| Recently, the mortality and morbidity of colorectal cancer have been rising steadily. Although surgical treatment combined with chemotherapy improved the prognosis, the survival rate remained unsatisfied for early diagnosis could not achieved for most of the patients. To discover the risk factors that related to the initiation and progression of colorectal cancer and early intervention may be a promising method to reduce the incidence of colorectal cancer and personalized treatment.Experts raised the theory of "tissue organization field", emphasizing the role of tissue microstructure and cellular microenvironment in tumor genesis. The abnormality of tissue organization field leads interruption to the communication between cells or cell and mesenchyme, which results in the uncontrolled proliferation.The balance between promotion and inhibition effects in inflammatory microinvirenment determind the development of the tumor. All the tumor stages, including initiation, promotion and grogression are closely associated with the development and function of immune system.T helper17cells (Thl7), the major source of Interleukin-17, is a recently defined lineage of CD4+effector T cells. As an important part of tumor inflammatory immunal microenvironment, Th17cells become the research focus in tumor genesis. They present with different function in different immune background and different tumor type.Epithelial-mesenchymal transition (EMT) is a key link in the progression of colorectal cancer. Inflammatory immunal microenvironment is closely related to tumor EMT, while the relationship between Th17cells and tumor EMT is not clarified.In this research, we investigated the variation tendency of T helper17cells in different stages of colorectal cancer, accessed the prognostic value of Th17cells for colorectal cancer. Discussed the impacts of Th17cells on the initiation and grogression of colorectal cancer. In the meanwhile, we built the steadily IL-17overexpressing colorectal cancer cell strain, tried to investigate the possible connection between Th17cells and the EMT of tumor cells on cellular and molecular levels. Providing the basic for futher research in vitro and in vivo.Part I The Variation Tendency of T Helper17Cells in Different Stages of Colorectal CancerObjectives:To clarify the variation tendency of T helper17cells in different stages of colorectal cancer.Methods:We collectted the peripheral blood sample of20healthy volunteer and20colorectal cancer patients respectively. Then measured the concerntration of IL-17and proportion of Th17cells in CD4+cells through ELISA and flow cytometry. Dealed the pathologically comfirmed normal epithelium, adenoma tissue, early cancer tissue and advanced cancer tissue with immunohistochenmical staining technique.20samples in each group. Counted the Th17cells (IL-17+) that infiltrated into the tissue.Results:The concerntration of IL-17and proportion of Th17cells in CD4+cells were elevated in the peripheral blood of colorectal cancer patients (p<0.05). In the classic sequence of "normal epithelium-adenoma-early cancer-advanced cancer", Th17cells that infiltrated in the tissue increased gradually. The Th17cells indensity in the cancer tissue was significantly higher than that in normal or adenoma tissue, and the Th17cells indensity in the advanced cancer tissue was significantly higher than that in early cancer tissue (p<0.05)Conclusions:Th17cells and its major cytokine, IL-17, were closely related to the initiation and progression of colorectal cancer.Part II A Study on the Association between T Helper17Cells and Prognosis of Colorectal CancerObjectives:To discuss the relationship between indensity of infiltrating Th17cells and the clinicopathological features of colorectal cancer, and access its value in prognosis of colorectal cancer.Methods:Tissue microassay was constructed with the tissues from90 patients who underwent radical colonectomy in Zhongshan Hospital from May2007to April2008. The microassay was dealed with immunohistochenmical staining. The indensity of infiltrating Th17cells in the tissue was measured. The samples falled into the group of high Th17cells indensity or low Th17cells indensity. The relationships between indensity of infiltrating Th17cells and the clinicopathological features as well as prognosis of colorectal cancer were analyzed.Results:More Th17cells were detected in the cases with vascular invasion (p=0.031). The survival curve analysis indicated that the accumulated survival rate of Th17high group was significantly low than that of Th17low group (P=0.001). Advanced TNM stage (p=0.009), poor differentiation (p=0.036) and high infiltrating Th17cell indensity (p=0.017) were indenpent prognostic factors for colorectal cancer.Conclusions:The infiltrating Th17cell indensity in tumor tissue was indenpent prognostic factor for colorectal cancer. It was negatively correlated with the prognosis of colorectal cancer.Part III A Pilot Research on the Mechanism of Promotion Effect of T Helper17Cells in the Initiation and Progression of Colorectal CancerObjectives:To build the steadily IL-17overexpressing colorectal cancer cell strain, and to explore the mechanism of promotion effect of T Helper17Cells in the initiation and progression of colorectal cancer.Methods:Built the steadily IL-17overexpressing colorectal cancer cell strain through transfection. Evaluated the variation of the abilities of anti-apoptosis, proliferation and invasion when effeced by the overexpressed IL-17by flow cytometry, CCK8test and Transwell test, respectively. Analyzed the relationships between expression of E-cad and prognosis of colorectal cancer as well as the indensity of infiltrating Th17cells. Investigated the possible connection between Th17cells and the EMT of tumor cells. RT-PCR was applied to test the expression level of biological marker of EMT in the IL-17overexpressing cells.Results:We obtained the IL-17overexpressing SW480. Flow cytometry, CCK8test and Transwell test indicated that when impacted by IL-17, abilities of anti-apoptosis, proliferation and invasion were significantly improved. The survival analysis showed that the expression of E-cad in tissue was positively correlated with the prognosis of colorectal cancer (p=0.006), and it was an independent prognostic factor for colorectal cancer (p=0.033). The expression of E-cad was negatively correlated with the indensity of infiltrating Th17cells (r=-0.267, p=0.011).indicating Th17cells was closely related with the EMT of colorectal cancer. The biological markers for EMT were tested by RT-PCR, in the IL-17overexpressing cells, the expression of E-cad was downregulated, while the expressions of Snail,3-catenin and Vimentin were upregulated (p<0.05), no significant change of the expressions of N-cadherin and Twist was observed (p>0.05)Conclusions:Thl7cells promoted the initiation and grogression of colorectal cancer, the promotion might be carried out by encouraging the EMT of the colorectal cancer. |
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