Syndecan-1 expression in human glioma is correlated with advanced tumor progression and poor prognosis

Syndecan-1 has been implicated in tumorigenesis and progression of various human malignancies. Recent studies have demonstrated that syndecan-1 may have a different function and biological activity depending on the specific tumor type. Therefore, the aim of this study was to investigate the clinical significance of syndecan-1 in human gliomas. One hundred and sixteen glioma patients (26 World Health Organization (WHO) grade I, 30 WHO grade II, 30 WHO grade III, and 30 WHO grade IV) and 15 normal brain specimens acquired from 15 patients undergoing surgery for epilepsy as control were collected. Immunohistochemistry assay, quantitative real-time PCR and Western blot analysis were carried out to detect the expression of syndecan-1 at gene and protein levels in glioma samples with different WHO grades. Syndecan-1 gene and protein levels were both higher in glioma tissues compared to controls (both P < 0.001). In addition, its expression levels increased with ascending tumor WHO grades according to the results of immunohistochemistry assay, quantitative real-time PCR and Western blot analysis. Moreover, the survival rate of syndecan-1-positive patients was significantly lower than that of syndecan-1-negative patients (P = 0.006). We further confirmed that the increased expression of syndecan-1 was an independent prognostic indicator in glioma by multivariate analysis (P = 0.01). Our data suggest for the first time that the increased expression of syndecan-1 at gene and protein levels is correlated with advanced tumor progression and poor outcome in patients with glioma. Syndecan-1 might serve as a potential prognosis predictor of this dismal tumor.     

Identification of miRNA-Mediated Core Gene Module for Glioma Patient Prediction by Integrating High-Throughput miRNA,mRNA Expression and Pathway Structure

The prognosis of glioma patients is usually poor, especially in patients with glioblastoma (World Health Organization (WHO) grade IV). The regulatory functions of microRNA (miRNA) on genes have important implications in glioma cell survival. However, there are not many studies that have investigated glioma survival by integrating miRNAs and genes while also considering pathway structure. In this study, we performed sample-matched miRNA and mRNA expression profilings to systematically analyze glioma patient survival. During this analytical process, we developed pathway-based random walk to identify a glioma core miRNA-gene module, simultaneously considering pathway structure information and multi-level involvement of miRNAs and genes. The core miRNA-gene module we identified was comprised of four apparent sub-modules; all four sub-modules displayed a significant correlation with patient survival in the testing set (P-values≤0.001). Notably, one sub-module that consisted of 6 miRNAs and 26 genes also correlated with survival time in the high-grade subgroup (WHO grade III and IV), P-value = 0.0062. Furthermore, the 26-gene expression signature from this sub-module had robust predictive power in four independent, publicly available glioma datasets. Our findings suggested that the expression signatures, which were identified by integration of miRNA and gene level, were closely associated with overall survival among the glioma patients with various grades.     

Effect of Change in Spindle Structure on Proliferation Inhibition of Osteosarcoma Cells and Osteoblast under Simulated Microgravity during Incubation in Rotating Bioreactor

In order to study the effect of microgravity on the proliferation of mammalian osteosarcoma cells and osteoblasts, the changes in cell proliferation, spindle structure, expression of MAD2 or BUB1, and effect of MAD2 or BUB1 on the inhibition of cell proliferation is investigated by keeping mammalian osteosarcoma cells and osteoblasts under simulated microgravity in a rotating wall vessel (2D-RWVS) bioreactor. Experimental results indicate that the effect of microgravity on proliferation inhibition, incidence of multipolar spindles, and expression of MAD2 or BUB1 increases with the extension of treatment time. And multipolar cells enter mitosis after MAD2 or BUB1 is knocked down, which leads to the decrease in DNA content, and decrease the accumulation of cells within multipolar spindles. It can therefore be concluded that simulated microgravity can alter the structure of spindle microtubules, and stimulate the formation of multipolar spindles together with multicentrosomes, which causes the overexpression of SAC proteins to block the abnormal cells in metaphase, thereby inhibiting cell proliferation. By clarifying the relationship between cell proliferation inhibition, spindle structure and SAC changes under simulated microgravity, the molecular mechanism and morphology basis of proliferation inhibition induced by microgravity is revealed, which will give experiment and theoretical evidence for the mechanism of space bone loss and some other space medicine problems.     

