Effect Of Lutein On High Expression Of Antioxidant Enzymes In Human Colon Cancer HT29 Cells Via P38MAPK Pathway

Objective:Colon cancer is one of the most common malignant tumors of the digestive tract.At present,the morbidity and mortality rate in our country ranks the forth in a variety of malignancies,ranking the third in the incidence of gastrointestinal cancer.The majority of patients in the diagnosis is already in the late because of its characteristic of the incidence of hidden.Now the main treatment program for colon cancer is still a simplification,that is only to rely on surgical resection,matching with the help of radiotherapy and chemotherapy drug.However,the treatment results are far away from satisfactory,colon cancer patients who has taken in colon resection for 5 years only have 50% survival rate,with a high risk of metastasis and recurrence.The frequent application of a variety of radiotherapy and chemotherapy drugs to the patient's psychology not only brought intolerable pain,but also caused irreparable permanent damage to the body.Therefore,it is of great theoretical and practical significance to find a high-efficiency,low-toxicity and plant-derived compound for the prevention and treatment of colon cancer,it is also significance to reduce the adverse reactions of radiotherapy and chemotherapy including improve the survival rate of patients.Lutein,as an antioxidant plant compound,can help the body to quench the singlet oxygen,scavenge active oxygen free radicals and prevent from its invasion of biofilms,delaying the aging of the body,and can fight against oxidation stress,inflammation and angiogenesis.It has important practical significance for the prevention of all kinds of tumor,including skin cancer,stomach cancer,prostate cancer,esophageal cancer,breast cancer and so on.The aim of this study is to investigate the mechanism of lutein in inhibiting the development of tumor by significantly increasingly up-regulating the expression of antioxidant enzymes in human colon cancer HT29 cells.Method:1.The growth curve of HT29 cells was measured by CCK-8 method for 8 days.2.Inhibitory effect of different concentrations of lutein on proliferation of HT29cells.The proliferation of HT29 cells was detected by CCK-8 method when the concentrations of lutein at 40,80,120 and 160 mg · L-1 for 24,48 and 72 h,respectively.3.Effects of different concentrations of lutein on cell cycle distribution of HT29 cells.Cell cycle distribution of HT29 cells were examined by flow cytometry after 48 h with different concentrations of lutein in with HT29 cells.4.Effects of different concentrations of lutein on m RNA and protein expression of HO-1,NQO1,SOD2 in HT29 cells.The expression of HO-1,NQO1 and SOD2 m RNA in the cells was detected by RT-q PCR after 48 h of different concentrations of lutein dealing with HT29 cells.Western blot was used to detect the protein expression level of HO-1,NQO1,SOD2 in HT29 cells.5.The effect of p38 MAPK pathway on the expression of HO-1,NQO1,SOD2 m RNA and protein in the downstream of Nrf2/ARE pathway in HT29 cells was studied.The m RNA expression level of HO-1,NQO1 and SOD2 in the cells was detected by RT-q PCR and the protein expression level of HO-1,NQO1 and SOD2 in the cells was detected by Western blot for 48 h incubating with lutein on the HT29 cells after being dealed with SB203580 for 1 h.Result:1.The results of CCK 8 showed that lutein significantly inhibited the proliferation of human colon cancer HT29 cells,and there was time-effect and dose-effect relationship(P <0.01).2.The results of flow cytometry showed that the cell cycle of HT29 cells was significantly blocked after treated with different concentrations of lutein for 48 h,and the number of cells in G0 / G1 phase was significantly increased,indicating that lutein can block the growth of HT29 cell in G0 / G1 phase.3.The results of RT-q PCR and Western blot dealing with HT29 cell with different concentrations of lutein after 48 h indicated that lutein could simultaneously up-regulate the expression of HO-1,NQO1 and SOD2 at the m RNA transcription level and protein expression level compared with the negative group(P <0.05).4.The expression level of HO-1,NQO1 and SOD2 m RNA and protein expression level in the lutein+ SB203580 group were significantly lower than that of the onlylutein group(P<0.05),indicating that the inhibitor SB203580 significantly inhibited the upregulation of lutein inducing the m RNA and protein levels of HO-1,NQO1 and SOD2 in the cells.Conclusion:Lutein has a significant inhibitory effect on the proliferation of human colon cancer HT29 cells,which may be mediated by blocking the HT29 cell cycle,acting on the p38 MAPK pathway to mediate Nrf2 / ARE at gene transcription levels and protein expression levels and upregulating the expression of antioxidant enzymes HO-1,NQO1,SOD2 m RNA and protein in the downstream of the Nrf2 / ARE signal pathway,and further inhibit the development of tumor.