Paneth cells respond to inflammation and SCF/c-Kit signaling by acquiring stem-like features and contributing to tissue regeneration

The experiment aims to analyze the differences in expressed genes in Paneth cells upon acute intestinal inflammation compared to Paneth cells from non-inflamed control mice 8-12 week old Lgr5-EGFP-IRES-CreERT2 mice were subjected to 3% DSS in the drinking water for seven days. Three days after removal of DSS, intestinal tissues were dissected, intestinal crypts isolated and single cell digested. Paneth cells were isolated from single cell suspension by FACS according to previously established gates (Roth et al., 2012). RNA was isolated from isolated Paneth cells using TRIzolTM Reagent (ThermoFisher Scientific) according to the manufacturer’s instructions. RNA quality and quantity was evaluated on a 2100 Bio-analyzer (Agilent) using the Agilent RNA 6000 Pico Kit (Agilent Technologies). For RNA sequencing analysis, RNA samples were further processed by GATC-Biotech (Konstanz, Germany) according to the INVIEWTM Transcriptome Explore library preparation protocol (Illumina based). Water treated mice were used as controls.