The effects of CD131 on experimental murine colitis

To explore the possible mechanisms that CD131 involved in experimental murine colitis, we subjected the colonic tissues from control and DSS-treated, wt and CD131-deficient, mice to RNA-seq. C57BL/6J mice (wt) were obtained from Beijing HFK Bioscience Co., Ltd., China. CD131-deficient mice (Csf2rb+/-) on a C57BL/6J background were purchased from Shanghai Model Organisms Center, Inc., China (C57BL/6JSmoc-Csf2rbem1Smoc, #NM-KO-190787). All mice were bred in specific-pathogen-free (SPF) animal facility, and were 8-12 weeks of age at the time of euthanasia. All animal studies were performed in an age- and sex-matched manner and littermate controls were adopted. All protocols were approved by the Institutional Animal Care and Use Committee. Acute Dextran sulfate sodium (DSS)-induced colitis was established by subjecting 8-10-wk-old SPF mice at least 18 g of weight to 2% (wt/vol) DSS (M.Wt. 36,000-50,000 Da; #160110, MP Biomedicals, USA) in the drinking water for 7 d. The DSS solutions were freshly prepared daily, and the volume of leftover DSS solution was measured the second day to exclude the possibility that changes in colitis activity were due to differences in DSS consumption. No difference in water consumption was observed between CD131-deficient and wt mice. For RNA-seq, total RNA was isolated from homogenized murine colonic tissues using TRIzol reagent (Invitrogen, USA). RNA quality was assessed using RNase-free agarose gel electrophoresis on a 2100 Bioanalyzer (Agilent, USA). Afterwards, the mRNA was enriched with Oligo(dT) beads. Subsequently, the enriched mRNA was fragmented by using fragmentation buffer and reverse transcribed into double-stranded cDNA by using NEBNext Ultra RNA Library Prep Kit for Illumina (New England Biolabs, USA) as per the manufacturer's instructions. The resulting ds cDNA was sequentially purified, end-repaired, adenine base-added, and ligated to Illumina sequencing adapters. The ligation reaction was purified with AMPure XP Beads; while the adapter-ligated cDNA was subjected to PCR enrichment and again purification. Finally, the resulting cDNA library was sequenced on a NovaSeq 6000 Sequencing System (Illumina, USA). RNA sequencing and original data analyses were done with the assistance of Guangzhou Gene Denovo Biotechnology Co., Ltd., China.