Clinical characteristics and outcomes of primary intracranial alveolar soft-part sarcoma: A case report.

BACKGROUND: Primary intracranial alveolar soft-part sarcoma (PIASPS) is a rare malignancy. We aimed to investigate the clinical profiles and outcomes for PIASPS. CASE SUMMARY: We firstly reported five consecutive cases from our institute. Then, the cases from previous studies were pooled and analyzed to delineate the characteristics of this disease. Our cohort included two males and three females. The median age was 21-years-old (range: 8-54-years-old). All the patients received surgical treatment. Gross total resection (GTR), radiotherapy, and chemotherapy were administered in 3 patients, 4 patients, and 1 patient, respectively. After a median follow-up of 36 mo, tumor progression was noticed in 4 patients; and 3 patients died of the disease. Pooled data (n = 14) contained 5 males and 9 females with a median age of 19 years. The log-rank tests showed that GTR (P = 0.011) could prolong progression-free survival, and radiotherapy (P < 0.001) resulted in longer overall survival. CONCLUSION: Patients with PIASPS suffer from poor outcomes. Surgical treatment is the first choice, and GTR should be achieved when the tumor is feasible. Patients with PIASPS benefit from radiotherapy, which should be considered as a part of treatment therapies.

Long-term outcome in meningiomas involving the major dural sinuses with combined therapy of subtotal resection and early postoperative gamma knife radiosurgery.

INTRODUCTION: Total resection of meningiomas involving the major dural sinuses (MIMDS) is still challenging for neurosurgeons. Gamma knife radiosurgery (GKRS) was shown to have a high probability of tumor control. The current study evaluated the clinical outcomes of patients who underwent subtotal resection alone or in combination with postoperative GKRS for the treatment of WHO grade I MIMDS. METHODS: From January 2006 to December 2016, 204 patients with MIMDS underwent Simpson IV subtotal resection in Wuhan Union Hospital. In 151 patients, no additional treatment was performed, while the tumor remnant was treated with GKRS in 53 patients. All patients were monitored with regular MR follow-ups. We retrospectively reviewed the clinical data, radiological characteristics, and outcomes of these 204 patients. Progression-free survival (PFS) was determined by Kaplan-Meier analysis. Related factors were determined by univariate Cox regression analyses. RESULTS: The mean follow-up period was 75.5 months. The tumor recurrence/progression rates were 13.9% in the microsurgery group and 3.8% in the combined therapy group (p = 0.045). The 5- and 10- year progression-free survival (PFS) rates were 92.3 and 80.7%, respectively, in the microsurgery group and 100.0 and 88.5% in the combined therapy group. Treatment approach was found to be an independent prognostic factor for tumor recurrence/progression in the univariable analyses (p=0.04). CONCLUSIONS: Compared with microsurgery alone, targeted Simpson grade IV resection combined with early gamma knife treatment resulted in longer progression-free survival without increased complications for WHO grade I MIMDS.

Using arterial-venous analysis to characterize cancer metabolic consumption in patients.

Understanding tumor metabolism holds the promise of new insights into cancer biology, diagnosis and treatment. To assess human cancer metabolism, here we report a method to collect intra-operative samples of blood from an artery directly upstream and a vein directly downstream of a brain tumor, as well as samples from dorsal pedal veins of the same patients. After performing targeted metabolomic analysis, we characterize the metabolites consumed and produced by gliomas in vivo by comparing the arterial supply and venous drainage. N-acetylornithine, D-glucose, putrescine, and L-acetylcarnitine are consumed in relatively large amounts by gliomas. Conversely, L-glutamine, agmatine, and uridine 5-monophosphate are produced in relatively large amounts by gliomas. Further we verify that D-2-hydroxyglutarate (D-2HG) is high in venous plasma from patients with isocitrate dehydrogenases1 (IDH1) mutations. Through these paired comparisons, we can exclude the interpatient variation that is present in plasma samples usually taken from the cubital vein.

