Entry, Capture and Presentation of Blood-Borne Antigens and Effects of TCR Ligand Density on Negative Selection and Treg Induction of CD4SP Thymocytes
Date of Award
Doctor of Philosophy (PhD)
The thymus employs two primary mechanisms to maintain self-tolerance: negative selection and the generation of regulatory T-cells (Treg). Both of these processes require the effective presentation of self-antigens whether derived intra-thymically or from the periphery. Here, we used Hen egg-white lysozyme (HEL) as a model antigen administered i.v. to examine the entry, presentation and effect of blood-borne antigens on thymic T-cell selection. Previous studies on Treg generation and negative selection have shown a requirement for high avidity T-cell receptor (TCR)/peptide-MHC (pMHC) interaction with agonist ligands presented on thymic antigen presenting cells (APC). However, the quantitative differences between signals that induce deletion of self-reactive T-cells by negative selection and those that induce differentiation of self-reactive T-cells into Treg remain elusive. Additionally, the contribution of different APC in determining whether thymocytes undergo negative selection or become Treg is unclear, especially in the case of blood-de rived antigens.
We found that thymic architecture modulates the amount of antigen that enters the thymus and consequently the level of pMHC density presented. There was size-dependent entry of proteins into the thymus. Less than 0.5 % of the injected HEL entered the thymus and presentation resulted in Treg induction and/or negative selection in TCR transgenic mice specific for the major and minor epitopes of HEL that depended on peptide-MHC density levels. Both the range of antigen dose and pMHC complexes that led to negative selection compared to Treg induction for both T-cells was distinct although there was some overlap. More than 25 pMHC complexes per DC were estimated to initiate negative selection with even lower pMHC complexes inducing Treg. Thus, despite the limiting amounts of antigen that entered, the efficiency of thymic antigen presentation resulted in a remarkable effect on thymocyte selection.
Despite disproportionate uptake, blood-borne HEL was presented primarily by Sirpα+ and CD8α+ conventional thymic dendritic cells (cDC), with minimal presentation by other DC subsets and thymic epithelial cells ( TEC ). Both cDC subsets induced Treg and negative selection to in vivo captured HEL to a similar extent. Further, in Ba tf3-/-mice, which have significantly reduced CD8α+ cDC but an intact Sirpα+ cDC population, the presentation of blood-borne HEL was unaffected suggesting that CD8α+ cDC cells are dispensable. There was minimal contribution of peripheral APC, particularly blood Sirpα+ cDC, in the entry and presentation of blood-borne HEL. These data suggest that the effects on thymocytes are mediated primarily by cDC localized within the thymus rather than circulating APC.
Together, our results emphasize the sensitivity of both negative selection and Treg induction to pMHC density levels ensuring that central tolerance will be effective regardless of limitations imposed by local antigen expression.
Chair and Committee
Emil R. Unanue
Paul M. Allen, Chyi-Song Hsieh, Kenneth M. Murphy, Barry Sleckman, Wojciech Swat
Atibalentja, Danielle Françoise, "Entry, Capture and Presentation of Blood-Borne Antigens and Effects of TCR Ligand Density on Negative Selection and Treg Induction of CD4SP Thymocytes" (2010). Arts & Sciences Electronic Theses and Dissertations. 72.