Inflammation-induced S100a8/a9Activates Id3and Promotes Colorectal Tumorigenesis

Objective:Colorectal cancer is one of the most common malignant tumor of alimentary tract, and the occurrence and development of colorectal cancer is closely associated with ulcerative colitis. While, the pathogenesis of colitis-associated cancer (CAC) is still unclear.Ca2+-binding proteins S100family members S100a8and S100a9are released in abundance in non-resolving inflammation such as inflammatory bowel disease and rheumatoid arthritis. Strong up-regulation of S100a8/a9was found in gastric, breast and bladder cancer. And it is becoming increasingly clear that S100a8/a9involved in tumor cell growth and metastasis.In our previous study, we employed a global view and a time-course manner to dissect the molecular events of CAC mouse model. Then we noted that the expression levels of S100a8and S100a9were dramatically increased from inflammation to cancer compared to normal tissue. These results led us to speculate S100a8and S100a9are key molecules in the development of CAC. Therefore, we focused on the pathogenesis of inflammation-induced S100a8/a9in colorectal tumorigenesis, which is expected to provide new targets for tumor biological therapy.Methods:The expression of S100a8and S100a9in the CAC mouse model and in the human colorectal adenoma, colorectal carcinoma, ulcerative colitis and normal colorectal tissue were detected by immunohistochemical staining, GEO DataSets analysis and qRT-PCR. The mechanism of S100a8/a9’expression and secretion in macrophage RAW264.7was verified by ELISA and qRT-PCR. The effects of S100a8and S100a9protein on cell proliferation and migration were detected by flow cytometry and Transwell assay. And the changes of gene expression profile in CT26.WT cells induced by S100a8and S100a9stimulation were assayed by Affymetrix gene array, and then the biological functions of key pathways and genes were verified.Results:S100a8and S100a9are up-regulated in the whole process of inflammation-linked cancer. The gene expression levels of S100a8and S100a9from the GEO database were significantly up-regulated in colorectal adenoma, colorectal carcinoma and ulcerative colitis compared to normal cohorts. In addition, from58adjacent normal colorectal tissues and104CRC specimens, we revealed that the expression levels of S100a8and S100a9were higher in CRC than in adjacent normal colorectal mucosa. And the results suggested that there were close correlations between the expression levels of S100a8and tumor metastasis and differentiation.LPS increased the expression and secretion of S100a8and S100a9significantly in the macrophage RAW264.7. Pretreated with either p38MAPK or PI3K inhibitor reversed the LPS’s induction on S100a8, while pretreated with NF-κB inhibitor and p38MAPK inhibitor reversed the LPS’s induction on S100a9. We also found that S100a8and S100a9can promote the recruitment of macrophages RAW264.7and promote the proliferation of colon cancer cell CT26.WT.To determine the mechanisms by which S100a8/a9promoted CAC tumorigenesis, we used the Affymetrix gene expression microarray system to study the different expressed genes induced by S100a8and S100a9recombinant protein stimulation. The results showed that TGF-β signaling pathway and its downstream gene, especially for Id3, were activated significantly by both S100a8and S100a9stimulation. We also discovered that S100a8/a9induced the expression of Id3through Akt-TGF-β/Smad5signaling. And Id3can inhibit p21expression and then promote cell-cycle progression.Conclusion:S100a8and S100a9are up-regulated in CAC mouse model and human colorectal cancer specimens, which play an important role in the colitis-associated cancer. Besides, S100a8and S100a9promote the recruitment of macrophages RAW264.7and promote the proliferation of colon cancer cell CT26.WT. In addition, S100a8/a9promotes the proliferation and cell-cycle progression of colon cancer cell through Aktl-TGF-β/Smad5-Id3-p21signaling axis.9figures,3tables,55...