Inhibition of tumor growth and angiogenesis by 2-(4-aminophenyl) benzothiazole in orthotopicglioma C6 rat model.

In the present study antitumor effect of 2-(4-aminophenyl) benzothiazole (BTZ) was evaluated against human U251 and rat C6 glioma cell lines using MTT assay. It was observed that BTZ exhibited significant antitumor effect with IC50 of 3.5 and 4 µM against human U251 and rat C6 glioma cells respectively. To gain in-depth insights about the antitumor effect of BTZ, glioma xenograft rat model was prepared. The rats were treated with 10 mg and 15 mg/kg body weight doses of BTZ daily for 21 days after C6 cell administration. Treatment of the rats with BTZ reduced the tumor volume to 12% compared to 100% in the untreated rats. TUNEL assay showed a remarkable increase in the proportion of apoptotic cells in the BTZ treated rats than those in the untreated rats. The increase in the population of apoptotic cells was 23-fold compared to control. Immuno-histological staining revealed marked reduction (16%) in the proportion of CD31-stained vessels in the BTZ treated rats than those of the untreated rats. These changes were accompanied with decreased transcript levels of vascular endothelial growth factor (VEGF) and the VEGF receptor Flt1 as well as ERK1/2 and matrix metalloproteinase-2 (MMP2). Moreover, BTZ altered the expression of several cell cycle control proteins. While as pRb protein expression decreased, E2F1 remained unaltered and cyclin D1 protein and p53 expression was enhanced. Taken together, the results indicate that BTZ is a potent inhibitor of glioma cell proliferation in vivo and exerts its effects on cell cycle control and angiogenesis related proteins.

Classification and Protection of Peritumoral Draining Veins of Parasagittal and Falcine Meningiomas.

OBJECTIVE: To investigate the importance and types of peritumoral draining veins in parasagittal and falcine meningiomas and administer corresponding protective strategies during surgery according to these different types to improve tumor resection rate and maximize the protection of neurologic functions. METHODS: The clinical information of 156 patients with parasagittal and falcine meningiomas who were admitted at the Neurosurgery Department of our hospital was collected and retrospectively analyzed. All patients underwent pathologic classification, magnetic resonance imaging scanning and enhancement, and magnetic resonance venography examinations. RESULTS: Among these patients, 113 (72.4%) had Simpson grade I and II resection, whereas 43 patients (27.6%) had Simpson grade III and IV resection and underwent postoperative adjuvant gamma knife surgery. Karnofsky Performance Status evaluation was carried out at 1 week after surgery. In total, 69 patients (44.3%) improved, 66 patients (42.3%) had no changes, and 21 patients (13.4%) had worsened conditions (7 patients had hemiplegia, 5 patients had aphasia, 4 patients had decreased vision, and 5 patients had ataxia). There were no deaths. According to the 2016 World Health Organization pathologic grading, 131 patients (84%) were grade I, 22 patients (14%) were grade II, and 3 patients were grade III (2%). Furthermore, 105 patients were followed up for 1-4 years. There were 11 cases of recurrence. CONCLUSIONS: The classification and evaluation of peritumoral draining veins through preoperative-combined magnetic resonance venography can be used as a guide in determining the corresponding protective strategy during surgery. This can significantly improve the tumor resection rate and decrease the postoperative disability rate.

Effect of microRNA-128 on cisplatin resistance of glioma SHG-44 cells by targeting JAG1.

