We recently demonstrated that ageing also impedes development of transplantation tolerance. Unlike their young counterparts (8-12 weeks of age) aged male recipients (greater than 12 months of age) transplanted with a full MHC-mismatched heart are resistant to tolerance mediated by anti-CD45RB antibody. Surprisingly, either chemical or surgical castration restored tolerance induction to levels observed using young recipients. Based on the strong impact of endocrine modulation on transplant tolerance, we explored the impact of ageing and castration on the immune system.

METHODS: For cytokine staining, cells were stimulated in Complete Medium and either with or without LPS for 5 hours. ELISPOT assays detected IFN-g production after 4 hrs and 72 hrs of culture, which distinguish memory from naive T cells. Graft survival between experimental groups was compared using Kaplan-Meier survival curves and Wilcoxon statistics. Other differences between experimental groups were analyzed using the Student's t test. P values less than 0.05 were considered statistically significant.

RESULTS: Here we report a significant increase in the percentage of T cells that produce interferon-γ (IFN-γ) in aged male versus young male animals and that the overall increase in IFN-γ production was due to an expansion of IFN-γ-producing memory T cells in aged animals. We observed no significant difference in percentage of IFN-g by naive T cells in young and old recipients. In contrast to IFN-γ production, we did not observe differences in IL-10 expression in young versus old male mice. We hypothesized that endocrine modulation would diminish the elevated levels of IFN-γ production in aged recipients, however, we observed no significant reduction in the percentage of IFN-γ+ T cells upon castration. Furthermore, we neutralized interferon-γ by antibody and did not observe an effect on graft survival. Diminished IL-7 in aged individuals may also contribute to age-dependent resistance. In young mice, we observed that neutralization of IL-7 resulted in rapid rejection of skin grafts, similar to rejection observed in aged mice (p<0.05).

CONCLUSIONS: We conclude that while elevated levels of interferon-γ serves as a marker of tolerance resistance in aged mice, other as yet to be identified factors are responsible for its cause. Defining these additional factors, such as IL-7, may be relevant to design of tolerogenic strategies for aged recipients.

CITATION INFORMATION: Kim J, Stott R, Deirawan H, Kojima L, Washburn L, Zhao G, Yeh H, Markmann J. Ageing as a Barrier to Transplant Tolerance. Am J Transplant. 2016;16 (suppl 3).

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