Experimental Study On The Induction Of Transplantation Tolerance To Cardiac Allografts By Intrathymic Inoculation And The Changes Of Cell Hemorheology

Background: The allograft heart transplantation has been developed to become an effective therapy for terminal heart disease. At present, donor heart preservation, surgical technique and the infection after transplantation have not been the major factors of the failure of transplantation. However the side effects of non-specific immune suppression and the high fee for service are all serious problems need to be settled. Induction of transplantation tolerance seems to be an ideal route to avoid this problem. As a result of the recent success of organ transplantation, the number of potential recipients for allogeneic organs has outgrown the number of available donors by more than twofold, and continues to increase. Alternatives to allogeneic organ transplantation, such as the use of xenogeneic donors, have therefore engendered considerable interest. Although current immunosuppressive protocols provide satisfactory short- and medium-term results after allotransplantation, these regimens are unlikely to be sufficient to prevent rejection of xenografts. A simple increase in the amount of non-specific immunosuppression is not a feasible option for successful xenografting. Therefore, induction of tolerance might be essential for the successful application of transplantation.The induction of transplantation tolerance by intrathymic inoculation has succeeded in heart, kidney and pancreas transplantation in rats. But in many studies, the acute rejection occurred to some extent even the induction was successful. Some researchers think that this phenomenon has relations with the survival lymphocytes of receptor, and how to eliminate these lymphocytes perhaps is the key for the induction. While, the reports about big animals are few. There are many studies in respect of immune tolerance of rodent. However, the rodent's vascular endothelial cell does not express the MHC antigen. The mechanism of acute rejection in rodents is different with human being. Considering the foreground of clinic application, the study about big animals should be paid more regard.Many researches have indicated that there were many phenomena of microcirculation obstruction occurred during acute rejection of heart transplantation, such as, erythrocyte and leucocyte assembled around the cardiac muscle and vessel endothelial cell, many erythrocytes assembled in the blood vessel, leucocyte and platelet adhered to the vasal endothelium and lacuna vasorum was narrowed and blocked, etc. These phenomena implied that the turbulence of cell hemorheology maybe play an important role in the course of acute rejection after heart transplantation. It is not clear of the action mechanism and the relation between the cell hemorheology and acute rejection.So, we chose the pigs as experiment objects. Firstly, the recipient pigs were pretreated with x-ray whole body irradiation. And then, induced the allogenic cardiac transplantation tolerance by intrathymic inoculation of donor spleen cells of pigs, and activated the survival lymphocytes of recipient apoptosis by mainline the donor lymphocytes later. Observe the survival days and detect the mixed lymphocyte reaction (MLR). To search for the possibility to induce allogenic cardiac transplantation tolerance in pigs by intrathymic inoculation of donor spleen cells and mainline the donor lymphocytes. While, observe the changes of cell hemorheology after heart transplantation and investigate the relationship between acute rejection and cell hemorheology.The whole work consists of three parts as follows:Part one: Establishment of a pig model of abdominal heterotopic heart transplantation. According to Ono and Cheng's model, we made some technique improvements. Heterotopic heart was transplanted using an end-to-side anastomosis between the donor's pulmonary artery and the recipient's abdominal inferior vena cava, and between the donor's aorta and the recipient's abdominal aorta. Some technical problems were discussed.Part two: The recipient pigs were pretreated with x-ray whole body irradiation. To search for the possibility to induce allogenic cardiac transplantation tolerance in pigs by intrathymic inoculation of donor spleen cells and to activate the survival lymphocytes of recipient apoptosis by mainline the donor lymphocytes. Animals were divided into 4 groups, in which the blank group was without any treatment except the allogenic cardiac transplantation, control group was pretreated with 2 Gy x-ray irradiation, experimental group A was pretreated with whole body irradiation and inoculated intrathymicaUy with donor spleen cells, and experimental group B was mainlined the donor lymphocytes seven days after inoculated intrathymically. In these three pretreatment groups, the changes of the peripheral blood lymphocytes and mixed lymphocyte reaction (MLR) were observed to test any alterations of immune functions. The allogenic cardiac transplantation was performed 14 days after pretreatment, and transplant survival times was observed. It was found that after 48 hours of irradiation, the number of lymphocytes was significantly decreased in comparison with that before irradiation. In the experimental group 2 weeks after intrathymic inoculation of spleen cells in recipient pigs, the stimulatory effect in MLR showed a tendency to reduce, while that with single irradiation showed no difference before and after pretreatment. The survival time of the donor cardiac grafts in the experimental group B (MST 9.6 days) was clearly prolonged in comparison with those of blank group (MST 4.2 days), control group (MST 4. 6 days) and experimental group A (MST 7.6 days). And there was no significant difference between blank and single irradiation group. These experimental results indicate that the intrathymic inoculation with donor spleen cells and pretreatment with whole body irradiation of the recipient out bred pigs could induce a transient immune suppression, and mainline the donor lymphocytes seven days after inoculated intrathymically could intensify this immune suppression, but not attain to the so called permanent immune tolerance as observed in small animals.Part three: The erythrocyte deformability index (EDI), erythrocyte aggregation index (EAI), and leucocyte deformability index (LDI) were measured at before irradiation, after irradiation 1,7,14d, and post transplantation 1,3,5,7 and 9d. It was found that the LDI of control group and experimental group (A,B) was decreased at 1st day after irradiation, and recovered normal quickly at 7st day after irradiation. The EAI of blank group and control group was significantly increased at 1st and 3th day, while the EDI and LDI were significantly decreased. The EAI of experimental group A was significantly increased at 5st and 7th day, while the EDI and LDI were significantly decreased. The EAI of experimental group B was significantly increased at 9st day, while the EDI and LDI were significantly decreased. These experimental results indicate that pretreatment with whole body irradiation of the recipient out bred pigs could induce a transient LDI decline. The EAI, EDI and LDI have no significant changes during lighter rejection, but they are exacerbated significantly when a more severe rejection occurs.(The software program SPSS13.0 was sed to deal with the experiment data)...