This current study intends to investigate the effect of microRNA-128 (miR-128) on cisplatin (DDP) resistance in glioma SHG-44 cells. SHG-44/DDP cells were transfected with miR-128 antisense oligonucleotide (ASO) and assigned into blank, resistance, NC, anti-miR-128, miR-128 mimic, si-JAG1, and anti-miR-128 + si-JAG1 groups. qRT-PCR and Western blotting were employed for determining expression of miR-128, JAG1, Bax and Bcl-2. MTT assay, Giemsa staining, and flow cytometry were applied to detect DDP resistance, cellular morphology, and cell cycle, respectively. JAG1 is targeted and negatively regulated by miR-128. In in vitro experiments, compared with the blank group, the rest groups exhibited declined miR-28 and Bax expression, lowered cell inhibition rate and apoptosis rate, but elevated JAG1 and Bcl-2 expression with cells arrested in the S phase. Compared with the resistance group, the anti-miR-128 group showed decreasedBax expression along with a lowered cell inhibition rate and apoptosis rate, but increased JAG1 and Bcl-2 expression with reduced cells arrested in the S phase; while the miR-128 mimic group showed an opposite trend; the si-JAG1 group showed decreased Bcl-2 expression and reduced cells in the S phase. In in vivo experiments, compared with the resistance group, the tumor growth rate, tumor volume, and weight as well as JAG1 expression accelerated in the anti-miR-128 group; whereas the miR-128 mimic and si-JAG1 groups exhibited an opposite trend. Our findings demonstrated that miR-128 ASO transfection might down-regulate the expression of miR-128 in SHG-44/DDP and up-regulate the DDP resistance in SHG-44/DDP cells, providing a potential treatment target for glioma.

Inhibitor of Nicotinamide Phosphoribosyltransferase Sensitizes Glioblastoma Cells to Temozolomide via Activating ROS/JNK Signaling Pathway.

Overcoming temozolomide (TMZ) resistance is a great challenge in glioblastoma (GBM) treatment. Nicotinamide phosphoribosyltransferase (NAMPT) is a rate-limiting enzyme in the biosynthesis of nicotinamide adenine dinucleotide and has a crucial role in cancer cell metabolism. In this study, we investigated whether FK866 and CHS828, two specific NAMPT inhibitors, could sensitize GBM cells to TMZ. Low doses of FK866 and CHS828 (5 nM and 10 nM, resp.) alone did not significantly decrease cell viability in U251-MG and T98 GBM cells. However, they significantly increased the antitumor action of TMZ in these cells. In U251-MG cells, administration of NAMPT inhibitors increased the TMZ (100 µM)-induced apoptosis and LDH release from GBM cells. NAMPT inhibitors remarkably enhanced the activities of caspase-1, caspase-3, and caspase-9. Moreover, NAMPT inhibitors increased reactive oxygen species (ROS) production and superoxide anion level but reduced the SOD activity and total antioxidative capacity in GBM cells. Treatment of NAMPT inhibitors increased phosphorylation of c-Jun and JNK. Administration of JNK inhibitor SP600125 or ROS scavenger tocopherol with TMZ and NAMPT inhibitors substantially attenuated the sensitization of NAMPT inhibitor on TMZ antitumor action. Our data indicate a potential value of NAMPT inhibitors in combined use with TMZ for GBM treatment.

SIRT6 suppresses glioma cell growth via induction of apoptosis, inhibition of oxidative stress and suppression of JAK2/STAT3 signaling pathway activation.

Sirtuin 6 (SIRT6) is a member of the mammalian NAD+­dependent deacetylase sirtuin family that acts to maintain genomic stability and to repress genes. SIRT6 has recently been reported to be a tumor suppressor that controls cancer metabolism, although this effect of SIRT6 is still in dispute. Moreover, the role of SIRT6 in glioma is largely unknown. In the present study, we found that overexpression of SIRT6 using an adenovirus inhibited glioma cell growth and induced marked cell injury in two glioma cell lines (U87­MG and T98G). Fluorescent terminal deoxyribonucleotidyl transferase (TdT)­mediated biotin­16­dUTP nick­end labelling (TUNEL) assay showed that SIRT6 overexpression induced obvious apoptosis in the T98G glioma cells. Immunoblotting and immunofluorescent staining demonstrated that SIRT6 overexpression promoted the mitochondrial-to­nuclear translocation of apoptosis­inducing factor (AIF), a potent apoptosis inducer. Moreover, we found that SIRT6 overexpression largely reduced oxidative stress and suppressed the activation of the JAK2/STAT3 signaling pathway in glioma cells. Finally, we showed that SIRT6 mRNA and protein levels in human glioblastoma multiforme tissues were significantly lower than the levels in peritumor tissues. In summary, our data suggest that SIRT6 suppresses glioma cell growth via induction of apoptosis, inhibition of oxidative stress and inhibition of the activation of the JAK2/STAT3 signaling pathway. These results indicate that SIRT6 may be a promising therapeutic target for glioma treatment.

