Umbilical cord-derived mesenchymal stromal/stem cells expressing IL-24 induce apoptosis in gliomas.

Although much progress has been made in the treatment of gliomas, the prognosis for patients with gliomas is still very poor. Stem cell-based therapies may be promising options for glioma treatment. Recently, many studies have reported that umbilical cord-derived mesenchymal stromal/stem cells (UC-MSCs) are ideal gene vehicles for tumor gene therapy. Interleukin 24 (IL-24) is a pleiotropic immunoregulatory cytokine that has an apoptotic effect on many kinds of tumor cells and can inhibit the growth of tumors specifically without damaging normal cells. In this study, we investigated UC-MSCs as a vehicle for the targeted delivery of IL-24 to tumor sites. UC-MSCs were transduced with lentiviral vectors carrying green fluorescent protein (GFP) or IL-24 complementary DNA. The results indicated that UC-MSCs could selectively migrate to glioma cells in vitro and in vivo. Injection of IL-24-UC-MSCs significantly suppressed tumor growth of glioma xenografts. The restrictive efficacy of IL-24-UC-MSCs was associated with the inhibition of proliferation as well as the induction of apoptosis in tumor cells. These findings indicate that UC-MSC-based IL-24 gene therapy may be able to suppress the growth of glioma xenografts, thereby suggesting possible future therapeutic use in the treatment of gliomas.

Up-regulation of MCM3 Relates to Neuronal Apoptosis After Traumatic Brain Injury in Adult Rats.

Minichromosome maintenance complex component 3, one of the minichromosome maintenance proteins, functions as a part of pre-replication complex to initiate DNA replication in eukaryotes. Minichromosome maintenance complex component 3 (MCM3) was mainly implied in cell proliferation and tumorigenesis. In addition, MCM3 might play an important role in neuronal apoptosis. However, the functions of MCM3 in central nervous system are still with limited acquaintance. In this study, we performed a traumatic brain injury (TBI) model in adult rats. Western blot and immunohistochemistry staining showed up-regulation of MCM3 in the peritrauma brain cortex. The expression patterns of active caspase-3 and Bax, Bcl-2 were parallel with that of MCM3. Immunofluorescent staining and terminal deoxynucleotidyl transferase-mediated biotinylated-dUTP nick-end labeling suggested that MCM3 was involved in neuronal apoptosis. In conclusion, our data indicated that MCM3 might play an important role in neuronal apoptosis following TBI. Further understanding of these insights could serve as the basis for broadening the therapeutic scope against TBI.

Expression of CDC5L is associated with tumor progression in gliomas.

Cell division cycle 5-like (CDC5L) protein is a cell cycle regulator of the G2/M transition and has been reported to participate in the catalytic step of pre-messenger RNA (mRNA) splicing and DNA damage repair. Recently, it was also found to act as a candidate oncogene in osteosarcoma and cervical tumors. However, the role of CDC5L expression in tumor biology was still unclear. Here, we analyzed the expression and clinical significance of CDC5L in gliomas. The expression of CDC5L in fresh glioma tissues and paraffin-embedded slices was evaluated by western blot and immunohistochemistry, respectively. We found that CDC5L was highly expressed in glioma tissues. The expression of CDC5L was significantly associated with glioma pathology grade and Ki-67 expression. Univariate and multivariate analyses showed that high CDC5L expression was an independent prognostic factor for glioma patients' survival. To determine whether CDC5L could regulate the proliferation of glioma cells, we transfected glioma cells with interfering RNA target CDC5L, then investigated cell proliferation with cell counting kit (CCK)-8, flow cytometry assays and colony formation analyses. Our results indicated that knockdown of CDC5L would inhibit proliferation of glioma cells. Besides, reduced expression of CDC5L could induce the apoptosis of glioma cells. These findings suggested that CDC5L might play an important role in glioma and thus be a promising therapeutic target of glioma.

CHD1L Regulates Cell Cycle, Apoptosis, and Migration in Glioma.

Chromodomain helicase/ATPase DNA binding protein 1-like (CHD1L) gene is a newly identified oncogene located at Chr1q21 and it is amplified in many solid tumors. In this study, we intended to investigate the clinical significance of CHD1L expression in human glioma and its biological function in glioma cells. Western blot and immunohistochemistry analysis showed that CHD1L was overexpressed in glioma tissues and glioma cell lines. In addition, the expression level of CHD1L was positively correlated with glioma pathological grade and Ki-67 expression. Kaplan-Meier curve indicated that high expression of CHD1L may result in poor prognosis of glioma patients. Accordingly, suppression of CHD1L in glioma cells was shown to induce cell cycle arrest and increase apoptosis. In addition, knockdown of CHD1L significantly accelerated migration and invasion ability of glioma cells. Together our findings suggest that CHD1L is involved in the progression of glioma and may be a novel target for further therapy.

Nucleostemin promotes the proliferation of human glioma via Wnt/ß-Catenin pathway.

