BLOCKING CELL MICROTUBULE ASSEMBLY INHIBITS THE ALLOIMMUNE RESPONSE IN-VITRO AND PROLONGS RENAL-ALLOGRAFT SURVIVAL BY INHIBITION OF TH1 ANDSPARING OF TH2 CELL-FUNCTION IN-VIVO

Colchicine inhibits cell microtubule assembly by binding to and preven ting the polymerization of tubulin monomers. Although there are data t o indicate that colchicine inhibits a variety of cell-mediated immune responses, the effects and mechanisms of inhibiting cell microtubule a ssembly on the alloimmune response have not been thoroughly investigat ed. It has recently been shown that colchicine prevents acute rejectio n and promotes the long-term survival of rat renal allografts. In this study, the effects and mechanisms of inhibiting cell microtubule asse mbly by colchicine on the alloimmune response in vitro and in vivo wer e examined. First, the effects of colchicine on T lymphocyte response to alloantigen in vitro were tested. In the standard one-way mixed lym phocyte response (MLR), responder Lewis rat lymph node cells were cult ured with irradiated Brown-Norway stimulators. Colchicine inhibited th e MLR in a dose-dependent manner, with 100% inhibition at a concentrat ion of 25 ng/mL (6.25 x 10(-8) M) and 50% inhibition at a concentratio n of approximately 5 to 10 ng/mL. Colchicine also inhibited the genera tion of cytotoxic T lymphocytes as well as cytotoxic T cell effector f unction in vitro in a dose-dependent fashion. Second, detailed immunoh istologic studies of renal allografts harvested from unmodified contro l (acutely rejecting) and colchicine-treated rats (Day 15 or 30) were performed. These studies showed that grafts from colchicine-treated an imals had significantly fewer mononuclear cell infiltrates and less ed ema, and moderately decreased deposition of immunoglobulin M, C3, and fibrin, as compared with acutely rejecting control grafts. In addition , grafts from colchicine-treated rats lacked up-regulation of activati on markers, including interleukin (IL)-2 receptor, major histocompatib ility complex class II, and intercellular adhesion molecule-1, and sho wed essentially no expression of the Th1-type activation and inflammat ory cytokines IL-2, interferon-gamma, and tumor necrosis factor-alpha, respectively. By contrast, these grafts showed markedly increased lab eling of intragraft mononuclear cells and associated endothelial cells for the Th2-related cytokine IL-4 and preservation of IL-6 expression , as compared with rejecting controls. These data indicate that blocki ng cell microtubule assembly down-regulates the alloimmune response in vitro, prevents acute rejection, and effects prolonged renal allograf t survival by the selective inhibition of Th1 and the sparing of Th2 c ell function in vivo. Understanding the mechanisms of how inhibiting c ell microtubule assembly leads to such a state of immune deviation may lead to the development of novel therapies in organ transplantation a s well as in autoimmune disease.