Efficacy of single-layer continuous suture of the posterior wall in anastomosis involving a difficult location of the digestive tract.

Surgery for digestive tract disease predominantly consists of reconstruction and anastomosis. Due to the difficult location, anastomosis is extremely challenging and the risk of complication increases accordingly. Traditional manual anastomosis and the application of a stapling device are insufficient. Therefore, the aim of this study was to investigate the feasibility and safety of a novel manual method in a difficult anastomotic location, consisting of a single-layer continuous suture in the posterior wall. In total, 15 beagle dogs were included in the study; eight underwent surgery with the novel manual method for reconstruction and anastomosis of the digestive tract, while seven underwent surgery with the stapler device as a control. The subsequent postoperative complications were observed and, three months later, the anastomotic ports were excised, and the pathological formation and morphological changes were evaluated. No statistically significant differences were identified between the total (50.0 vs. 57.1%; P=0.782) and anastomotic (0.0 vs. 28.6%; P=0.200) complication rates in the manual suture and staple suture groups, respectively. Compared with the control group, the operative expenditure was lower in the manual group (1726.7±33.5 vs. 2135.7±43.1 renminbi; P=0.001), the diameter of the anastomotic port was larger in the manual group (3.04±0.07 vs. 2.24±0.25 cm; P=0.004) and the thickness of the anastomotic port (in cm) was thinner in the manual group (2.94±0.06 vs. 5.07±0.85; P=0.002). Furthermore, the pathological formation of the anastomositic port in the manual group was improved. The results of the current study suggest single-layer continuous suture of the posterior wall in anastomosis of the digestive tract to be a novel method with feasibility and safety, particularly in difficult anastomotic locations.

Dendritic cell-based vaccine for the treatment of malignant glioma: a systematic review.

OBJECTIVE: Glioblastoma multiforme (GBM) has a poor prognosis. The purpose of this systematic review and meta-analysis was to analyze the outcomes of clinical trials which compared immunotherapy with conventional therapy for the treatment of malignant gliomas. METHODS: PubMed, Cochrane and Google Scholar databases were searched for relevant studies. The 2-year survival rate was used to evaluate effectiveness of immunotherapy. RESULTS: Of 171 studies identified, six comparative trials were included in the systematic review. Immunotherapy was associated with a significantly longer OS and 2-year survival compared to conventional therapy. CONCLUSION: Immunotherapy may improve the survival of patients with GBM.

Human IP10-scFv and DC-induced CTL synergistically inhibit the growth of glioma in a xenograft model.

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed in glioma cells, and the EGFRvIII-specific dendritic cell (DC)-induced tumor antigen-specific CD8(+) cytotoxic T lymphocytes (CTLs) may hold promise in cancer immunotherapy. Interferon (IFN)-γ-inducible protein (IP)-10 (IP-10) is a potent inhibitor of angiogenesis and can recruit CXCR3(+) T cells, including CD8(+) T cells, which are important for the control of tumor growth. In this study, we assessed if the combination of IP10-EGFRvIIIscFv with DC-induced CTLs would improve the therapeutic antitumor efficacy. IP10-scFv was generated by linking the human IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv with a (Gly4Ser)3 flexible linker, purified by affinity chromatography, and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. DCs were isolated from human peripheral blood monocyte cells and pulsed with EGFRvIII-peptide, then co-cultured with autologous CD8(+) T cells. BALB/c-nu mice were inoculated with human glioma U87-EGFRvIII cells in the brain and treated intracranially with IP10-scFv and/or intravenously with DC-induced CTLs for evaluating the therapeutic effect. Treatment with both IP10-scFv and EGFRvIII peptide-pulsed, DC-induced CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, which was accompanied by the inhibition of tumor angiogenesis and enhancement of cytotoxicity, thereby increasing the numbers of brain-infiltrating lymphocytes (BILs) and prolonging the residence time of CTLs in the tumor.

