An amphiregulin reporter mouse enables transcriptional and clonal expansion analysis of reparative lung Treg cells
Treg cells play critical roles in tissue repair processes, certain of which are mediated by production of the growth factor amphiregulin (Areg). We developed a knock-in mouse model for detection of Areg production in live Treg cells, inserting a Thy1a (Thy1.1) construct into the endogenous Areg locus, such that for every transcript of Areg that is translated, a Thy1.1 transcript is also translated and trafficked separately to the cell surface. We used this mouse model to sort for Thy1.1- or Thy1.1+ (Areg-producing or -non-producing) lung Treg cells in models of lung damage (influenza A virus infection and bleomycin acute lung injury), and used them for assessment of gene expression and clonal expansion of these populations. Overall design: Foxp3GFP mice were administered bleomycin (1 mg/kg). At 15 days post-instillation (dpi) (bleomycin), lung Treg cells were sorted into 4 separate pools (from 2-4 mice each), each pool was split between 2 wells, then paired wells were treated with IL-2, IL-7, and either IgG control or 4-1BB agonistic antibody (clone 3H3) (10 µg/ml) for 4h. Treg cells were then washed out of wells and lysed in Trizol for bulk RNA-seq.