Clinical significance of 4-1BBL protein expression in human glioma tissues

AIM: To investigate the expression of 4-1BBL(CD137L) protein in human glioma tissues, and to analyze its clinical significance. METHODS: The expression levels of 4-1BBL protein in 82 human glioma specimens were measured by the method of immunohistochemistry. The correlation of 4-1BBL protein level with histopathologic features and survival time was analyzed. RESULTS: 4-1BBL protein positive cells were observed in human brain astrocytic tumors, which were not found in meningioma and normal brain tissues. In our study, the expression of 4-1BBL protein in 3 cases of normal brain tissues was negative. The expression of 4-1BBL protein in 31 cases of glioma specimens were positive, and total positive rate was 37.8% (31/82). The expression of 4-1BBL protein was related to the survival time and age of the patients, but not related to the pathological grade of glioma. Statistically, the patients with positive expression of 4-1BBL protein had longer survival time (P<0.05). CONCLUSION: Expression of 4-1BBL protein may have reference value to predict the prognosis of patients with glioma.     

Research progress of glioma cell origin

Glioma, a common primary brain tumor, is considered to be incurable, and easy to relapse. The median survival of the patients with glioma after treatment is only 15 to 19 months. The high heterogeneity of glioma is the main reason leading to poor clinical treatment, and the underlying mechanism is closely related to the cell origin of glioma. Therefore, to elucidate the cell origin of glioma is important to further study the mechanism of tumor formation and develop effective treatment programs. For a long time, studies have attempted to speculate on the cell origin of glioma based on the morphological characteristics, but agreements still haven't been reached. This review focuses on the most probable origin cells, such as neural stem cells, astrocytes, oligodendrocyte progenitor cells and neurons.     

Effect of Vaccinium vitis procyanidin on regulation of glioma cell growth

AIM: To explore the effect of Vaccinium vitis procyanidin on the growth of glioma cells. METHODS: Glioma C6 cells were cultured and divided into control and 10, 20 and 40 μg/L Vaccinium vitis procyanidin groups. The influence of Vaccinium vitis procyanidin on the growth of C6 cells was measured by MTT assay and the observation under inverted microscope. The apoptotic rate was detected by Annexin V/PI staining. The protein expression of Bcl-2 and Bax was determined by immunocytochemistry. The protein levels of Bcl-2, Bax and caspase-3 were also examined by Western blotting. RESULTS: The growth of C6 glioma cells was inhibited by Vaccinium vitis procyanidin at concentrations of 10, 20 and 40 μg/L. The growth was significantly inhibited in 40 μg/L Vaccinium vitis procyanidin group at 24 h and 48 h, and in 20 and 40 μg/L Vaccinium vitis procyanidin groups at 72 h (P<0.01). The density of the cells was decreased when the concentration of Vaccinium vitis procyanidin increased. The apoptotic rate was increased when the concentration of Vaccinium vitis procyanidin increased either. The expression of Bcl-2 was decreased and Bax was increased after 10, 20 and 40 μg/L Vaccinium vitis procyanidin treatments. The ratio of Bax/Bcl-2 was increased when the dose of Vaccinium vitis procyanidin increased (P<0.05 or P<0.01). The expression of Bcl-2 was decreased (P<0.01), and Bax and caspase-3 were increased after 10, 20 and 40 μg/L Vaccinium vitis procyanidin treatments. The ratio of Bax/Bcl-2 was increased when the dose of Vaccinium vitis procyanidin increased (P<0.01). CONCLUSION: Vaccinium vitis procyanidin inhibits the growth of glioma cells by down-regulating Bcl-2 protein and up-regulating Bax protein to activate caspase-3, thus inducing apoptosis.     

Expression of B7-H4 in human glioma tissues and its clinical significance

AIM: To investigate the expression of B7-H4 in human glioma tissues and its clinical significance. METHODS: The histological staging of 150 cases of human glioma tissues was determined by HE staining. Immunohistochemistry was performed to study the protein level of B7-H4 in 150 human glioma specimens. Furthermore, the relationships between the expression level of B7-H4 and clinicopathological parameters, as well as patients survival rate, were analyzed. RESULTS: HE staining result indicated that there were 12 cases staged as stage I, 50 cases stage II, 39 cases stage III and 49 cases stage IV in 150 glioma specimens. Ninety-seven cases highly expressed B7-H4 in total 150 glioma samples with a 64.7% high expression rate. The histological grade of the tissues with high B7-H4 expression was mostly III~IV. There were 53 cases with low B7-H4 expression in the total 150 glioma patients with a 35.3% low expression rate. The histological grade of the tissues with low B7-H4 expression was mostly I~II. The B7-H4 expression was related to the age of the patients (P<0.01) and the pathological grade of glioma (P<0.01), but not related to the location of glioma (P>0.05). Kaplan-Meier survival curves demonstrated that increased B7-H4 expression was associated with shorter overall survival time (P<0.01). Multivariate Cox regression analysis revealed that B7-H4, age, sex and pathological grade were independent prognostic factors in glioma patients. CONCLUSION: B7-H4 is expressed in most of glioma tissues. B7-H4 may be a novel prognostic biomarker and a new target of molecular therapy for gliomas.     