Single-layer continuous suture contributes to the reduction of surgical complications in digestive tract anastomosis involving special anatomical locations.

The key point of digestive cancer surgery is reconstruction and anastomosis of the digestive tract. Traditional anastomoses involve double-layer interrupted suturing, manually or using a surgical stapler. In special anatomical locations, however, suturing may become increasingly difficult and the complication rate increases accordingly. In this study, we aimed to investigate the feasibility and safety of a new manual suturing method, the single-layer continuous suture in the posterior wall of the anastomosis. Between January, 2007 and August, 2012, 101 patients with digestive cancer underwent surgery in Xi'an Gaoxin Hospital. Of those patients, 27 underwent surgery with the new manual method and the remaining 74 underwent surgery using traditional methods of anastomosis of the digestive tract. Surgical time, intraoperative blood loss, drainage duration, complications, blood tests, postoperative quality of life (QOL) and overall expenditure were recorded and analyzed. No significant differences were observed in surgical time, intraoperative blood loss, temperature, blood tests and postoperative QOL between the two groups. However, compared with the control group, the new manual suture group exhibited a lower surgical complication rate (7.40 vs. 31.08%; P=0.018), lower blood transfusion volume (274.07±419.33 vs. 646.67±1,146.06 ml; P=0.053), shorter postoperative hospital stay (14.60±4.19 vs. 17.60±6.29 days; P=0.038) and lower overall expenditure (3,509.85±768.68 vs. 6,141.83±308.90 renminbi; P=0.001). Our results suggested that single-layer continuous suturing for the anastomosis of the digestive tract is feasible and safe and may contribute to the reduction of surgical complications and overall expenditure.

A novel recombinant protein of IP10-EGFRvIIIscFv and CD8(+) cytotoxic T lymphocytes synergistically inhibits the growth of implanted glioma in mice.

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed on glioma cells and has been thought to be an excellent target molecule for immunotherapy. IP-10 is a potent chemokine and can recruit CXCR3(+) T cells, including CD8(+) T cells that are important for the control of tumor growth. This study is aimed at investigating the therapeutic efficacy of a novel fusion protein of IP10-EGFRvIIIscFv (IP10-scFv) in combination with glioma lysate-pulsed DCs-activated CD8(+) cytotoxic T lymphocytes (CTLs) in a mouse model of glioma. A plasmid of pET-IP10-scFv was generated by linking mouse IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv, a (Gly4Ser)3 flexible linker and a His-tag. The recombinant IP10-scFv in E. coli was purified by affinity chromatography and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. C57BL/6 mice were inoculated with mouse glioma GL261 cells in the brain and treated intracranially with IP10-scFv and/or intravenously with CTL for evaluating the therapeutic effect. The glioma-specific immune responses were examined. The IP10-scFv retained anti-EGFRvIII immunoreactivity and IP-10-like chemotactic activity. Treatment with both IP10-scFv and CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, accompanied by increasing the numbers of brain-infiltrating lymphocytes (BILs) and the frequency of CXCR3(+)CD8(+) T cells, enhancing glioma-specific IFN-γ responses and cytotoxicity, and promoting glioma cell apoptosis in mice. Our novel data indicate that IP10-scFv and CTL have synergistic therapeutic effects on inhibiting the growth of mouse glioma in vivo.

