Study Of The Functional Role Of LRR1 In Breast Cancer And Its Mechanism

Background and ObjectivesBreast cancer is one of the three most common tumors globally,while breast cancer incidence in China increases year by year,and breast cancer has become a severe disease threatening women’s health and life.Currently,it is believed that overexpression and activation of multiple oncogenes are the leading causes of breast cancer radiotherapy resistance,infiltration,metastasis,and progression.Exploring new mechanisms of breast cancer occurrence and development and finding new intervention ideas are significant issues that need to be urgently addressed in basic and clinical research on breast cancer.LRR1 is a leucine repeat sequence that is mainly involved in protein modification and processing.It has been found that LRR1 can also participate in the immune response as a stimulatory signal of 41 BB.Besides,recent studies have shown that LRR1 can inhibit cell cycle inhibitors（CKI）,promoting cell cycle and proliferation.In this work,we propose to explore the role and mechanismof LRR1 in breast cancer cells using cell biology and molecular biology techniques.Methods and MaterialsHuman breast cancer cell lines MDA-MB231,HCC70,and primary breast cancer cells were cultured,and lentivirus-encapsulated RNAi-LRR1,overexpressed LRR1,and empty vector sequences were first transfected into human breast cancer cells,followed by the antibiotic screening of stably transfected breast cancer cells.Fluorescence and PCR were performed to detect whether LRR1 was stably transfected and observe whether the transfection efficiency met the experimental criteria.The effects of downregulation and overexpression of LRR1 on proliferation,infiltration,migration,invasion,cycle,apoptosis,and autophagy of breast cancer cells were investigated.Subsequently,combined with the bioinformatic database UCSC XENA data,the differences in LRR1 expression in human breast cancer and normal tissues adjacent to cancer and its effects on survival and prognosis were analyzed.Our experiments used continuous cell culture and microscopic counting to detect cell growth;Ed U assay to detect cell DNA replication and proliferation;Transwell assay to detect cell migration;Matrigel Transwell assay to detect cell invasion;TUNEL and JC-1 assay to detect cell apoptosis.Flow cytometry was used to detect cell cycle;Western Blot was used to detect protein expression and explore possible signaling mechanisms;UCSC XENA database was used to detect differential expression of LRR1 and survival analysis.Statistical analysisEach experiment was repeated more than three times for each cell line,and three secondary wells were set up for each experiment.Statistical analysis was performed using Graph Pad Prism8.0 software for image data collation.Statistical significance was considered when P<0.05.Results1.LRR1 positively regulates the growth of breast cancer cells.In breast cancer cell lines,overexpression of LRR1 resulted in increased cell growth,DNA replication and proliferation,migration and invasion in cytological assays.2.Down-regulation of LRR1 revealed that,in contrast to overexpression,cell proliferation,migration,invasion and cycling were all inhibited to varying degrees.3.LRR1 can regulate the proliferation,migration and invasion of breast cancer through the activation of the AKT-mTOR pathway.4.A search of the bioinformatics database UCSC XENA database showed that LRR1 was highly expressed in human breast cancer tissues and lowly expressed in normal breast tissues adjacent to cancer,and survival analysis revealed that breast cancer patients with high expression of LRR1 had shorter survival than the low expression group（p<0.05）.Conclusions1.LRR1 is highly expressed in breast cancer and promotes proliferation,cycle,migration and invasion of breast cancer cells.2.Downregulation of LRR1 was associated with inhibition of proliferation,cycling,migration and invasion of breast cancer cells and increased apoptosis.3.LRR1 promotes the proliferation,cycle,migration and invasion of breast cancer cells through activation of the AKT-mTOR signaling pathway.