Characterization of MAD2B and other mitotic spindle checkpoint genes.

Aneuploidy is a characteristic of the majority of human cancers, and recent work has suggested that mitotic checkpoint defects play a role in its development. To further explore this issue, we isolated a novel human gene, MAD2B (MAD2L2), which is homologous to the spindle checkpoint gene MAD2 (MAD2L1). We determined the chromosomal localization of it and other spindle checkpoint genes, including MAD1L1, MAD2, BUB3, TTK (MPS1L1), and CDC20. In addition, we resolved the genomic intron-exon structure of the human BUB1 gene. We then searched for mutations in these genes in a panel of 19 aneuploid colorectal tumors. No new mutations were identified, suggesting that genes yet to be discovered are responsible for most of the checkpoint defects observed in aneuploid cancers.     

Expression of Tumor-Associated Macrophage in Progression of Human Glioma

The aim of this study is to investigate the expression of tumor-associated macrophages (TAMs) M1, M2 phenotypic in human glioma tissues, and to explore the clinical significance and prognostic value of TAMs in glioma patients. A total of 50 glioma samples were obtained from patients diagnosed in our hospital from 2007 to 2010. Clinical follow-up was conducted via return visits and telephone interviews after discharge. Progression free survival (PFS) was calculated based on tumor progression by MRI and CT examination from the primary operation. Overall survival (OS) time was calculated from the initial surgical operation date to end date of follow-up or death. Kaplan–Meier methodology was used to evaluate the survival of patients and log-rank test for comparing differences between groups. The expression levels of CD16 and CD206 were investigated in the 4 μm serial paraffin sections by immunohistochemistry. M1-type macrophages filtrated in all the grades of glioma samples, and the lower expression level was associated with high grade glioma. A negative correlation was found between WHO pathological grades and the expression of M1-type macrophages by Spearman correlation analysis. M2-type macrophages filtrated in all the grades of glioma samples with the higher expression level associated with high grade glioma. A positive correlation was found between WHO pathological grades and the expression of M2-type macrophages by Spearman correlation analysis. The PFS and OS among patients with high levels of M1-type macrophages (CD16+++) were significantly higher than those with less expression. The PFS and OS among patients with high levels of M2-type macrophages (CD206+++) were significantly lower than those with low expression. M1-type macrophages may inhibit the tumor growth and improve the therapeutic outcome of glioma patients. M2 ratios are associated with tumor proliferation and poor prognosis. TAMs phenotypes of glioma samples are the potential biomarkers in assessing the degree of malignancy, tumor invasion, and patient prognosis in clinic.     

Spindle Assembly Checkpoint Protein Dynamics Reveal Conserved and Unsuspected Roles in Plant Cell Division

Background

In eukaryotes, the spindle assembly checkpoint (SAC) ensures that chromosomes undergoing mitosis do not segregate until they are properly attached to the microtubules of the spindle.

Methodology/Principal Findings

We investigated the mechanism underlying this surveillance mechanism in plants, by characterising the orthogolous SAC proteins BUBR1, BUB3 and MAD2 from Arabidopsis. We showed that the cell cycle-regulated BUBR1, BUB3.1 and MAD2 proteins interacted physically with each other. Furthermore, BUBR1 and MAD2 interacted specifically at chromocenters. Following SAC activation by global defects in spindle assembly, these three interacting partners localised to unattached kinetochores. In addition, in cases of ‘wait anaphase’, plant SAC proteins were associated with both kinetochores and kinetochore microtubules. Unexpectedly, BUB3.1 was also found in the phragmoplast midline during the final step of cell division in plants.

Conclusions/Significance

We conclude that plant BUBR1, BUB3.1 and MAD2 proteins may have the SAC protein functions conserved from yeast to humans. The association of BUB3.1 with both unattached kinetochore and phragmoplast suggests that in plant, BUB3.1 may have other roles beyond the spindle assembly checkpoint itself. Finally, this study of the SAC dynamics pinpoints uncharacterised roles of this surveillance mechanism in plant cell division.     

