A novel biological assay to detect the active form of TGF-β in urine to monitor renal allograft rejection

Abstract

Background. Transforming growth factor-β (TGF-β) plays an important role in renal fibrosis. Measurement of the concentration of the active form of TGF-β particularly in urine may help our understanding of the mechanism of chronic allograft nephropathy and could be used as a diagnostic tool. However, TGF-β release and activation are complex and, consequently, there is currently no accurate way to measure TGF-β activity. Methods. TGF-β-sensitive BL41 cells were stably transfected with a reporter plasmid harboring a synthetic TGF-β-inducible DNA sequence upstream from the luciferase gene. Cells were incubated with urine samples from normal donors or transplanted recipients with or without patent nephropathy, and the active form of TGF-β was determined as luciferase activity. Results. We have established a cell line which expresses luciferase activity in response to active TGF-β in a dose-dependent manner. Moreover, the use of a histone deacetylase inhibitor greatly increased sensitivity to TGF-β and also stabilized luciferase inductibility. This test is highly specific to active TGF-β. Detectable levels of TGF-β were found in urine from patients with renal dysfunction due to acute or chronic renal allograft rejection (P < 0.001), but not in that from patients with stable, correctly functional kidneys. Conclusion. We describe a highly sensitive and specific assay for active TGF-β. We also show that, in cases of renal allograft, TGF-β expression is highly and significantly correlated with acute or chronic rejections. © 2005 by the International Society of Nephrology.