Role of TGF-β1 in production of fibronectin in vascular smooth muscle cells cultured under high-phosphate conditions

Background: Hyperphosphatemia is associated with up-regulation of the extracellular matrix formation in vascular smooth muscle cells (VSMCs) and increased risk of cardiovascular disease. In the present study, the role of transforming growth factor-beta 1 (TGF-beta 1) in the production of fibronectin (FN) in vascular smooth muscle cells in high-phosphate environments was evaluated. Methods: Rat VSMCs were stimulated by high levels of phosphate or TGF-beta 1 in vitro. Levels of FN, TGF-beta 1, and TGF-beta 1 type I receptor (T beta RI) proteins were measured by Western blot and immunostaining. Levels of active TGF-beta 1 in the supernatant of the culture medium were detected by ELISA. Results: Production of FN was increased after VSMCs were incubated under high-phosphate conditions (2.5 mmol/L) for 12 hours. TGF-beta 1 (1 ng/mL) increased FN levels in cells as early as 3 hours after the start of treatment. Both TGF-beta 1 and T beta RI were significantly up-regulated after 3-6 hours of stimulation with high phosphate. When VSMCs were pretreated with TGF-beta 1 neutralization antibody (10 mu g/mL) for 30 minutes, induction of FN stimulated by high levels of phosphate was largely attenuated. Conclusion: TGF-beta 1 plays a critical role in the pathogenesis of high-phosphate-induced FN production in VSMCs in vitro.