Background/Purpose: Most rheumatoid arthritis (RA) and juvenile idiopathic arthritis (JIA) patients experience episodic arthritis flares and require life-long medications to control their disease. We observed that arthritis tends to flare repeatedly in the same joints, even after decades of remission, displaying joint-specific memory. We have identified tissue resident memory T cells (TRM) in arthritic joints and demonstrated in murine models that these long-lived T_RM remain in synovium during remission and act as key mediators of arthritis flares. However, strategies to model synovial TRM development and function in a human context are lacking.

Methods: To recapitulate the synovial stromal structure in a 3D organoid, we encapsulated fibroblast-like synoviocytes isolated from synovial biopsies from human RA donors with human umbilical vein endothelial cells in Matrigel. These cells were seeded on a polyHEMA-coated low-attachment plate and cultured in a 1:1 mixture of complete RPMI media and human endothelial cell growth media. CD8 memory T cells from healthy donors were isolated from peripheral blood and co-cultured with the synovial organoid for 2-3 weeks. Where indicated, TNF was added to the culture media for the first 3-5 days to simulate inflammation. Cells were dissociated from the organoids and assayed for TRM surface phenotype by flow cytometry. T cells were evaluated for their ability to take up free fatty acids compared to other T cell populations, reflective of a shift toward fatty acid metabolism. To assess the propensity of TRM to remain in tissues, the capacity of organoid-derived T cells to migrate across a transwell membrane in response to tissue egress signal CCL21 was also tested.

Results: Culturing CD8 memory T cells isolated from human blood in 3D synovial organoids, particularly with TNF, supported development of CD8 cells with T_RM surface phenotype (CD45RO+CD62L-CCR7-HLADR-CD25-CD103+CD49a+). These TRM-like cells have enhanced free fatty acid uptake compared to other T cell subtypes, consistent with a shift in metabolic activity seen in TRM. Unlike central memory T cells extracted from synovial organoids and naïve T cells isolated from blood, TRM differentiated in the synovial organoids did not migrate across the transwell membrane to CCL21, confirming this key sessile feature of TRM in tissues.

Conclusion: We developed a novel model for studying human TRM by differentiating TRM within 3D synovial organoids composed of stromal cells from human RA synovium. This model system will be valuable in gaining insights to synovial TRM biology including factors that may drive human synovial TRM development and function.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/generation-of-human-resident-memory-t-cells-in-3d-synovial-organoid-model/