The first report of three-way macrochimerism in a patient receiving multivisceral transplantation following loss of an initial liver-intestine transplant

Introduction: The success of intestinal transplantation is compromised by high rejection rates. Our previous studies demonstrated that macrochimerism (≥4% peak level of donor T cells) in recipient blood and slower replacement of donor T cells in the intestinal graft by recipient are associated with less rejection. Intestinal graft-derived hematopoietic stem and progenitor cells (HSPCs) contribute to persistent multi-lineage chimerism in blood and may promote immune tolerance. Long-term blood T cell chimerism in re-transplantation has not been explored. Methods: We longitudinally monitored chimerism in peripheral blood and intestinal allograft intraepithelial lymphocytes (IEL) and lamina propria lymphocytes (LPL) of a patient who rejected a liver-intestinal transplant (LITx) and subsequently received a multivisceral transplant (MVTx). HLA typing information for the recipient and two donors was used to identify HLA class I allele-specific antibodies that discriminate cell origin by multicolor flow cytometry. Immunosuppression regimen included anti-thymocyte globulin, tacrolimus, and methylprednisolone. Results: The patient was diagnosed with necrotizing enterocolitis and received LITx at 20 months of age from a 3-month-old donor (Donor 1). Mild rejection occurred (based on pathologic scoring scheme) on postoperative day 15 (POD15), progressing to severe rejection on POD19. Graft loss occurred on POD786 with donor chimerism of T cells in blood never surpassing peak of 2.36% on POD9. Meanwhile, recipient T cell chimerism in graft surpassed 95% by POD19. He received MVTx from a 19-month-old donor (Donor 2) on POD786. 4 episodes of mild rejection and 1 of mild-moderate rejection occurred over the next 5 years. Blood macrochimerism from Donor 2 peaked at 29.9% on POD20. Concomitantly, macrochimerism from Donor 1 peaked at 8.98% on POD48. The majority of these circulating Donor 1 cells were CD8+ (13.3%), not CD4+ T cells (0%). Recipient T cells were slower to repopulate graft this time in both ileal IELs and LPLs. Recipient T cell chimerism in graft did not surpass 25% up to POD251, but gradually increased to 99.4% in IELs and 93.9% in LPLs by POD377, and then remained between 80% and 100% through POD1004 (last timepoint with data). Recent thymic emigrant (RTE) [CD45RA+CCR7+CD31+] analysis revealed CD4+ RTEs only from recipient and Donor 2. CD8+ RTEs, however, were detected from recipient, Donor 2, and Donor 1 from POD5 onward. Conclusions: For the first time, we report development of persistent triple blood T cell chimerism in our patient, with both donors exhibiting macrochimerism in recipient after re-transplantation, associated with improved clinical outcomes compared to initial LITx. The RTE phenotypes of circulating T cells from both donors, especially Donor 1 more than 2 years post 1st LITx, are in line with our previous demonstration that intestinal graft-derived HSPCs promote long-term mixed chimerism in blood and may induce tolerance.