Biological and Clinical Significance of MAD2L1 and BUB1, Genes Frequently Appearing in Expression Signatures for Breast Cancer Prognosis

To investigate the biologic relevance and clinical implication of genes involved in multiple gene expression signatures for breast cancer prognosis, we identified 16 published gene expression signatures, and selected two genes, MAD2L1 and BUB1. These genes appeared in 5 signatures and were involved in cell-cycle regulation. We analyzed the expression of these genes in relation to tumor features and disease outcomes. In vitro experiments were also performed in two breast cancer cell lines, MDA-MB-231 and MDA-MB-468, to assess cell proliferation, migration and invasion after knocking down the expression of these genes. High expression of these genes was found to be associated with aggressive tumors and poor disease-free survival of 203 breast cancer patients in our study, and the association with survival was confirmed in an online database consisting of 914 patients. In vitro experiments demonstrated that lowering the expression of these genes by siRNAs reduced tumor cell growth and inhibited cell migration and invasion. Our investigation suggests that MAD2L1 and BUB1 may play important roles in breast cancer progression, and measuring the expression of these genes may assist the prediction of breast cancer prognosis.     

Conditional targeting of MAD1 to kinetochores is sufficient to reactivate the spindle assembly checkpoint in metaphase

Fidelity of chromosome segregation is monitored by the spindle assembly checkpoint (SAC). Key components of the SAC include MAD1, MAD2, BUB1, BUB3, BUBR1, and MPS1. These proteins accumulate on kinetochores in early prometaphase but are displaced when chromosomes attach to microtubules and/or biorient on the mitotic spindle. As a result, stable attachment of the final chromosome satisfies the SAC, permitting activation of the anaphase promoting complex/cyclosome (APC/C) and subsequent anaphase onset. SAC satisfaction is reversible, however, as addition of taxol during metaphase stops cyclin B1 degradation by the APC/C. We now show that targeting MAD1 to kinetochores during metaphase is sufficient to reestablish SAC activity after initial silencing. Using rapamycin-induced heterodimerization of FKBP-MAD1 to FRB-MIS12 and live monitoring of cyclin B1 degradation, we show that timed relocalization of MAD1 during metaphase can stop cyclin B1 degradation without affecting chromosome-spindle attachments. APC/C inhibition represented true SAC reactivation, as FKBP-MAD1 required an intact MAD2-interaction motif and MPS1 activity to accomplish this. Our data show that MAD1 kinetochore localization dictates SAC activity and imply that SAC regulatory mechanisms downstream of MAD1 remain functional in metaphase.     

High Cytoplasmic FOXO1 and pFOXO1 Expression in Astrocytomas Are Associated with Worse Surgical Outcome

FOXO1 is at a convergence point of receptor tyrosine kinase (RTK) signaling, which is one of the three core pathways implicated in glioblastoma. It was recently shown that FOXO1 can effectively induce glioma cell death and inhibit tumor growth through cell cycle arrest and apoptosis. We therefore evaluated FOXO1 and pFOXO1 protein expression in 181 primary astrocytoma samples and 16 normal brain samples. Astrocytoma samples expressed higher cytoplasmic FOXO1 and pFOXO1 than normal brain samples. Nuclear pFOXO1 level was significantly higher than nuclear FOXO1 in astrocytomas. High cytoplasmic FOXO1 expression was associated with older onset age (P = 0.001) and higher WHO grade (P = 0.001). The trend was also observed between cytoplasmic pFOXO1 expression and WHO grade although not significant. Univariate survival analysis showed that both high cytoplasmic FOXO1 and pFOXO1 expression indicated a significantly shorter median overall survival and progression-free survival. Multivariate survival analysis revealed cytoplasmic FOXO1 expression, cytoplasmic pFOXO1 expression, WHO grade, gender, extent of resection and radiotherapy to be independent prognostic factors for overall survival and progression-free survival. Thus, our data suggested that cytoplasmic FOXO1 and pFOXO1 expression may serve as valuable prognostic variables in astrocytomas and may have significant implications for the development and application of targeted therapy.     

Identification of COL6A1 as a differentially expressed gene in human astrocytomas

Diffuse infiltrating gliomas are the most common tumors of the central nervous system. Gliomas are classified by the WHO according to their histopathological and clinical characteristics into four classes: grade I (pilocytic astrocytoma), grade II (diffuse astrocytoma), grade III (anaplastic astrocytoma), and grade IV (glioblastoma multiforme). Several genes have already been correlated with astrocytomas, but many others are yet to be uncovered. By analyzing the public SAGE data from 21 patients, comprising low malignant grade astrocytomas and glioblastomas, we found COL6A1 to be differentially expressed, confirming this finding by real time RT-PCR in 66 surgical samples. To the best of our knowledge, COL6A1 has never been described in gliomas. The expression of this gene has significantly different means when normal glia is compared with low-grade astrocytomas (grades I and II) and high-grade astrocytomas (grades III and IV), with a tendency to be greater in higher grade samples, thus rendering it a powerful tumor marker.     

