Dendritic cells and the 60kDa heat shock protein (Hsp60): strategies for immunoregulation in the mouse immune system / Células dendríticas e a proteína de choque térmico 60KDA (HSP60): estratégias para imunorregulação do sistema imune murino

AUTOR(ES)
DATA DE PUBLICAÇÃO

2008

RESUMO

The induction of tolerance to allotransplant without the help of immunosupressive drugs is one of the major challenges in immunology. Considering the tolerogenic potential of dendritic cells and the immunoregulatory activity of Hsp60, our objective was to determine the capacity of dendritic cells (DCs) of inducing immunoregulation through the interaction with Hsp60, in vitro, and tolerance to the skin allograft, in the murine system. For this, we have generated three types of DCs derived from bone marrow of Balb/c mice: immature DCs (iDCs and IL-10 iDCs treated with IL-10) and mature DCs (mDCs). The DCs were characterized as to their: (i) morphology, (ii) immunophenotype, (iii) immature phenotype stability; (iv) spontaneous cytokine production; (v) induction of allogeneic LT proliferation. The three types of DCs generated in this work (mDCs, iDCs and IL-10 iDCs) have presented immunophenotypic and functional differences in relation to the stimulation of allogeneic proliferative response. The mDCs presented the highest expression of coestimulatory molecules and the strongest induction of allogeneic T lymphocyte proliferation. The IL-10 iDCs presented stability of the immature phenotype after LPS challenge and showed low capacity of inducting allogeneic TL proliferation. We did not observe a differential profile in relation to the spontaneous cytokine production by all three types of DCs in the period analyzed. However, TNF- was the most frequent cytokine detected in the three types of DCs, especially in the mDCs, while IL-6 was mostly produced by mDCs, and IL-10 by IL-10 iDCs. After the characterization of the different DCs, these cells were treated with the fragments of Hsp60 for 24 hours. We evaluated if these fragments had the capacity of modifying in the DCs: (i) the expression of coestimulatory molecules; (ii) the production of cytokines; (iii) the capacity of inducing and inhibiting proliferation and cytokine in autologous cocultures; (iv) the capacity of inhibiting proliferation and cytokine production in response to CD3 stimulation; (v) the capacity of inducing tolerance to skin allotransplantation in mice by the injection of DCs treated with fragments of Hsp60 or by the injection of the antibody DEC205-N3. The interaction of Hsp60 fragments with the different DCs induced a modification in the expression of coestimulatory molecules and in the production of cytokines. Some peptides stood out as predominately inductors of immunoregulatory cytokines (IL-10 and TGF-; peptides p277 and N7), and others as predominately inductors of pro-inflammatory cytokines (TNF-, IFN- and IL-12; peptides I8 and I2). In addition, the peptide N7 was the greatest inhibitor of the expression of coestimulatory molecules, while the protein CHsp60 at times increased, and at times decreased this expression. The peptide N7 presented a dominant effect in the inhibition of autoreactivity of TLs directed to the DCs, both in proliferative response and in the production of cytokines, especially inflammatory ones. Some fragments of Hsp60 (C-Hsp60, p277 and especially the peptide N7) were capable of inhibiting proliferation and production of inflammatory cytokines in cocultures of TLs with DCs stimulated with CD3 antibody. Despite the dual functional activity of Hsp60 in the interaction with DCs, in vitro, in this work we observed a predominance of immunoregulation. We highlight that the peptide N7 presented the most immunoregulatory profile. Our data suggest the involvement of multiple mechanisms of action for the immunoregulatory activity of Hsp60 in the interaction with DCs. Although our protocols have not induced tolerance to the skin allograft in mice, we have observed that the animals injected with the IL-10 iDCS treated either with p277 or N7 presented increased allograft survival (16 and 17 days versus 14 days). Therefore, we believe that these protocols for tolerance induction can be optimized for the use in murine models, aiming future applications in the clinic in transplants and auto-immune diseases

ASSUNTO(S)

células dendríticas tolerância hsp60 hsp60 Îmunorregulação tolerance transplante dendritic cells transplantation imunoregulation dec205 dec205 peptides peptídeos

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