The Expression And Biological Function Of PTBP1 In Colon Cancer

BackgroundColon cancer as a disease of digestive tract tumor has a high incidence rate, excepting lung cancer and breast cancer. Among various kinds of tumor disease, the mortality of colon cancer is higher. There are varieties of factors inducing colon cancer, such as environment, diet and genetics. The things of organism self also lead to cancer, such as the activation of proto-oncogenes or inhibition of tumor suppressor genes. The complex inducing reasons of colon cancer revealed the mechanism of colon cancer morbidity is more complex. So, to clarify the function of proto-oncogenes or tumor suppressor genes in colonic cancer occurrence is very helpful for the treatment of colon cancer.As a member of the family of heterogeneous nuclear rib nucleoproteins,PTBP1 can bind to RNA through four different binding areas. As binding to RNA, PTBP1 has the functions of changing the RNA structure and regulating alternative splicing of RNA. PTBP1 shuttles between the nucleus and cytoplasm when cells stimulated by outside pressure such as virus infection and toxic exposure. PTBP1 exerts some functions in the cell nucleus, such as regulating alternative splicing of RNA and enhancing the transcription of genes through combining with UTRS. In the cytoplasm, PTBP1 has functions of increasing the stability of the m RNA through combine with 3’UTR, regulating the m RNA positioning and transduction. PTBP1 can enhance the B cell function by regulating insulin secretion, regulate cell cycle and apoptosis related proteins expression, farther more affect the cell division and differentiation, and other important life activities. PTBP1 have a certain relationship with embryonic development, neurons differentiation, type 2 diabetes and cancer.Many researches have shown that PTBP1 are highly expressed in breast and ovarian cancer. Suppression of PTBP1 can obviously reduce the proliferation ability of breast and ovarian cancer cells.But contrary to this conclusion, in non-small cell lung cancer cells, overexpression of PTBP1 have the ability of inhibiting cell proliferation. So far, PTBP1’s role in colon tumorigenesis is still unclear. As a high incidence of colon cancer, this paper aims to investigate the biological function of PTBP1 in colon cancer occurrence and development, which is useful for the treatment of colon cancer.Chapter I The expression of PTBP1 in colon cancer with Tissue MicroarrayObjectiveTo detect PTBP1 protein levels in human colon adjacent tissues and colon cancer tissues with tissue microarray.Methods1. Slected 90 cases of colon cancer and embedded them with paraffin.2. Made tissue microarray with 90 cases of embedding paraffin.3. Analalied tissue microarray.ResultPTBP1 was higher expressed in colon cancer tissues than in cancer adjacent tissues; PTBP1 overexpression was associated with colon cancer lymphatic metastasis and staging.ConclusionPTBP1 overexpression is a character of colon cancer and can be a biomarker in colon cancer lymphatic metastasis and staging.Chapter II The influence of PTBP1 in biobehavior of colon cancer tumorigenesisObjectiveTo analyze the expression of PTBP1 in different colon normal and cancer cells. Inhibited the PTBP1 protein levels in colon cancer cells then observe the changes in cell viability, cell cycle and apoptosis.Methods1.Expression of PTBP1 in FHC/HCT116/SW480/HCT8 cells was measured by Western blot.2.Inhibited the expression of PTBP1 in HCT116 and SW480 cells with PTBP1 si RNA and then measured the cell viability by MTS agent and proliferation by Ed U assay.3.HCT116 and SW480 cells were firstly transfected with PTBP1 si RNA. Measured the cell cycle changes of PTBP1 knockdown cells by Flow cytometry analysis.4.Measured apoptosis of PTBP1 knockdown cells by Flow cytometry analysis and Hoechst33258 agent. HCT116 and SW480 cells were firstly transfected with PTBP1 si RNA.5.HCT116 cells were firstly transfected with PTBP1 si RNA. Measured the cell viability of PTBP1 knockdown HCT116 cell exposed to 5-FU, Cisplatin and Doxorubicin for 48 h by MTS agent.Result1.The m RNA and protein levels of PTBP1 were highest in HCT116 cells.2.The cell viability and proliferation were decreased when PTBP1 silenced in HCT116 and SW480 cells.3.The cell cycle was blocked in S phase when PTBP1 silenced in HCT116 and SW480 cells.4.The number of apoptotic cells was increased when PTBP1 silenced in HCT116 and SW480 cells.5.The sensitivity of PTBP1 knockdown HCT116 cells to 5-FU, Cisplatin and Doxorubicin was increased compeared to control group.ConclusionPTBP1 can regulate the growth of tumor cells. Inhibiting PTBP1 expression can reduce the growth of tumor cells, the reason for this is that PTBP1 knockdown can decrease cell proliferation, induce cell cycle arrest and apoptosis. Therefore, PTBP1 is a typical oncogenes in colon tumorgenesis.Chapter The mechanism of PTBP1 function in colon cancer cellsⅢObjectiveTo investigate the molecular mechanism of PTBP1 regulating HCT116 cell viability, proliferation, cell cycle and apoptosis.Methods1.Measured the p85/p-AKT protein levels in HCT116 cells transfected with NC si RNA and PTBP1 si RNA by Western blot.2.Measured the proteins associated with cell cycle(cyclin D1/CDK4) in HCT116 cells transfected with NC si RNA and PTBP1 si RNA by Western blot.3.Measured the proteins associated with apoptosis(p53/cyto C/Bcl-2/Bax/PARP1/Caspase3) in HCT116 cells transfected with NC si RNA and PTBP1 si RNA by Western blot.Result1.The expressions of p85/p-AKT were decreased following the PTBP1 silence in colon cancer cells.2.The expression of cyclin D1 was decreased following the PTBP1 silence in colon cancer cells.3.The expression of Bcl-2 was decreased and Bax/PARP1/caspase3/p53/cyto C was increased following the PTBP1 silence in colon cancer cells.ConclusionSupression of PTBP1 can inhibited the expression of proliferation associated proteins such as p-AKT/p85, cell cycle protein cyclin D1, apoptosis protein Bcl-2, increased the expression of apoptosis proteins such as p53/cyto C/Bax and activated PARP1/Caspase3.