Differential Expression of Circular RNAs in Glioblastoma Multiforme and Its Correlation with Prognosis

OBJECTIVE: The present study aimed to explore the expression profiles of circular RNAs (circRNAs) in glioblastoma multiforme (GBM) in an attempt to identify potential core genes in the pathogenesis of this tumor. METHODS: Differentially expressed circRNAs were screened between tumor tissues from five GBM patients and five normal brain samples using Illumina Hiseq. Bioinformatics analysis was used to analyze their potential function. CircBRAF was further detected in different WHO grades glioma tissues and normal brain tissues. Kaplan-Meier curves and multivariate Cox's analysis were used to analyze the association between circBRAF expression level and prognosis of glioma patients. RESULTS: A total of 1411 differentially expressed circRNAs were identified in GBM patients including 206 upregulated circRNAs and 1205 downregulated circRNAs. Differential expression of circRNAs was closely associated with the biological process and molecular function. The downregulated circRNAs were mainly associated with ErbB and Neurotrophin signaling pathways. Moreover, the expression level of circBRAF in normal brain tissues was significantly higher than that in glioma tissues (P < .001). CircBRAF was significantly lower in glioma patients with high pathological grade (WHO III & IV) than those with low grade (WHO I & II) (P < .001). Cox analysis revealed that high circBRAF expression was an independent biomarker for predicting good progression-free survival and overall survival in glioma patients (HR = 0.413, 95% CI 0.201-0.849; HR = 0.299, 95% CI 0.135-0.661; respectively). CONCLUSION: The present study identified a profile of dysregulated circRNAs in GBM. Bioinformatics analysis showed that dysregulated circRNAs might be associated with tumorigenesis and development of GBM. In addition, circBRAF could severe as a biomarker for predicting pathological grade and prognosis in glioma patients.     

Increased expression of microRNA-9 predicts an unfavorable prognosis in human glioma

microRNA-9 (miR-9) has been found to be upregulated along with tumor progression of gliomas by microarray-based expression profiling, and also be strongly linked to glioblastoma subtypes. However, its prognostic value in glioma is still elusive. miR-9 expression in human gliomas and nonneoplastic brain tissues was measured by real-time quantitative RT-PCR assay. miR-9 expression in glioma tissues was significantly higher than that in corresponding nonneoplastic brain tissues (P < 0.001). The increased expression of miR-9 was more frequently observed in glioma tissues with high WHO grade than those with low WHO grade tissues (P = 0.001). The expression levels of miR-9 in glioma tissues with low Karnofsky performance score (KPS) were also significantly higher than those with high KPS (P = 0.008). Moreover, the overall survival of glioma patients with high miR-9 expression was obviously lower than that with low miR-9 expression (P < 0.001). Multivariate analysis further showed that high miR-9 expression was an independent prognostic factor for overall survival in glioma patients (P = 0.01). More importantly, the subgroup analyses indicated that the overall survival of glioma patients with high WHO grade (III–IV) was significantly worse for high miR-9 expression group than for low miR-9 expression group (P < 0.001), but no significant difference was found for patients with low WHO grade (I–II). These findings suggest for the first time that the increased expression of miR-9 may play an important role in tumor progression in human gliomas. miR-9 might be a useful marker for predicting the clinical outcome of glioma patients, especially for advanced subtypes.     

Hyperplasia of Pericytes Is One of the Main Characteristics of Microvascular Architecture in Malignant Glioma

Objectives

To investigate the role of pericytes in constructing the malformed microvessels (MVs) and participating microvascular architecture heterogeneity of glioma.

Methods

Forty human glioma tissue samples (WHO grade II-IV) were included in present study. Observation of blood vessel patterns, quantitative analysis of endothelial cells (ECs)- and pericyte-labeled MVs and comparison between malignant grades based on single- or double-immunohistochemical staining. The MV number density (MVND), microvascular pericyte number density (MPND), and microvascular pericyte area density (MPAD) were calculated. The expression of PDGFβ was also scored after immunostaining.

