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Our studies were aimed to explore the effect and mechanism of the inhibition of the formation of vasculogenic mimicry (VM) in human glioblastoma cells by Xihuang pill (XHP) medicated serum through regulating the hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor A (VEGFA)/vascular endothelial growth factor receptor 2 (VEGFR2) signaling pathway. The medicated serum of XHP was prepared by gavage for 7 days to male SD rats (approval number of animal experiment ethics: 202105A051). The hypoxia model of U251 cells was established using 200 μmol·L-1 of CoCl2. After treatment with XHP-medicated serum, cell viability and proliferation of U251 cells were detected by CCK-8 and cell cloning experiment. Cell apoptosis and cell cycle of U251 cells were determined by flow cytometry. Cell migration and invasion were evaluated by wound healing and Transwell invasion assay. The formation of VM was assessed by three-dimensional cell culture of U251 cells. The protein expression levels of HIF-1α, VEGFA, VEGFR2, phosphorylated-VEGFR2 (p-VEGFR2), vascular endothelial-cadherin (VE-cadherin), Eph receptor tyrosine kinases A2 (EphA2), matrix metalloproteinase 2 (MMP2), matrix metalloproteinase 14 (MMP14) and laminin γ2 in U251 cells were detected by Western blot. The results showed that 10% XHP-medicated serum had little effect on the cell viability, proliferation, apoptosis and cell cycle of U251 cells under hypoxia. Compared with the model group, 10% XHP-medicated serum at 1.0, 1.5 and 2.0 h significantly decreased the migration rate (P < 0.01) and the number of invading U251 cells (P < 0.01). 10% XHP-medicated serum at 2.0 h significantly suppressed the formation of VM tubular structures in U251 cells under the condition of hypoxia (P < 0.01). Western blot experiment showed that 10% XHP-medicated serum significantly down-regulated the expression of HIF-1α, VEGFA, phospho-VEGFR2, VE-cadherin, EphA2 and MMP14 proteins (P < 0.05). In conclusion, XHP could inhibit the formation of VM in human glioblastoma U251 cells to suppress the angiogenesis by down-regulating the HIF-1α/VEGFA/VEGFR2 signaling pathway.
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OBJECTIVE@#To investigate the inhibitory effect of agkistrodon halys venom antitumor component-I (AHVAC-I) on vasculogenic mimicry (VM) formation in triple-negative breast cancer MDA-MB-231 cells and explore its possible mechanism.@*METHODS@#CCK8 assay was used to determine the optimal concentration of AHVAC-I for cell treatment based on its halfinhibitory concentration (IC50). MDA-MB-231 cells were treated with different concentrations of AHVAC-I or 5-Fu, and the changes in vasomimetic capacity of the cells were examined using Matrigel assay. The expression levels of matrix metalloproteinase-2 (MMP2) and MMP9 in the treated cells were detected using quantitative PCR and Western blotting.@*RESULTS@#Compared with the control treatment with culture medium, treatment with 5, 10 and 20 μg/mL AHVAC-I significantly reduced vasomimetic ability of MDA-MB-231 cells in a dose-dependent manner (P < 0.01). MMP2 supplementation obviously restored the vasomimetic ability of the cells inhibited by AHVAC-I.@*CONCLUSION@#AHVAC-I inhibits VM formation in triplenegative breast cancer cells in vitro by down-regulating MMP2 production.
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Animals , Humans , Agkistrodon/metabolism , Cell Line, Tumor , Healthy Life Expectancy , Matrix Metalloproteinase 2/metabolism , Neovascularization, Pathologic/metabolism , Triple Negative Breast Neoplasms/metabolism , VenomsABSTRACT
Objective To investigate the relation between vasculogenic mimicry (VM) and MIG-7 in osteosarcoma, as well as their roles in the prognosis, and to establish a model for predicting the prognosis of osteosarcoma. Methods VM was identified by CD31/PAS double-staining in 156 cases of AJCC stage Ⅱ extremity osteosarcoma. Tumor samples were also immunohistochemically stained for MIG-7 to determine whether it was associated with the occurrence of VM. Univariate and multivariate Cox regression analyses were used to identify prognostic factors and a prognostic nomogram for predicting 3- and 5-year OS and MFS was constructed. C-index and calibration curves were used to verify the predictive accuracy of the model. Results The MIG-7 expression in osteosarcoma tissues was associated with VM formation, but MIG-7 expression was not associated with gender, age, AJCCⅡA/ⅡB stage, tumor location, surgical type or histological response to pre-operative chemotherapy. Survival analysis showed that MIG-7 expression, VM and pre-operative chemotherapy were identified as three independent prognostic factors. The value of C-index in nomogram was greater than 0.7. The predicted calibration curve was similar to the standard curve. Conclusion MIG-7 accelerates the progression of osteosarcoma by promoting VM formation, and may also affect prognosis through other mechanisms. The nomogram could afford accurate prognosis prediction and individualized diagnosis and treatment for osteosarcoma patients.
