The Role And Mechanism Of ALKBH5 In Proneural-mesenchymal Transition In Human Glioma

Objective: The purpose of this study was to investigate the role and mechanism of a-ketoglutarate-dependent dioxygenase alk B homolog 5(ALKBH5)in invasion and preneural-mesenchymal transition of human glioma cells.Methods:(1)Using CGGA(Chinese Glioma Genome Altas)and The Human Protein Atlas,the relationships between ALKBH5 expression in glioma tissues and clinic pathological classification,molecular typing and prognosis were firstly studied;(2)q RT-PCR and Western blot were used to detect the differential expression of ALKBH5 in the four types of human glioma cell lines;(3)sh-ALKBH5,sh-EGFP plasmids or 3×Flag-ALKBH5,Vector plasmids were transfected into glioma cells;CCK8 and cloning formation assay were used to detect the ability of proliferation and clone formation of the above glioma cells,and Transwell assay was used to detect the migration and invasion ability in glioma cells;Also,Western blot was used to detect the marker expression of preneural-mesenchymal transition,including E-cadherin,N-cadherin,OLIG2,YKL40,Vimentin and Met,and AMPK/m TOR signaling related proteins.Results:(1)The ALKBH5 expression was associated with the high-grade gliomas,and higher in mesenchymal gliomas than that in preneural type,and negatively correlated with the overall survival time of patients.(2)sh-ALKBH5 could significantly inhibit the expression of ALKBH5 in U87 MG and U251 MG cells compared with sh-EGFP;and 3×Flag-ALKBH5 could significantly enhance its expression in LN229 and SW1783 cells compared with vector;(3)ALKBH5knockdown inhibited the abilities of proliferation,migration and invasion in U87 MG and U251 MG cells,up-regulated the expression of preneural markers E-cadherin and OLIG2,and down-regulated the expression of mesenchymal markers N-cadherin,MET and YKL40,and p-AMPK and p-TSC2 were up-regulated,while p-m TOR and p-Raptor were down-regulated;Wheras ALKBH5 overexpression resulted in contrary changes in LN229 and SW1783 cells.Conclusions:(1)ALKBH5 expression in gliomas is positively correlated with pathological grading and negatively correlated with overall survival time of the patients.(2)ALKBH5 enhances the proliferation,migration and invasion of glioma cell,and promotes the preneural-mesenchymal transition of glioma cells.(3)ALKBH5affects the preneural-mesenchymal transition of glioma cells probably through the AMPK/m TOR pathway.