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Temozolomide (TMZ) is a standard chemotherapeutic drug used in the treatment of glioblastoma multiforme (GBM); however, resistance to this drug is common. SRY-Box 9 (SOX9) expression is associated with a poor prognosis of patients with GBM and with resistance to TMZ. Therefore, the aim of this study was to examine the effects of SOX9 inhibition on the sensitivity of glioma cells to TMZ treatment. We knocked down the expression of SOX9 (SOX9KD) via lentiviral infection in two glioblastoma (U87 and U251) cell lines, and the cells were then subjected to gene microarray, Gene Ontology and KEGG analysis pathway, all of which revealed a close association between SOX9 and the carbonic anhydrase 9 (CA9) gene. The TMZ-mediated apoptosis of glioma cells was significantly increased in the cells in the SOX9KD group. The potential underlying mechanism involved the downregulation of SOX9 and CA9 expression, which in turn decreased Akt phosphorylation, downregulated BCL‑2 expression, and upregulated BAX expression, as assessed by western blot analysis and RT-qPCR. The effects were found to be substantially enhanced in the cells in the SOX9KD group treated with TMZ. Subsequently, considering the association between SOX9 and CA9, the effects of CA9 inhibition, using a CA9 inhibitor (U‑104), on the chemosensitivity of glioma cells to TMZ were assessed. The results revealed that the use of U‑104 + TMZ effectively induced glioma cell death, compared to treatment with TMZ alone. The underlying mechanisms were similar to those observed with the silencing of SOX9 in the TMZ-treated glioma cells. On the whole, the findings of this study establish the SOX9/CA9-mediated oncogenic pathway in glioma, the inhibition of which enhances the sensitivity of glioma cells to TMZ treatment, and thus highlights the value of developing small molecules or antibodies against the SOX9/CA9 pathway, for combination therapy with TMZ, in the more efficient management of glioma.

The present study aimed to investigate the location, expression and clinical significance of Iroquois homeobox gene (IRX1) in human glioma. The expression of IRX1 gene in glioma cell lines (U87, U373, LN229 and T98G) and normal brain tissue was detected via reverse transcription-polymerase chain reaction. The IRX1 protein in fresh glioma specimens, with the adjacent normal brain tissue, was quantified through western blotting. The archived glioma only specimens from the present hospital and glioma specimens with adjacent normal brain tissue, from Alenabio biotechnology, were subjected to immunohistochemistry and tissue microarray analysis, respectively. The Kaplan-Meier method was employed to assess the correlation between the IRX1 level and the overall survival time of the patients. IRX1 gene was demonstrated to be expressed at varying levels in U373, LN229 and T98G cells, however not in U87 cells and normal brain tissue. Western blotting revealed increased IRX1 expression in glioma tissue compared with adjacent normal brain tissue. Furthermore, a direct correlation was observed between the IRX1 expression and the clinical glioma grade, with a significant difference in the gene expression between high grade and low grade glioma (P<0.05). Notably, IRX1 was identified to be localized to the cytoplasm in the adjacent normal brain and World Health Organization grade I glioma, whereas was identified to be present in the nucleus in higher grade glioma. In addition to being established as a significant prognostic variable, IRX1 expression was positively correlated with the overall survival of glioma patients. IRX1 gene may therefore exhibit an oncogenic role in glioma condition, and thus may be of clinical importance as a future therapeutic target.