Results

In grade II glioma, most of tumor MVs were the thin-wall CD34⁺ vessels with near normal morphology. In addition to thin-wall CD34⁺ MVs, more thick-wall MVs were found in grade III glioma, which often showed α-SMA positive. Most of MVs in grade IV glioma were in the form of plexus, curled cell cords and glomeruloid microvascular proliferation while the α-SMA⁺ cells were the main components. The MVs usually showed disordered arrangement, loose connection and active cell proliferation as shown by Ki67 and α-SMA coexpression. With the increase of glioma grades, the α-SMA⁺ MVND, CD34⁺ MVND and MPND were significantly augmented although the increase of CD34⁺ MVND but not MPAD was statistically insignificant between grade III and IV. It was interesting that some vessel-like structures only consist of α-SMA⁺ cells, assuming the guiding role of pericytes in angiogenesis. The expression level of PDGFβ was upregulated and directly correlated with the MPND in different glioma grades.

Conclusion

Hyperplasia of pericytes was one of the significant characteristics of malignant glioma and locally proliferated pericytes were the main constituent of MVs in high grade glioma. The pathological characteristics of pericytes could be used as indexes of malignant grades of glioma.     

Glioma IL13Rα2 Is Associated with Mesenchymal Signature Gene Expression and Poor Patient Prognosis

A major challenge for successful immunotherapy against glioma is the identification and characterization of validated targets. We have taken a bioinformatics approach towards understanding the biological context of IL-13 receptor α2 (IL13Rα2) expression in brain tumors, and its functional significance for patient survival. Querying multiple gene expression databases, we show that IL13Rα2 expression increases with glioma malignancy grade, and expression for high-grade tumors is bimodal, with approximately 58% of WHO grade IV gliomas over-expressing this receptor. By several measures, IL13Rα2 expression in patient samples and low-passage primary glioma lines most consistently correlates with the expression of signature genes defining mesenchymal subclass tumors and negatively correlates with proneural signature genes as defined by two studies. Positive associations were also noted with proliferative signature genes, whereas no consistent associations were found with either classical or neural signature genes. Probing the potential functional consequences of this mesenchymal association through IPA analysis suggests that IL13Rα2 expression is associated with activation of proinflammatory and immune pathways characteristic of mesenchymal subclass tumors. In addition, survival analyses indicate that IL13Rα2 over-expression is associated with poor patient prognosis, a single gene correlation ranking IL13Rα2 in the top ~1% of total gene expression probes with regard to survival association with WHO IV gliomas. This study better defines the functional consequences of IL13Rα2 expression by demonstrating association with mesenchymal signature gene expression and poor patient prognosis. It thus highlights the utility of IL13Rα2 as a therapeutic target, and helps define patient populations most likely to respond to immunotherapy in present and future clinical trials.     

脑胶质瘤组织miR-211、miR-374、miR-510表达水平与临床病理特征及预后的关系研究

摘要 目的：探讨脑胶质瘤组织微小RNA（miR）-211、miR-374、miR-510表达水平与临床病理特征及预后的关系。方法：选择2013年8月至2015年8月我院诊治的83例脑胶质瘤患者作为研究对象,选择同期由于脑外伤在我院行内减压术切除的正常脑组织样本31份作为对照样本。采用荧光定量PCR检测miR-211、miR-374、miR-510表达水平,生存分析采用Kaplan-Meier法,应用Cox比例风险回归模型分析预后的影响因素。结果：与正常脑组织相比,脑胶质瘤组织中miR-211、miR-374表达水平明显下降,miR-510表达水平明显升高（P<0.05）。脑胶质瘤组织miR-211、miR-374、miR-510表达均与WHO分级和卡氏功能状态量表(KPS)评分有关（P<0.05）。miR-211、miR-374低表达患者的5年总生存率明显低于高表达患者,miR-510低表达患者的5年总生存率明显高于高表达患者（P<0.05）。WHO分级、KPS评分、miR-211、miR-374和miR-510表达是脑胶质瘤患者预后的影响因素（P<0.05）。结论：脑胶质瘤组织中miR-211和miR-374表达下调,而miR-510表达上调,miR-211、miR-374和miR-510表达均与WHO分级、KPS评分和预后相关,检测miR-211、miR-374和miR-51在脑胶质瘤患者的诊断和治疗中具有一定临床意义。