MiR-375/SLC7A11 Axis Regulates Proliferation And Invasion In Oral Squamous Cell Carcinoma

Research backgroundOral squamous cell carcinomas(OSCC),also known as oral cancer,are the most common malignant tumors of head and neck,and are characterized by epithelial aggressive tumors of different degrees of squamous differentiation and an inclination of early lymphatic metastasis.Clinically,the main therapy for OSCC is surgery assisted by radiotherapy and/or chemotherapy.However,its cure rate is not satisfactory.With the continuous development of modern biotechnology,people keep deepening their understanding of the mechanism of genesis and development of tumors and realize that tumors are caused by change of gene levels.Hence,to explore the mechanism of genesis and development of OSCC from levels of genes,transcription and protein function etc.can identify molecular markers of OSCC and carry out targeted therapy on the basis of the tumor regulation mechanism,which is of great significance for increasing its five-year survival rate and reducing its mortality.The microRNA is a type of non-coding RNA with short nucleotide sequences that plays an important role in regulating genesis and development of tumors.It was found which miR-375 shows differential expression in various cancers and serves as a tumor suppressor gene.In addition,SLC7A11 is a gene encoding functional subunit xCT of cystine/glutamate transporter.It was found which SLC7A11 shows high expression in numerous malignant tumors.Few studies have paid attention to miR-375 and SLC7A11 and their interaction in OSCC.Therefore,this study expected to provide new ideas for research on pathogenesis of OSCC and treatment of OSCC by studying expression of miR-375 and SLC7A11 in OSCC and their roles in regulating genesis and development of OSCC.ObjectiveThis study detected expression of miR-375 in tumor tissue of OSCC patients and discussed influences of miR-375 expression on proliferation and invasion of oral cancer cells.In addition,by mechanism research,it explored whether miR-375 signals regulate SLC7A11 expression in oral cancer cells and affect biological behavior of oral cancer through SLC7A11.This study helped to understand effects of microRNA in the mechanism of genesis and development of OSCC and provided molecular biomarkers for early diagnosis of oral cancer and an experimental basis for molecular targeting treatment.Methods(1)40 samples of oral cancer and related para-carcinoma tissue were collected.Culture of cell lines of oral cancer,FaDu,SCC-25,CAL-27 and Tca8113,was conducted.RT-qPCR were applied to detection of expression of miR-375 in these samples and four cell lines respectively.Correlation between expression change of miR-375 and SLC7A11 in clinical samples and clinical parameters of patients were analyzed.(2)Target genes of miR-375 were analyzed with bioinformatics software.RT-qPCR and western blot were used to test expression of mRNA and protein of SLC7A11.(3)The Dual-Luciferase Reporter Assay Kit was used to verify that whether miR-375 regulate the 3'UTR area of SLC7A11.RT-qPCR were applied to detection of SLC7A11 mRNA expression in CAL-27 and Tca8113 cells after transfection of miR-375 mimics.RT-qPCR were used to test expression of miR-375 and SLC7A11 in transfected cells.Cell proliferation after transfection was detected through MTT and CFC(colony-forming cell)assays.Detection of cell movement and invasion was conducted through wound healing test and Transwell chambers.Cell apoptosis and cell cycles were tested through flow cytometry.Results(1)Detection results of RT-qPCR showed that expression of miR-375 in tissue of oral cancer and in FaDu,SCC-25,CAL-27 and Tca8113 cells were significantly lower than that in the para-carcinoma tissues and normal oral cell(P<0.005),(2)Expression of SLC7A11 in tissue of oral cancer was higher than that in the para-carcinoma tissue(P<0.05).According to detection of RT-qPCR and western blot,expression of miR-375 in FaDu,SCC-25,CAL-27 and Tca8113 was significantly higher than that in normal oral cells(P<0.05).(3)SLC7A11 was evaluated as the target gene of miR-375.After transfection of miR-375 mimics,detection results of RT-qPCR indicated successful culture of CAL-27 and Tca8113 cell lines with overexpression of miR-375.After transfection of miR-375 mimics,proliferation rates of CAL-27 and Tca8113 cells and numbers of invasion cells were significantly lower than those in the control group(P<0.05).Results of wound healing tests showed that the healing capability of the experimental group was lower than that of the control group and the apoptosis rate of the experimental group was higher than that of the control group(P<0.05).According to cell cycle tests,the cell proportion in the experimental group was higher than that in the control group during G1/G0 phase(P<0.05),but significantly lower than that in the control group during S and G2 phases.In addition,after transfection of miR-375 mimics,functional change of CAL-27 and Tca8113 cells with overexpression of SLC7A11 was rescued to some extent.ConclusionMiR-375 is of expression in clinical samples of OSCC and cell lines of oral cancer and SLC7A11 is high expression in clinical samples of OSCC and cell lines of oral cancer.Their expression amounts are closely related to pathological grading,clinical stages and lymphatic metastasis.MiR-3 75 can affect a series of biological processes,such as proliferation,invasion and apoptosis of cells of oral cancer and cell cycle regulation,by inhibiting expression of SLC7A11 protein.