Acylglycerol kinase (AGK) regulates various cellular processes involved into tumorigenesis and tumor progression. To investigate involvement of AGK in human gliomas, here, quantitative real-time polymerase chain reaction, Western blot, and immunohistochemistry analyses were performed to respectively detect the expression of AGK mRNA and protein in glioma and nonneoplastic brain tissue specimens. Then, the associations of AGK expression with various clinicopathological characteristics and patients' prognosis were statistically evaluated. Moreover, the effects of siRNA-mediated AGK knockdown on glioma cell proliferation, migration, and invasion were respectively assessed via Cell Counting Kit-8 and Transwell assays in vitro. As a result, AGK expression, at both mRNA and protein levels, were markedly up-regulated in glioma tissues compared with nonneoplastic brain tissues (both P < .001). In addition, high AGK expression was significantly associated with the grade of malignancy and poor prognosis in glioma patients (all P < .05). Importantly, Cox regression model of multivariate analysis identified AGK expression as an independent prognostic factor for glioma patients (P = .03). Furthermore, silencing the expression of AGK dramatically suppressed cell proliferation, migration, and invasion of glioma cells in vitro (all P < .05). In conclusion, AGK up-regulation may be involved into glioma development and progression, highlighting its prognostic value for the treatment of patients with this malignancy. Further loss-of-function experiments suggest that AGK might play an important role in the viability and motility of glioma cells, implying its potentials as an attractive therapeutic target for this tumor., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Gliomas make up about 80% of all malignant brain tumors, and cause serious public health problem. Genetic factors and environmental factors jointly caused the development of gliomas, and understanding of the genetic basis is a key component of preventive oncology. However, most genetic factors underlying carcinogenesis of gliomas remain largely unclear. In current study, we systematically evaluated whether genetic variants of SOX9 gene, a transcription factor that plays a central role in the development and differentiation of tumors, contribute to susceptibility of gliomas among Chinese population using a two-stage, case-control study. Results showed that SOX9 rs1042667 was significant associated with increased gliomas risk after adjusted by age, gender, family history of cancer, smoking status and alcohol status (Allele C vs A: OR=1.25; 95% CI=1.11-1.40; P=1.2×10-4). Compared with the carriers of genotype AA, both those of genotype AC (OR=1.37; 95% CI=1.13-1.66) and CC (OR=1.53; 95% CI=1.22-1.91) had significantly increased gliomas risk. This should be the first genetic association study which aims to evaluated the association between genetic variants of SOX9 and susceptibility of gliomas. Additional functional and association studies with different ethnic groups included are needed to further confirm our results.

Grb2-associated binder 1 (Gab1) and Gab2 play important roles in cancer cell signaling. In particular, it has been demonstrated that the upregulation of Gab2 may be correlated with the World Health Organization (WHO) grade of gliomas and that patients with high Gab2 expression levels exhibited shorter survival time. However, the prognostic value of combined expression of Gab1 and Gab2 has not been explored. Gab1 and Gab2 expression in human gliomas and non-neoplastic brain tissues was measured by immunohistochemistry. Both the expression levels of Gab1 and Gab2 proteins in glioma tissues were significantly higher than those in non-neoplastic brain tissues (both P < 0.001). In addition, the overexpression of Gab1 and Gab2 proteins were both significantly associated with advanced WHO grades (both P < 0.001) and low KPS (both P = 0.01). Moreover, the overall survival of patients with high Gab1 protein expression or high Gab2 protein expression was obviously lower than those with low expressions (both P < 0.001). Notably, glioma patients with combined overexpression of Gab1 and Gab2 proteins (Gab1-high/Gab2-high) had shortest overall survival (P < 0.001). Furthermore, multivariate analysis showed that Gab1 expression (P = 0.01), Gab2 expression (P = 0.02), and combined expression of Gab1 and Gab2 (Gab1/Gab2, P = 0.006) were all independent prognostic factors for overall survival in glioma patients. Gab1 and Gab2 proteins are differentially expressed in glioma patients and closely correlated with the biological behavior of this malignancy. Combination of Gab1 and Gab2 expression may represent a promising biomarker for prognostication of human gliomas.

