The Experimental Studies Of Combined Application Of CTLA4-Ig And Infusion Of Donor Bone Marrow Cells Induce Immunotolerance To Murine Cardiac Allograft

Objective:To explore whether combined application of CTLA4-Ig and infusion of donor bone marrow cells can induce immune tolerance to murine cardiac allograft and its mechanism. Methods: 1,Developing murine heterotopic heart transplantation model.SPF mice were used as recipients and donors in the experiments.2%Chloral Hydrate was used as the anaesthetic.Surgical procedures was performed under 16×magnification.The donor hearts remained ascending aorta and pulmonary artery,which were cut before their bifurcation.Other blood vessels of the donor hearts were ligated.The aorta of the donor was anastomosed to the recipient abdominal aorta by end-to side fashion with 11-0 sutures.The pulmonary artery was also anastomosed to the recipient inferior vena cava by the same way.The blood circulating way of the donor heart is: abdominal aorta of the recipient→aorta of the donor→coronary artery of the donor→coronary vena of the donor→right atria of the donor→right ventricle of the donor→pulmonary artery of the donor→inferior vena cavaof the recipient. 2,Grouping of the experimental animals. The experimental animals were classified four groups. The recipients were infused donor bone marrow cells or PBS through the portal vein intra operation,and injected CTLA4-Ig or PBS into abdominal cavity 48 hours postoperation. Group A:14 cases were infused PBS 1 ml.through the portal vein in the operation, and infused PBS 0.3 ml into abdominal cavity 48 hours postoperation. Group B: 14 cases were infused 3×10~7 donor bone marrow cells ,and injected CTLA4-Ig 0.15 mg. Group C: 14 cases were infused PBS 1 ml, and injected PBS 0.3 ml into abdominal cavity 48 hours postoperation. Group D: 14 cases were infused 3×10~7 donor bone marrow cells, and injected CTLA4-Ig 0.15 mg. 3,Treatment after heart transplantation. ⑴Touched the heart beating to confirm whether the donor hearts were still alive every day after heart transplantation. Registered the survive time of the donor hearts. ⑵Five cases from every group were killed 8 days after heart transplantation.To investigate histopathology of the donor hearts allografts. ⑶Observed the Delayed Type Hypersensitivity of the recipients to the antigen from the third kind of mouse 24 days after heart transplantation. ⑷Observed the Delayed Type Hypersensitivity of the recipients to the antigen from the donor 41 days after heart transplantation. ⑸Skin graft between the donor mice and the recipient mice were engaged 48 days after heart transplantation.observe the survive time of the grafted skin,and graded the grafted skin every 7 days . Results: 1,56 cases heterotopic heart transplantation were performed.50 cases were successfully performed.Successful rate of surgery was 89.28%.The operation condition among four groups was similar. 2,Survive time of mice's heart allografts.Group A: 9.0±1.4 days;Group B: 16.8±1.8 days.Group C: 9.7±2.1 days.Group D: 79.6±23.7 days.The survive time of mice's heart allografts in group D was the longest.There were significantly difference between group D and group A,group B,group C,respectively(P<0.01). 3,Histopathology changes:Cardiac allografts from group A and C developed severe cellular rejection with severe infiltrating lymphocytes,myocyte injury and necrosis,interstitial edema and hemorrhage,which had acute rejection of grade Ⅲor Ⅳ. Cardiac allografts from group B had acute rejection of grade Ⅱor Ⅲ. Cardiac allografts from group D were characterized by sparse lymphocytes infiltration,mild endocarditis and vasculitis and preserved myocardial architecture,which had acute rejection of grade Ⅰ. There were significantly difference between group D and group A,group C,respectively(P<0.01).4,The Delayed Type Hypersensitivity of the recipients to the antigen from the thrd kind of mouse:Every recipient had normal response to the antigen from the thrd kind of mouse. There were not significantly among four groups. 5,The Delayed Type Hypersensitivity of the recipients to the donor antigen: The recipients from group D had low response to the donor ant...