Optimal T-cell response requires two signals, the TCR signal provided by antigen-MHC complex as well as costimulatory signals provided by costimulatory molecules expression on APC. To investigate the antigen-presenting function of IKK2dn-transfected DC, a mixed lymphocyte reaction

was preformed by co-culturing different number of MMC-treated Adv-IKK2dn-infected Lewis DC and fixed number (1 × 106) of BN T cells, selleck products using MMC-treated uninfected Lewis DC and control virus-infected Lewis DC as controls. T-cell proliferation was measured by MTT assay, and results are presented as stimulation index. Results indicated that different Adv-IKK2 infection could significantly suppress Lewis DC-induced BN T-cell proliferation (Fig. 3A). DC infected by over 50 MOI Adv-IKK2 are compatible with uninfected immature DC in terms of their capacity to stimulate allogenic T-cell proliferation. These results also indicated that 50 MOI Adv-IKK2 infection is sufficient to inhibit DC maturation and suppress their ability to stimulate alloreactive T-cell proliferation. Further, we used 50 MOI Adv-IKK2dn-infected Lewis DC loaded with BN antigen PI3K inhibitor and studied their ability to stimulate Lewis T-cell proliferation, without alloantigen-loaded

IKK2dn-transfected DC, uninfected immature DC with or without alloantigen loaded were used as controls. Results indicated that IKK2dn transfection significantly suppressed the ability of alloantigen-loaded DC-induced syngeneic T-cell proliferation (Fig. 3B). To understand the mechanism of IKK2dn transfection suppressed alloreactive T-cell proliferation, we tested the cytokine production in the supernatant of the mixed lymphocyte cultures.

We found that the IL-10 production was markedly increased in Adv-IKK2dn-DC co-cultured group in comparison with uninfected and control virus-infected DC co-cultured groups. In contrast, the IFNγ production was significantly lower in Adv-IKK2dn-infected DC and uninfected DC co-cultured however groups than control virus-infected group; there is no statistical difference between Adv-IKK2dn-DC and uninfected immature DC groups in terms of their IFNγ production (Fig. 3C,D). In vitro studies indicated that Adv-IKK2dn-infected DC have the potential to suppress anti-alloimmune response. To investigate whether IKK2dn-DC had a tolerogenic potential in vivo, 1 × 107 uninfected immature DC, Adv-IKK2dn-DC, and AdV-0-DC from LW rats loaded with BN antigen were infused into naive LW rats 7 days before kidney transplantation, and no immunosuppressive drugs were used during the study. Their survival was monitored everyday after transplantation. Results indicated that in Adv-IKK2dn-DC-treated group the survival time was prolonged significantly in comparison with untreated, uninfected DC treated, and Adv-0-DC treated, as well as Wister groups (Fig. 4). The detailed rat number and survival time in each group were described in Table 1.