The Impact Of Rapamycin Combined With Regulatory T Cell On The Cardiac Graft And Anti-tumor Immunity Of Mice

ObjectiveTo study the effects of short-term use of rapamycin combined with regulatory T cell(Treg)on the long time survival of allogeneic mice cardiac transplant,as well as to explore its affect on the anti-tumor immunity of recipient.MethodsMurine Treg were purified from recipients’ spleen by magnetic activated cell sorting(MACS),and expanded by CD3/CD28 monoclone antibody immunomagnetic beads and 2000 U/ml Recombinant Murine IL-2 ex vivo,and the purity was tested by FACS.Allogeneic mice cardiac transplant models were established(H-2~b to H-2~d),and the mice were divided into three groups: control group(transplant only),Rap group,Rap + Treg group.In Rapa group,mice were treated with rapamycin(1mg/kg,ip)for 14 consecutive days,mice in Rap + Treg group were receive the same treatment,and simultaneously 1′107 Tregs were adoptive transferred through tail vein on the day of transplant.Meanwhile the syngeneic transplant group was set up(H-2~d to H-2~d).Allograft survival was monitored daily and the graft was harvested on the indicated day and histological evaluated.In the experiment of recipient’s anti-tumor immunity,similar three group of cardiac transplant models were established(H-2~b to H-2~d),and B16-F10 cells(donor derived)were transferred through tail vein,another three groups of transplanted mice(H-2~d to H-2~b)were also transferred B16-F10 cells(recipient derived).Two weeks later,the tumor nodules of the lung were compared.ResultsThe purity of CD4~+CD25~+Foxp3~+ Treg before expansion was 86.68%±0.02%(n=5),while after expansion,the number was 30~40 times as large as which before expansion,and the purity of CD4+CD25+Foxp3+ Treg was 84.58%±0.03%,there was no significant difference.The median survival time(MST)of the graft in the control group was 7 days,which prolonged to 15 days in Rap group,and significantly prolonged to 93 days in Rap + Treg group,and histology analysis of long time survival grafts showed lymphocytes infiltration and chronic vasculopathy.For donor derived tumor,no tumor nodule in the control group,and significantly increased to 15±8(Rap group)and 14±7(Rap + Treg group),with no significantly difference between the later two groups;for recipient derived tumor,the tumor nodules in Rap + Treg group were 146±12,which were significantly elevated compared with control group(70±12)and Rapa group(28±9).ConclusionShort-term use of low dose rapamycin combined with Treg can significantly prolong the survival of mice transplanted heart,but cannot inhibit tumor progression of the recipient.