The Construction Of ANCR And EZH₂ Gene Overexpression And Knockdown Cell Lines In Salivary Adenoid Carcinoma?SACC?

EMT(Epithelial-Mesenchymal transition)is to point to in certain pathological or physiological condition,with the polarity of epithelioid cells to ectomesenchymal cells has the ability to migrate into the process?Emt process can lead to decreased expression of molecules in epithelial surface,loss of cell polarity,and changes in cytoskeletal markers;Markers have been lost,such as E-cadherin(e-cad),along with increased expression of interstitial markers and increased anti-apoptosis.salivary adenoid cystic carcinoma,SACC,as a common head and neck malignancy,is derived from epithelial-derived highly malignant salivary gland neoplasms of the tubal reserve cells.The typical clinical and biological features are slow growth,but the local invasion is strong,highly neurotropic and lung metastasis.Although complete surgical resection and adjuvant radiotherapy have been shown to improve long-term survival,the prognosis is still poor.In recent years,with the domestic and foreign scholars of head and neck tumor epithelial mesenchymal transition(EMT)in-depth research.With the development of genomics,epigenetics,molecular biology and other biological sciences as well as the application of high-throughput sequencing,a variety of non-coding mas,growth and transcription factors,signaling pathways and epigenetic modification enzymes are gradually unveiled in SACCAmong them,EZH2 is an important protein with increased expression in various cancers,and its high expression is positively correlated with the development,metastasis and poor prognosis of cancer,while overexpression of EZH2 in normal cells can increase the canceration and proliferation capacity of cells.In cancer,EZH2 can regulate the occurrence,development and metastasis of cancer at multiple levels.As an important histone methyltransferase,EZH2 regulates target genes at the epigenetic modification level by inhibiting gene transcription.EZH2 is recruited into the promoter region of epidermal protein marker by relevant transcription factors,and methylated to inhibit its transcription expression.EZH2 is also a key epigenetic modifier that induces and promotes EMT,and many of its target genes are tumor suppressor genes that inhibit cancer metastasis,which promotes the occurrence of tumor cell EMT.At the same time,EZH2 is capable of transcriptional inhibition(histone methylation modification)of multiple tumor suppressor genes,leading to the development and metastasis of cancer.LncRNA is closely related to head and neck cancer,breast cancer,lung cancer,liver cancer,colon cancer,stomach cancer,ovarian cancer and other cancers.Many of the same lncrnas have similar important functions in different malignant tumors,including unlimited proliferation,resistance to apoptosis,induction of tumor angiogenesis,immortalization of cells,enhancement of invasion and metastasis,and escape growth inhibition.ANCR,as an LncRNA,it was initially found to inhibit the differentiation of epidermal progenitor cells.During the differentiation of progenitor cells,ANCR expression was significantly down-regulated,while ANCR knockdown could significantly promote the occurrence of differentiation and promote the expression of some differentiation related genes.In breast cancer,ANCR was found to promote EZH2 degradation by binding to EZH2,inhibit the invasion and migration of breast cancer cells,and inhibit the tumorigenic ability of breast cancer cells and remote metastasis of breast cancer.However,whether it plays the same regulatory role in adenoid cystic carcinoma of salivary glands remains to be further studied.Our experiment was verified by transformation,monoclonal replication and small extraction,we successfully extract pwpxld-ancr expression plasmid,pwpxld-ezh2 plasmid,shANCR#1,shANCR#2 plasmid,shEZH2#1,shEZH2#2 plasmid,plkko.1/pWPXLd plasmid andsecond-generation packaging plasmid(PAX,PMD2G)?ANCR and EZH2 gene overexpression(pwpxld-ancr expression plasmid,pwpxld-ezh2 plasmid)and knockdown(shANCR#1,shANCR#2 plasmid,shEZH2#1,shEZH2 plasmid)were produced in 293T cells by lentivirus packaging,cell infection and other cell biology experimental techniques.Sacc-lm and sacc-83 were recovered by PEI transfection.Western blot protein level,real-time transcription level and flow apoptosis analysis molecular level were used to detect the effect of virus infection.In this study,sacc-lm-ANCR,sacc--83-ANCR,sacc-83-EZH2,acc-lm-shEZH2#1,sacc-lm-shEZH2#2,sacc-83-shANCR#1,sacc-83-shANCR#2 and sacc-83-EZH2 cell lines were successfully constructed.Subsequent functional tests,in vivo experiments and other further in-depth mechanism experiments lay the foundation.