4EGI-1 Affect Biological Behavior Of Human Glioma U251 Cells In Vitro

Glioma is the most common primary intracranial tumor, originating from the glial cells. According to classification of the central nervous system tumor from WHO, glioma was divided into level I to IV. The level III- IV belongs to malignant glioma, accounting for 77.5% of all glioma. Study shows that the incidence of malignant glioma increased at 1.2% annually. Malignant glioma mainly cause increasing intracranial pressure and nerve dysfunction, which seriously threaten patient safety, reduce the patient's life quality. The main treatment for the malignant glioma is surgical resection and combined with radiotherapy, chemotherapy and other measures according to the result of pathologic examination. New treatment methods, such as immune therapy, gene therapy and molecular target therapy are also gradually applied to the treatment of malignant glioma. As images, anesthesia, stereotaxi, navigation, surveillance, radiation, chemotherapy and drugs develop, the prognosis of patients with malignant glioma is still not optimistic. Data shows that mortality rate of our brain and CNS malignant tumor is 2.91/100 thousand, the ninth of top ten high death tumor and the proportion of 2.39%. The high death and morbidity and high recurrence rate of malignant glioma, seriously threaten human health. Therefore, it is important to find effective means of treatment of malignant glioma clinically, which would relieve the pain of the masses of glioma.In eukaryotes, the cap-translation initiation eIFs has played a key role in transcription, translation of mRNA and the process of protein biosynthesis. And eIF4A, eIF4E, eIF4G compose eIF4F compounds, starting with 5 'cap structure mRNA translation process. Studies have shown that eIF4F compounds can regulate various abnormal expression of genes, closely related to the tumor. And the specific performance are to promote proliferation and inhibit apoptosis, induce resistance of radiation and chemotherapy, and accelerate of tumor tissue vascularization etc. And successful assembly of eIF4F complex depends on phosphorylation levels of eIF4E binding protein 4E-BPs. Low level of phosphorylation blocks eIF4E, thus blocking the success of eIF4F complex assembly. High phosphorylation level of 4E-BPs does not block eIF4E, activating the eIF4F complex.Mitochondria plays an important role in cell metabolism, redox balance and cell death. In addition, increased mitochondrial stress reaction has been conformed highly with the aggressiveness of tumors and poor prognosis. When the mitochondrial stress response exceeds the cellular capacity it would trigger cell toxicity and inhibit cancer progression. The specific performance includes the swelling of mitochondria membrane, opening of membrane channel relieve apoptosis factors such as cytochrome C and decreased ATP synthesis and supply. Induction of mitochondrial dysfunction as a target in tumor treatment has made more and more attention in recent years. A number of factors were confirmed to affect mitochondrial function and inhibit biological behavior of tumor cell in vivo and in vitro.4EGI-1, a small molecular compound, can mimic the function of the 4E-BPs, which disrupt the eIF4G derived peptide to combine with eIF4E, then conform combination of eIF4E/4EGI-1 compounds and block eIF4F formation. Studies show that in many human tumor cell lines, 4EGI-1 inhibits cap-translation and inhibit cell proliferation, induce cell apoptosis. This small molecule compounds, as gene therapy target for eIF4F complex of malignant tumor, has potential application prospect. However the role of 4EGI-1 for tumor cell proliferation and apoptosis and related mechanism has not been confirmed.Therefore we treated U251 with cap-translation inhibitor 4EGI-1 and we observed eIF4F compound by Western bolt, cell apoptosis by TUNEL staining and flow cytometry, cell viability by MTS test, MMP change by Rh123 staining, ROS generation by DCFH-DA, cytotoxicity by LDH assay, O2 consumption rate by Clarke-electrode, mt DNA and transcription by Real-Time PCR, mitochondrial morphology by mitochondrial fluorescence staining, mitochondrial dynamic protein change by Western blot and observed biological behavior of human glioma U251 cells by Si-Drp-1 and mdivi treatment.Our conclusions are:1. In U251 glioma cells, 4EGI-1 blocked eIF4F compound by affecting the eIF4G rather than eIF4E and 4E-BPs.2. In U251 glioma cells, 4EGI-1 induce cell apoptosis and suppress cell proliferation by inducing oxidative stress reaction.3. In U251 glioma cells, 4EGI-1 can reduce mitochondrial respiratory activity but not affect the mitochondrial genesis.4. In U251 glioma cells, 4EGI-1 exert anti-tumor effect via affecting mitochondrial dynamic protein expression and mitochondrial fission protein Drp-1 makes a key role.