Malignant gliomas are the most common central nervous system tumors and the molecular mechanism driving their development and recurrence is still largely unknown, limiting the treatment of this disease. Here, we show that restoring the expression of miR-218, a microRNA commonly downregulated in glioma, dramatically reduces the migration, invasion, and proliferation of glioma cells. Quantitative reverse transcription PCR and Western blotting analysis revealed that expression of the stem cell-promoting oncogene Bmi1 was decreased after overexpression of miR-218 in glioma cells. Mechanistic investigations defined Bmi1 as a functional downstream target of miR-218 through which miR-218 ablated cell migration and proliferation. We documented that miR-218 also blocked the self-renewal of glioma stem-like cells, consistent with the suggested role of Bmi1 in stem cell growth. Finally, we showed that miR-218 regulated a broad range of genes involved in glioma cell development, including Wnt pathways that suppress glioma cell stem-like qualities. Taken together, our findings reveal miR-218 as a tumor suppressor that prevents migration, invasion, proliferation, and stem-like qualities in glioma cells.

Chromodomain helicase DNA-binding protein 5 (CHD5), a member of the CHD family, is involved in key cellular processes including chromatin remodeling, cell cycle regulation, and cellular adhesion. Recent studies have demonstrated that CHD5 is the product of a novel tumor suppressor gene and is implicated in certain tumor types. However, the clinicopathological significance of CHD5 expression in human malignant gliomas remains unclear. To address this problem, CHD5 expression in human gliomas and non-neoplastic brain tissues was measured using real-time quantitative polymerase chain reaction (RT-PCR) assay, Western blot, and immunohistochemistry. The association of CHD5 immunostaining with clinicopathological factors or prognosis of glioma patients was statistically analyzed. Genetic and protein expression of CHD5 were downregulated in glioma tissues compared to corresponding non-neoplastic brain tissues (both p<0.001). Additionally, decreased expression of CHD5 in glioma was significantly associated with pathological grade (p=0.007); high pathological grade was associated with low CHD5 expression. Loss of CHD5 protein expression was also significantly correlated with a low Karnofsky performance scale score (p=0.01). Moreover, overall survival of patients with low CHD5 protein expression was dramatically shorter than those of patients with high CHD5 protein expression (p=0.003). Multivariate Cox regression analysis indicated that CHD5 expression was an independent prognostic factor for patients with gliomas (p=0.01). In conclusion, these data offer convincing evidence for the first time that CHD5 might act as a tumor suppressor in glioma, may act as a regulator of aggressive development, and is a candidate prognostic marker for this malignancy., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Background & Aim: Human kinesin superfamily proteins (KIFs) are a conserved class of microtubule-dependent molecular motor proteins with adenosine triphosphatase activity and motion characteristics. As a member of KIFs, KIF14 plays an important role in the regulation of cell cycle and mitotic progression. Deregulation of KIF14 has been found in several human malignancies and also has been demonstrated to be involved in tumor progression and related to patient survival. The aim of this study was to investigate the clinicopathological significance of KIF14 expression in glioma., Methods: Real-time quantitative RT-PCR assay was performed to detect KIF14 mRNA expression, and Western blot and immunohistochemistry analyses were performed to detect KIF14 protein expression in human gliomas and non-neoplastic brain tissues, respectively. Then, the association of KIF14 immunostaining with clinicopathological factors and prognosis of glioma patients was also statistically analyzed., Results: KIF14 mRNA and protein expression were respectively increased 5.5- and 4.2-fold on average in glioma tissues relative to non-neoplastic brain tissues (both P < 0.001). Additionally, both KIF14 mRNA and protein expression increased with ascending pathological grade. Then, the high KIF14 immunostaining in glioma tissues was significantly associated with advanced pathological grade (P = 0.008), low Karnofsky performance score (KPS) (P = 0.02), high mitotic index (P = 0.005) and Ki-67 index (P = 0.008). Furthermore, both univariate and multivariate Cox regression analyses determined that KIF14 overexpression effectively predicted decreased overall survival in patients with gliomas., Conclusions: These findings offer the first convinced evidence that KIF14 expression in gliomas is tumor-specific and increased in more aggressive tumors. KIF14 might function as a candidate prognostic marker for human gliomas., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Aim: Matrix metalloproteinase (MMP)-14 and MMP-19 have been demonstrated to play an important role in the development of human gliomas. However, their prognostic values are not clear. The aim of this study was to investigate whether co-expression of MMP-14 and MMP-19 has prognostic relevance in human gliomas., Methods: Immunohistochemistry and western blot were used to investigate the expression of MMP-14 and MMP-19 proteins in 128 patients with gliomas., Results: The expression levels of MMP-14 and MMP-19 proteins in glioma tissues were both significantly higher (both P < 0.001) than those in non-neoplastic brain tissues according to the immunohistochemistry analysis, which was confirmed by the western blot analysis. Additionally, the overexpression of either MMP-14 or MMP-19 was significantly associated with the advanced WHO grade (both P = 0.02), the low Karnofsky performance score (KPS) (P = 0.008 and 0.01, respectively) and the poor overall survival (both P = 0.01). Moreover, the Multivariate Cox proportional-hazards regression analysis revealed that the increased expressions of MMP-14 and MMP-19 were both independent prognostic factors for poor overall survival (both P = 0.02). Furthermore, the co-expression of MMP-14 and MMP-19 was additively and more significantly (P = 0.006) associated with adverse prognosis in patients with gliomas than respective expression of MMP-14 and MMP-19., Conclusions: These findings indicated for the first time that the co-expression of MMP-14 and MMP-19 is significantly correlated with prognosis in glioma patients, suggesting that the co-expression of these proteins may be used as both an early diagnostic and independent prognostic marker.

