Identifying Tissue-Resident Memory T Cells in Graft-Versus-Host Disease

Tissue-resident memory T cells (T_RM) 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 T_RM cells can be identified by the expression of CD69 and αE integrin, CD103. However, CD69⁺CD103^- T_RM 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, T_RM may develop.

We first explored T_RM 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 T_RM 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^- T_RM 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 T_RM identity of TS1 cells based on their transcriptional and migratory profiles and these data will be presented.

Alloreactive T_RM were also identified in the B6 (H-2^b) into 129 (H-2^b) 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 K^b-restricted miHA LTFNYRNL derived from H60, which can be tracked with tetramers (Tet^H60). 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⁺Tet^H60+ 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⁺Tet^H60+ 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 T_RM phenotype.

Using two different murine models, we found GVHD-inducing T cells with T_RM phenotypes. Future experiments will confirm the T_RM 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.