The Role Of Drp1-induced Mitochondrial Fission In Hypoxia-induced Migration Of Human Glioblastoma U251 Cell And Its Underlying Mechanism

Objective:In the present study,we focused on the role of Drp1-induced mitochondrial fission in hypoxia-induced migration of human glioblastoma U251cell and its underlying mechanism.Method:In our study,we chose the human glioblastoma cell line（U251）as our research object.The three gas incubator（37℃,1%O₂,5%CO₂,and 94%N₂）was used to mimic hypoxia.In order to change the activity of intracellular Drp1,we using those method,like using Drp1 inhibitor Mdivi-1,knockdown Drp1 or dominant negative Drp1-K38A.In order to observed the morphological changes of mitochondrial,the Mito-DsRed plasmid was transfected into cells,and the morphological was observed under the fluorescence microscope.Both scratch and Transwell assays were used to detecting the migration ability of U251 cells.Intracellular ROS was labeled by DCFH-DA fluorescent probe and quantitative the ROS levels by flow cytometry.The expressing of p-IκBαand N-cadherin were examined by western blot assay,which respectively activate NF-kB signaling pathway and promote epithelial-mesenchymal transition.Results:The mitochondrial morphology results show that mitochondrial fission was increased after hypoxia treatment,while pretreatment with Drp1 inhibitor Midivi-1or silenicing Drp1 by siRNA or dominant negative Drp1-K38A efficiently attenuated hypoxia-induced mitochondrial fission.Both scratch and Transwell assays indicated that hypoxia significantly enhanced the migration of U251,while pretreatment with Drp1 inhibitor Midivi-1 or knockdown Drp1 efficiently attenuated hypoxia-induced the migration of U251.The intracellular ROS production assay results show that intracellular ROS was increased after hypoxia treatment.In contrast,pretreatment with Midivi-1 or knockdown Drp1 significantly inhibited hypoxia-induced intracellular ROS increment.The expressing of p-IκBαand N-cadherin were upregulated under hypoxia condition,while pretreatment with Midivi-1 or knockdown Drp1 or dominant negative Drp1-K38A significantly decreased.Conclusion:Drp1-induced mitochondrial fission primarily by promoting the level of ROS that produced by mitochondrial pathway,and activate NF-κB pathway and promote epithelial-mesenchymal transition,thereby enhanced the migration of glioblastoma cells.In a word,this study will provide new therapeutic targets and theoretical foundation for inhibiting the migration and invasion of glioblastoma cells.