A novel biological assay to detect the active form of TGF-beta in urine to monitor renal allograft rejection.
BACKGROUND
Transforming growth factor-beta (TGF-beta) plays an important role in renal fibrosis. Measurement of the concentration of the active form of TGF-beta particularly in urine may help our understanding of the mechanism of chronic allograft nephropathy and could be used as a diagnostic tool. However, TGF-beta release and activation are complex and, consequently, there is currently no accurate way to measure TGF-beta activity.


METHODS
TGF-beta-sensitive BL41 cells were stably transfected with a reporter plasmid harboring a synthetic TGF-beta-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-beta was determined as luciferase activity.


RESULTS
We have established a cell line which expresses luciferase activity in response to active TGF-beta in a dose-dependent manner. Moreover, the use of a histone deacetylase inhibitor greatly increased sensitivity to TGF-beta and also stabilized luciferase inductibility. This test is highly specific to active TGF-beta. Detectable levels of TGF-beta 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.


CONCLUSIONS
We describe a highly sensitive and specific assay for active TGF-beta. We also show that, in cases of renal allograft, TGF-beta expression is highly and significantly correlated with acute or chronic rejections.
Rogier
Edith
E
INSERM Unité 542, Hôpital Paul Brousse, Villejuif, France.
Durrbach
Antoine
A
Abecassis
Lucile
L
Ferlicot
Sophie
S
Snanoudj
Renaud
R
Baudreuil
Severine
S
Arzouk
Nadia
N
Vazquez
Aime
A
Charpentier
Bernard
B
Bourgeade
Marie-Françoise
MF
D000328:Adult; D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002460:Cell Line; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005069:Evaluation Studies as Topic; D015870:Gene Expression / Q000187:drug effects; D006084:Graft Rejection / Q000175:diagnosis* / Q000652:urine*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007668:Kidney / Q000166:cytology; D007674:Kidney Diseases / Q000175:diagnosis / Q000652:urine; D016030:Kidney Transplantation; D008156:Luciferases / Q000235:genetics; D008875:Middle Aged; D012680:Sensitivity and Specificity; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000652:urine*; D014184:Transplantation, Homologous
D004791:Enzyme Inhibitors; D006877:Hydroxamic Acids; D016212:Transforming Growth Factor beta; C012589:trichostatin A; D008156:Luciferases