Senescence Of Colorectal Cancer Cells Induced By Millet Peroxidase And Its Application

Cell sensescence is ubiquitous phenomenon in biology.aging occurs mainly by the activation of proto-oncogenes,reactive oxygen species and DNA damage reagents factors,phenotypes such as nuclear enlargement,loss of fibrillar protein B1,and increase of B-galactoglase activity also occur,which also causes cell cycle arrest and activation of p53-p21,p16-p Rb and other signaling pathways.Tumor cells that break out of the bondage of aging,break through the boundary of Hayflick,and can proliferate indefinitely.Therefore,inducing the aging of tumor cells is a potential means to treat cancer.Colorectal cancer is one of the most common cancers in developed and developing countries.One of its cellular characteristics is reduced cell senescence,which is a powerful tumor suppressive mechanism.Induced cell senescence can inhibit tumor cell grouth.Fluorouracil drugs is the core of the treatment of colorectal cancer drugs,but the formation of tumor reduced susceptibility and resistance to chemotherapy is the important factors influencing the treatment effect,will induce drugs and tumor therapy combined with aging,not only can increase the tumor sensitivity to drugs,but also weaken the resistance of death of tumor cells.Proso millet peroxidase(PmPOD)used in this project is obtained from millet bran.Previous studies have shown that it can promote dependent programmed Necroptosis of HCT116 and HT29 cells.It has been proved that PmPOD,as a DNA damage agent,can cause DNA damage and cell cycle arrest of colorectal cancer cells,which is also the main reason for Necroptosis of HCT116 and HT29 cells induced by PmPOD.Whether DNA damage induced by PmPOD in HCT116 and HT29 cells can promote the senescence of colorectal cancer cells needs further study.This topic is explored from the following two aspects:MTT,immunofluorescence,SA-β-gal staining,fluorescence quantitative PCR and other methods were used to explore whether PmPOD can induce the aging of colorectal cancer cells.Results show that:PmPOD inhibit colon cancer cells activity in concentration dependent manner,observed by the SA-β-Gal dyeing blue in senescent cells,aging markers Lamin B1 degradation occurs,the nucleus,genes involved in aging m RNA and protein expression level of increase,these studies indicate that PmPOD induced aging HCT116 cells.Secondly,si RNA p53 and p53 overexpression were used to study the relationship between p53 and PmPOD-induced cell senescence by transfection technology.The results showed that the degradation of Lamin B1 induced by PmPOD was regulated by p53,and the SA-β-gal staining was not directly related to p53.Finally,the relationship between PmPOD-induced aging and DNA damage was explored through experiments.The results showed that PmPOD could degrade PARP-1 in colorectal cancer cells and decrease the content of NAD~+in the cells,which indicated that the reduced activity of PARP-1 weakened the DNA damage repair effect,and then promoted the aging of tumor cells.The combined action of 5-fluorouracil and PmPOD was conducted to explore the potential application of PmPOD-induced tumor cell aging in enhancing chemotherapeutic drug sensitivity.In this study,SA-β-gal,Flow cytometry,Western blot and other techniques were used to detect the mechanism of PmPOD and 5-Fu combined action on HCT116cells.The results showed that the combined action of PmPOD and 5-Fu reduced the aging effect of PmPOD induced colorectal cancer cells,and promoted the necrosis of HCT116 cells.