LTi-like type 3 innate lymphoid cells are associated with the transient induction of tertiary lymphoid structures in inflamed pulmonary tissue

Abstract

Abstract Tertiary lymphoid structures (TLS) are ectopic lymphoid aggregates observed in inflamed tissues. Their presence in pulmonary tissue has been associated with good prognosis in patients with lung cancer. Although TLS formation shares common features with secondary lymphoid organogenesis, the mechanisms underlying this process are yet to be fully elucidated. Presence of ILC3 in the vicinity of tumor-associated TLS correlates with the density of lymphoid structures. We hypothesized that RORγt+ ILC3 play a major role in TLS induction in inflamed lungs, as do RORγt+ fetal lymphoid tissue inducer cells (LTi) during lymph node formation. We developed a murine model of pulmonary inflammation induced by LPS intranasal instillation that triggers TLS formation. ILC3 presence in lungs was observed with a specific flow cytometry antibody panel. Three RORgt+ILC3 subsets (NCR− CCR6−, NCR−CCR6+, NCR+CCR6−) were detected. In particular, NCR−CCR6+ LTi-like cells represented about 25% of CD117+CD127+ILC (ILC2/ILC3) and less than 0.04% among CD45+ cells. Analyses of ILC subsets at different time points following LPS instillation showed an increase in the number and % (up to 0.59%) of ILC3 among CD45+ cells, as well as an increase in the % of LTi-like ILC3 among CD45+ cells, that peaked around day 10 (from 0.02% up to 0.15%). Quantification of TLS by histochemistry showed that they are induced in lung tissue in an LPS dose-dependent manner. TLS density increases between days 3 and 10 and decreases thereafter. They were no longer detected at day 25, while LTi-like ILC3 % returned to baseline. Gene expression analysis and functional assays of sorted LTi-like ILC3 are essential to determine if these cells are key players in TLS induction. Supported by grants from La Ligue Nationale contre le cancer (TAHG21042) and Fondation ARC (R19232DD).