*Purpose: To elucidate the mechanism of cytotoxicity in FK506-treated Jurkat T cells, signal transduction pathway of TNF-related events was studied.

*Methods: Viability of Jurkat T cells was measure by MTT assay. The catalytic activation of caspase-3 and caspase-9 proteases was determined by digestion of fluorogenic biosubstrates and western blot with anti-caspase-3 and anti-caspase-9 antibodies. The levels of m-RNA and proteins for p53, Bax, PUMA, Proline oxidase, TRAIL(TNF- related apoptosis inducing ligand), TRAIL-R1(DR 4), TRAIL-R2(DR 5), Fas, Fas-L, TNF-α , IL-6, and NF-κB were measured by RT-PCR and western blot with specific antibodies. Also we further examined the localization of TRAIL- family proteins using by fluorescent microscope with specific TRAIL- family antibodies.

*Results: FK 506 decreased the viability of Jurkat T cells concentration- and time-dependently along with catalytic activation of caspase-3 and caspase-9, p53 phosphorylation, and changes in expression levels of Bax, PUMA, and Proline oxidase protein. It caused an increase in expression of TRAIL, TRAIL-R1(DR 4), TRAIL-R2(DR 5), Fas, and Fas-L in the levels of m-RNA and proteins of Jurkat T cells. Furthermore, FK 506 increased extracellular release of TNF-α and IL-6 cytokines in Jurkat T cells. It also induced the transactivation of NF-κB through the dephosphrylation of Ser 486 residues in Jurkat t cells.

*Conclusions: These results suggest that FK 506 induces apoptotic death of Jurkat cells through activation of caspase family protease, Bcl-2 family protein-related mitochondrial dysfunction, activation of TNF-related death-receptors.

« Back to 2020 American Transplant Congress