Effects of Heat-Treatment and Cold-Rolling on Mechanical Properties and Impact Failure Resistance of New Al 6082 Aluminum Alloy by Continuous Casting Direct Rolling Process.

Al 6082 aluminum alloy has excellent corrosion resistance, strength, and formability. However, owing to the recrystallization effect of a hot working process, coarse grains form easily in this material, which reduces its strength and service life. The novel continuous casting direct rolling (CCDR) method can prevent the deterioration of this material. Thus, we used CCDR Al 6082 aluminum alloy as the research material in this study. By subjecting a CCDR Al 6082 aluminum alloy to heat treatment (T4 and T6) and cold rolling, the influence of recrystallization effect on its mechanical properties and on impact failure resistance were explored. The results demonstrated that the specimen subjected to T4 heat treatment had a higher elongation and that the specimen subjected to T6 heat treatment had a higher strength. After cold rolling, the hardness and strength of the specimens subjected to different heat treatments (coded T4R4 and T6R4) increased because of the work's hardening effect. Moreover, the elongations of both specimens decreased, but they were higher than the industrial standard (>10%). The strength of specimen T6R4 was higher (up to 400 MPa) than specimen T4R4. Moreover, relative to specimen T4R4, specimen T6R4 had greater tensile and Charpy impact failure toughness.

The effect of luteinizing hormone changes in GnRH antagonist protocol on the outcome of controlled ovarian hyperstimulation and embryo transfer.

BACKGROUD: To investigate the effect of Luteinizing hormone (LH) level changes on the outcomes of controlled ovarian hyperstimulation (COH) and embryo transfer (ET) in gonadotropin-releasing hormone antagonist (GnRH-ant) protocol. METHODS: A total of 721 patients undergoing GnRH-ant protocol COH for the first IVF/ICSI cycles were retrospectively analyzed. COH process were divided into 2 stages, before (stage 1) and after (stage 2) the GnRH-ant initiation, and each with 5 groups basing on LH levels: LH decreased more than 50% (groups A1, A2), decreased 25-50% (groups B1, B2), change less than 25% (groups C1, C2), increased 25-50% (groups D1, D2), and increased more than 50% (groups E1, E2). RESULTS: There were no significant differences among groups of stage1 regarding COH and ET outcomes. For stage 2, the more obvious the decrease of LH level, the more the number of oocytes retrieved, mature oocytes, fertilized oocytes, embryos cleavaged and the numbers of embryo available (P < 0.05), but without significant differences regarding ET outcomes. We also found the freeze-all rate in Group A2 was higher (P < 0.001). CONCLUSION: LH level changes before GnRH-ant addition were not related to COH and ET outcomes. LH level changes after the addition of GnRH-ant were related to the outcome of COH, and no significant differences were found relating to ET outcomes.

Malignant glomus tumor of the intestinal ileum with multiorgan metastases: A case report and review of literature.

BACKGROUND: Glomus tumors (GTs) are rare mesenchymal neoplastic lesions derived from cells of the glomus body. GTs rarely occurs in the visceral organs, where there may be few or no glomus bodies, and the majority of GTs are benign, rarely demonstrating aggressive or malignant behavior and histological features. CASE SUMMARY: We report a patient with malignant GTs of the intestinal ileum with multiorgan metastases who was admitted due to moderate anemia. Capsule endoscopy revealed a bleeding mass in the intestinal ileum, and the patient underwent segmental ileal resection through laparoscopic surgery. The histopathological and immunohistochemical diagnoses were consistent with malignant GT. Long-term follow-up showed that the GT had metastasized to multiple organs such as the colon, brain, and possibly the lung. CONCLUSION: This case was characterized by the highest degree of malignancy and by multiorgan metastases, and it was the first case of intestinal GT uncovered by capsule endoscopy.

Prognostic value of the 2018 FIGO staging system for cervical cancer patients with surgical risk factors.

