The Thymic Microenvironment Differentially Regulates Development and Trafficking of Invariant NKT Cell Sublineages

Abstract

The regulatory role of the thymic microenvironment during trafficking and differentiation of the invariant NKT (iNKT) cell lineage remains poorly understood. In this study, we show that fractalkine receptor expression marks emigrating subpopulations of the NKT1, NKT2, and NKT17 sublineages in the thymus and peripheral organs of naive mice. Moreover, NKT1 sublineage cells can be subdivided into two subsets, namely NKT1a and NKT1b, which exhibit distinct developmental and tissue-specific distribution profiles. More specifically, development and trafficking of the NKT1a subset are selectively dependent upon lymphotoxin (LT)α1β2-LTβ receptor–dependent differentiation of thymic stroma, whereas the NKT1b, NKT2, and NKT17 sublineages are not. Furthermore, we identify a potential cellular source for LTα1β2 during thymic organogenesis, marked by expression of IL-7Rα, which promotes differentiation of the NKT1a subset in a noncell-autonomous manner. Collectively, we propose a mechanism by which thymic differentiation and retention of the NKT1 sublineage are developmentally coupled to LTα1β2-LTβ receptor–dependent thymic organogenesis.

NKT sublineage specification and survival requires the ubiquitin-modifying enzyme TNFAIP3/A20

“…1 D ), with the highest comparative A20 GFP MFI detected within splenic iNKT cells. In contrast, A20 GFP expression within NKT1, 2, and 17 sublineages ( Lee et al, 2013 ; Drennan et al, 2014 ) was comparable in all organs examined ( Fig. 1 E ), with the highest overall expression detected within splenic and popliteal LN (PLN) NKT sublineages, respectively.…”

“…NKT1, NKT2, and NKT17 sublineage cells are characterized by differential expression of the transcription factors T-bet, GATA-3, and RORγt and produce the cytokines IFN-γ, IL-4, and IL-17, respectively. Furthermore, NKT1 sublineage cells can be subdivided into two subsets, defined as NKT1 a and NKT1 b , which exhibit distinct developmental and tissue-specific distribution profiles ( Drennan et al, 2014 ). An analysis of iNKT cell development in CD4-cre;A20 F/F mice using the aforementioned alternate NKT cell developmental scheme revealed a selective reduction in NKT1 a thymocytes in CD4-cre;A20 F/F thymi relative to controls ( Fig.…”

“…PCR products were amplified with the Fast Start SYBR Green Master mix (Roche) and detected with a LightCycler 480 System (Roche). Samples were normalized using qBasePlus (Biogazelle NV) against at least three of the following genes: Rpl13a , Eif4b , B2m , Actb , or Gapdh as described previously ( Drennan et al, 2014 ).…”

Induction of antiinflammatory purinergic signaling in activated human iNKT cells

“…These findings relate to an interesting feature of iNKT cells -their ability to produce diverse cytokines depending in part on environmental cues. Although subsets of NKT1, NKT2, and NKT17 cells that differentiate in the thymus have been defined in mice (44), iNKT cell subsets are not well defined in humans. Rather, the majority of circulating human iNKT cells produce multiple cytokines, and the ratio of Th1-/ Th2-type cytokines is known to be influenced by several factors.…”

An Optimized Method for Isolating and Expanding Invariant Natural Killer T Cells from Mouse Spleen

“…Unfortunately FACS sorters are expensive and they are not always available in every laboratory. Nevertheless, although the current protocol has been optimized to isolate and culture murine iNKT cells, the enrichment steps described herein can also be used to isolate and study iNKT recent thymic emigrants (RTEs) 16 and can be adapted to enrich for and characterize other rare T cell populations by FACS.…”