A total of 140 healthy SPF grade BALB/c male mice, aged 6-8 weeks and weighing 18-22 g, were provided by Hunan Shrek Jingda Experimental Animal Co., Ltd., license No.: SCXK (Hunan,China) 2016-0002. Raised in the barrier system of the Animal experiment Center of Dali University, the temperature is 20-26 °С, the humidity is 40-70 %, and the pressure difference is 25-50 Pa, and the light and shade alternated at 12 h. All experimental studies are in line with the guiding principles of animal research of the China Ethics Committee.
Drugs and reagents
Dextran sulfate sodium (Shenzhen Regent Co., Ltd., batch number: SLBP0889V) oxalazine sodium (Tianjin Lisheng Pharmaceutical Co., Ltd., batch number: 1608005); Red blood cell lysate (Thermo Fisher Co., Ltd., batch number: 4338657); PBS (Solarbio Co., Ltd., batch number: 11310220);Fetal bovine serum (Zhejiang Tianhang Biotechnology Co., Ltd., batch number: 20161013); Rat anti mouse CD3 FITC (batch number: 6053673), Rat anti mouse CD4 PE (batch number: 7023677), Rat anti mouse CD8 APC (batch number: 7081511) and Rat anti mouse CD25 Percp-cy5.5 (batch number: 7032624) are all from BD Co., Ltd.; Occult blood kit (batch number: 20160824) and Mouse IL-1β ELISA kit（batch number: 20170405）are all from Nanjing Jiancheng Bioengineering Research Institute and so on.
Electronic analytical balance (model: AL204- IC, Mettler-Toledo Instruments Co., Ltd.); Macro camera (model: DIGITAICAMERA D7100, Nikon); Medical image analysis system (model: BI-2000; Chengdu Tai Meng Technology Co., Ltd.); Flow cytometry (model: FACS Calibur; BD USA); Enzyme labeling instrument (model: SpectraMax M5 position Devices) and so on.
Preparation of main reagents
DSS (3%) configuration: The 120 g DSS was dissolved in 4 L hot-pressed ultra-pure water to obtain 3 % DSS solution, which was then prepared at that time.
The preparation of PBS buffer (PH=7.4): NaCl 8.0 g/KCl 0.2 g/Na2HPO4·12H2O 1.44 g/KH2PO4 0.24 g. Put the weighing reagent into the flask, add ultra-pure water to 1000 ml, and shake well, then sterilize with high pressure steam (121 °С , 15 min). Storing at 4 °С and set aside.
Preparation of spleen single cell suspension: The fresh spleen of mice was placed on a 200-mesh cell sieve and ground into a single cell suspension with the core of a 5 ml syringe. The cell suspension was collected and centrifuged, and the supernatant was discarded. Adding 1 ml red blood cell lysate, after the red blood cell is fully lysed, add 3 % FCS-PBS of 4 ml to wash twice, then discard the supernatant, and add RPMI-1640 complete culture medium to re-suspend spleen single cells. The cell suspension was stained with 0.4 % trypan blue staining, and the cell concentration was adjusted to 107/mL.
Establishment of experimental model
One hundred and forty healthy male BALB/c mice were fed adaptively for one week. Except for the normal group, the mice were given 3 % (w/v) DSS aqueous solution to drink freely for 7 days. After the establishment of the model, the mice were fed with hot pressure sterilized ultra-pure water for 10 days, and the whole process takes a total of 17 days .
Grouping and administration
Eighteen mice were randomly selected as the normal group, and the rest of the model mice were evaluated by disease activity index (DAI) at the 7th day. According to the DAI score, the model mice with very mild inflammation were excluded (Very mild inflammation at 0-3, mild inflammation at 4-6, moderate inflammation at 7-9, and severe inflammation at 10-12). The rest of the mice were randomly divided into model group and oxalazine group (600 mg·Kg-1) according to the severity of inflammation, with 24 mice in each group (The number of mice killed during modeling was not included in the model group). At the end of the model, each group was given corresponding drugs according to 0.1 ml·kg-1·d-1, while the model group and normal group were given 0.1 ml·kg-1·d-1 saline once a day for 10 day. The mice in the normal group and the model group were scored by DAI at 0, 1, 4, 7, 8, 10, 14 and 17 day respectively, and the drug group was scored by DAI at 8, 11, 14 and 17 day after the experiment. Except for the normal group, 6 mice were randomly killed on the day of DAI score in other groups  (In the normal group, the mice were killed at 0,7 and 17 day to observe the effect of environment on the experiment).
General status observation
The changes of body mass, drinking water, diet and fecal characteristics of mice in each group were observed and recorded during the experiment.
Measurement of colonic length and calculation of organ index in mice
The mice were killed after taking blood from their eyeballs. The liver, spleen, lung, thymus and colon were taken, and the organ mass was weighed. The organ index (organ index = organ mass (mg)/mouse body weight (g) ) was calculated. The intestines were dissected longitudinally along the mesenteric edge of mice, the intestinal contents were cleared, the colon was weighed and the length of colon was measured.
Disease activity Index (DAI) score
According to the standard of Hamamoto et al. , the weight of mice was weighed, the characteristics of feces and occult blood in feces were observed. According to the score of Table 1, the scores of weight loss, fecal traits and fecal occult blood were summed up, and the disease activity index of each mouse was calculated to evaluate the disease activity.
The removed colon was laid flat on white paper, and the colonic mucosal injury was observed and scored according to Ekstr ö m GM standard  and Luk standard  (Table 2).
Observation of colonic pathomorphology and HS score
"Swiss rolls" is a routine method for histological and immunohistochemical staining of intestinal tissue: the longitudinally sectioned colon is cut in half, wrapped, fixed and preserved in 10% neutral formalin solution for HE staining. The pathological sections were observed under light microscope according to the standard of Ekstr ö m GM et al.  (Table 3), and the colon HS score was performed.
Determination of lymphocyte subsets
100 μL spleen cell suspension (106 spleen cells) was added to the flow tube, and the corresponding mixed fluorescent antibody was added for FACS Calibur detection.The ratio of T lymphocytes (CD3+) to single spleen cells and the proportion of Th cells (CD3+CD4+CD8-), Tc cells (CD3+CD4-CD8+) and Treg cells (CD3+CD4+CD25+) to T lymphocytes in spleen were analyzed.
Detection of 1 β content
The blood extracted from eyeballs of mice was placed at 4 ℃ for 4 h, and the supernatant was obtained by centrifugation at 3000 rpm (4 ℃) for 10 min. The other half of the longitudinally dissected colon was made into 10% homogenate in an ice bath, and the supernatant was obtained by centrifugation at 3000 rpm (4 ℃) for 10 min. According to the instructions, the contents of related biochemical factors in serum and colonic mucosa were determined by ELISA method, and the data were calculated by ELISACalc.
SPSS 23.0 and GraphPad Prism 5.0 were used to describe and analyze the data. Rank sum test is used to compare the rank data, and the concentrated trend and discrete trend are expressed by median (M) and quartile (QR) spacing, respectively. The measurement data are expressed as , the data with normal distribution and uniform variance are analyzed by t-test and one-way ANOVA, and the data that are not consistent with normal distribution are tested by rank sum test. The continuous data were analyzed by repeated analysis of variance and univariate analysis of variance. LSD test was used for pairwise comparison between groups. P<0.05 was used as the standard of statistically significant difference.