2.1 Animal studies
Male C57BL/6 mice (6-8 weeks old, 20 ± 2 g) were purchased from Guangdong Experimental Animal Center (license number: SCXK (YUE) 2018-0002). All animal experiments were conducted according to Guangzhou University of Chinese Medicine’s relevant principles (approved No. 20201013002). All the mice were raised in the SPF feeding room with a 12 h light/ 12 h dark cycle and free water intake. The first animal experiments were designed as follows: after one week of adaptive feeding, they were randomly divided into five groups: control group (distilled water for 7 days), DSS group (4% DSS for 7days), and TBⅡ-treated groups with 4% DSS for 7days (25, 50 and 100 mg·kg-1) (Zhao et al., 2016), ten mice were used per group. The body weight, fecal status and hematochezia of all mice were observed every day, and scored by an experimenter who was unaware of the scheme according to the standard scoring system. Therefore, the disease activity index (DAI) was calculated as ((body weight loss score + feces status score + hematochezia score )/3).
For the second animal experiments, the mice were randomly divided into three groups: control group (distilled water for 12 weeks), CAC group was given azoxymethane (AOM) 10 mg·kg-1 on the first day. What’s more, the second, fifth and eighth weeks were given 2% DSS and the remaining weeks were given distilled water. TBⅡ-treated group was treated the same as the CAC group, plus oral administration TBⅡ (100 mg·kg-1) for 12 weeks. Body weight was recorded during the experiment, the number and size of tumors were also recorded.
Male NLRP3-/- mice (C57BL/6 background, #021302, purchased from Jackson Laboratory (Bar Harbor, ME, USA)) and wide type mice (WT) were randomly assigned to eight groups: WT-control group (distilled water for 7 days), WT-DSS group (4% DSS for 7days), WT-MCC950-treated groups with 4% DSS for 7days (10 mg·kg-1), WT-TBⅡ-treated groups with 4% DSS for 7days (100 mg·kg-1), NLRP3-/--control group (distilled water for 7 days), NLRP3-/--DSS group (4% DSS for 7days), NLRP3-/--MCC950-treated groups with 4% DSS for 7days (10 mg·kg-1), and NLRP3-/--TBⅡ-treated groups with 4% DSS for 7days (100 mg·kg-1). During the treatment, weight loss, stool consistency, and fecal blood were also recorded.