Experimental Study Of Significance And Function Of HDAC9 Expression In Osteosarcoma

Background and purpose Increasing studies have demonstrated that altered expression of histone deacetylases(HDACs)act a pivotal part in the tumorigenesis through up-regulation or down-regulation of key genes trapped in cell proliferation,cell-cycle regulation and apoptosis.And accumulative researches showed that HDAC9 was apparently up-regulated in a variety of solid tumors,which was of great significance in the occurrence and progress of tumors.However,its effects in osteosarcoma have not yet been elucidated.The purpose of our objective is to explore the expression of HDAC9 in osteosarcoma tissue and osteosarcoma cell lines;to explore the function,the biological behaviors and mechanism of HDAC9 gene in osteosarcoma.Methods:(1)Real-time quantitative PCR and Western-blot methods were used to measure the level of HDAC9 expression in osteosarcoma tissues and matched normal bone tissues in 30 patients,and the evaluated the clinical significance of HDAC9 in osteosarcoma.(2)Expression levels of HDAC9 in MG-63 and U-2OScells were detected by real-time quantitative PCR and Western-blot assay;After constructing HDAC9 adenovirus vector,we tranfected MG-63 and U-2OS cells with HDAC9 and the Negative Control vector.The HDAC9 expression in MG-63 and U-2OS cells after transfected were detected by real-time quantitative PCR and Western-blot assay.MTS assay was used to analyze osteosarcoma cell proliferation after transfected in vitro,and invasion assays were analyzed by Transwell invasion chambers.(3)Expression levels of p53 in MG-63 and U-2OS cells were measured by using real-time quantitative PCR and Western-blot assay before and after transfection.And the relationship between HDAC9 and p53 was veritfied by chromatin immunoprecipitation(Chip)assay.Results:1.HDAC9 is upregulated in osteosarcoma tissues than those in adjacent normal bone tissues.Over expression of HDAC9 is related to lymph node metastasis of osteosarcoma patients,while it has no to do with their gender,age,tumor locations and between two groups.2.Overexpression of HDAC9 in MG-63 and U-2OS cells promoted cell proliferation and invasion;while,inhibiting HDAC9 gene lead toinhibition of osteosarcoma cells proliferation and invasion;3.The tumor suppressor gene p53 expression was directly and negatively regulate by HDAC9,and HDAC9 epigenetically repressed p53 transcription through binding to its proximal promoter region.Conclusion:1.Upregulated of HDAC9 in osteosarcoma tissues.Between the expression of HDAC9 and distant metastasis in osteosarcoma tissues exist a positive correlation,while there was no to do with their sex,age,tumor location and differentiation degree and the other parameters.Which show that upregulated expression of HDAC9 potentially contributes to human osteosarcoma distant metastasis and may acts a prognostic factor for osteosarcoma patients;2.Overexpression of HDAC9 in MG-63 and U-2OS cells promoted cell proliferation and invasion;while,down-regulation HDAC9 lead to inhibition of cell proliferation and invasion of osteosarcoma cells.It indicates that the overexpression of HDAC9 may act a pivotal part in osteosarcoma cell function.3.The expression of HDAC9 was conversely correlated with tumor suppressor gene p53 m RNA and protein level in human MG-63 and U-2OS cells.Overexpression of HDAC9 in osteosarcoma take part in the development process of osteosarcoma via negatively control downstream expression of target genes p53.Therefore,HDAC9 might become a candidate biomarker in osteosarcoma diagnosis or treatment in gene therapy.