LONGTERM CARDIAC ALLOGRAFT SURVIVAL IN VAV1 KNOCKOUT MICE

Abstract

O331* Aims: T cells are key players in the immune response leading to graft rejection. The guanine nucleotide exchange factor Vav1 mediates TCR and CD28-triggered activation and proliferation of T lymphocytes by activating small GTPases such as Rac1. To assess the involvement of Vav1 in graft rejection, we performed heart transplantation experiments in Vav1 knockout (ko) mice. Moreover, immune responses in Vav1-deficient mice were characterized in vivo and in vitro. Methods: BALB/c cardiac allografts are transplanted heterotopically into Vav1 ko C57B/6 recipient mice. Graft survival is monitored by palpation of heart beat. At the end of the study graft histology is performed. The effect of Vav1 deficiency on T cell-dependent antibody formation is assessed by i.p. immunization with DNP-KLH. Ex vivo immune assays include one-way mixed lymphocyte reaction (MLR) with irradiated BALB/c splenocytes as stimulators as well as FACS-based analysis of activation marker expression on lymphocyte subsets in whole blood and spleen. Results: In the absence of immunosuppressive treatment, the median survival time of cardiac allografts in Vav1-deficient recipient mice was more than 100 days as compared to 7 days in WT recipients. Terminal immunohistological evaluation of grafts revealed scattered infiltrates of T and B cells as well as of perivascular macrophages which was associated with signs of remodeling of vessels in grafts surviving 100 days post transplantation. In the DNP-KLH mouse model the anti-DNP titer is largely suppressed by cyclosporine. In contrast Vav1-deficient mice are capable of T cell-dependent antibody formation indicating that Vav1-dependent and CsA-inhibited pathways are different. In the one-way MLR assay the proliferation was markedly decreased using various ratios of BALB/c and Vav1-deficient splenocytes. Stimulation of Vav1-deficient lymphocytes from spleen and whole blood with ConA, anti-CD3 or anti-CD3/anti-CD28 resulted in markedly reduced expression of the activation marker CD69 as compared to WT cells. This defect was observed with CD4+, CD8+ and B220+ lymphocytes. Conclusions: We show for the first time that absence of Vav1 in recipient mice prevents allograft rejection. Vav1 deficiency was associated with defects in T cell activation.