Enzymatic/mechanical digestion outperforms explant culture for isolation of tissue resident memory t cells from human lungs

Abstract

Abstract Tissue resident memory T cells (TRM) comprise a distinct population which persists in the tissue after infection. Lung TRM help to coordinate early immune responses to infections of the respiratory tract, but sampling of the relevant anatomical sites in humans is logistically difficult and not easily standardized. We used fresh human tissues to compare methods for organ dissociation and lymphocyte enrichment with regard to T cell phenotype and viability. The two main approaches to tissue processing were explant culture and combined enzymatic/mechanical digestion. While cytokine supplementation was able to induce T cell egress from intact respiratory samples, the majority of TRM remained in situ and the viability of all cells was compromised after three days of ex vivo culture. More T cells were recovered after IL-2 supplementation than with IL-7, but it remains unclear whether IL-2 is inducing proliferation in vitro. Enzymatic and mechanical digestion did not appear to alter the antigenic phenotype of the lymphocyte population and yielded enriched and purified cellular samples that were suitable for downstream flow cytometric or transcriptomic assays. Results suggest that minimizing processing time and using a targeted enzymatic cocktail are important considerations for working with human lung TRM. Supported by NIH grant GR531572.