Objective: To determine the predictive value of the 2018 International Federation of Gynecology and Obstetrics (FIGO) staging system for cervical cancer patients with surgical risk factors. Methods: Data of 662 cervical cancer patients (stages IB and IIA) with surgical risk factors treated at Zhejiang Cancer Hospital between 2008 and 2011 were retrospectively reviewed. Univariate log-rank test and multivariate Cox regression models were adopted to evaluate the relationship between 2018 FIGO stage and survival. Results: On re-staging of patients, 17.3%, 44.5%, 25.4%, and 37.1% of the patients with FIGO 2009 stage IB1, IB2, IIA1, and IIA2, respectively, were upgraded to FIGO 2018 IIIC1P stage, and 2.1%, 3.0%, 3.1%, and 2.1% patients, respectively, were upgraded to IIIC2P stage. The 5-year overall survival (OS) rates of patients with FIGO 2018 stage IB1, IB2, IB3, IIA1, IIA2, IIIC1P, and IIIC2P were 95.3%, 95.1%, 90.4%, 92.4%, 86.4%, 81.9%, and 56.3%, respectively. The 5-year progression-free survival (PFS) rates were 94.0%, 91.0%, 88.5%, 91.4%, 86.4%, 79.5%, and 43.8%, respectively. The 5-year OS rates of patients with 1-2 positive pelvic lymph nodes (PLNs) and those with >2 positive PLNs were 86.0% and 73.7%, respectively, and the 5-year PFS rates were 84.2% and 70.2%, respectively. OS and PFS of patients with 1-2 positive PLNs in stage IIIC1P were similar to those of patients in stage IIA2 without lymph node metastasis, but significantly better than those of patients with >2 positive PLNs. Multivariate analysis showed FIGO 2018 stage to be an independent prognostic factor for OS and PFS. Conclusion: The 2018 FIGO staging system for cervical cancer appears to be useful for predicting prognosis of patients with risk factors after radical surgery. Survival of stage IIA1 patients is better than that of stage IB3 patients. Stage IIIC1 is not homogenous; survival in stage IIIC1P depends on the number of positive PLNs.

A comprehensive analysis of the factors of positive pelvic lymph nodes on survival of cervical cancer patients with 2018 FIGO stage IIIC1p.

Objective: To evaluate the factors associated with positive pelvic lymph nodes (LNs) on the survival of patients with 2018 FIGO stage IIIC1p cervical cancer. Methods: We retrospectively analyzed 155 patients with pelvic lymph node metastasis (LNM) confirmed by pathology after radical resection of cervical cancer treated at Zhejiang Cancer Hospital, China, between March 2008 and October 2011. We analyzed the influence of the factors associated with positive pelvic LNs on the survival of patients. Results: The 5-year progress-free survival (PFS) and overall survival (OS) of patients were 78.1% and 81.9%, respectively. The 5-year PFS and OS of patients with more than 2 LNM were worse compared with patients with 1 or 2 LNM (68.4% vs 83.7%, p=0.013; 72.4% vs 87.6%, p=0.017, respectively). The 5-year PFS and OS of patients with more than 2 LNM sites were worse than that of patients with 1 or 2 LNM sites (60.0% vs 82.4%, p=0.008; 70.0% vs 84.8%, p=0.045, respectively). The 5-year PFS and OS of patients with common iliac LNM was poorer than that of patients without common iliac LNM (60.7% vs 81.9%, p=0.008; 67.9% vs 85.0%, p=0.020, respectively). Compared with other patients, the survival of patients with these three factors (more than 2 LNM, more than 2 LNM sites, and common iliac LNM) was the worst (p<0.05). Conclusion: More than 2 LNM, more than 2 LNM sites, and common iliac LNM were predictive factors of poor survival in stage IIIC1p cervical cancer patients. Survival of patients with stage IIIC1p cervical cancer declined with increasing presence of such factors. The combined evaluation of the factors associated with positive pelvic LNs is a more comprehensive and pragmatic approach in evaluating the prognosis of cervical cancer.

Cisplatin concurrent chemoradiotherapy vs adjuvant radiation in stage IB/IIA cervical cancer with intermediate risk factors, treated with radical surgery: a retrospective study.