Nucleostemin, nucleolar guanosine triphosphate (GTP)-binding protein 3, is a member of the MMR1/HSR1 GTP-binding protein family. The important roles of nucleostemin in self-renewal, cell cycle regulation, apoptosis, and cell proliferation of various cancer types as been shown. Nevertheless, its expression and potential functions in human glioma is still unclear. In the present study, we demonstrated that up-regulation of nucleostemin was tightly related to poor 5-year-survival ratios. In serum-starved and re-feeding models of U251 and U373MG, we observed the rising expression of nucleostemin and p-ß-Catenin (p-Tyr645) were accompanied with cell proliferation markers (cyclin D1 and proliferating cell nuclear antigen (PCNA)). Employing nucleostemin-depletion models, we found down-regulated nucleostemin and p-ß-Catenin. The flow cytometry analysis proved the weakened cell proliferation. Moreover, we detected the translocation of ß-Catenin into the nucleus was impaired, meaning the inhibition of the Wnt/ß-Catenin pathway. Taken together, we identified a positive correlation between up-regulation of nucleostemin and human glioma cell proliferation and that knocking-down nucleostemin alleviated glioma proliferation by reducing ß-Catenin transportation into the nucleus. All results suggested that nucleostemin might accelerate human glioma proliferation via the Wnt/ß-Catenin pathway.

Overexpression of DSPP promotes the proliferation and migration of astrocytes.

Astrocytes are activated after central nervous system (CNS) injury, such as spinal cord injury (SCI). Activated astrocytes can form glial scar to block nerve regeneration. Dentin sialophosphoprotein (DSPP), a member of the SIBLING (Small integrin-binding ligand N-linked glycoproteins) family, has been reported to contribute to the proliferation and migration of different types of tumor cells, including glioma. However, the functions of DSPP in reactive astrocytes after CNS injury remain unknown. In this study, starvation-serum stimulation model in astrocytes was conducted to explore this issue. Our results showed that DSPP expression was increased in reactive astrocytes comparing to normal ones. Meanwhile, up-regulation of DSPP was accompanied with PCNA and GFAP. To explore the role of DSPP in astrocytes, we overexpressed DSPP with recombinant GFP-DSPP plasmid and the results showed that overexpression of DSPP could promote the proliferation and migration of the cells, the important characteristics of reactive astrocytes. In addition, overexpression of DSPP obviously increased the activation of Akt/mTOR pathway in astrocytes. Taken together, we demonstrated that DSPP may play a key role in the proliferation and migration of astrocytes, suggesting that targeting DSPP might be a promising therapeutic strategy for treating CNS injury which characterized by glia scar formation.

Knockdown of DSPP inhibits the migration and invasion of glioma cells.

Dentin sialophosphoprotein (DSPP) is a member of the SIBLING (Small integrin-binding ligand N-linked glycoproteins) family of phosphoglycoproteins and has been proved to contribute to the migration of a variety of solid tumor cells. However, whether DSPP participates in the pathogenic process of glioma remains unknown. In this study, we aimed to investigate the expression and biological function of DSPP in human glioma cells. We demonstrated through Western blot that DSPP is overexpressed in glioma tissues comparing to normal brain tissues. To investigate the role of DSPP in glioma carcinogenesis, we reduced the DSPP expression by small interfering RNA (siRNA) and found that DSPP silencing significantly inhibited the migration and invasion of glioma cells, the critical characteristics of glioma. Furthermore, we showed that DSPP down-regulation significantly decreased the activation of the AKT/mTOR/p70S6K pathway in glioma cells. Taken together, these findings indicate that knockdown of DSPP inhibits glioma cells migration and invasion, suggesting that targeting DSPP might be a potentially effective therapeutic strategy for treating glioma.

High expression of adenylate cyclase-associated protein 1 accelerates the proliferation, migration and invasion of neural glioma cells.

Adenylate cyclase-associated protein 1 (CAP1), a conserved member of cyclase-associated proteins was reported to be associated with the proliferation, migration or invasion of the tumors of pancreas, breast and liver, and was involved in astrocyte proliferation after acute Traumatic Brain Injury (TBI). In this study, we sought to investigate the character of CAP1 in the pathological process of human glioma by detecting human glioma specimens and cell lines. 43 of 100 specimens showed high expression of CAP1 via immunohistochemistry. With statistics analysis, we found out the expression level of CAP1 was correlated with the WHO grades of human glioma and was great positively related to Ki-67 (p<0.01). In vitro, silencing CAP1 in U251 and U87MG, the glioma cell lines with the relatively higher expression of CAP1, induced the proliferation of the cells significantly retarded, migration and invasion as well. Obviously, our results indicated that CAP1 participated in the molecular pathological process of glioma indeed, and in a certain sense, CAP1 might be a potential and promising molecular target for glioma diagnosis and therapies in the future.

Expression and clinical role of RBQ3 in gliomas.

