. *P < 0.05.

• 

  Fig. 3 The ratio of the expression of VEGF120/β-actin was increased in HG 6, 24, 72 hour groups than NG group, and decreased in HTz 24 and 72 hours than HG 24 and 72 hour groups, respectively. *P < 0.05.

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  Fig. 4 The effect of troglitazone on the TGF-β mRNA expression according to incubation time and glucose concentration of the media. NG, normal glucose concentration of media (5.5 mM); HG, high glucose concentration of media (30 mM); HTz, high glucose concentration of media treated with troglitazone (glucose 30 mM and troglitazone 20 µM); Mann, mannitol.

• 

  Fig. 5 The ratio of the expression of TGF-β/β-actin was increased in HG 6, 24 and 72 hours than NG group. Troglitazone had no effect on the expression of TGF-β mRNA in mesangial cells. *P < 0.05.

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  Fig. 6 Western blot analysis of VEGF protein production. NG, normal glucose concentration of media (5.5 mM), HG; high glucose concentration (30 mM); HTz, high glucose concentration treated with troglitazone (glucose 30 mM and troglitazone 20 µM); Mann, mannitol.

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