PURPOSE: To determine if postoperative cisplatin concurrent chemoradiotherapy (CCRT) improves the outcome in stage IA/IIB cervical cancer patients with intermediate risk factors, when compared with radiation therapy (RT) alone, and identify the potential eligible populations for this treatment. PATIENTS AND METHODS: We reviewed medical records of 1,240 patients with stage IA/IIB cervical cancer who underwent radical hysterectomy and pelvic lymphadenectomy in our hospital between January 2008 and December 2011. Of the 1,240 patients, 436 displayed 1 or more intermediate risk factors. Of these, we screened 306 patients who underwent RT only or CCRT. We analyzed the effects of CCRT on survival and prognosis. RESULTS: The 5-year progress-free survival (PFS) in the CCRT group was superior to that in the RT-only group (96.0% vs 89.0%, respectively; P=0.031). The 5-year overall survivals (OSs) were not different between the 2 groups (P=0.141). Compared with RT-only group, CCRT did not improve PFS or OS in patients with 1 risk factor, large tumor size, or deep stromal invasion (P>0.05). Compared with RT-only group, CCRT improved PFS (97.9% vs 82.8%; P=0.017) but did not increase OS (97.9% vs 89.7%; P=0.109) in patients with lymphovascular space invasion plus deep stromal invasion/large tumor size. OS (92.3% vs 70.6%; P=0.048) and PFS (92.3% vs 64.7%; P=0.020) in the CCRT group were superior to those in the RT-only group with 3 risk factors. Compared with RT-only group, CCRT was an independent prognostic factor for favorable PFS (hazard ratio [HR] =0.238; 95% CI =0.0827-0.697, P=0.009) and OS (HR =0.192; 95% CI =0.069-0.533, P=0.002). CONCLUSION: Postoperative CCRT improved survival in stage IA/IIB cervical cancer patients with intermediate risk factors. Patients with 2 or more intermediate risk factors, including lymphovascular space invasion, may benefit from CCRT.

Activation of PAR-1/NADPH oxidase/ROS signaling pathways is crucial for the thrombin-induced sFlt-1 production in extravillous trophoblasts: possible involvement in the pathogenesis of preeclampsia.

BACKGROUNDS/AIMS: Preeclampsia was characterized by excessive thrombin generation in placentas and previous researches showed that thrombin could enhance soluble Fms-like tyrosine kinase 1 (sFlt-1) expression in first trimester trophoblasts. However, the detailed mechanism for the sFlt-1 over-production induced by thrombin was largely unknown. The purpose of this study was to explore the possible signaling pathway of thrombin-induced sFlt-1 production in extravillous trophoblasts (EVT). METHODS: An EVT cell line (HRT-8/SVneo) was treated with various concentrations of thrombin. The mRNA expression and protein secretion of sFlt-1 in EVT were detected with real-time polymerase chain reaction and ELISA, respectively. The levels of intracellular reactive oxygen species (ROS) production were determined by DCFH-DA. RESULTS: Exposure of EVT to thrombin induced increased intracellular ROS generation and overexpression of sFlt-1 at both mRNA and protein levels in a dose dependent manner. Short interfering RNA (siRNA) directed against PAR-1 or apocynin (an inhibitor of NADPH oxidase) could decrease the intracellular ROS generation and subsequently suppressed the production of sFlt-1 at mRNA and protein levels. CONCLUSIONS: Our results suggested that thrombin increased sFlt-1 production in EVT via the PAR-1 /NADPH oxidase /ROS signaling pathway. This also highlights the PAR-1 / NADPH oxidase / ROS pathway might be a potential therapeutic target for the prevention of preeclampsia in the future.

Interaction between NBS1 and the mTOR/Rictor/SIN1 complex through specific domains.

Nijmegen breakage syndrome (NBS) is a chromosomal-instability syndrome. The NBS gene product, NBS1 (p95 or nibrin), is a part of the Mre11-Rad50-NBS1 complex. SIN1 is a component of the mTOR/Rictor/SIN1 complex mediating the activation of Akt. Here we show that NBS1 interacted with mTOR, Rictor, and SIN1. The specific domains of mTOR, Rictor, or SIN1 interacted with the internal domain (a.a. 221-402) of NBS1. Sucrose density gradient showed that NBS1 was located in the same fractions as the mTOR/Rictor/SIN1 complex. Knockdown of NBS1 decreased the levels of phosphorylated Akt and its downstream targets. Ionizing radiation (IR) increased the NBS1 levels and activated Akt activity. These results demonstrate that NBS1 interacts with the mTOR/Rictor/SIN1 complex through the a.a. 221-402 domain and contributes to the activation of Akt activity.

