The Anti-colorectal Tumor Effect And Mechanism Of Tea Polysaccharides On Tumor Biological Behavior In Tumor Inflammatory Microenvironment

The occurrence and development of tumor cells not only depend on themselves but also depend on the "soil" where the cells lived the tumor microenvironment.Two main cores which form the tumor microenvironment are the immunity and inflammation.The immune cells and immune molecules mediate the connection between the immunity and inflammation.Furthermore,they maintain a balance between the function of defense and the control of the excessive inflammation.The tranformation of macrophage phenotype in the tumor inflammatory microenvironment and the release of inflammatory factor were the key steps of theinflammatory related tumor's development.Currently,discovering and targeting thebioactive elements of plants that can resist tumor development in the tumor inflammatory microenvironment is a research hotspotChina is known as the hometown of tea tree(Camellia sinensis L.O.Kuntze),which is a major tea producer globally.Tea Polysaccharides(TPS)are the complex polysaccharides which possess biological activities in tea leaves.They are a kind ofacidic polysaccharides or acid glycoprotein combined with proteins.They showed good biological function,e.g.anti-virus,anti-inflammation,hypoglycemic,anti-tumor,immunity enhancement.Meanwhile,they have very few side effects to the body.The present study aimed at the influence and the mechanism that TPS worked on the biological behaviors of the colonic tumor cells in the inflammatory microenvironment.Furthermore,it focuses on making meaningful attempts and providing useful preliminary experimental data,therefore establish the scientific basis for TPS as functional foods and new pharmaceutical foods.This study discussed the bioactivities of TPS in preventing the colon tumor and the related mechanisms in vivo and in vitro.We targeted tumor inflammatory microenvironment and focused on the signal pathways IL-6/STAT3 to explore the anti-tumor activity of TPS and the molecular mechanisms.The research contents and results of this paper are mainly summarized as followed:To study the preventive and therapeutic effects of TPS on colon cancer in a mouse model of AOM/DSS induced colitis-associated colon cancer.The animals were intraperitoneally injected with AOM[10 mg/kg initial body weight(BW)],followed by one week interval and 7 days of 2%DSS in the drinking water,and then were subjected to two more 2%DSS cycles with 14-day interval of each.TPS at the dose of 50 mg/kg,100 mg/kg and 200 mg/kg BW were orally gavaged from Week 4,with AOM/DSS group as control.At the end of the 16th week,the blood samples of mice and organs such as colon,liver,spleen and thymus were collected.We observed the preventive.effect and treatment effect of TPS on colon cancer by using microscope and eyes,and megascopic tumors were calculated and surveyed with a caliper The results showed that:compared with the colitis-associated cancer(CAC)model group,the TPS groups showed a significant.inhibition of tumor in a dose-dependently.The weight of liver,spleen and thymus index showed that TPS had no obvious toxic effect to the organs.2.The effects of TPS on cell proliferation and apoptosis using TUNEL and Ki-67 immunohistochemical staining technique in the colon tissues of mice.The results showed that TPS significantly promoted the apoptosis of mice's colon tumor cells and restrained their proliferation.We also explored immunohistochemical and Western Blot to investigate the protein expression of cyclin D1,MMP-2 and MMP-9.The results showed TPS could significantly reduce these proteins' levels.The results of flow cytometry showed that TPS could lead the block of the CT26 cells in G0/G1 period and then restrained the cell proliferation.Meanwhile,the TPS group showed that the rate of cell apoptosis was obviously increased in a dose-dependent increase.3.To do the research from the aspect of inflammatory microenvironment,the ELISA method was used to detect the degree of homogenized inflammatory cytokines in the mice's colonic mucosal tissues The HE-staining and immunofluorescence technique were used to observe the inflammation and infiltration degree of the immune cells in the colonic tumor.The ELISA detection results showed that TPS could significantly decrease the degree of pro-inflammatory cytokines TNF-??IFN-??IL-6 and IL-2.At the same time,TPS could increase the concentration of anti-inflammatory cytokine IL-10.HE-staining results find that the aggregation of inflammatory cells in the colonic tumor was significantly restrained by TPS.By using Iimunofluorescence assay,we found that the infiltration degree of macrophage,B lymphocytes and MDSC in the tumor area were obviously restrained by TPS,and the inflammatory response caused by inflammatory cells was relieved.In addition,we focused on the change of the inflammatory signal pathway IL-6/STAT3.Detected by Western Blot,it was found that the phosphorylation activation of STAT3 and the expression levels of downstream target proteins were obviously restrained by TPS.TPS could control the development of colon tumor by restraining the inflammatory response in the tumor microenvironment.4.We established an in vitro non-contact co-culture system with macrophages,and observed the influence of the biologic behavior that TPS worked on the tumor cells CT26.Western Blot detected the phosphorylation level of STAT3 and the protein expression of downstream target proteins.The results showed that,TPS could affectthe ability of proliferation,migration and invasion of tumor cells by restraining the activation of STAT3 and the expression of downstream genes IL-6/STAT3.