| Objective: Massive and complex tissue defects,caused by congenital or aquired factors,such as trauma,tumor resection,et al,are still challenging.Conventional reconstructive strategy can only cover the wound,without restoration of aesthetic and function.Composite tissue allotransplantation(CTA),defined as transplantation of tissues including different components,could provide an ideal solution to complex tissue defects.The major factor hindering the wide application of CTA is still the immunological rejetion after sugery.The life-long and high dose use of immunosuppressants can result severe complications,such as opportunistic infection,tumor,and drug toxicity.PG490 is the main active component of the Chinese herb Thunder Vine.Recently,it has been introduced to the field of transplantation,and was verified to be effective in prolonging the survival time of allotransplants when combined with conventional immunosupressants.However,its role in CTA has not been elucidated.This study will focus on the immunosuppressive effect of PG490 on CTA and its potential function of inducing tolerance via chimerism enhancement.Methods: Part 1: Transplantation of a 2cm×2cm free flap from the donor groin area based on superficial inferior epigastric artery from BN to F344.The recipients were then randomly divided into 4 groups and treated with CsA comibined with or without PG490.The mean survival time of the flap was observed,and the immunological rejection assessment and change of Treg were monitored.Also the cytokines relevant to DC maturation were evaluated post transplanation.Part 2: Establish a new CTA model of heterotopic allotransplantation of whole hind limb from BN to Lewis,and were divided into control group,in which,the rats received only vehicles,and the FK506 group,0.5mg/kg/d,i.p.Also,Lewis-to-Lewis transplantation was performed to control.FCM was applied to detect the chimerism level of recipient peripheral blood after surgery.Part 3: The established heterotopic hind limb models in part 2 were used and were further divided into 8 groups,which were treated with different doses of FK506 combined with or without single dose of PG490.The mean surivival time and the pathological classification of the transplants were evaluated.Aslo,the T lymphocyte immunosupression and inflammatory cytokines in perpheral blood were determined by FCM and ELISA.Furthermore,the mixed chimerism level,the apoptosis of transplanted bone marrow cells and the apoptotic relevant genes were detected.Results: The MST of free transplanted groin flap could be obviously prolonged by combination treatment of PG490 and CsA.And the pathological rejection response was reduced.The underline mechanism might be the regulation of Treg and inhibition of DC maturation related cytokine expression.A new heterotopic whole hind limb transplantation model was succefully established for CTA immunological rejection research.Compared to conventional orthotopic model,this model is much simpler,timesaving,lower incidence of complications,and the process of rejection was identical and stable.The chimerism could be monitored by FCM as long as the recipient accepted the graft.PG490 could decrease the required dose of FK506 and reduce the side effects of immunosupressants.Meanwhile,the expression of inflammatory cytokines was decreased and the inhibition of T cells was enhanced by PG490 combined treatment in the peripheral blood of recipient.In addition,PG490 could increase the mixed chimerism level might by inhibiting the apoptosis of bone marrow cells via regulation of bcl-2 and Bax of transplanted bones,which might contributing to the acceptance of allgrafts.Conclusion: PG490,as an adjunctive agent for immunosuppressive protocol after CTA,can reduce the required dose of conventional immunosupressants and decrease the associated side effects.Furthermore,PG490 can increase the chimerism level of recipients after limb allotransplantation,which involves vascularized bone marrow.It might contribute to the graft acceptance and donor specific tolerance induction. |
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