Identifying Tissue-Resident Memory T Cells in Graft-Versus-Host Disease
Authors: Sacirbegovic Faruk , Zhu Jieqing , Liu Jinling , Rosenberger Sarah , Shlomchik Mark J , Shlomchik Warren D 
Published: October 2019

Abstract
Tissue-resident memory T cells (TRM) are a newly described subset of transcriptionally-distinct memory CD4 and CD8 cells that persist in barrier and non-barrier tissues. They are non-circulating, able to facilitate the recruitment of circulating effector cells and elicit rapid recall responses. The majority of TRM cells can be identified by the expression of CD69 and αE integrin, CD103. However, CD69+CD103- TRM cells have also been described.
In graft-vs-host disease (GVHD), alloreactive effector T cells enter GVHD target tissues and mediate tissue damage through direct and indirect mechanisms.The recruitment of effector T cells into tissues is in general not dependent on the tissue expressing the target antigen; and even if a target antigen is available in a tissue, there could be niches free of presented alloantigen. We therefore hypothesized that even in GVHD where alloantigen is ubiquitous, TRM may develop.
We first explored TRM formation in a CD4 T cell receptor (TCR) transgenic (Tg) GVHD system wherein donor BALB/c RAG2-/- TS1 TCR Tg T cells target the S1 peptide derived from HA, which is expressed ubiquitously in BALB/c RAG2-/- HA104 mice. In this model, GVHD is induced by <1000 TS1 cells and is manifest by weight loss, death, and TS1-infiltrative pathology of the skin, liver, small bowel and colon. We harvested tissues from GVHD mice at days 21 and 28 post-transplant and quantitated TS1 cells with a TRM phenotype. At day 21, a fraction of TS1 cells expressed CD69+CD103+ (as % of total TS1 cells) in the epidermis (2.8% ± 1.2), dermis (11.3% ±7.9), colon (11.5% ±4.5) and small intestine (SI) intraepithelial (IEL) (33.4% ±5.8) and lamina propria (LP) (6.8% ±0.8). At day 28, CD69+CD103+ TS1 cells (% of total TS1) were present in the epidermis (13.6% ± 1.9), dermis (15.9% ± 7.7), colon (30.8% ± 6.7) and the SI IEL (69.1% ± 9.2) and SI LP (18.7 ±3.2). While there were no CD69+CD103+ TS1 in the spleen, bone marrow (BM) or liver (days 21 and 28), a small number (1.5% ±0.7, day 21; 1.7% ± 0.3, day 28) were found in the mesenteric lymph node (mLN). CD103- TRM have been described in the liver and secondary lymphoid organs. Consistent with this, CD69+CD103- TS1 cells (% of total TS1) were found in the liver (24.6% ±6.3, day21; 31.5% ±3.1, day 28), BM (23.1% ±3.1, day 21; 35.4% ±3.2, day 28), spleen (1.8% ±0.6, day 21; 9.1% ±0.7, day 28) and mLN (9.9% ±4.5, day 21; 23.5% ± 3.7, day 28). We are currently confirming the TRM identity of TS1 cells based on their transcriptional and migratory profiles and these data will be presented.
Alloreactive TRM were also identified in the B6 (H-2b) into 129 (H-2b) MHC-matched, multiple minor histocompatibility antigen (miHA)-mismatched model in which GVHD is induced by a mix of CD4 and CD8 cells. A fraction of CD8 cells target the Kb-restricted miHA LTFNYRNL derived from H60, which can be tracked with tetramers (TetH60). At day 22 post-transplant, CD69+CD103+ CD4 cells (% of total CD4 T cells) were found in the epidermis (22.9% ±1.2), dermis (19.7% ±2.8), colon (9.4% ±3.1), SI IEL (26.5% ±2.6), SI LP (16.1% ± 3.7) and mLN (6.9% ±3.0). CD8+TetH60+ T cells (% of total CD8T cells) were detected in the epidermis (7.5% ± 3.5), dermis (10.6% ± 6.7), colon (6.9% ± 2.7), SI IEL (8.9% ±6.2), SI LP (10.2% ±3.8), mLN (3.0 ± 0.7) and spleen (9.0% ±2.9). A fraction of CD8+TetH60+ cells in the dermis (7.3% ±1.9), colon (18.1% ±12.2), SI IEL (50.1% ±4.3), SI LP (52.6% ±13.6), and mLN (6.3% ± 0.8) were CD69+CD103+, suggesting that alloreactive H60-directed CD8 T cells acquired a TRM phenotype.
Using two different murine models, we found GVHD-inducing T cells with TRM phenotypes. Future experiments will confirm the TRM identity of these cells and will determine their importance in the maintenance of GVHD, perhaps by serving as a reservoir of cells that maintain GVHD locally.

Disclosures
No relevant conflicts of interest to declare.