Dominant negative form of IκB kinases 2-transfected recipient immature dendritic cells induce CD4 + CD25-T cells with tolerogenic properties

Abstract

Objective To investigate the characteristics and mechanisms of the immune tolerance of CD4 + CD25-T cells induced by recipient-derived immature dendritic cells (imDC) transfected with dominant negative form of IκB kinases 2 (IKK2dn) and loaded with donor antigens.Methods ImDC were obtained from recipient rats' (Lewis) bone marrow,transfected with IKK2dn at 200 multiplicities of infection (MOI) (200 MOI) for 2 days,loaded with donor antigens for 48 h,and cultured with Lewis (LW) lymphocytes in primary mixed lymphocyte reaction (MLR) for 72 h.Thereafter,the CD4 + CD25-T cells were purified from primary MLR and incubated in secondary coculture MLR with LW lymphocytes for another 72 h.The T lymphocyte proliferation in vitro was measured by MTT and the expression of serum interleukin (IL)-2,IL-10,transforming growth factor-β (TGF-β) and interferon (IFN)-γ was detected by using ELISA.Results Western blotting revealed high and stable expression of IKK2dn gene in imDC transfected with IKK2dn gene.In contrast to the untransfected imDC group (72.20 ±3.30)％ and the recipient Adv0 transfected imDC group (63.70 ± 1.80)％,the CD4 +T cells induced by recipient imDC transfected with IKK2dn contained a very low percentage of CD25 (18.50 ± 1.40) ％ (P ＜ 0.05).As compared with the CD4 +T cells induced by Adv0-DC (0.419 ± 0.014),the CD4 + CD25-T cells induced by Adv-IKK2dn-DC could inhibit recipient T cells proliferation (0.044 ± 0.006) (P ＜ 0.05).The CD4 + CD25-T cells were able to completely suppress the recipient T cells proliferation at high ratio (0.044 ± 0.006),but at low ratio the suppressive effect was no longer evident (0.106 ±0.006) (P ＜0.05).There was a stronger secretion of IL-10 and TGF-β (P ＜ 0.05) and a weaker secretion of IL-2 and IFN-γ(P ＜ 0.05) in the supematants in MLR.Conclusion Recipient-derived imDC transfected with IKK2dn and loaded with donor antigen could induce Lewis rat T lymphocytes into CD4 + CD25-T cells.This kind of CD4 + CD25-T cells could inhibit T-cell response and induce immune tolerance. Key words: Dendritic cells; IκB kinases 2; Lymphocyte reaction; CD4; T cells