Exploring the role of infection experienced lung stromal cells following re-challenge with influenza A virus

Abstract

The concept that stromal cells are permanently altered by inflammatory responses is termed ‘trained immunity’. We hypothesise that Influenza A virus (IAV) infections influence future immune responses via changes to lung stromal cells, enabling rapid communication with immune cells following subsequent infection. To address this, mice were infected with IAV (H1N1) for 30 days and subsequently re‑challenged with IAV (H3N2) for 2 days. The transcriptional profiles of FACS sorted lung fibroblasts and epithelial cells were identified by NanoString assay. Biological functions were investigated by flow cytometry. Putative upstream regulators were compared with publicly available RNA-seq and ChIP-seq datasets. The location of altered lung stromal and immune cells was determined using RNAscope and immunohistochemistry. Genes involved in T cell communication were significantly upregulated in lung fibroblasts and epithelial cells following re‑challenge with IAV (day 32). Fibroblasts displayed enrichment in biological processes regulating T cell activation, while positive regulation of cytokine production was enriched in epithelial cells. Rapid upregulation of genes involved in antigen processing and presentation (Tap1, Cd74) was detected at day 32 in both cell types. The transcription factor, SpiB, was identified as a shared upstream regulator of these genes. At d30, SpiB+ airway epithelial cells were only found adjacent to immune cell clusters (T/B cells); clusters were retained following IAV re‑challenge. Rapid changes in immunomodulatory molecules by infection experienced stromal cells, suggest enhanced communication with lung T cells following a subsequent infection.