SOX9 belongs to the SOX (Sry-related high-mobility group box) family and acts as a transcription factor that plays a central role in the development and differentiation of multiple cell lineages. Recent studies have demonstrated that SOX9 is required for the carcinogenesis in several cancer types. The aim of this study was to investigate the clinicopathological significance of SOX9 expression in human malignant glioma. SOX9 mRNA expression was detected by real-time quantitative RT-PCR assay in glioma and nonneoplastic brain tissues. Then, the association of SOX9 mRNA expression with clinicopathological factors or prognosis of glioma patients was statistically analyzed. In addition, the small interfering RNA was used to knockdown SOX9 expression in a glioma cell line and to analyze the effects of SOX9 inhibition on cell growth, cell cycle and apoptosis of glioma cell line. The expression level of SOX9 mRNA in glioma tissues was significantly higher than that in corresponding nonneoplastic brain tissues (P < 0.001). In addition, a high level of SOX9 mRNA expression was significantly more common in glioma tissues with advanced WHO grade than those with low grade (P = 0.02). The increased expression of SOX9 mRNA was also significantly correlated with low Karnofsky performance score (P = 0.008). Meanwhile, the disease-free and overall survival rates of patients with high SOX9 mRNA expression were obviously lower than those of patients with low SOX9 mRNA expression (both P = 0.01). Multivariate analysis showed that high SOX9 mRNA expression was an independent prognostic factor for glioma patients (P = 0.02). Moreover, the down-regulation of SOX9 could inhibit the cell growth, induce the cell arrest in G2/M phase of cell cycle and enhance the apoptosis in glioma cells. Our data suggest for the first time that the over-expression of SOX9 mRNA is closely associated with poor clinical outcome of patients with malignant gliomas, and targeting SOX9 may be a novel therapeutic strategy for this tumor.

