| Objective:Tumor formation is an extremely complex process of abnormal cell proliferation,and every step of its formation is affected by many factors,including the tumor microenvironment and cancer stem cells.Brain gliomas are the most common in craniocerebral tumors,and high-grade gliomas have a short survival time and a high recurrence rate.Current treatment programs have limited efficacy.At present,there is a critical need to develop novel therapeutic strategies to prolong the life of patients,so it is particularly important to study the formation and progression mechanism of glioma.This study aimed to explore the role of collagen I and fibronectin of extracellular matrix in glioma formation and progression.Methods:The expression of type Ⅰ collagen and fibronectin in tumor tissues from glioma patients was examined by immunohistochemical staining.Then a three-dimensional I collagen/fibronectin culture system was established to simulate the tumor microenvironment in vivo.The correlations and molecular mechanism between collagen/fibronectin and glioma progression were analyzed by LN229 and T98 G glioma cells in vitro.The level of integrin αvβ3 protein in glioma cells was detected by Western blot.The immunofluorescence assay was used to detect the expression of integrin αvβ3 in glioma tissues from patients and PI3K/AKT signal associated proteins expression in glioma in vitro.In order to explore the function of PI3K/AKT signal pathway related proteins,The proliferation and colony formation ability of glioma cells were detected by CCK-8,Image J image analysis software and plate cloning formation experiment.The tumorigenesis assays of LN229 and T98 G glioma cells and the anticancer effect of integrin inhibitors in vivo were studied by xenograft glioma mice model.Results:The the expression of type Ⅰ collagen and fibronectin in glioma tissues were found significantly increased in high-grade gliomas from patients compared with low-grade gliomas by immunohistochemical staining,and the expression of type Ⅰ collagen and fibronectin was highly correlated.The addition of fibronectin to 3D type Ⅰ collagen culture model can significantly promote the growth of glioma cells,which indicates that type Ⅰ collagen and fibronectin synergistically transmit pro-survival signals to glioma cells.Glioma cells cultured in a 3D type Ⅰ collagen/fibronectin culture system were significantly enhanced in proliferation,colony formation,and tumorigenesis,indicating that glioma cells acquired tumorigenic potential and sand revealed strengthened proliferative characteristics.The level of αvβ3 protein in 3D Ⅰ collagen/fibronectin culture system was significantly higher than that in conventional culture flask,which indicated that Ⅰ collagen/fibronectin was closely associated with αvβ3.The integrin αvβ3 in the glioma tissues from patients was detected by immunofluorescence,and it was found that it has higher expression in high-grade gliomas than low-grade gliomas.The expression of PI3K or AKT proteins decreased significantly in cultured glioma cells treated with αvβ3 inhibitors,and cells proliferation,colony formation and tumorigenic ability of tumor cells were inhibited,indicating that αvβ3 transmitted signals through the PI3K/AKT signaling pathway.Similarly,LN229 glioma cells treated with PI3K or AKT inhibitors significantly inhibited tumor cell proliferation,colony formation,and tumorigenesis,and significantly decreased expression of SOX2,suggesting that SOX2 is a downstream regulator of the PI3K/AKT.Conclusion:We demonstrated that type Ⅰ collagen and fibronectin could collaborate to promote glioma progression through PI3K/AKT/SOX2 signaling pathway.Type I collagen and fibronectin synergistically regulate the stem cell characteristics of glioma cells,inducing sustained tumor growth and recurrence.Blockade of the upstream molecular integrin αvβ3 revealed improved outcome in glioma therapy,which provide new insights for eradicating tumors and reducing glioma cancer relapse. |
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