Long-Term Follow-up of Donor Chimerism and Tolerance After Human Liver Transplantation.
Authors: Grande Silvia , Diaz Rosa Maria Ayala , Gilsanz Florinda , Moreno Enrique , Martinez Joaquin 
Published: October 2019

Abstract
Abstract 3534
Poster Board III-471
The quantification of hematopoietic chimerism after allogeneic stem cell transplantation is an important tool for monitoring post-transplant outcome. This phenomenon has also been observed in solid organ transplantation and in Liver Transplantation (LT) and Starzl and Zinkernagel hypothesized that organ engraftment is a form of chimerism-dependent partial tolerance. The aim of this study was to quantify peripheral Donor Chimerism (DC) in liver recipient and to analyze its association with graft and recipient outcome.

Patients
A total of 42 LT patients and their respective donors took part in this study. 226 samples were studied. 78 were pretransplant samples from both the recipient and the donor, and 148 were post-transplantation samples.

Methods
DNA from serum samples was used for all post-transplantation studies. DC was carried out by quantitative Real–Time PCRs (qPCR) to detect bi-allelic single nucleotide polymorphisms (SNPs) using TM probes and insertion-deletion di-allelic polymorphisms using Hyb probes as polymorphic markers. DC levels were correlated with the recipients and donor characteristics as well as the overall survival (OS) and the rejection-free survival (RFS).The follow-up time post-transplantation to DC study was 46.9 months.

Results
An informative polymorphism between recipient and donor was found in 39 of 42 cases (93%). DC was detected in 111 of the 148 samples (75%) with a median of 1.27%. There was no significant correlation between DC and immunosuppressive therapy for each transplant recipient during the study, although there was an inverse correlation between immunosuppressor levels and the percentage of DC. No correlation was found between DC and characteristics of the recipient patient, that is, gender, liver disease, ABO group, and Rh blood group apart from age (adult and child median DC, 1.22% and 5.72% respectively; p=0.023). DC levels tended to be higher in the O blood group liver recipients than in the A blood group liver recipients (2.24% and 1.27% respectively; P=0.095). No relationship was found between DC and graft characteristics (gender, graft/donor gender match, partial liver graft, ABO blood group and ABO matching), although DC was significantly lower in Rh-negative grafts than in Rh-positive grafts (1.27% and 2.24% respectively, in univariate analyses; P=0.047). DC levels tended to be lower in LDLT compared with DDLT (1.23% and 2.24% respectively; P=0.067).
The median DC of all the patients was 1.27% versus the median DC from 27 samples in 12 patients with ongoing graft rejection, which was 0.39%. This difference was, however, not significant. The median DC levels in 55 samples from 12 patients with no apparent clinical events was 1.17% whereas in 43 samples from 9 patients with recurrent liver disease were 2.70%. The median DC in 11 samples from 3 patients with both recurrent disease and rejection was 5.47%. There was macrochimerism in 3 (33%) of the 9 patients with graft rejection, in 8 (80%) of the 10 patients with post-transplantation recurrent liver disease, and in 7 (53.8%) of the 13 patients with no significant clinical events. There was a tendency toward longer OS in patients with higher DC levels compared with those with lower DC levels (100% survival versus 89.7%), with a median follow-up of 44.92 versus 39.24 months respectively (p= 0.0988). The median RFS in recipients with lower DC was 44.48 months (range, 31.25-57.72 months), whereas the median RFS in recipients with higher DC was 55.18 months (44.17- 66.9 months; p=0.4439).

Conclusion
DC analysis by qPCR is feasible in most LT cases and has several advantages over previously employed methods such as flow cytometry, PCR/single-strand conformation polymorphism, FIS X/Y, and STR. The evolution of DC levels should be analyzed in accordance with the clinical outcome of the patient. Although we have studied only a relatively small number of patients, our results suggest that stable high DC levels, in the absence of other major clinical events, could be a marker of transplantation tolerance and may help to tailor immunosuppressive treatment.
Our results are consistent with recent studies that have confirmed the importance of DC, induced by donor hematopoietic stem cell transplantation, in the tolerance of solid graft transplantation.

Disclosures:
No relevant conflicts of interest to declare.