Study On The Chemotherapeutic Resistance Mechanism Of LRP Protein In Colorectal Cancer

Part One The expression of LRP in colorectal cancerObjective: We do scientific research so as to explicit the expression of LRP in colorectal cancer.Methods:1 Western blot and Real-time PCR were applied to detect the expression of LRP protein in colorectal cancer tissues and paired normal colorectal tissues.2 Immunohistochemical technique was used to detect the expression of LRP protein in the microarray of colorectal cancer tissues.Results:1.Western blot,RT-pcr showed that LRP protein expression is higher in colorectal cancer than in normal colorectal tissue.2.Tissue microarray showed that LRP protein expression is higher in colorectal cancer.Part Two Effect of LRP protein on biological characteristics of colorectal cancer cell lines and its mechanism of chemoresistanceObjective: We study the effect of LRP on the biology of colorectal cancer cells and explore the effect of LRP protein on chemosensitivity of colorectal cancer.Methods:1 We knocked down the expression of LRP and observed the growth and proliferation of colorectal cancer cells.2 We suppressed the expression of LRP and observed the invasion and migration of colorectal cancer cells.3 Compared to Colorectal cancer cell transfected with control siRNA,cells transfected with LRP specific siRNA were more sensitive to 5fluorouracil or not.4 SW480(p53 mutant cells),HCT116(p53 wild-type cells)were treated by DDP or 5 fluorouracil and wild type p53 plasmid were transfected.Western blot was used to detected the expression changes of LRP.5 Targetscan software was used to predict the upstream regulators of LRP.6 Luciferase Reporter and Western blot were used to verify the regulation of mi RNA on LRP.7 Western blot was used to verify that whether p53 could regulate the expression of LRP via mi RNA.Results:1.RTCA assay demonstrated that LRP low-expression could inhibit the proliferation of HCT116.2.We found that the invasion and migration of colorectal cancer cells were suppressed when we knocked down the expression of LRP(p<0.05).3.Inhibition of LRP expression could enhance the chemosensitivity of colorectal cancer cells to 5FU.4.The wild p53 could down-regulate the expression of LRP,while when the p53 mutated,the inhibition of LRP was lost.5.We predicted that mi R-34 a may regulate the expression of LRP through Targetscan software.6.Luciferase Reporter and Western blot showed that mi R-34 a could directly target the 3'UTR of LRP and regulate the expression of LRP.7.P53 could regulate the expression of LRP through mi R-34 a.Conclusions: In colorectal cancer tissues,the expression of LRP is higher.Inhibition of LRP expression can suppress the proliferation and clone formation ability of colorectal cancer cells,and suppress the invasion and distant metastasis of colorectal cancer cells.Inhibition of LRP expression could enhance the chemosensitivity of colorectal cancer cells to 5FU.In addition,p53 can regulate the expression of LRP through mi R-34 a,the wild type p53 could down-regulate the expression of LRP.However,when p53 mutated,it lost the ability to inhibit LRP and lead to the high-expression of LRP.