Looped host defense peptide CLP-19 binds to microtubules and inhibits surface expression of TLR4 on mouse macrophages.

The looped host defense peptide CLP-19 is derived from a highly functional core region of the Limulus anti-LPS factor and exerts robust anti-LPS activity by directly interacting with LPS in the extracellular space. We previously showed that prophylactic administration of CLP-19 even 20 h prior to LPS challenge might significantly increase the survival rate in a lethal endotoxin shock mouse model. Such an effect may be associated with immune regulation of CLP-19. To investigate the underlying mechanisms, peptide affinity chromatography, immunofluorescence, and Western blotting procedures were used to identify α- and ß-tubulin as direct and specific binding partners of CLP-19 in the mouse macrophage cell line RAW 264.7. Bioinformatic analysis using the AutoDock Vina molecular docking and PyMOL molecular graphics system predicted that CLP-19 would bind to the functional residues of both α- and ß-tubulin and would be located within the groove of microtubules. Tubulin polymerization assay revealed that CLP-19 might induce polymerization of microtubules and prevent depolymerization. The immunoregulatory effect of CLP-19 involving microtubules was investigated by flow cytometry, immunofluorescence, and Western blotting, which showed that CLP-19 prophylactic treatment of RAW 264.7 cells significantly inhibited LPS-induced surface expression of TLR4. Taken together, these results suggest that CLP-19 binding to microtubules disrupts the dynamic equilibrium of microtubules, reducing the efficacy of microtubule-dependent vesicular transport that would otherwise translocate TLR4 from the endoplasmic reticulum to the cell surface.

Connexin 43 reverses malignant phenotypes of glioma stem cells by modulating E-cadherin.

Malfunctioned gap junctional intercellular communication (GJIC) has been thought associated with malignant transformation of normal cells. However, the role of GJIC-related proteins such as connexins in sustaining the malignant behavior of cancer stem cells remains unclear. In this study, we obtained tumorspheres formed by glioma stem cells (GSCs) and adherent GSCs and then examined their GJIC. All GSCs showed reduced GJIC, and differentiated glioma cells had more gap junction-like structures than GSCs. GSCs expressed very low level of connexins, Cx43 in particular, which are key components of gap junction. We observed hypermethylation in the promoter of gap junction protein α1, which encodes Cx43 in GSCs. Reconstitution of Cx43 in GSCs inhibited their capacity of self-renewal, invasiveness, and tumorigenicity via influencing E-cadherin and its coding protein, which leads to changes in the expression of Wnt/ß-catenin targeting genes. Our results suggest that GSCs require the low expression of Cx43 for maintaining their malignant phenotype, and upregulation of Cx43 might be a potential strategy for treatment of malignant glioma.

Cyclic Limulus anti-lipopolysaccharide (LPS) factor-derived peptide CLP-19 antagonizes LPS function by blocking binding to LPS binding protein.

Inflammation and septic shock due to endotoxins from Gram-negative bacteria infection continue to pose significant challenges to human healthcare. It is, therefore, necessary to develop therapeutic strategies targeting endotoxins, such as lipopolysaccharide (LPS), to prevent their potentially systemic effects. Pathogenesis due to Gram-negative bacteria involves LPS binding to the host LPS-binding protein (LBP), causing detrimental downstream signaling cascades. Our previous study showed that CLP-19, a synthetic peptide derived from the Limulus anti-LPS factor (LALF), could effectively neutralize LPS toxicity; however, the detailed mechanisms underlying this anti-LPS effect remained unexplained. Thus, we carried out investigations to determine how the CLP-19 neutralizes LPS toxicity. CLP-19 was found to block LPS binding to LBP in a dose-dependent manner, as evidenced by competitive enzyme-linked immunosorbent assay (ELISA). In peripheral blood mononuclear cells, CLP-19 blocked LPS-induced phosphorylation of mitogen activated protein kinase (MAPK) signaling proteins p38, extracellular signal-regulating kinase (ERK)1/2 and c-Jun N-terminal kinase (JNK)1/2. Furthermore, CLP-19 potency in LPS antagonism in vitro and in vivo was directly associated with its ability to block the LPS-LBP interaction. Taken together, the results suggested that CLP-19's inhibitory effect on LPS-LBP binding and on the subsequent MAPK pathway signaling may be responsible for its anti-LPS mechanism. This peptide appears to represent a potential therapeutic agent for clinical treatment of sepsis.