Recent studies have demonstrated that the chemokine CCL20 and its receptor CCR6 may be involved in tumorigenesis, tumor progression and metastatic spread of various human malignancies. The aim of this study was to investigate the clinicopathological significance and prognostic value of CCL20 and CCR6 expression in human malignant glioma. CCL20 and CCR6 expression in human gliomas and nonneoplastic brain tissues was measured by immunohistochemistry. The association of CCL20 and CCR6 expression with clinicopathological factors or prognosis in glioma patients was statistically analyzed. The expression levels of CCL20 and CCR6 proteins were both up-regulated in glioma tissues. There was a significantly positive correlation between the expression of the two markers (r = 0.88; P < 0.001). In addition, the overexpressions of CCL20 and CCR6 were both detected in high-grade glioma tissues compared with those in low-grade tissues and increased with ascending tumor World Health Organization (WHO) grades (P = 0.006 and 0.008, respectively). The increased expressions of CCL20 and CCR6 proteins were also significantly correlated with low Karnofsky performance score (both P = 0.01). Moreover, univariate analysis found that CCL20 expression (P = 0.002), CCR6 expression (P = 0.002) and CCL20/CCR6 co-expression (P < 0.001) were all significantly associated with poor prognosis. In particular, glioma patients with CCL20/CCR6 co-expression have the shortest overall survival. Multivariate analysis further identified the expression levels of CCL20 and CCR6 to be independent prognostic factors. Our data suggest for the first time that CCL20 and CCR6 might play an important role in the regulation of aggressiveness in human gliomas. The up-regulation of CCL20 and CCR6 might be closely associated with poor clinical outcome of patients with gliomas.

FAT10, as a small ubiquitin-like modifier, plays an important role in various cellular processes, including mitosis, immune response, and apoptosis, the dys-regulation of which may arise tumorigenesis. Therefore, the aim of this study was to examine the expression of FAT10 at gene and protein levels in glioma samples with different WHO grades and its association with survival. One hundred and twenty-eight glioma specimens and 10 non-neoplastic brain tissues were collected. Immunohistochemistry assay, quantitative real-time PCR and Western blot analysis were carried out to investigate the expression of FAT10. Kaplan-Meier method and Cox's proportional hazards model were used in survival analysis. Immunohistochemistry showed that FAT10 protein was over-expressed in glioma tissues. FAT10 mRNA and protein levels were both higher in glioma compared to control on real-time PCR and Western blot analysis (both P < 0.001). Additionally, its expression levels increase from grade I to grade IV glioma according to the results of real-time PCR, immunohistochemistry analysis and Western blot. Moreover, the survival rate of FAT10-positive patients was significantly lower than that of FAT10-negative patients (P < 0.001). We further confirmed that the increased expression of FAT10 was a significant and independent prognostic indicator in glioma by multivariate analysis (P < 0.001). Our data provides convincing evidence for the first time that the increased expression of FAT10 at gene and protein levels is correlated with poor outcome in patients with glioma. FAT10 may promote the aggressiveness of glioma and may be a potential prognosis predictor of glioma.

Aim: The aberrant expression of regenerating islet-derived family member, 4 (Reg IV) has been found in various human cancers. However, the roles of Reg IV gene and its encoding product in human glioma have not been clearly understood. Therefore, the aim of this study was to investigate the clinicopathological significance of Reg IV expression in glioma., Methods: Reg IV mRNA and protein expression in human gliomas and non-neoplastic brain tissues were respectively detected by real-time quantitative RT-PCR assay, Western blot, and immunohistochemistry. The association of Reg IV immunostaining with clinicopathological factors and prognosis of glioma patients was also statistically analyzed., Results: Reg IV mRNA and protein expression levels in glioma tissues were both significantly higher than those in the corresponding non-neoplastic brain tissues (both P < 0.001). Additionally, the increased Reg IV immunostaining in glioma tissues was significantly associated with advanced pathological grade (P = 0.008). Reg IV protein up-regulation was also significantly correlated with low Karnofsky performance score (KPS) (P = 0.02). Moreover, the overall survival of patients with high Reg IV protein expression was dramatically shorter than those with low Reg IV protein expression (P < 0.001). Multivariate Cox regression analysis further confirmed that Reg IV expression was an independent prognostic factor for patients with gliomas (P = 0.008)., Conclusions: These convinced evidences suggest for the first time that Reg IV might accelerate disease progression and act as a candidate prognostic marker for gliomas., Virtual Slides: The virtual slide(s) for this article can be found here:http://www.diagnosticpathology.diagnomx.eu/vs/2145344361720706.

