The Role And Mechanism Of Immunity-Related Gtpase Family M Protein In The Proliferation And Autophagy Of Glioma

BACKGROUND & OBJECTIVE: Glioma is the most common intracranial malignancy,of which glioblastoma(GBM)is the most malignant.The brain tumor is derived from glial cells and is characterized by high proliferation rate and high invasiveness.At present,the standardization treatment is not effective and the patient's prognosis is poor.However,there is little research on the immune-related GTPase family M protein(IRGM)in the central nervous system.The biological function and mechanism of IRGM in glioma are discussed,which is the immunotherapy for glioma.Provide a new target.This study explores the role of IRGM in the survival of gliomas.METHODS: The glioma tissues were taken for immunohistochemistry to detect the expression of IRGM.The location and expression of IRGM protein were verified by immunofluorescence and western blot.The lentiviral vector was used to construct the UGM-overexpressing U-251 MG and U-343 MG cell lines.The effect of up-regulation of IRGM protein on proliferation of glioma was detected by colony formation assay and CCK-8assay.Western blot was used to detect the expression levels of PI3 K and Akt,autophagy-associated markers p62 and LC3,and the phosphorylation levels of the Akt protein amino acid residues Thr308 and Ser473.IRGM in glioma A172 cell line was knocked down by shRNA lentiviral vector,and IRGM protein knockdown was identified by Western blot and immunofluorescence.The knockdown cell line was used to establish a subcutaneous model of nude mice glioma to observe the tumor size,and the fluorescence intensity was detected by in vivo imaging analysis.RESULTS: Immunohistochemistry of the patient's glioma tissue revealed that the expression level of IRGM protein was up-regulated,and immunofluorescence revealed that the IRGM protein was localized in the nucleus.Then,the expression of IRGM protein in glioma cell line was verified by western blot.The UGM-overexpressing U-251 MG and U-343 MG cell lines were subsequently constructed using lentiviral vectors.Cloning experiments and CCK-8assays were used to detect the upregulation of IRGM protein and promote the proliferation of glioma.The expression of P62 protein in the overexpressing cell line was increased by Western blot,while the ratio of LC3-II and LC3-II to LC3-I was decreased.In addition,compared with the control group,the proliferation ability and colony forming ability of the glioma cell line after IRGM overexpression were significantly up-regulated.In terms of mechanism,the level of PI3 K protein in the IRGM overexpressing cell line was significantly up-regulated,and the phosphorylation levels of the amino acid residues Thr308 and Ser473 of the Akt protein were increased.It indicates that the level of Akt protein activated is significantly increased,and it is possible to influence the proliferative ability through this signaling pathway.Moreover,under nutritionally sufficient conditions,IRGM protein reduces the level of basal autophagy in gliomas.The use of knockdown cell lines to establish asubcutaneous model of nude mice glioma revealed that IRGM knockdown inhibited tumor growth.Conclusion: We have revealed that IRGM protein expression is significantly up-regulated in gliomas,and the function of IRGM on glioma proliferation and regulation of autophagy may be achieved by PI3K/Akt signaling pathway.