The Expression Of FOXJ2 In Glioma And The Role Of FOXJ2 In The Migration Of Human Glioma Cells

Objective:Gliomas are the most frequent primary brain tumors, accounting for >50% of all brain tumors. Despite multimodal therapy regimens, including neurosurgical resection, radiotherapy, and chemotherapy, the prognosis of glioma patients remains poor. Local recurrence is the main cause for the failure of treatment. Strong invasiveness and migration of human glioma cells is an important cause of local recurrence. According to previous studies, FOXJ2 appeared to be involved in positively regulating the progression of the cell cycle or aiding in tumorigenesis. For example,one study displayed that up-regulation of FOXJ2 might potentially be an effective therapeutic approach for the inhibition of cell migration and invasion of breast cancer. In this study, we investigated the expression of FOXJ2 in glioma cell lines and human glioma specimens and the relationship between clinical pathological factors in glioma. The expression of FOXJ2 and E-cadherin was detected to analysis the relationship and the influence on the migration of glioma cells. Abnormal expression of FOXJ2 was detected to explore the significance in the occurrence and the development process in glioma. Methods:1.From the histological level, the expression of FOXJ2 was detected in two normal brain tissue samples and seven paired glioma samples by Western Blot analysis. The expression of FOXJ2 and E-cadherin in 80 human glioma specimens by immunohistochemical method was studied using the Spearman rank correlation test. The statistical significance of the correlations between FOXJ2 and E-cadherin expression and the clinic pathologic features, such as age, gender, tumor location, tumor size, histological grade and so on,were analyzed using the χ2 test. Overall survival curves were calculated with the Kaplan–Meier method and were analyzed with the log-rank test.2.From cell and molecular levels, we examined the expression of FOXJ2 in five glioma cell lines and selected U87 cell line for Plasmid transfection. Glioma cells with high expression or knockdown the expression of FOXJ2 were selected as the research object. Western Blot and immunofluorescence analysis were used to test the expression of E-cadherin and Vimentin. The wound healing and trans-well assays were used to test the migration of glioma cells. Through various methods above, we investigated the potential role of FOXJ2 in the occurrence and development of glioma.Results:1. Both Western Blot and immunohistochemistry analysis indicated that the expression of FOXJ2 in normal brain tissue samples was significantly higher than in glioma. Moreover, its level is significantly less in anaplastic glioma(grade III) and glioblastoma multiforme(grade IV) than in low-grade glioma(grade II).The expression of FOXJ2 was significantly associated with histological grade(P=0.012). No notable correlation was observed between FOXJ2 low expression and patient’s gender(P=0.884), age(P=0.551), tumor location(P=0.869), type of surgery(P=0.330), vessel density(P = 0.103), tumor diameter(P = 0.755), or necrosis(P=0.684). Kaplan-Meier survival curve analysis showed that poor overall survival in glioma patients was significantly associated with low expression of FOXJ2(P<0.01).2. The expression level of FOXJ2 was higher in weakly invasive cells(H4), while it was lower in highly invasive cells(U251). We detected the expression of E-cadherin and Vimentin in U87 cells and U251 cells by Western Blot analysis, the result showed that the expression of E-cadherin was increased in U87 cells and decreased in U251 cells. Down-regulation of FOXJ2 was significantly correlated to the low expression of E-cadherin.A strongly positive correlation was observed between FOXJ2 and E-cadherin(r=0.303;P<0.01). The expression of Vimentin was opposite to E-cadherin.3. A FOXJ2 plasmid was transient transfected into U87 cells. The increased expression of FOXJ2 was accompanied by up-regulation of E-cadherin and down-regulation of Vimentin. This data was confirmed with Western Blot and immunofluorescence analysis. We also found that over expression of FOXJ2 could inhibit the migration of U87 cells by wound healing assays and trans-well assays.4. We knocked down the expression of FOXJ2 in U87 cells.We found that the FOXJ2 sh RNA2 most effectively suppressed the expression of FOXJ2 compared with that of control and the decreased expression of E-cadherin and an increased expression of Vimentin. The knockdown efficiency was verified using Western Blot and immunofluorescence analysis. Besides, we found that sh RNA2 could promote the migration of U87 cells by wound healing assays and trans-well assays. Conclusions:1. The expression of FOXJ2 decreases with the increasing of histological grade in glioma. Down-regulation of FOXJ2 was significantly associated with poor overall survival in glioma patients.2. The expression of FOXJ2 can influence the expression of E-cadherin and Vimentin, affects the process of EMT, which inhibits the migration of glioma cells.