Vagoglossopharyngeal neuralgia treated by microvascular decompression and glossopharyngeal rhizotomy: clinical results of 21 cases.

BACKGROUND: Microvascular decompression (MVD) and rhizotomy are all selected for treating vagoglossopharyngeal neuralgia (VGPN). Nonetheless, controversies still exist about their curative effect on VGPN. Here we evaluate the effectiveness of MVD together with rhizotomy of the glossopharyngeal nerve for the treatment of VGPN. METHODS: This study was carried out on 21 patients who were diagnosed with VGPN between the years 2005 and 2010. Patients underwent MVD and glossopharyngeal rhizotomy through a retromastoid keyhole approach. Surgical technique, operation results and complications were our particular concern. RESULTS: Eighteen (85.7%) of 21 patients experienced immediate and complete relief of pain after surgery. In the remaining 3 patients (14.3%), the pain faded away within the following week. No patient complained of dysphonia or dysphagia. All 21 patients reported no change in their outcome at follow-up. CONCLUSIONS: Intracranial vagoglossopharyngeal nerve MVD with glossopharyngeal rhizotomy is an effective and safe procedure to treat VGPN.

Effect of Nogo-A gene inhibition on dopamine release in PC12 cells.

OBJECTIVES: Reticulon proteins, which are localized in the endoplasmic reticulum, have recently been shown to be involved in hormone secretion, in particular RTN1 and RTN3. The aim of the present study was to examine the effects of Nogo-A gene expression knockdown by RNA interference (RNAi) on dopamine release in PC12 cells. METHODS: A small hairpin RNA (shRNA) eukaryotic expression vector, targeting the Nogo-A gene, was constructed and transfected into cultured PC12 cells by lipofecamine2000. Inhibition of Nogo-A gene expression was detected by semi-quantitative reverse transcription PCR and Western blot analysis. Following transfection, dopamine release was detected by high performance liquid chromatography. RESULTS: The pGenesil-1-Nogo-A-2 plasmid was identified by gene sequencing. After transfection of the recombinant vector in PC12 cells, Nogo-A gene expression was significantly inhibited (p<0.01). Compared with the empty vector control group, dopamine release significantly decreased within 48 hours after transfection. CONCLUSION: Results from this study suggest that Nogo-A might be involved in the mechanism of DA release in PC12 cells.

Generation of allo-restricted cytotoxic T lymphocytes against malignant glioma by artificial antigen-presenting cells.

The interleukin (IL) 13 receptor alpha2 (IL-13Ralpha2) is a glioma-restricted cell-surface epitope not otherwise detected within the central nervous system. This study reported a novel approach for targeting malignant glioma with IL-13Ralpha2-specific allo-restricted cytotoxic T cells (CTLs) induced from the peripheral blood lymphocytes (PBLs) of HLA-A2-negative healthy donors by multiple stimulations with artificial antigen-presenting cells (aAPCs) made by coating HLA-A2/pIL-13Ralpha2(345-354) tetrameric complexes, anti-CD28 antibody and CD83 molecules to cell-sized latex beads. The induced allo-restricted CTLs exhibited specific lysis against T2 cells pulsed with the peptide pIL-13Ralpha2(345-354) and HLA-A2+ glioma cells expressing IL-13Ralpha2(345-354), while HLA-A2- glioma cell lines that express IL-13Ralpha2(345-354) could not be recognized by the CTLs. The peptide-specific activity was inhibited by anti-HLA class I monoclonal antibody. These results suggested the induced allo-restricted CTLs specific for IL-13Ralpha2(345-354) peptide could be a potential target of specific immunotherapy for HLA-A2+ patients with malignant glioma.

Negative correlation of Nogo-A with the malignancy of oligodendroglial tumor.

