The Role And Mechanism Of NDUFC1 In Promoting Malignant Progression Of Glioma

ObjectiveThis study aims to explore the role and mechanism of NDUFCI in promoting the malignant progression of glioma.MethodThis research consists of two parts.In the first part,we studied the expression of NDUFC1 in human glioma cells.To explore the effect of NDUFC1 on the biological behaviors of glioma cells such as proliferation,migration,invasion and apoptosis.The second part explores the possible mechanism of NDUFC1’s effect on the function of glioma cells through the apoptotic protein antibody chip and western blotting.The expression of NDUFC1 in paraffin sections of 177 human gliomas and 24 normal brain tissues was detected by immunohistochemical method,and its clinical significance was further explored.RT-q PCR was used to detect the expression of NDUFC1 in human glioma cell lines and screen suitable cells.The NDUFC1 RNA interference lentiviral vector was constructed and transfected into SHG-44 cells and U251 cells to establish NDUFC1 knockdown SHG-44 cell lines and U251 cell lines.The transfection efficiency was detected by immunofluorescence experiment,RT-q PCR,western blot assay.By this basis,the glioma cells infected with lentivirus were divided into interference group(sh NDUFC1 group)and control group(sh Ctrl group).In each group,we used MTT cell assay to detect the proliferation ability of glioma cells,and the number of cells was counted by flow cytometry.Group glioma cell growth cycle and changes in cell apoptosis.In addition,To examine the ability of glioma cells in each group to migrate,we performed the Transwell Cell Migration and Cell Scratch Healing assay.In order to observe the effect of NDUFC1 knockdown on the growth of glioma cells in nude mice,we used a common subcutaneous tumor formation model in nude mice.The apoptotic protein antibody chip detection and Western blot verification of U251 cells infected with NDUFC1 interfering with lentivirus were carried out to explore the possible mechanism of NDUFC1 promoting the proliferation of glioma cells and inhibiting apoptosis.Results1.Immunohistochemistry was used to examine the difference in NDUFC1 expression levels between human glioma tissue samples and normal brain tissue samples.The results showed that NDUFC1 expression levels were significantly increased in human glioma tissues compared to normal brain tissues(P <0.001),while NDUFC1 expression levels were significantly different in human glioma tissues in different pathological data,such as pathological grade and tumor recurrence.The expression of NDUFC1 increases with the malignant degree of the patient’s tumor;the expression of NDUFC1 is positively correlated with tumor recurrence,that is,the expression of NDUFC1 gene is higher in patients with tumor recurrence.2.The SHG-44 and U251 cell lines were transfected with lentivirus to construct the NDUFC1 knockdown model.The proliferation and migration of glioma cells can be inhibited.And promote the apoptosis of glioma cells.It can obviously block the circulation process of glioma cells,and mainly block it in the G2 phase;It has a significant inhibitory effect on the growth of G2 glioma cells.It can significantly inhibit the growth of glioma cells in nude mice.3.Through the detection of apoptosis protein antibody chip on U251 cells infected with NDUFC1 interfering lentivirus,it was found that compared with the control group,the expression of apoptosis-related proteins in the sh NDUFC1 group.When P<0.05,and the number of differences exceeds 20%,the expressions of 5apoptosis-related proteins including Bad,BIM,Caspase3,p21,TRAILR-1,HSP60,Insulin Growth Factor Binding Protein-2(IGFBP-2)are up-regulated And soluble tumor necrosis factor-receptor 1(s TNF-R1)three kinds of apoptosis-related protein expression down-regulated.Western Blot further verified the expression of related pathway proteins for the above genes.After lentivirus infection,compared with the control group,the expression of knockdown histones P-ERK,CDK1,CDK6,PIK3 CA was down-regulated,and the expression of protein ERK did not change significantly.ConclusionThe expression level of NDUFCI was increased in human glioma compared to normal brain tissue.As the expression of NDUFCI increased,so did the pathological grade of the tumor and the risk of recurrence of the patient.NDUFCI may through the PI3K/AKT pathway to regulate the proliferation,invasion and migration of glioma cells,thereby playing a role in the occurrence and development of glioma.