Animal Modeling of ANIT-Induced Intrahepatic Cholestasis and Grouping

All SD rats were acclimated for seven days prior to the study and then randomly assigned into four groups with ten rats each. The ratio of males to females in each group was 1:1. Rats in the normal control group (control group) received a continuous intragastric administration of saline (0.9%) for six days. Rats in the group treated with ANIT (Sigma-Aldrich, St. Louis, MO, USA) (ANIT group) received a continuous intragastric administration of saline (0.9%) for six days and were treated with ANIT (50 mg/kg) once on the fifth day. Rats in the group treated with LDHJ granules (CR SANJIU, Shenzhen, China) (ANIT+LDHJ group) received a continuous intragastric administration of LDHJ granules (74 g/kg/d, the solvent was distilled water) for six days and were treated with ANIT (50 mg/kg) once on the fifth day. Rats in the group treated with UDCA (Meilunbio, Dalian, China) (ANIT+UDCA group) received a continuous intragastric administration of a UDCA (60 mg/kg/d) for six days and were treated with ANIT (50 mg/kg) once on the fifth day. Rats were fasted for 24 h after the last intragastric administration prior to sacrifice and then used in vivo experiments. The grouping information was illustrated as step 1 in Figure 1 .

Flow chart of modeling group. Step 1: Exploration of the effects of LDHJ granules on ICH and the possible mechanism; Step 2: Verification of the key role of CaSR in cholestasis-related hepatocyte apoptosis; Step 3: Selection of the active substances of LDHJ granules for cell experiments; Step 4: Confirmation of the protective role of three active substances of LDHJ granules on the cholestasis-related hepatocyte apoptosis through regulating CaSR.