OBJECTIVE: Nogo-A is an axon regeneration inhibitor, and its function in central nervous system (CNS) is still unknown. The present study is to explore the relationship between the expression of Nogo-A and the malignancy of oligodendroglial tumors in patients. METHODS: Tumor tissue samples with different malignancy grade were obtained from the hospitals. The samples used for detection had been diagnosed as oligodendroglial tumors (oligodendroglioma or anaplastic oligodendroglioma). The expression of Nogo-A was detected by immunohistochemistry and western-blot analysis. The correlation test between the Nogo-A expression and the morphological changes (the percentages of atypical cells and mitotic cells in the tumors) related to the malignancy of tumor tissues was performed. RESULTS: There was significant negative correlation between the Nogo-A expression and the morphological change of tumor tissues according to immunohistochemistry. Western-blot analysis also indicated that the gray value of Nogo-A protein band in the oligodendroglioma group was significantly higher than that in the anaplastic oligodendroglioma group. CONCLUSION: Nogo-A expression was negatively correlated with the malignancy grade of oligodendroglial tumors.

[Vascular endothelial growth factor shRNA mediated by pEGFP-H1 vector plasmid effectively inhibits glioma proliferation: an experimental study].

OBJECTIVE: To investigate the inhibitory effects of RNA silencing via plasmid-mediated vascular endothelial growth factor (VEGF) shRNA on proliferation of glioma cells in vitro and in vivo. METHODS: pEGFP-H1/VEGF vector plasmid containing enhanced green fluorescent protein (EGFP) gene and expressing VEGF shRNA was constructed. The EGFP expression was detected by fluorescent microscopy and flow cytometry. Glioma cells of the line U251 were cultured and divided into 3 groups to be transfected with blank vector, pEGFP/H1plasmid, and pEGFP-H1/VEGF respectively. RT-PCR was used to detect the mRNA expression of VEGF in the supernatants of the culture media. The protein expression of VEGF in the supernatant was detected by ELISA. The cell growth was observed with MTT method. Fifteen nude rats were randomly divided into 3 equal groups to be transplanted with U251 cells and pEGFP-H1, U251 cells and pEGFP-H1/VEGF, and U251 cells only as blank control group. The growth of glioma was observed every day. 30 days after the rats were killed and the tumors were taken out to be examined. RESULTS: Twenty-four hours after transplantation, fluorescence microscopy showed great numbers of U251 cells that expressed green fluorescence in both the pEGFP-H1 group and pEGFP-H1/VEGF group. Flow cytometry showed that the rates of green fluorescent protein positive cells were 68.37% and 65.29% in these 2 groups (P > 0.05). MTT method showed no significant difference in the effect on cell growth among the 3 groups (all P > 0.05). PCR showed that the VEGF mRNA expression was significantly inhibited in the pEGFP-H1/VEGF group in comparison with those in the blank control group and the group transfected with pEGFP-H1 (both P < 0.05). The concentration of VEGF protein of the U251 cells transfected with pEGFP-H1/VEGF was 530 ng/L +/- 118 ng/L, significantly lower than those of the control group (2571 ng/L +/- 572 ng/L) and the group transfected with pEGFP-H1 (2402 ng/L +/- 310 ng/L, by 77.9%) (both P < 0.01). Tumor could be touched 13 days after transplantation in the rats of pEGFP-H1/VEGF group, markedly later than in the other 2 groups. Thirty days after, the weight and volume of tumor were 0.5 g +/- 0.4 g and 401 mm(3) +/- 272 mm(3) respectively in the rats of pEGFP-H1/VEGF group, both significantly lower than those of the control group and pEGFP-H1 group (1.7 g +/- 0.4 g and 1573 mm(3) +/- 330 mm(3); and 1.5 g +/- 0.7 g and 1430 mm(3) +/- 382 mm(3)) (all P < 0.01). CONCLUSION: The successfully constructed shRNA targeting at VEGF efficiently decreases the VEGF expression of the glioma cells in vitro and suppresses the growth of glioma cells in vivo.