Shaoyao Decoction Prevents Colitis-Associated Colorectal Cancer Through Limiting IL-6/STAT3 Signaling Pathway

Objective: Through measuring IL-6, STAT3 and related molecules in colitis-associated colorectal cancer(CAC) model, we aim to demonstrate that Shaoyao decoction(SYD) prevents CAC through limiting IL-6/STAT3 signaling pathway and its downstream IL-17 A immune reaction and epithelial-mesenchymal transition(EMT).Methods: Male C57BL/6J mice aged 6 to 8 weeks were maintained in a specific pathogen-free(SPF) environment with food and water provided. They were randomly divided into three groups: 10 mice in control group, 30 in model group and 30 in SYD group. AOM(10mg/kg) was injected intraperitoneally on day 1. Seven days after the AOM injection, mice were given 2.5% DSS in the drinking water for 7 days and maintained on regular water for 14 days. Then they were subjected to two more cycles of 2.5% DSS treatment(7 days/cycle), each separated by 14 days of regular water. SYD(16g/kg) or equivalent normal saline was administered by gavage using a tube at day 1 and continued till harvest. The mice were killed by cervical dislocation at day 70. Serum samples were collected. The colons from the ileocecal junction to the anal verge were removed. The number of tumors in each mouse was counted. The size of each identified tumor was determined by measuring the largest diameter. Histological changes in the colonic tissues were examined by standard hematoxylin and eosin(H&E) staining. Scoring of colonic inflammation and proliferation were conducted in a blind method with a scoring system described previously. The levels of IL-6 and IL-17 A were measured by ELISA using commercial kits. The protein levels of STAT3, pSTAT3, Cyclin D1, Bcl-2 and Snail were determined by Western blot. The expressions of Ki-67, E-cadherin, N-cadherin, fibronectin and vimentin were examined by immunohistochemistry. The mRNA expressions of IL-6, STAT3, IL-17 A and Snail were quantified by Quantitative Real-time PCR. All data were presented as mean ± SD and analyzed using SPSS 19.0. Differences were compared with 2-independent samples T-test or one-way analysis of variance(ANOVA). A value of P < 0.05 was considered as statistically significant.Results: The number and size of tumors were decreased in SYD group. And the number of Ki-67-positive cells was found to be significantly decreased by SYD administration. SYD inhibited IL-6 expression and secretion, and subsequent STAT3 activation. It lowered the protein levels of Cyclin D1 and Bcl-2 significantly. Besides, SYD showed a marked reduction in the expression of IL-17 A. It significantly increased the expression of E-cadherin and decreased the expression of N-cadherin, fibronectin, vimentin and Snail.Conclusions: We demonstrate for the first time that SYD prevents CAC through limiting IL-6/STAT3 signaling pathway and its downstream IL-17 A immune reaction and EMT.