After the mice were sacrificed, gallbladder was removed. A total of 10 gallbladders per group were washed with D-Hanks solution, and cut into small pieces. One mL 0.25% Trypsin (Gibco, China) were added, and the mixture was shaken for 20 min at 37C. The mixture was passed through a 40 μm cell strainer to collect the filtrate. The gallbladder pieces were re-digested for another two times. All filtrates were centrifuged at 1500 rpm for 5 minutes, and the supernatants were removed. The cell pellets were then suspended in DMEM containing 10% FBS. The living single cells were transferred to cold scRNA-seq lysis buffer. The cells were re-suspended on concentration of 700~1,200 cell/μL and loaded on a Chromium Single Cell Instrument (10X Genomics, United States) to generate single cell gel beads in emulsion. GEMs were constructed for single cell separation according to the number of cells to be harvested. After GEMs were normally formed, GEMs were collected for reverse transcription in a PCR machine for labeling. The GEMs were oil-treated, and the amplified cDNA was purified by magnetic beads and then subjected to cDNA amplification and quality inspection. The 3ʹ Gene Expression Library was constructed with the quality-qualified cDNA. After fragmentation, adaptor ligation, sample index PCR, etc., the library is finally quantitatively examined. The final library pool was sequenced on the Illumina Novaseq6000 instrument using 150-base-pair paired-end reads.