The Effects of High Glucose, Angiotensin II and ACE Inhibitor on the Expression of TGFbeta mRNA in Cultured Human Mesangial Cells

Kang, Min Mo; Lee, So Young; Han, Sang Yup; Guahk, Jae Young; Jo, Sang Kyoung; Yoon, Jong Woo; Cho, Won Yong; Kim, Hyoung Kyu; Ihm, Chun Gyoo; Cha, Dae Ryong

Abstract

OBJECTIVE
Diabetic nephropathy is an important cause of end stage renal disease in Korea and associated with major morbidity and mortality. The precise pathogenic mechanism of this disease is still controversial, but it has been considered that multiple factors are contribute to the development and progression of diabetic nephropathy. One of these factors, renin-angiotensin system has been proven to be a major mediator of this disease via activation of angiotensin II, which has multiple functions such as induction of production of extracellular matrix protein and various intraglomerular cells, tubulointerstital component and increment of intraglomerular pressure. Transforming growth factor(TGFbeta) is a multifunctional cytokine with major profibrotic character, which stimulates the production of extracellular matrx(ECM) protein, inhibit the degradation of ECM and induce the interaction of mesangial cells with ECM via integrin receptors. This study was done to evaluate the role of angiotensin II and angiotensin converting enzyme inhibitor in expression of TGFbeta mRNA which is a main mediator in the pathogenesis of diabetic nephropathy.
METHODS
Human mesangial cells(MCs) were cultured by standard culture techniqne. For this study, cells in the 5th to 7th passage were used. To make a different glucose concentration in culture medium, normal(100mg/dl) or high glucose(450mg/dl) concentrations of D-glucose were added, and cultured in 17% heat inactivated fetal bovine serum. Angiotensin II and ACE inhibitor(captopril) were administered to the culture medium at final concentration of 10-6M. After 72 hours, MCs were harvested to measure the expression of TGFbeta mRNA. To measure the mRNA expression of TGFbeta in each condition, semi quantitative PCR was done and all results were corrected by beta-actin gene.
RESULTS
mRNA expression of TGFbeta was significantly increased in the high glucose medium(30 mM) compared to normal glucose medium(5.5mM) (3.82+/-0.465 vs 2.27+/-0.13, p<0.05). Administration of angiotensinII(10-6M) in high glucose medium induced a further increase in the TGFbeta expression to 4.29+/-0.476(p<0.05). AngiotensinII(10-6M) in normal glucose medium also showed a significant increase in TGFbeta expression as 3.40+/-1.88(p<0.05). Administration of ACE inhibitor(Captopril, 10-6M) in high glucose medium prevented the increse of TGFbeta expression(1.20+/-0.18 vs 3.82+/-0.465, p<0.05).
CONCLUSION
From these findings, it suggest that angiotensinII is an important mediator in the pathogenesis of diabetic nephropathy. ACE inhibitor may have a role in the progress of this disease via direct suppression of TGFbeta system as well as beneficial intraglomerular hemodynamic effect.