Long-term exposure of proximal tubular epithelial cells to glucose induces transforming growth factor-β1 synthesis via an autocrine PDGF loop

We have recently reported increased transforming growth factor (TGF)-beta1 gene transcription in proximal tubular cells within 12 hours of exposure to 25 mmol/L D-glucose, with a requirement for a second stimulus such as platelet-derived growth factor (PDGF) to increase its translation in short-term experiments. In the current study we investigated the effect on TGF-beta1 production of prolonged exposure of proximal tubular cells to high glucose concentrations. Enzyme-linked immunosorbent assay of cell culture supernatant showed significant increase in latent TGF-beta1 only after 7 days exposure to high glucose. Radiolabeling of glucose-stimulated cells with H-3 amino acids and subsequent immunoprecipitation of TGF-beta1 demonstrated de novo synthesis from day 5 of high glucose exposure onwards. Similarly, polysome analysis showed enhanced translation of TGF-beta mRNA after 4 or more days of high glucose exposure. TGF-beta1 synthesis, following addition of glucose, was inhibited by blockade of the PDGF-a receptor subunit. Glucose did not alter PDGF expression, nor expression of PDGF a-receptors. Activation of the receptor following addition of 25 min D-glucose could be demonstrated suggesting increased sensitivity to endogenous PDGF. Exposure to glucose activated p38MAP kinase, and inhibition of this activation abrogated both glucose induced TGF-beta1 transcriptional activation and TGF-beta1 synthesis. Inhibition of p38MAP kinase did not influence the effect of exogenous PDGF when cells were stimulated sequentially by glucose and PDGF. We postulate that glucose induces an early increase in TGF-beta1 transcription via activation of p38MAP kinase. In addition, glucose causes a late increase in PDGF-dependent TGF-beta1 translation by enhancing cellular sensitivity to PDGF. This provides a potential explanation for the clinical observation that prolonged poor glycemic control may contribute to progression of diabetic nephropathy.