Introduction: Glioma is one of the most aggressive human tumors; however, little is known about its genetic risk factors. The role of heredity is likely to be explained by combinations of common low-risk variants. Previous studies have indicated that more than 100 single nucleotide polymorphisms (SNPs) are associated with the risk of glioma., Methods: To further investigate how and to what extent these SNPs contribute to glioma susceptibility in a Chinese population, we analyzed 43 SNPs of 226 glioma patients and 254 normal people in order to evaluate the associations between SNPs and the risk of glioma., Results: Overall, we found three protective alleles for glioma in patients: the allele "G" of rs1801275 in the IL4R gene by allele model (odds ratio [OR], 0.71; 95% confidence interval [CI], 0.50-0.99; P=0.04) and dominant model (OR, 0.67; 95% CI, 0.46-0.99; P=0.04) analysis respectively, the allele "T" of rs17748 in the TREH gene by recessive model (OR, 0.48; 95% CI, 0.23-1.01; P=0.05) analysis, and the allele "G" of rs6470745 in CCDC26 gene by recessive model (OR, 0.48; 95% CI, 0.26-0.89; P=0.02) analysis., Conclusion: This study provides evidence for three glioma susceptibility genes - TREH, IL4R and CCDC26 - in a Chinese population; this may shed light on molecular markers of glioma susceptibility and could therefore be used as a diagnostic and prognostic marker for glioma patients in clinical study., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: Chloride intracellular channel 1 (CLIC1) is expressed ubiquitously in human tissues and is involved in the regulation of cell cycle, cell proliferation and differentiation. Recent studies have shown that CLIC1 is highly expressed in several human malignant tumors. However, its roles in human gliomas are still unclear. The aim of this study was to investigate the clinicopathological significance and prognostic value of CLIC1 expression in human gliomas., Methods: CLIC1 expression in human gliomas and nonneoplastic brain tissues was measured by real-time quantitative RT-PCR assay and immunohistochemistry. Its association with clinicopathological factors or prognosis in patients with gliomas was statistically analyzed., Results: The expression of CLIC1 at both mRNA and protein levels was significantly increased in high-grade (Grade III~IV) glioma tissues compared with that in low-grade (Grade I~II) and nonneoplastic brain tissues, and was up-regulated with ascending tumor World Health Organization (WHO) grades. The elevated expression of CLIC1 protein was also significantly correlated with low Karnofsky performance score (KPS) (P=0.008). Moreover, both univariate and multivariate analysis shown that high CLIC1 expression was significantly associated with poor prognosis in patients with gliomas (P<0.001 and P=0.01, respectively). In particular, the elevated CLIC1 expression also correlated with shorter overall survival in different glioma subgroups stratified according to the WHO grading., Conclusions: Our data provide the first evidence that CLIC1 expression might play an important role in the regulation of aggressiveness in human gliomas. The elevated expression of CLIC1 might represent a valuable prognostic marker for this disease.

Aim: To examine the expression of N-methylpurine-DNA glycosylase (MPG) gene and protein in glioma samples with different WHO grades and its association with patients' survival., Methods: Immunohistochemistry assay, quantitative real-time PCR and Western blot analysis were carried out to investigate the expression of MPG gene and protein in 128 glioma and 10 non-neoplastic brain tissues., Results: MPG gene expression level in glioma tissues was significantly higher than that in non-neoplastic brain tissues (P < 0.001). Immunohistochemistry also showed that MPG protein was over-expressed in glioma tissues, which was consistent with the resutls of Western blot analysis. Additionally, the expression levels of MPG gene and protein both increase from grade I to grade IV glioma according to the results of real-time PCR, immunohistochemistry and western blot analysis. Moreover, the survival rate of MPG-positive patients was significantly lower than that of MPG-negative patients (P < 0.001). We further confirmed that the over-expression of MPG was a significant and independent prognostic indicator in glioma by multivariate analysis (P < 0.001)., Conclusions: Our data showed the over-expression of MPG gene and protein in human gliomas, and also suggested for the first time that MPG be an unfavorable independent prognostic indicator for glioma patients.