TBXAS1 Promotes Proliferation And Migration Of Glioblastoma Through PI3K/AKT Signaling Pathway

Background:Glioblastoma is considered to be a common malignant tumor within the central nervous system.The 2-year survival rate for GBM patients was only 14.8%,thus patients with GBM have poor prognosis.At present,the therapeutic options remain limited,which mainly depend on neurosurgery,then supplemented by radiotherapy and chemotherapy,and temozolomide is the commonly used anti-tumor drugs.Due to the specificity growth location and rapid infiltrating growth,the modern medical treatment may not eradicate the full extent of the tumors.However,most patients have adverse prognosis and tumor recurrence during treatment,because the temozolamide is associated with significant side effects and drug resistance.In order to improve patient’s prognosis,it’s critical to discover the molecular mechanism and explore a new cure.Thromboxane A Synthase 1(TBXAS1)is an endoplasmic reticulum membrane protein which belongs to the P450 epoxygenase family.For a long time,people concerned the association between TBXAS1 and cardiovascular,renal,and inflammatory diseases.However,the potential role of TBXAS1 in tumor progression has been widespread considered in recent years.In this study,we mainly discussed the role of TBXAS1 on the development and progression of GBM,and explored possible molecular mechanisms,as well as provided the theoretical basis for pathogenesis and pharmaceutical of GBM.Objects:(1)To study the expression of TBXAS1 in GBM.(2)To discover the effect of TBXAS1 on proliferation and migration of GBM.(3)To explore the potential molecular mechanism of TBXAS1 in GBM.Methods:(1)Bioinformatics data analysis was conducted to analyze the correlation between TBXAS1 expression levels and the occurrence of GBM by using GEPIA,TCGA,CGGA,and GEO databases.survival analysis was performed from CGGA and TCGA databases in the high expression TBXAS1 of glioma group and the low expression TBXAS1 of glioma group.(2)The expression of TBXAS1 in patients samples and cell lines was detected with Western blot and immunological fluorescence assay,then we verified whether the experimental result were consistent with databases or not.(3)Construction of overexpression plasmid to upregulate the expression of TBXAS1 in GBM cells,the effects of overexpression of TBXAS1 were detected by using CCK8,transwell test and scratch test methods.(4)Using Picotamide,an inhibitor of TBXAS1,then we explored the effects of TBXAS1 on the proliferation,induction of cycle arrest,and migration of GBM cell lines C6 and DBTRG by CCK8 assay,cellular immunological fluorescence assay,plate cloning,flow cytometry,scratch test and transwell test.(5)The Western blot method was used to clarify the molecular mechanism of TBXAS1 on GBM.Results:(1)The expression level of TBXAS1 in glioma is significantly higher than that in normal brain tissue,and the expression level increases with the grade of glioma.The CGGA and TCGA database showed that there was also a correlation between the expression of TBXAS1 and survival,patients with high TBXAS1 expression frequently predict poor prognosis.(2)We used p EGFP-N2 and TBXAS1-OE plasmids to transfect the GBM cells DBTRG,then verified the results of successful construction of empty and overexpressed cell lines through q-PCR and Western blot assay.(3)Through the CCK8 it was found that overexpression of TBXAS1 can promote the proliferation of GBM cells;scratch and Transwell experiments manifested that overexpression of TBXAS1 can promote the migration ability of glioblastoma cells.(4)The CCK8 experiment found that Picotamide can inhibit the proliferation of GBM cells through dose-and time-depend manner,plate cloning experiments and cell immunofluorescence experiments demonstrated that Picotamide can inhibit the proliferation of GBM cells.(5)The Western blot and flow cytometry experiments showed that Picotamid can arrest cell cycle in G2/M phase to inhibit cell proliferation.(6)The transwell invasion and scratch migration test found that Picotamide can inhibit the migration of GBM cells.(7)The Western blot results showed that Picotamide can decrease the expression of p-PI3K/PI3 K,p-AKT/AKT proteins in DBTRG,which suggested that TBXAS1 may mediate cell proliferation and migration through the PI3K/AKT pathway.Conclusion:The expression of TBXAS1 in GBM is higher than that in normal tissues,Inhibition of TBXAS1 can block GBM cells in G2/M phase,resulting in inhibition of cell proliferation and migration.Moreover,TBXAS1 may accelerate the progression of GBM by promoting abnormal activation of the PI3K/AKT signaling pathway.