Transferrin-bound Yb2 uptake by U-87 MG cells and effect of Yb on proliferation of the cells

AIM:To investigate the role of transferrin/transferrin receptor system in transferrin-bound Yb₂ (Yb₂Tf) uptake by U-87 MG cells and the effect of transferrin-bound and -free Yb₂ on proliferation of U-87 MG cells.METHODS:Cell culture and ICP-MS measurement of Yb₂.RESULTS:Yb₂Tf uptake by U-87 MG cells increased with the concentrations of Yb₂Tf, and reached saturation as the concentration in the incubation medium was raised to about 2 μmol/L. Also, Yb₂ uptake by the cells increased with increase of the mole ratio (Yb₂: apoTf), reaching a maximum at 1.5 mole ratio. Yb₂Tf in 0.4 μmol/L significantly inhibited proliferation of U-87 MG cells, however, 10 μmol/L Yb³⁺ had no significant effect on proliferation of the cells.CONCLUSION:The uptake of Yb₂ by U-87 MG cells might be mediated by transferrin/transferrin receptor system. Transferrin-bound but not transferrin-free Yb₂ could significantly inhibit proliferation of U-87 MG cells.     

Inhibitory effect of different target-side antisense oligonucleotides on bFGF expression in SWO-38 cells

AIM:Three different antisense oligonucleotides complementary to basic fibroblast growth factor (bFGF) mRNA were compared in inhibitory effect on gene targeted expression.METHODS:After transfecting bFGF antisense oligonucleotides (asODN) into SWO-38 cells by lipofectin, the proliferation of cells was identified by MTT method, apoptosis was examined by flow cytometric cell cycle analysis and the expression levers of bFGF were detected by Western-blotting.RESULTS:There were 49%, 33%, 51% inhibition of cell growth and 35%, 27%, 18% cell apoptosis after asODN1, asODN2 and asODN3 treatment.In addition, the decrease in bFGF protein was 63%, 42%, 11%, respectively.CONCLUSION:The data suggeste that asODN1 is a potent target to bFGF mRNA, which inhibits cell growth and induces apoptosis in SWO-38 cells.     

Effect of VEGF over-expressed C6 glioma cells on the expression of Flk-1and Flt-1in cocultured microvascular endothelial cells

AIM: To study the effects of VEGF over-expressed C6 glioma cells on the expression of Flk-1and Flt-1in cocultured microvascular endothelial cells using the rat hepatic cells BRL 3A as control. METHODS: Cocultured systems of rat pulmonary microvascular endothelial cells with C6 and endothelial cells with BRL 3A were established. Immunocytochemical method was used to investigate the expression change of Flk-1and Flt-1protein in cocultured microvascular endothelial cells. The expression of Flt-1and Flk-1mRNA was analyzed by RT-PCR and Northern blot. RESULTS: The microvascular endothelial cells cocultured with C6 showed increased expression of Flk-1and Flt-1protein (P <0.05), while that cocultured with BRL 3A, showed decreased expression of Flt-1and Flk-1protein(P <0.01). The results of RT-PCR and Northern blot showed that the Flk-1, Flt-1mRNA of microvascular endothelial cells were markedly up-regulated after coculture with C6 glioma cells (P <0.01), while the endothelial cells cocultured with BRL 3A had down-regulated expression of Flk-1, Flt-1mRNA (P <0.01). CONCLUSION: VEGF over-expressed C6 glioma cells may markedly up-regulate the expression of Flt-1and Flk-1in cocultured microvascular endothelial cells. These results suggest that this effect could be one of the important mechanisms in glioma angiogenesis in vivo.     

Experimental study of antitumor immunity of DC/C6 fusion vaccine

AIM:To study the anti-tumor effect and mechanism of fusion vaccine on DCs and C6 glioma cells. METHODS:PEG was used to fuse DCs with C6 glioma cells. Immunofluorescence with GFAP-FITC was used to identify the DC/C6 fusion cells. Rat brain glioma models were made by stereotactic technique. After 5 days of inoculation of C6, 107 fusion cells were injected through tail vein in group A. The same number of DCs and the same volume of PBS were used in group B and group C. The survival time of rats in these three groups was analyzed by Log-rank survival analysis. Tumor samples were checked by HE staining and immunohistochemical staining with CD8Mcab. RESULTS:Positive result of GFAP-FITC immunofluorescence was observed in DC/C6 fusion cells. The Log-rank survival analysis showed that statistically significant difference in group A was observed compared to that in group B and group C (P<0.01). Tumor sample stained with HE showed that many inflammatory cells infiltrated in tumor tissues. The result of immunohistochemical staining with CD8Mcab was positive. CONCLUSION:DC/C6 fusion cells had the ability of antigen presentation and activating T lymphocytes effectively. CD8⁺T lymphocytes play an important role in antitumor immunity.     

Effects of lentivirus-mediated RWDD3 silencing on proliferation and invasion of human glioma U251 cells

AIM: To investigate the effect of RWDD3 gene silencing on the biological characteristics of human glioma U251 cells.METHODS: A lentiviral vector expressing RWDD3 shRNA was constructed and transfeeted into the U251 cells. The expression of RWDD3 at mRNA and protein levels was detected by real-time PCR and Western blot, respectively. The cell activity was determined by MTT assay. The colony formation ability was detected by the colony formation assay. The cell proliferation ability was detected by BrdU incorporation assay. The cell invasion and migration were evaluated by Transwell assay. Flow cytometry was used to monitor the changes of cell cycle distribution and apoptosis.RESULTS: Recombinant lentivirus was successfully transfected into U251 cells. Compared with the cells transfected with the scrambled shRNA and control cells, the cell activity, colony formation ability, and the invasive and migratory activities were inhibited, the cell cycle was arrested in G₀/G₁ phase, and the apoptosis was increased in the U251 cells transfected with RWDD3 shRNA(P<0.05).CONCLUSION: RWDD3 plays a vital role in proliferation and invasion of glioma cells. It may serve as a potential target of gene therapy for glioma.