The Study of Induction of Immune Tolerance and Enhancement of Immune Reconstitution by Thymus Transplantation in Murine with H-2 Haploidentical Bone Marrow Transplantation
Authors: Fan Zhiping , Liu Qifa , Zhang Yu , Xu Dan , Wei Yongqiang 
Published: October 2019

Abstract
Abstract 1895

Background and Objectives:
Graft versus host disease (GVHD) and opportunistic infection, which remain the main complications after allogeneic hematopoietic stem cell transplantation (allo-HSCT), lead to the high transplant-related mortality. Nowadays, the key to prolong disease free survival and enhance quality of life of patients is to induce immune torlerance for donor T cells to recipient antigen and enhance T cells reconstitution after allo-HSCT. Recently, researchers found that solid organs transplantation combined with thymus transplantation could decrease the rejection of solid organ apparently and promote T cells reconstitution after solid organ transplantation. In the present study, we established C57BL/6 to CB6F1murine H-2 haploidentical bone marrow transplantation (BMT) combined with the same fetal thymus transplantation model to modify the donor and recipient's T cells, to see wether this method could lower the incidence of GVHD, improve long-term survival and enhance the reconstitution of T cells after transplantation.

Methods:
24 CB6F1 murine were divided into two groups randomly: thymus transplantation group (experimental group, EG) and non-thymus transplantation group (control group, CG). All CB6F1 murine were treated with H-2 haploidentical BMT, and EG murine were cotransplanted with H-2 haploidentical fetal thymus under the left kidney capsule simultaneously. GVHD scores and long term survival of murine were observed and evaluated. The reconstitution of CD3+CD4+T and CD3+CD8+T cells of peripheral blood at 1, 2, 3 month after transplantation were detected by flow cytometry. Mixed lymphocyte reactivity (MLR) was used to dectect the response of splenocytes from CB6F1 murine grafted with C57BL/6 fetal thymus tissues to donor C57BL/6 murine splenoctyes.

Results:
After treated with TBI+CY conditioning regimen and administered methylprednisolone orally intermittent after transplantation, H-2 haploidentical C57BL/6 fetal thymus could grow continuously under the left kidney capsule of the CB6F1 murine. At the 3d month after transplantation, thymus tissue was densely populated with thymocytes and the thymic stromal structure was normal. Immunohistochemistry analysis demonstrated that the transplanted thymus was from C57BL/6 murine. The GVHD score of EG and CG murine has no significant difference during the first 12 weeks after transplantation, but the GVHD score of EG was lower apparently than CG from 13 weeks after transplantation (P=0.000). After the discontinuation of methylprednisolone, EG murine had moderate non-lethal GVHD, but CG murine had severe lethal GVHD. At day 110 after transplantation, there were still 5 mice alive (42%) in EG, but there were none in CG. The incidence of cumulative survival between the two group differed significantly (P=0.010). The percentage of CD3+CD4+T and CD3+CD8+T cells of EG and CG had no significant differences at 1, 2, 3 month after transplantation in H-2 haploidentical transplantation or syngeneic transplantation. But the percentage of CD3+CD4+T and CD3+CD8+T cells appeared to increase as time went by in H-2 haploidentical transplantation or syngeneic transplantation. In vitro one-way MLR, splenocytes from CB6F1 murine grafted with C57BL/6 fetal thymus tissues showed a lower response to donor C57BL/6 murine splenoctyes, thus brought about immune tolerance to donor, but showed a strong response to BABL/c and KM murine splenoctyes.

Conclusions:
The severity of GVHD and the mortality were decreased and the cumulative survival were prolonged significantly in recipient CB6F1 murine after treated with H-2 haploidentical BMT and cotransplanted with the same donor strain murine's fetal thymus. Thymus transplantation could induce immune tolerance to donor specific antigens. The data of the present study could not demonstrate H-2 haploidentical fetal thymus transplantation can enhance T cell reconstitution after H-2 haploidentical BMT.

Disclosures:
No relevant conflicts of interest to declare.