The Effect Of Cytoskeletal Protein Coronin3 On The Invasive Ability Of Glioblastoma And Preliminary Study Of Its Mechanism

Objective This study,we investigated the effects of different concentrations of TGF-βon the induction of epithelial-mesenchymal transition in glioma T98G cells and the expression of cytoskeletal protein coronin3 in invasive ability of glioma cells T98G.Methods ①Under the condition of in vitro cell culture,coronin3 overexpression empty plasmid group,coronin3 overexpression group,wild type T98G,coronin3 silencing empty plasmid group,coronin3 silencing group were transwell cell migration assay and transwell cell invasion assay(including matrigel).The effects of cytoskeletal protein coronin3 on migration and invasion of glioma T98G cells were investigated by comparison between groups.②Under in vitro cell culture conditions,T98G cells in logarithmic growth phase were inoculated into culture flasks,106 per flask.When the cells grew to 70%-80%confluence,serum-free starvation for 24 hours,then TGF-β was added.The final concentration was Ong/nl,10ng/ml,20ng/ml,30ng/ml.After 48 hours of culture,the cell proteins were collected.Western blot was used to identify the expression of epithelial marker cadherin(E-cadherin),interstitial marker vimentin.Results ①Cell migration assay:coronin3 overexpression empty plasmid group,coronin3 overexpression group,wild type T98G group,coronin3 silencing empty plasmid group,coronin3 silencing group,five groups,coronin3 overexpression empty plasmid group through the small chamber membrane cells(99.80±9.41),the number of cells passing through the ventricular membrane of the coronin3 overexpression group was(125.20±11.77),and the number of cells passing through the ventricular membrane of the wild type T98G group was(103.40±7.50),and the coronin3 silencing empty plasmid group passed through the small compartment membrane.The number of cells was(98.20±8.34),and the number of cells in the coronin3 silencing group passing through the ventricular membrane was(74.20±8.22).The two groups were compared with each other,P<0,05 indicated that the difference was statistically significant.The coronin3 overexpression empty plasmid group and the coronin3 overexpression group were compared,and P<0.05 was found.The difference was statistically significant.The coronin3 silencing empty plasmid group and the coronin3 silencing group were compared,and P<0.05 was obtained.The coronin3 overexpression empty plasmid group,coronin3 overexpression group,coronin3 silencing empty plasmid group,coronin3 silencing group and wild type T98G group were compared,and the coronin3 overexpression group,coronin3 silencing group and wild type T98G group were obtained.Between the two,P<0.05,the difference was statistically significant;coronin3 overexpression empty plasmid group,coronin3 silent empty plasmid group and wild type T98G group,P>0.05,the difference was not statistically significant.②Cell invasion assay:Coronin3 overexpression empty plasmid group,coronin3 overexpression group,wild type T98G group,coronin3 silencing empty plasmid group,coronin3 silencing group,five groups,coronin3 overexpression empty plasmid group through the matrigel membrane cells(77.00±8.30),the number of cells in the coronin3 overexpression group passing through the Matrigel membrane was(135.60±10.69),and the number of cells in the wild-type T98G group passing through the Matrigel membrane was(84.60±6.80),and the coronin3 silencing empty plasmid group passed through.The number of cells in the Matrigel membrane was(81.80±7.46),and the number of cells in the coronin3 silencing group passing through the Matrigel membrane was(48.00±9.21).The two groups were compared with each other,P<0.05 indicated that the difference was statistically significant.The coronin3 overexpression empty plasmid group and the coronin3 overexpression group were compared,and P<0.01 was found.The difference was statistically significant.The coronin3 silencing empty plasmid group and the coronin3 silencing group were compared,and P<0.01 was obtained.The difference was statistically significant.Significance;coronin3 overexpression empty plasmid group,coronin3 overexpression group,coronin3 silencing empty plasmid group,coronin3 silencing group and wild type T98G group were compared,and the coronin3 overexpression group,coronin3 silencing group and wild type T98G group were obtained.P<0.01,the difference was statistically significant;coronin3 overexpression empty plasmid group,coronin3 silent empty plasmid group and wild type T98G group,P>0.05,the difference was not statistically significant.③After 48 hours of treatment with wild-type T98G cells with different concentrations of TGF-β,the relative expression level of epithelial marker cadherin decreased gradually with the increase of TGF-β concentration.The concentration of T98G cells in each concentration of TGF-β cells The relative expression levels of cadherin were 37.2%(P>0.05),64.7%(P<0.05),and 76.4%(P<0.05),respectively,compared with the control group(0 ng/ml).After 48 hours of treatment with wild-type T98G cells with different concentrations of TGF-β,the relative expression level of the interstitial marker vimentin gradually increased with the increase of TGF-β concentration,and the vimentin in T98G cells of each concentration of TGF-p group.The relative expression level was 9%(P>0.05),154%(P<0.05),and 363%(P<0.01)compared with the blank control group(0 ng/ml).Compared with the blank control group(Ong/ml),the morphology of T98G cells in the 20ng/ml TGF-β group changed significantly,and the cells became more slender,stretched,long fusiform,and intercellular junctions.Loose and present a more dispersed growth pattern.The wild-type T98G group and the wild-type T98G group(including TGF-β)were used for cell migration experiments.The number of cells passing through the small chamber membrane of the wild-type T98G group was(91.40±6.65),and the wild-type T98G group(including TGF-β)was worn.The number of cells passing through the small chamber membrane was(117.60±8.73),and the two were compared,P<0.05,and the difference was statistically significant.Conclusion①Increased expression of coronin3 can promote the migration and invasion of glioma T98G cells.②To a certain extent,TGF-β can induce epithelial-mesenchymal transition(EMT)in glioma T98G cells in a concentration-dependent manner.