Attenuation of the Expression of Fibrogenic Molecules by Transforming Growth Factor-beta1 Antisense in Cultured Rat Mesangial Cells.
- Author:
Gyu Tae SHIN
1
;
Heungsoo KIM
Author Information
1. Department of Nephrology, Ajou University School of Medicine, Suwon, Korea. gtshin@ajou.ac.kr
- Publication Type:In Vitro ; Original Article
- Keywords:
Adenoviral vector;
TGF-beta1 antisense;
Mesangial cells
- MeSH:
Adenoviridae;
Animals;
Collagen;
Cytokines;
Cytomegalovirus;
Diabetic Nephropathies;
DNA;
Extracellular Matrix;
Fibronectins;
Fibrosis;
Gene Expression;
Genetic Vectors;
Glucose;
Kidney;
Mesangial Cells*;
Models, Animal;
Polymerase Chain Reaction;
Rats*;
Renal Insufficiency, Chronic;
RNA, Messenger;
Transforming Growth Factor beta1
- From:Korean Journal of Nephrology
2003;22(4):349-357
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
BACKGROUND: The diabetic milieu augments the expression of transforming growth factor-beta1 (TGF-beta1) in mesangial cells, which, in turn, stimulates the accumulation of extracellular matrix in glomeruli. Accordingly, mesangial cells have been shown to produce increased amount of TGF-beta1 when exposed to high glucose concentrations in vitro. METHODS: In the present study, we examined the effects of antisense TGF-beta1 gene transfer on the expressions of several cytokines implicated in kidney fibrosis. The DNA fragment containing bases -162 to +168 of rat TGF-beta1 sequence was isolated by reverse transcription-polymerase chain reaction (RT- PCR) from rat kidney tissue. This PCR product was ligated in the xbaI-KpnI restriction sites of a shuttle vector in forward and reverse orientations and replication defective recombinant adenoviral vectors containing either the sense or antisense TGF-beta1 genes under the control of a cytomegalovirus promoter - rAdTGFbeta1S and rAdTGFbeta1AS, respectively - were generated. Rat mesangial cells infected with recombinant adenoviruses were grown in normal (NG, 100 mg/dL) or high glucose (HG, 450 mg/dL) media for six days, and mRNA expressions of the monolayers were evaluated by RT-PCR. RESULTS: An adenoviral vector containing the antisense TGF-beta1 gene significantly downregulated the gene expressions of TGF-beta1, collagen, fibronectin and PDGF-B in cultured rat mesangial cells. CONCLUSION: The expression of fibrogenic molecules were significantly attenuated by adenoviral vectors with antisense TGF-beta1, and such results should be confirmed in the future study of animal models of diabetic nephropathy or chronic kidney diseases.
