A. The purpose of this study is to immunocharacterize the first reported case of intestinal transplant (ITx) operational tolerance, in parallel with a stable ITx cohort and a healthy control cohort.

B. An immunophenotype of our TOL (operational tolerant patient) was generated by collecting blood and intestinal samples. Demographically corresponding samples of matched control cohorts from seven stable ITx patients on immunosuppression and eight healthy males were studied in tandem and analyzed with polychromatic flow cytometry (PFC) and immunohistochemistry (IHC).

C. TOL is a 20 year-old male recipient of an isolated ITx for pseudo-obstruction seven years ago and who, due to non-compliance, has been off all immunosuppressive medication for over three years and without rejection. His course was complicated by graft-versus host disease (GvHD) six months post-op, thus we initially hypothesized that his tolerance was secondary to chimerism. However, repeated genotyping of his peripheral blood has consistently showed no evidence of chimerism, suggesting instead peripheral tolerance. In order to test this theory, his allograft lymphocytes were analyzed via PFC revealing 7.6% CD3⁺ T cells which is lower than in the stable ITx cohort at 12.3%. TOL had a comparable level of pro-inflammatory CCR6⁺ Th17 cells (20%) along with the stable ITx cohort (24%), suggesting the capacity to mount healthy anti-microbial immunoresponses. TOL's allograft T cell analysis reveals 1.65% and 5.10% of CD4⁺ and CD8⁺ effector memory T cells (CD45RO⁺ CD62L^–) versus 83.1% and 58.0% in the stable ITx cohort, respectively. IHC analysis of TOL's allograft demonstrates an average of 70.8 cell count per 5 HPFs at 20x magnification of FoxP3⁺ stained Treg cells versus 29.1 in the stable ITx cohort. In blood, CD4⁺ T cell subsets analysis revealed a significant presence in naïve CD4⁺CD62L⁺ (p=0.0014) T cells and a significant decrease in memory CD4⁺CD45RO⁺ (p= 0.0049) T cells in TOL and the healthy cohort compared to the stable ITx patients. Importantly, we found that TOL had 4.82% FoxP3⁺CD4⁺ Tregs in his blood, which is comparable to 4.03% in the healthy cohort and significantly higher than 2.37% in the ITx cohort (p=0.0351).

D. Operational tolerance in ITx is a proven phenomenon characterized by a significant presence of Tregs, which have been shown to aid in tolerance, and an overall balanced and competent immunological phenotype that aligns closer to that of healthy patients.

CITATION INFORMATION: Aguirre O., Houlihan B., Fishbein T., Kroemer A. Immunomonitoring of Operational Tolerance in Intestinal Transplantation Am J Transplant. 2017;17 (suppl 3).

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