The chemokine CXCL12 and its receptor CXCR4 promote glioma stem cell-mediated VEGF production and tumour angiogenesis via PI3K/AKT signalling.

Chemokines and their receptors are actively involved in inflammation, immune responses, and cancer development. Here we report the detection of CD133(+) glioma stem-like cells (GSCs) co-expressing a chemokine receptor CXCR4 in human primary glioma tissues. These GSCs were located in areas adjacent to tumour vascular capillaries, suggesting an association between GSCs and tumour angiogenesis. To test this hypothesis, we isolated CD133(+) GSCs from surgical specimens of human primary gliomas and glioma cell lines. As compared to CD133(-) cells, CD133(+) GSCs expressed significantly higher levels of CXCR4 mRNA and protein, and migrated more efficiently in response to the CXCR4 ligand CXCL12. In addition, CXCL12 induced vascular endothelial growth factor (VEGF) production by CD133(+) GSCs via activation of the PI3K/AKT signalling pathway. Furthermore, knocking down of CXCR4 using RNA interference or inhibition of CXCR4 function by an antagonist AMD3100 not only reduced VEGF production by CD133(+) GSCs in vitro, but also attenuated the growth and angiogenesis of tumour xenografts in vivo formed by CD133(+) GSCs in SCID mice. These results indicate that CXCL12 and its receptor CXCR4 promote GSC-initiated glioma growth and angiogenesis by stimulating VEGF production.

[Association between endometrial cancer and metabolic syndrome].

OBJECTIVE: To study the association between endometrioid uterine carcinomas and metabolic syndrome (MS). METHODS: A retrospective study was conducted on 123 patients who were admitted in Department of Gynecology Oncology, Zhejiang Cancer Hospital (study group) and 90 healthy women (control group) with matching age from Jan. 2005 to Mar. 2009. The general conditions [including age, whether menopausal, body mass index (BMI)]; the risk factors for MS [including waist circumference, fasting plasma glucose, triglycerides (TG), high-density lipoprotein (HDL) and systolic and diastolic blood pressure] were analyzed. The clinical stage, histological type, and pathology differentiated degree of study group with or without MS were also analyzed by univariate analysis and Cox proportional hazards models. RESULTS: (1) The univariate survival analysis shown that there were no significant difference with age in two groups [(54.3 +/- 0.6) vs. (54.2 +/- 0.9) years; P > 0.05], while the rate of menopausal, BMI (> or = 25 kg/m(2)), the cases coupled with MS, the size of waist circumference (> 80 cm), the level of fasting plasma glucose (> or = 5.6 mmol/L), TG (> 1.7 mmol/L) and abnormal systolic and diastolic blood pressure in study group were higher than those in control group (67.5% vs. 48.9%, 45.5% vs. 23.3%, 43.9% vs. 18.9%, 50.4% vs. 27.8%, 53.7% vs. 21.1%, 40.7% vs. 21.1% and 40.7% vs. 25.6%, respectively, all P < 0.05). The percentage of HDL (< 1.30 mmol/L) was higher in study group than that in control group (63.4% vs. 32.2%, P < 0.05). (2) There were not significant difference for the clinical stage, pathological type, grades between patients with or without MS in study group (P > 0.05). (3) The Logistic multivariate survival analysis shown that central obesity, higher TG, lower HDL and abnormal plasma glucose were independent risk factors for endometrioid uterine carcinomas coupled with MS (P < 0.05). CONCLUSION: Metabolic syndrome is marginally associated with an increased risk of endometrioid uterine carcinomas, which may be the new point to screen, prevention and treatment endometrioid uterine carcinomas.

Glial activation involvement in neuronal death by Japanese encephalitis virus infection.

