Changes And Significance Of Lysophosphatidylcholine Acyltransferase 1 During Radiotherapy For Colorectal Cancer

Objective:Colorectal cancer has a high incidence rate in clinical tumors.Nowadays,the main treatment options for colorectal cancer are surgery combined with local radiotherapy and systemic chemotherapy.Despite advances in radiation therapy technology,the existence of resistance to radiation therapy is still an important cause of metastasis,recurrence,radiotherapy failure,and poor prognosis in patients with malignant tumors.Therefore,increasing radiation sensitivity is an important challenge.Lysophosphatidylcholine acyltransferase 1(LPCAT1)is an important enzyme in the phospholipid metabolism and occurs in colon cancer cells.The purpose of this study is to investigate the effect of LPCAT1 and combination radiotherapy on the spread of colorectal cancer line and to investigate the role and value of LPCAT1 in radiotherapy.Methods:We cultured the colorectal cancer cells SW620,SW480,and HCT116.Detect the expression level of LPCAT1 in three cell types by polymerase chain reaction(RT-PCR)and Western blotting,and select cell lines with higher LPCAT1 expression levels.By using lentivirus transfection technology to knock down the expression of LPCAT1 in cells,stable transfected knockdown group cells and negative control group cells were obtained.Cells in the knockdown group and the negative control group were irradiated with 6 Gy radiation,and the negative control combined radiotherapy group and knockdown combined radiotherapy group were obtained and set.The CCK-8 experiment was used to detect the cell viability of the negative control group,knockdown group,negative control combined with radiotherapy group,and knockdown combined with radiotherapy group.The clone formation experiment further detected the changes in the proliferation ability of the four groups of cells and plotted the cell survival curve.Subsequently,We used RT-PCR and Western blot to have detections of the relative expression of LPCAT1 in the cells of the knockdown combined radiotherapy group and the negative control combined radiotherapy group,to determine whether there is any change in the expression of LPCAT1 in the cells after radiotherapy.In order to further explore the impact of knockdown LPCAT1 combined with radiotherapy on radiosensitivity and the specific mechanism of colorectal cancer cells,the protein samples from the negative control group,knockdown group,negative control combined with radiotherapy group,and knockdown combined with radiotherapy group were collected for proteomic testing and bioinformatics analysis,and proteins related to the research topic were screened.RT-PCR was used for further validation in vitro experiments.The experimental data obtained were statistically and analytically processed using statistical software SPSS 23.0.The data measured between the two groups were analyzed by t-test and the comparison between several groups was made by variation analysis.Results:1.The relative content of LPCAT1 m RNA and protein in colorectal cells HCT116 was higher.Through lentivirus transfection technology,LPCAT1 expression in HCT116 cells can be downregulated,and colorectal cancer cells with knockdown LPCAT1 expression can be successfully constructed.2.Compared with the negative control group,the knockdown group had a decrease in cell viability,clonal colonies,and proliferative capacity.Under the effect of combined radiation therapy,the proliferation ability of cells further decreased(P<0.05).3.By detecting the content of LPCAT1 in cells after radiotherapy,the results showed that the content of LPCAT1 m RNA and protein in negative control cells decreased after irradiation(P<0.01).While the expression of LPCAT1 in the cells of the knockdown combined with radiotherapy group further decreased(P<0.0001).4.Proteomic techniques were used to analyze the intracellular differential proteins in each group,and the differentially expressed proteins were subjected to KEGG functional classification and functional enrichment analysis.CCNB1 was selected as the key protein for differential downregulation.Further RT-PCR validation found that compared with the negative control group,the relative expression of CCNB1 m RNA in the negative control combined with radiotherapy group and the knockdown combined with radiotherapy group decreased(P<0.0001);The m RNA of CCNB1 in the negative control combined with radiotherapy group was higher than that in the knockdown combined with radiotherapy group(P<0.01).Conclusion:Knocking down LPCAT1 can inhibit the proliferation ability of HCT116 cells,and knocking down LPCAT1 combined with radiotherapy can further reduce the proliferation ability of cells.After radiotherapy for colorectal cancer cells,the expression of LPCAT1 in the cells may decrease.Knockdown of LPCAT1 may increase the radiosensitivity of colorectal cancer cells by promoting the reduction of CCNB1.LPCAT1 may be a potential target for regulating radiosensitivity in colorectal cancer.