The immunological characteristics of CD8+ CD28- suppressor T cells induced by siRNA technique

Objective To induce CD8+ CD28- suppressor T cells(CD8+Ts)by DC with MHCⅠ expression by siRNA,under the stimulator of allograft antigen and then investigate the immunological characteristics of Ts.Methods After the isolation and cultivation of DCs from femoral bone of SI)rats.the optimal sequence of MHCⅠ siRNA was designed and constructed,and the MHC Ⅰ siRNA was transfected into DCs.The IX3s were stimulated by mesenteric lymphatic tissue fluid of Wistar rats.and then co-cultured with CD8+ Ts isolated from SD rat spleen.SD rat CD8+T cells were ohtained by magnetic separation method.Cell proliferation was assayed after SD rat spleen cells loading Wistar rat mesenteric lymph node tissue antigen reacted with different amount of Ts.After SD rat spleen cells were stimulated by Wismr rat's mesenteric lymph node and reacted with OVA respectively.different amounts of Ts were added.Gell proliferation of the spleen lymphocytes in different groups was assayed, rrIL-2 was added to the mixed lymphocyte culture system containing Wistar rat's mesenteric lymph node tissue antigen, SD rat's Ts and SD rat spleen lymphocytes, then the influence of the IL-2 on the function of SD rat Ts was observed. The expression of TGF-β and IFN-γ mRNA was examined by Real time PCR, and CD25 was detected by Real time-PCR and flow cytometry in CD8+ CD28- T cells group. Results Ts had inhibitory influence on the mixed lymphocytes proliferation between SD rat' s spleen lymphocytes and Wistar rat's mesenteric lymph node tissue, but exerted no efforts on the mixed lymphocytes proliferation between OVA and SD rat's spleen lymphocytes. After addition of rrIL-2 into the mixed lymphocyte culture system containing Wistar rat mesenterie lymph node tissue antigen and Ts, there was no significant increase in the mixed lymphocytes proliferation of SD rat spleen lymphocytes (P＞0. 05). The expression levels of TGF-βand IFN-γ mRNA were higher in CD8 + CD28- Ts group than in CD8 + CD28+ Ts group and CD8+ Ts group (P＜0. 01, P＜0. 05). The expression of CD25(IL-2R) was lower in CD8+ CD28- Ts group than in CD8 + CD28+ Ts group and CD8 + Ts group (P ＜ 0. 05). Conclusion The CD8+ CD28- suppressor T cells can be induced by DC interfered MHC-I expression under the stimulator of allograft antigen. Ts has characteristics of suppressor-effector in vitro, which cannot be reversed when the rrIL-2 exists, and the immune regulatory effect of Ts has antigen specificity. Key words: RNA interference; T-lymphocytes, suppressor-effector;  Immunosuppression