TNF-? Promoted Glioblastoma Cell Mitochondrial Apoptosis Via Augmenting Mitochondrial Fission And Repressing MAPK-ERK-YAP Signaling Pathways

Glioblastoma multiforme(GBM)is the most common and fatal brain tumor.Its poor prognosis with a median survival of less than one year and inevitable recurrence 32-36 weeks after surgery makes it a heavy burden on the society.There are few effective therapeutic strategies against GBM.Besides surgery,irradiation and chemotherapy,scientists also attempt to develop new biological therapies.Several approaches have been attempted to induce the death of glial cells,especially TNFa-based therapy.A gene delivery strategy to induce TNF-a overexpression was demonstrated to increase the apoptotic index of glioblastoma cells.The effectiveness of the TNF-?-based therapy was later validated by several clinical studies.Ample in vivo and in vitro evidence potentially implies that TNF-? considerably augments the apoptosis of glioblastoma cells.This information indicates that TNF-?-based therapy is a promising tool for the treatment of glioblastoma.However,the molecular mechanisms of TNF-? involved in glioblastoma cell death have not been fully described.Mitochondria control an array of subcellular functions,such as energy metabolism,ROS production,cell proliferation,calcium balance and cell death,the dysfunction of which would induce pathological conditions and diseases.Recently,mitochondrial fission has been thought to be the early feature of mitochondrial abnormalities and promotes the death of several kinds of tumors.Whether TNF-? induce apoptosis of glioblastoma cells via mitochondrial fission remains to be studied.Using human glioblastoma cell line A172 as the model,we first proved that TNF-?could induce the apoptosis of glioblastoma cells via the classic caspase 3-dependent pathway.Mechanistically,we proved that TNF-? induced the apoptosis of glioblastoma cells via mitochondrial fission.After TNF-? treatment,mitochondria become small,roundish fragments that are characteristic of mitochondrial fission.The average length of mitochondria also became shorter.The levels of Drpl,Mff and Fisl,the key elements in executing mitochondrial fission were upregulated,while the levels Mfn2 and Opal,the inhibitors of mitochondrial fission were downregulated in response to TNF-a treatment.This mitochondrial fission further induced mitochondrial damage in glioblastoma cells.TNF-? treatment significantly increased ROS production and decreased cellular antioxidants such as GSH,SOD and GPX in glioblastoma cells,inducing less ATP production.After TNF-? treatment,the protein expression of mitochondrial respiratory complex was also significantly repressed.The mitochondrial potential was reduced,and the opening rate of the mitochondrial permeability transition pore(mPTP)was increased.The excessive opening of mPTP further facilitated mitochondrial pro-apoptotic cyt-c translocation into the cytoplasm where cyt-c interacted with and activated caspase-9.Accordingly,the expression levels of mitochondrial pro-apoptotic proteins such as Bax and Bad were upregulated while the levels of anti-apoptotic proteins such as Bcl-2 and x-IAP were downregulated,which eventually induced the mitochondrial apoptosis.Lastly,we showed that TNF-? induced mitochondrial fission and apoptosis of glioblastoma cells via MAPK-ERK-YAP signaling pathways.When YAP was overexpressed in glioblastoma cells,TNF-? treatment-induced mitochondrial fission and apoptosis was reversed.The morphology change,the mitochondrial length reduction,the ROS level elevation,the cyt-c translocation and release,the caspase-9 activation,the cell death and LDH secretion induced by TNF-? were all inhibited.In summary,here we showed that TNF-? could induce the mitochondrial fission and apoptosis of glioblastoma cells via MAPK-ERK-YAP signaling pathways,providing potential molecular basis for TNF-?-based glioblastoma therapy.