Proliferation makes a substantive contribution to the maintenance of airway resident memory T-cell subsets in young pigs

Tissue-resident memory (TRM) T cells play an important role in protection against respiratory infection but whether this memory is maintained by long-lived or dividing cells remains controversial. To address the rate of division of lung TRM T cells, deuterium-enriched water was administered orally to young pigs to label dividing lymphocytes. T-cell subsets were separated from blood, lymph nodes, and airways [bronchoalveolar lavage (BAL)], the latter comprising almost exclusively TRM. We show that, as in other species, circulating memory T-cell subsets divide more rapidly than naïve T cells. Rates of labelling of memory subsets were similar in blood and lymph nodes, consistent with the rapid and free exchange. Strikingly, the fraction of label in BAL was similar to those in blood/lymph nodes after 5-21 days of labelling, suggesting replacement with recently divided cells, but this was preceded at Day 2 by a phase when labelling was lower in BAL than blood/lymph node in some memory subsets. Our data exclude long-lived TRM as the source of BAL memory cells leaving three possible hypotheses: blood/airway exchange, <i>in situ</i> proliferation, or proliferation in the lung interstitium followed by migration to BAL. When considered in the context of other information, we favour the latter interpretation. These results indicate the dynamic nature of memory in the lung and have implications for harnessing immune responses against respiratory pathogens.