| Background Severe war trauma, limb tumor resection, congenital malformation and other factors lead to the damaging or missing of limbs, which is a big problem in clinical orthopedics reconstruction. In particular, physiological functions of human upper limbs and hands are complex and more difficult to complete reconstruction. Traditional autologous flap transplantations not only cause secondary damage for the patients, but, are also helpless for the reconstruction of large defects. Meanwhile, limb defects seriously affect the patient’s ability to take care of themselves, and impose heavy psychosocial burdens on them. At present, vascularized osteomyocutaneous composite allotransplantation(VCA) is recognized as the best solution to solve this problem.However, since the grafts of VCA contain a large number of highly antigenic skin, nerves, muscles, bones, and other tissues, it usually requires a long-term, large doses and various types of immunosuppressive drugs to prevent acute or chronic rejection after surgery, which is followed by heavy economic burdens and serious side effects of the drugs. Furthermore, as the grafts of VCA are usually exposed within the sight of the patients, they may change the dosage of the drugs according to their own experience. These have resulted in the poor medication compliance of some patients and the unsatisfactory survival time of some grafts. Therefore, the method to induce the immune tolerance of the grafts, reduce the dosage of immunosuppressants or even completely disable immunosuppressants, decrease the incidence of rejection, prolong the survival time of grafts are still the key points and difficulties of VCA research. Currently, the liver or renal transplantation combined with bone marrow transplantation to establish the chimera is the ideal way to induce the immune tolerance of allogeneic transplant, which has acquired good results in clinical practice. However, this method is rarely applied to VCA, and their specific mechanism of action is poorly understood.In view of this, our study attempts to take the method that FK506 combined with bone marrow transplantation in miniature swine model of the VCA. We use 100 cGym body irradiation and 700 cGym thymic irradiation preoperatively to reduce the number of T cells in peripheral blood. The aim is to observe the rejection of allogenic composite tissue allografts and to explore its possible mechanism.Objective To improve and establish the miniature swine model of VCA which is suitable for the test design. Improve perfect technique of the acquisition, separation, purification, cryopreservation and revitalization of donor-derived vertebral bone marrow cells, to provide a reference for future clinical application. Before surgery, receptors received 100 cGy total body irradiation and 700 cGy thymic irradiation, and then received FK506 combined with donor bone marrow transplantation consenquently after the VCA transplantation. The survival time of VCA grafts were observed. Meanwhile, the skin of the transplanted grafts is monitored by histopathology to compare the rejection reactions of each group. The regulatory T cells proportional change of receptors is detected by flow cytometry.Methods The receptors were randomly divided into three groups: Group A(simple immunosuppressive drug group), Group B(immunosuppressive drugs combined with bone marrow transplantation), Group C(autologous heterotopic transplant as control). Preparation of modified swine groin embedded model of VCA, all receptors were underwent 100 cGy total body irradiation and 700 cGy thymic irradiation before surgery. The grafts and the general condition of receptors were observed after surgery, and the graft survival curves were plotted. On postoperative 3d, 7d, 14 d, 21 d, one month, two month, etc, skin grafts were taken for histopathological examination. During the 7th day before operative, the first month, the second month after operation, and so on, the changes of Treg ratio in peripheral blood were detected by flow cytometry.Results The grafts in group A were necrotic at postoperation day 7,9,10 due to immune rejection, The grafts in Group B survived to the postoperative day 80,91,114(P<0.05), the grafts in Group C still survived well until the date of observation. Compared with Group B, the HE staining and transmission electron microscopy observation showed that the structural of the skin was integrated, and lymphocytic infiltration was less, and immune rejection grade was significantly lower in Group B.Conclusion With all receptors underwent 100 cGy total body irradiation and 700 cGy thymic irradiation before surgery, FK506 combined with bone marrow transplants can prolong the survival time of grafts and reduce immune rejection. Graft skin biopsy is a reliable tool for diagnosis of rejection. |
PDF Full Text Request