Japanese encephalitis is characterized by profound neuronal destruction/dysfunction and concomitant microgliosis/astrogliosis. Although substantial activation of glia is observed in Japanese encephalitis virus (JEV)-induced Japanese encephalitis, the inflammatory responses and consequences of astrocytes and microglial activation after JEV infection are not fully understood. In this study, infection of cultured neurons/glia with JEV caused neuronal death and glial activation, as evidenced by morphological transformation, increased cell proliferation and elevated tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6 and RANTES (regulated upon activation, normal T-cell expressed and secreted) production. Replication-competent JEV caused all glial responses and neurotoxicity. However, replication-incompetent JEV lost these abilities, except for the ability to change microglial morphology. The bystander damage caused by activated glia also contributed to JEV-associated neurotoxicity. Microglia underwent morphological changes, increased cell proliferation and elevated TNF-alpha, IL-1beta, IL-6 and RANTES expression in response to JEV infection. In contrast, IL-6 and RANTES expression, but no apparent morphological changes, proliferation or TNF-alpha/IL-1beta expression, was demonstrated in JEV-infected astrocytes. Supernatants of JEV-infected microglia, but not JEV-infected astrocytes, induced glial activation and triggered neuronal death. Antibody neutralization studies revealed that TNF-alpha and IL-1beta, but not RANTES or IL-6, released by activated microglia appeared to play roles in JEV-associated neurotoxicity. In conclusion, following JEV infection, neuronal death was accompanied by concomitant microgliosis and astrogliosis, and neurotoxic mediators released by JEV-activated microglia, rather than by JEV-activated astrocytes, had the ability to amplify the microglial response and cause neuronal death.

A novel approach to the identification and enrichment of cancer stem cells from a cultured human glioma cell line.

Enrichment of cancer stem cells for studies of carcinogenesis remains a difficult issue. We hypothesized that the unique features of cancer stem cells (CSCs) may allow formation of their colonies in vitro with distinct morphology. We therefore investigated the possibility to use morphological diversity of colonies to identify and enrich CSCs from cultured malignant human glioma cells. We found that a small proportion of the cells from a human glioma cell line U251 formed tight and round-shaped colonies in culture. Most cells in such colonies were capable of self-renewal, generating tumor spheres and differentiating into lineages with markers for neurons, astrocytes and oligodendrocytes. In addition, several neural stem cell-related genes were highly expressed by tumor cells in those tight colonies. Our results thus demonstrate a novel approach to the identification and enrichment of CSCs based on unique morphology of their colonies formed in vitro.

Downregulating FPR restrains xenograft tumors by impairing the angiogenic potential and invasive capability of malignant glioma cells.

G-protein-coupled formylpeptide receptor (FPR) has recently been found to be functionally expressed in gliomas and are probably involved in their malignant biological behavior. In an attempt to explore the therapeutic significance of FPRs, we used wild-type human glioblastoma cells (U87), the corresponding FPR short-interfering RNA transfected (siRNA U87) cells, and mock-transfected U87 cells (mock U87) to establish xenografts in mice brains. Compared to wild-type and mock transfected cells, siRNA U87 cells formed smaller and more well-differentiated xenografts with fewer mitotic figures and more glial filaments within their cytoplasm. The density of microvessels, which presented as a nearly normal morphous, was also decreased significantly in FPR knockdown cells. Moreover, fewer invasive foci could be observed in the xenografts derived from siRNA U87 cells, which also showed a poor migratory capacity in vitro. We suggest that decreased VEGF and MMP-2/-9 expression might be a possible mechanism for the decreasing angiogenic potential and invasive capability of U87 cells after FPR knockdown. Functional FPR might be essential for sustaining the growth and aggressive phenotype of gliomas, and could therefore be a potential therapeutic target.

[Aluminum-resistance of AHA1 transgenic Arabidopsis thaliana: physiological analysis].

As one of the major genes encoding plasma membrane H+ -ATPase, AHA1 gene plays an important role in regulating plant development and resistance to adverse stress. Taking AHA1 transgenic and wild type Arabidopsis thaliana as test plants, the nutrient uptake, resistance to oxidative stress, and organic acid secretion of the plants under aluminum (Al) stress were examined. The results showed that Al decreased the uptake of nitrogen (N), potassium (K), calcium (Ca) and magnesium (Mg), but increased the phosphorus (P) uptake by A. thaliana roots. AHA1 transgenic plant could accumulate more P and less Al than wild type plant. Al stress induced the increase of plant SOD and POD activities, but no significant difference was observed between AHA1 transgenic and wild type A. thaliana. Al triggered the secretion of organic acids significantly, and AHA1 transgenic plant secreted more organic acids than wild type plant. Vanadate, an inhibitor of plasma membrane H+ -ATPase, could inhibit the secretion of organic acids significantly, while Zn2+ and Mg2+ could promote the Al-induced secretion, and partially improve the inhibitory effects triggered by vanadate. It was suggested that AHA1 transgenic A. thaliana could increase its Al resistance via enhanced P uptake and organic acid secretion.

