Date: Tuesday, June 5, 2018
Session Time: 2:30pm-4:00pm
Presentation Time: 3:18pm-3:30pm
Location: Room 602/603/604
Background: Human CD4+ T cell allo-immunity plays pivotal roles during cellular and humoral allograft rejection. Our previous analysis showed that allo-reactive precursors were present in both naïve and memory CD4+T cells. It is unclear their distribution within T follicular (TFH:CD45RO+CXCR5+) and T conventional (TCONV:CD45RO+CXCR5–) subsets. To address this question, we measured their allo-specific polarization, proliferation, and TCR clonal repertoire.
Methods: FACS-sorted TCONV or TFH cells from 6 healthy control (HC) were used to set up CFSE-MLR assays with autologous monocytes pulsed with allogeneic PBMC lysates. Proliferation and cytokine production (IFN-γ, IL-10, IL-17a, IL-21) were assessed by multi-parameter flow cytometry. In addition, DNA from selected in vitro co-cultures was isolated and the TCR-CDR3 regions were profiled by PCR amplification followed by high throughput sequencing using the immunoSEQ platform (Adaptive Biotechnologies).
Results: Indirect allo-stimulation induced low proliferation, low effector cytokines (IFN-γ/IL-17a/IL-21) and significant high IL-10 responses within TFH and TCONV cells (considered naïve response) in all subjects but one, who displayed low IL-10 secretion paralleled by high IFN-γ/IL-21 in TFH and high IFN-γ/IL-17a in TCONV cells, indicating a memory allogeneic response. ImmunoSEQ was performed on one naïve and one who displayed functional memory. Although resting TFH cells from both subjects displayed a more diverse (polyclonal) TCR repertoire compared to TCONV, allo-stimulation triggered significant TCR clonotype expansion in both TFH and TCONV only in the HC who showed allo-reactive memory responses. Moreover, resting TCONV cells contained most alloreactive TCR clonotypes that correlated positively with expanded allo-reactive TCR clonotypes after allo-stimulation. Two out of 38 TCR expanded clones were shared between TFH and TCONV after in vitro allo-specific stimulation, suggesting potential intraclonal diversity.
Conclusion: These comprehensive analyses of human circulating TFH and TCONV cells reflect the heterogeneity of allo-reactivity among humans. In addition, monitoring of circulating allo-reactive TFH and TCONV cells may be important for identifying patients with pre-formed donor-reactive memory responses (effector cytokines and high TCR clonotype expansion) that may be deleterious post-transplant.
CITATION INFORMATION: Macedo C., Yamada M., Gu X., Suchoroski M., Hamm D., Kaplan M., Metes D. Allogeneic T Follicular and T Conventional Responses Assessed by Proliferation, Cytokine Production, and TCR Clonality Am J Transplant. 2017;17 (suppl 3).
To cite this abstract in AMA style:Macedo C, Yamada M, Gu X, Suchoroski M, Hamm D, Kaplan M, Metes D. Allogeneic T Follicular and T Conventional Responses Assessed by Proliferation, Cytokine Production, and TCR Clonality [abstract]. https://atcmeetingabstracts.com/abstract/allogeneic-t-follicular-and-t-conventional-responses-assessed-by-proliferation-cytokine-production-and-tcr-clonality/. Accessed July 8, 2020.
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