RNA-seq of mus liver portal area macrophages
Mouse livers were perfused with 0.5 mg/mL collagenase IV and 20 μg/mL DNase I to obtain vascular-enriched and non-vascular-enriched liver tissue. The vascular-enriched liver tissue was further digested with 0.5 mg/mL collagenase IV, 5 mg/mL collagenase I, 5 mg/mL Liberase TL, and 20 μg/mL DNase I. Vascular-enriched liver tissue was used for LPAMs sorting, while non-vascular-enriched liver tissue was used for sorting liver KCs, and LCMs, using the Moflo XDP cell sorter (Beckman). Sorted LPAMs, LCMs, and KCs were then collected to isolate total RNA using TRIzol (Thermo, Carlsbad, US) according to the manufacturer's instructions. RNA integrity and quantity were assessed using Agilent 2100 (Agilent Technologies) and Nanodrop spectrophotometer (Thermo). The KC-DigitalTM Stranded mRNA Library Prep Kit (Seqhealth Technology Co., Ltd. Wuhan, China) was use for RNA sequencing library preparation. In brief, the library products were sequenced on a DNBSEQ-T7 platform (MGI), and the data were mapped to the GRCm38 mouse reference genome.