Cultured tubule cells from TGF-β1 null mice exhibit impaired hypertrophy and fibronectin expression in high glucose

Background. To firmly establish the role of the transforming growth factor-β1 (TGF-β1) isoform in the pathophysiology of diabetic tubulointerstitial hypertrophy and fibrosis, we examined how the total absence of TGF-β1 would alter the effect of high glucose on cellular hypertrophy and matrix expression in tubuloepithelial cells cultured from TGF-β1 null mice. Methods. Primary tubule cell cultures, obtained from kidneys of TGF-β1 knockout mice and their wild-type littermates, were treated with exogenous TGF-β1 or high glucose. The TGF-β system was characterized at the ligand and receptor levels using Northern and Western blotting. Cellular hypertrophy and growth were assessed by thymidine incorporation, cell counting, leucine incorporation, and protein content. Fibronectin expression was assessed by Northern analysis and enzyme-linked immunosorbent assay (ELISA). Results. Knockout cells did not express TGF-β1 but did express TGF-β2, TGF-β3, and TGF-β type I and type II receptors. Exogenous TGF-β1 down-regulated the ligand-binding type II receptor but up-regulated type I receptor expression. Knockout cells proliferated more rapidly than wild-type cells, but restoring TGF-β1 to knockout cells slowed their proliferation. In wildtype cells, high glucose caused cellular hypertrophy, evidenced by greater leucine incorporation and protein content along with decreased thymidine incorporation. High glucose also increased fibronectin message and protein. However, in knockout cells, high glucose failed to induce hypertrophy and was severely limited in its capacity to stimulate fibronectin. Conclusion. In tubular epithelial cells, TGF-β1 mediates the hypertrophic and fibronectin-stimulatory effects of high glucose, confirming the role of the TGF-β1 isoform in the pathogenesis of diabetic tubular hypertrophy and fibronectin overexpression.