Study On The Mechanism Of Long Non-Coding RNAPTAR Regulating The Migration And Invasion Of Ovarian Cancer Through MicroRNA-101

Background and purposeOvarian cancer is not uncommon in female malignancies,most of which develop into abdominal cavity metastases.Epithelial-mesenchymal transition(EMT)plays a key role in tumor cell metastasis.However,it is not clear how long non-codingRNA(LncRNA)participates in the EMT process and affects cell invasion and metastasis in Ov Ca.However,it is not yet clear how LncRNA affects the EMT process through its downstream target genes and how it affects the invasion and metastasis of ovarian cancer cells.In this study,we intend to clarify the mechanism by which LncRNA PTAR participates in the regulation of EMT and aggressive metastasis of ovarian cancer.MethodsWe used the Cancer Genome Atlas Database(TCGA)to screened and verified the differences of LncRNA PTAR,hsa-miR-101-3p and ZEB1 in ovarian cancer and their relationships.In vivo,nude mice were injected with ovarian cancer cell line SKOV3 cells for 7 days and then injected with lentiviral LncRNA PTAR(sh-PTAR)intraperitoneally to construct a xenograft tumor model.Four weeks later,the tumor weight and size was compared while the expression of EMT specific markers was detected by immunohistochemistry.In vitro,the migration and invasion ability of ovarian cancer cells was evaluated by Wound Healing and Transwell test.Western-blot and qRT-PCR were used to detect the expression levels of EMT-related markers in vivo and in vitro.Luciferase experiment was used to detect the target binding relationship between PTAR and miR-101,miR-101 and ZEB1.Results(1)Based on our previous bioinformatics analysis,we constructed the interstitial ovarian cancer ceRNA regulatory network and screened out differentially expressed LncRNAs,miRNAs and mRNAs.It is noteworthy that LncRNA PTAR and ZEB1 are highly expressed and hsa-miR-101-3p is low expressed in ovarian cancer.In GEPIA database,LncRNA PTAR was up-regulated in ovarian cancer samples compared with normal tissue samples.TCGA database showed that hsa-miR-101-3p was negatively correlated with LncRNA PTAR、ZEB1,and LncRNA PTAR was positively correlated with ZEB1.High expression of hsa-miR-101-3p improves long-term survival in patients with ovarian cancer compared with low expression in starbase.(2)Compared with the blank group sh-Scramble,the size and number of tumor tissues in mice transfected with sh-PTAR were significantly reduced,while the expression level of epithelial cell marker protein E-cadherin was significantly reduced and the mesenchymal cell marker proteins FN1,vimentin and ZEB1 decreased.(3)PTAR and PTAR+miR-101 were transfected into ovarian cancer cell lines SKOV3 and A2780 cells,respectively.The results showed that the PTAR group promoted the migration and invasion of SKOV3 and A2780 cells(P <0.05),but PTAR+miR-101 reversed the migration and invasion caused by PTAR overexpression;at the same time,PTAR inhibited the epithelial cell marker protein and promoted the expression of mesenchymal cell marker protein,but it was reversed by the miR-101co-transfected PTAR group(P <0.05).On the contrary,TGF-β1 promotes cell migration and invasion.Knocking down PTAR can inhibit the migration and invasion induced by TGF-β1,but cancelled by miR-101 inhibitor(AMO-101)(P <0.05);Reducing PTAR promoted epithelial cell marker protein and inhibited the expression of mesenchymal cell marker protein,but reversed by AMO-101(P <0.05).(4)Luciferase clarified the target binding relationship between PTAR and miR-101(P <0.05).(5)miR-101 was transfected into ovarian cancer cell lines SKOV3 and A2780 cells and TGF-β1 induced cell migration and invasion.The results showed that miR-101 could inhibit the migration and invasion promoted by TGF-β1(P <0.05).miR-101 promoted the epithelial cell marker protein induced by TGF-β1 and inhibited the expression of mesenchymal cell marker protein(P <0.01).At the same time,miR-101 up-regulates the level of ZEB1 protein and mRNA,while AMO-101down-regulates the protein expression of ZEB1.In addition,Luciferase proved that miR-101 can target ZEB1.Conclusions(1)LncRNA PTAR was highly expressed in ovarian cancer.(2)LncRNA PTAR promoted EMT and invasive metastasis of ovarian cancer.(3)LncRNA PTAR promoted the occurrence and development of ovarian cancer by competitively binding with miR-101 to regulate ZEB1.