| Background:Osteosarcoma(OS)is a relatively common malignant bone tumor with an increasing incidence trend.OS cells are prone to spread,and metastasis is the cause of death in most patients.Compared with other malignant tumors,OS has its unique characteristics,including the complexity and instability of the genomic background,as well as a large number of mutations driven genes.At the molecular level,there are many unknown factors in the mechanisms leading to malignant tumor metastasis,and there is currently little potential biological information about driving OS metastasis.Therefore,the purpose of this study is to elucidate the relevant mechanisms driving the metastasis of OS at the genetic level,in order to develop more specific treatment methods and reduce the mortality rate of OS patients.method:Based on the Gene Expression Omnibus(GEO)database,we obtained the gene expression profile of OS metastasis related tissues.Weighted gene co-expression network analysis(WGCNA)was used to identify the modules with the most significant differences.Analysis of the function and pathway enrichment of differentialy expressed genes(DEGs)can identify the functional nodes in which certain genes participate.The obtained gene sequence map is used to construct protein-protein interaction networks(PPI)and search for corresponding hub genes.Knocking down the core gene Leucine Proline-enriched Proteoglycan 1(LEPRE1)to explore the impact on human Osteosarcoma cells.Targeted downregulation the expression of LEPRE1 gene in human osteosarcoma cells through siRNA technology and detect the mRNA and protein expression of related genes by RT qPCR and Western blot.The proliferation ability of Osteosarcoma cells after down-regulation of related genes was detected by CCK-8 test and plate cell cloning test.Wound healing assay and Transwell test were used to detect the migration and invasion ability of Osteosarcoma cells.Result:We conducted WGCNA correlation analysis on the GSE21257 dataset and found the grey module with the most significant genetic differences.After analyzing the metastatic and non-metastatic samples in another data set GSE99671,we finally identified five core genes related to Osteosarcoma metastasis(LEPRE1,LTF,ELANE,LCN2,PLOD1).We confirmed the hypothesis through further experiments,and we found that they were closely related to distant metastasis of bone malignant tumor.The enrichment analysis of differential genes showed that the genes related to OS metastasis were mainly located in extracellular regions and specific granule lumen.Its molecular functions mainly include oxygen transporter activity,peroxidase activity and haptoglobin binding.The biological processes involved mainly include defense response to bacterium,carbon dioxide transport,oxygen transport and erythrocyte differentiation.It is suggested that the occurrence and metastasis of tumors are closely related to these mechanisms involved.After interfering with the expression of LEPRE1 in human OS cells(143B and MG63),the results of CCK-8 and plate clone formation assay showed that the proliferative activity and cloning ability of OS cells were inhibited.Wound healing assay and Transwell assay further showed that the migration and invasion ability of OS cells was also inhibited after knocking down the expression of LEPRE1 gene.Conclusion:The genes we screened through bioinformatics are closely related to the metastasis of OS.Our further experiments demonstrate that knocking down the LEPRE1 gene can inhibit the proliferation,migration,and invasion of OS cells.The above studies suggest that LEPRE1 may be an important potential target for OS cells. |
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