RBQ3, also known as RBBP5 (RB-binding protein 5), was an RB-binding protein. Besides, it was one of core components of MLL1 (mixed lineage leukemia 1), which were required for H3K4 methyltransferase activity. MLL1 dysfunction was found to be associated with the progression of some cancers such as acute leukemias. However, the precise role of RBQ3 in tumor progression remains obscure. In this study, we explored the expression and clinical role of RBQ3 in gliomas. Our results showed that RBQ3 was significantly upregulated in clinical glioma specimens by Western blot and immunohistochemistry. Moreover, its level was significantly associated with the pathology grades. High RBQ3 expression was suggested to be an independent prognostic factor for glioma patients' survival by univariate and multivariate analyses. Serum starvation and refeeding assay indicated that the expression of RBQ3 increased 8h after serum-stimulation, together with percentage of cells at S phase. In addition, knockdown of RBQ inhibited U87-MG cell proliferation with CCK8 kit, flow cytometry assays and colony formation analyses; while the depletion of RBQ3 induced the apoptosis of U87-MG cells. All the findings suggested that RBQ3 might play an important role in glioma, and RBQ3 inhibitors might be novel anti-tumor agents.

Overexpression of CCT8 and its significance for tumor cell proliferation, migration and invasion in glioma.

Overexpression of chaperonin containing t-complex polypeptide 1 (TCP1), or CCT, has been reported in various classes of malignancies. However, little is known about the expression of t-complex protein subunits TCP1theta (CCT8) in gliomas. In this study, the expression of CCT8 protein was detected using blotting analysis and immunohistochemistry. CCT8 was found to be overexpressed in gliomas and to correlate with the WHO grade of gliomas. To further investigate the biological function of CCT8 in gliomas, CCT8-silenced U87 glioblastoma multiforme (GBM) and U251MG cells were constructed using a small interference RNA (siRNA) sequence. The knockdown effect of CCT8 on proliferation and invasion in these cells was analyzed using the CCK8, flow cytometry cycle, scratch, transwell invasion and fluorescence assays. Compared with the controls, the glioma cells expressing CCT8-siRNA exhibited a significantly decreased proliferation and invasion capacity, as well as a dysregulated cell cytoskeleton. This study showed that high CCT8 protein expression might be related to poor outcome of glioma, and that CCT8 regulates the proliferation and invasion of glioblastomas.

The role of FoxJ2 in the migration of human glioma cells.

Previous studies have demonstrated that FoxJ2 (forkhead box J2) is a member of Forkhead Box transcription factors and acts as an important prognostic indicator in human breast cancer. Our study aimed to assess the expression and function in human glioma. Western blot analysis and immunohistochemistry were performed in human glioma tissues. Low FoxJ2 expression was observed in 80 samples and its level was correlated with the grade of malignancy. A strongly positive correlation was observed between FoxJ2 and E-cadherin. Overexpression of FoxJ2 increased E-cadherin expression and decreased vimentin expression. The wound healing and transwell assays showed that overexpression of FoxJ2 significantly inhibited their migration in U87 cells. Consistent with this, knockdown of FoxJ2 promoted cellular motility. In a word, FoxJ2 suppressed cell migration and invasion in glioma, which might be a potential novel molecular targeted therapy for surgery and immune treatment.

Knockdown of PFTK1 Inhibits the Migration of Glioma Cells.

The prognosis of glioma patients is generally poor, so it is urgent to find out the underlying molecular mechanisms. PFTK1 is a member of cyclin-dependent kinases (Cdks) family and has been reported to contribute to tumor migration and invasion. In this study, we aimed to explore the expression and function in human glioma. Western blot and immunohistochemistry were used to evaluate the expression of PFTK1. PFTK1 expression was higher in glioma tissues compared with normal brain tissues, and its level was associated with the WHO grade in Western blot analysis. The suppression of PFTK1 expression by RNA interference was shown to inhibit the migration of glioma cells. Knockdown of PFTK1 increases E-cadherin expression and decreases vimentin expression. These data show that PFTK1 may participate in the pathogenic process of glioma, suggesting that PFTK1 can become a potential therapeutic strategy for gastric cancer.

Knocking down the expression of SYF2 inhibits the proliferation of glioma cells.

SYF2 is thought to be a cell cycle regulator at the G1/S transition, which encodes a nuclear protein that interacts with cyclin D-type binding-protein 1. In the present study, we investigated the role of SYF2 in human glioma progression. Immunohistochemical and Western blot analyses were performed in human glioma tissues. High SYF2 expression (located in cell nuclei) was observed in 80 samples, and its level was correlated with the grade of malignancy. A strongly positive correlation was observed between SYF2 and Ki-67 expression (P < 0.01). More importantly, high expression of SYF2 was associated with a poor outcome. In vitro, after the release of U87 cell lines from serum starvation, the expression of SYF2 was upregulated, as well as PCNA and cyclin D1. In addition, knockdown of SYF2 by small interfering RNA transfection diminished the expression of PCNA, cyclin D1 and arrested cell growth at G1 phase. These results indicate that SYF2 in glioma is essential for cell proliferation; thus, targeting SYF2 or its downstream targets may lead to novel therapies for glioblastomas.