Isolation and characterization of cancer stem cells from a human glioblastoma cell line U87.

A variety of malignant cancers have been found to contain a subpopulation of stem cell-like tumor cells, or cancer stem cells (CSCs). However, the existence of CSCs in U87, a most commonly used glioma cell line, is still controversial. In this study, we demonstrate that U87 cell line contained a fraction of tumor cells that could form tumor spheres and were enriched by progressively increasing the concentration of serum-free neural stem cell medium with or without low dose vincristine. These cells possessed the ability of self-renewal and multipotency, the defined characteristics of CSCs. Moreover, the tumors formed by the secondary spheres displayed typical histological features of human glioblastoma, including cellular pleomorphism, pseudopalisades surrounding necrosis, hyperchromatic nuclei, high density of microvessels and invasion to the brain parenchyma. These results indicate that gradually increasing the concentration of serum-free neural stem cell culture medium with or without vincristine is a simple and effective method for isolation of CSCs to study the initiation and progression of human glioblastoma.

Unique proteomic features induced by a potential antiglioma agent, Nordy (dl-nordihydroguaiaretic acid), in glioma cells.

Nordy is a chirally synthesized compound of a natural lipoxygenase inhibitor nordihydroguaiaretic acid. In this study, we found that Nordy inhibited the growth of human glioma cell lines in vitro and their tumorigenicity in mice. In addition, Nordy promoted differentiation of highly malignant human glioma cells. Investigation into the mechanistic basis of Nordy activities revealed that it altered the pattern of protein expression profiles in tumor cells. By using 2-DE, we found that in human glioma cell lines, at least six proteins were down-regulated after Nordy treatment, while four proteins were elevated in the same cells. Among the six down-regulated proteins, microsequencing with MALDI TOF MS confirmed the identity of five: proliferation-associated gene A (PAG-A), alternative splicing factor-3 (ASF-3), beta-galactoside binding lectin, eukaryotic translation initiation factor 5A (eIF-5A), and coffilin-1 (nonmuscle). Four up-regulated proteins were GST-pi, glyceraldehyde-3-phosphate dehydrogenase, alpha-enolase, and cyclophilin. All these proteins have been reported to participate in key cellular functions including proliferation, metabolism, differentiation, apoptosis, and gene transcription. Our results suggest that Nordy may constitute a promising drug lead for the development of novel antitumor agents targeting proteins that control tumor cell function at multiple levels.

Production of angiogenic factors by human glioblastoma cells following activation of the G-protein coupled formylpeptide receptor FPR.

Activation of the formylpeptide receptor (FPR), a G-protein-coupled receptor, by its chemotactic peptide ligand N-formylmethionyl-leucyl-phenylalanine (fMLF) promotes the directional migration and survival of human glioblastoma cells. fMLF also stimulates glioblastoma cells to produce biologically active VEGF, an important angiogenic factor involved in tumor progression. In this study, we examined the capacity of FPR to regulate the production of another angiogenic factor, the chemokine IL-8 (CXCL8), in addition to its demonstrated ability to induce VEGF secretion by malignant glioma cells. We showed that the human glioblastoma cell line U87 secreted considerable levels of IL-8 (CXCL8) upon stimulation by the FPR agonist peptide fMLF. Tumor cells transfected with small interference (si)RNA targeting FPR failed to produce IL-8 as well as VEGF in response to fMLF. Glioblastoma cells bearing FPR siRNA exhibited reduced rate of tumorigenicity in nude mice and tumors formed by such tumor cells showed less active angiogenesis and lower level expression of both IL-8 and VEGF. These results suggest that FPR plays an important role in the angiogenesis of human malignant gliomas through increasing the production of angiogenic factors by FPR positive tumor cells.