Role Of HER2/PI3K/AKT Signaling Pathway In Proliferation And Migration Of Maligant Meningioma Cell Line

Objective:To investigate whether HER2 gene affects the proliferation and migration of malignant meningioma cells through PI3K/AKT signaling pathway.Methods:1、 IOMM-Lee maligant meningioma cell lines were transfected by silencing and overexpression of HER2 gene HER2 plamid with lentivirus vectors.The experiment was divided into five groups: the first group was untreated(blank),the second group was negative control of silence(NC-sh),the third group was the silence of HER2 gene(HER2-sh),the fourth group was negative control of overexpressing(NC-ox),the fifth group was the overexpressing of HER2gene(HER2-ox).The expression of HER-2 gene was evaluated by q-PCR and Western blot.2、 The CCK8 experiment,was to investgate the proliferation ability of different groups.Cell migration analysis were investgated by Matrigel migration assay.Cell cycle and apoptosis assays by flow cytometry were to investgate the cell cycles.3、 Western blot.were used to detect the expression of HER2、PI3K、AKT、p-AKT、m TOR、p-m TOR、S6、p-S6、NF-κB、Cyclin D1 and VEGF protein.4、 To apply with different concentrations of PI3 K inhibitors LY294002 、m TOR inhibitors KU-0063794(final concentration of 20、60、100、5、10、20 μmol/L)after 24、48、72h of the culture medium,CCK8、Transwell were used to detect malignant meningioma cell proliferation and migration;Cell cycle assays by flow cytometry were to investgate the cell cycles;Westen blot test protein expression of HER2、 PI3K、AKT、P-AKT、m TOR、P-m TOR、S6、P-S6、NF-κB、Cyclin D1 and VEGF.Results:1、 the plasmid of silencing and the overexpression the HER2 were established successfμlly.The level of m RNA and protein of HER2 reached the requirements of each groups.2、 The proliferation and migration of the meningioma cells with a down-regμlation of HER-2 by sh RNA humanly were significantly reduced.While the proliferation and migration of the human meningioma cells with a up-regμlation of HER-2 by sh RNA were significantly enhanced.The influence of HER2 gene in the cell cycle was associated with G0/G1-phase to S-phase.3、 The influence of down-regμlation and up-regμlation the HER2 gene take effects on PI3K、p-AKT、m TOR、p-m TOR、P-S6、NF-κB、Cyclin D1 in PI3K/AKT signal pathway,but not on AKT、S6、VEGF.4、 To apply with different concentrations of PI3 K inhibitors LY294002 、m TOR inhibitors KU-0063794(final concentration of 20、60、100、5、10、20 μmol/L)after 24,48,72 h of the culture medium,CCK8 was used to test the cell proliferation of different groups with different incubation time,then find out the best inhibition concentration and action time of PI3 K inhibitors LY294002 、 m TOR inhibitors KU-0063794 were 100 μmol/L 24h、20μmol/L 24h;The proliferation and migration of the meningioma cells with the LY294002、KU-0063794 were significantly reduced after 24 h.;LY294002,KU0063794 influence the change of cell cycle in G0 / G1-S;The protein expression of the cell with the different concentration of PI3 K inhibitors LY294002 on HER2、PI3K、p-AKT、m TOR、P-m TOR、S6、P-S6、NF-κB、Cyclin D1、VEGF in PI3K/AKT signal pathway were significantly decreased with the increasing of inhibitor concentration,but not on AKT、m TOR;The protein expression of the cell with the different concentration of KU-0063794 on HER2、PI3K、AKT、p-AKT、m TOR、p-m TOR、S6、P-S6、NF-κB、Cyclin D1、VEGF in PI3K/AKT signal pathway were significantly decreased with the increasing of inhibitor concentration.Conclusions:1、 HER2 gene can promote the proliferation and migration of malignant meningioma cells.2、 HER2 gene can take effects on the PI3K/AKT signaling pathways in malignant meningioma cells.