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Vasculogenic mimicry (VM) plays an important role in human glioma progression and resistance to antiangiogenic therapy as a compensatory neovascularization mechanism in malignant tumors. Caveolin-1 (Cav-1) has been found to contribute to VM formation. However, it remains largely unknown whether Cav-1 expression correlates with VM in glioma. In this study, we examined CAV-1 expression levels and VM in human glioma cell lines and in 94 human gliomas with different grades of malignancy, and present Cox proportional hazards regression. The molecular role of Cav-1 in glioma cells was investigated using quantitative polymerase chain reaction (qRT-PCR) assays, western blotting, CCK-8 assays, and tubule formation assays. Cav-1 expression and VM formation were positively correlated with each other and both were closely associated with glioma development and progression as evidenced by the presence of cystic tumor, shortened survival time, and advanced-stage glioma in glioma patients with Cav-1 overexpression/increased VM formation. Cav-1 promoted U251 glioma cell proliferation and VM formation in a Matrigel-based 3D culture model. VM-associated factors including hypoxia-inducible factor 1α (HIF-1α) and p-Akt was significantly elevated by Cav-1 overexpression but suppressed by siCav-1 in U251 cells. Collectively, our study identified Cav-1 as an important regulator of glioma cell proliferation and VM formation, contributing to glioma development and progression.
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Humans , Caveolin 1/genetics , Glioma , Cell Line, Tumor , Cell Proliferation , Neovascularization, PathologicABSTRACT
Vasculogenic mimicry (VM) is a new tumor angiogenesis mode independent of endothelial cells and an important component of tumor microcirculation. The formation mechanism of VM in glioma is complex and variable. Various molecules and signal pathways (such as hypoxia induction factor and matrix metalloproteasefamily) interact in the formation process, to jointly regulate the formation of VM. The in-depth study of molecular mechanism can provide a theoretical basis for drug research and development against VM formation.
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@#AIM: To investigate the effects of doxycycline(DOX)on vasculogenic mimicry(VM)in human pterygium fibroblasts(HPFs)and its molecular mechanisms.<p>METHODS: Primary cultured HPFs were identified by Vimentin and CK through immunocytochemical staining. HPFs were divided into control group and DOX group including low, medium and high concentrations(50, 100, 200mg/L). The activity and migration of HPFs were detected by cell counting kit-8(CCK-8)and wound healing assay. The density of VM was observed by three-dimensional cell culture and periodic acid schiff(PAS)staining and compared the differences of VM formation in each group. Western blot was used to analyze the expression of matrix metalloproteinase-9(MMP-9)and vascular endothelial growth factor(VEGF).<p>RESULTS:Immunocytochemical staining results showed that the cells were spindle shaped, meanwhile, they were positive for Vimentin and negative for CK, which were consistent with the characteristics of fibroblasts. Compared with the control group, the cell activity, mobility, VM density and the expression of MMP-9 and VEGF proteins in the DOX group were significantly decreased(<i>P</i><0.05). Compared among different concentrations of DOX groups, the differences were statistically significant(<i>P</i><0.05). Correlation analysis indicated that VM density formed by HPFs was significantly positively correlated with the protein expression of MMP-9 and VEGF(<i>r</i>=0.949, 0.960, all <i>P</i><0.05).<p>CONCLUSION: DOX can inhabit HPFs activity, migration, VM density by reducing the expression of MMP-9 and VEGF, suggesting that MMP-9 and VEGF may be the molecular mechanisms of VM formation in pterygium.
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Objective To evaluate the potential anti-osteosarcoma effects of Paris polyphylla ethanol extract (PPEE) and investigate its underlying mechanisms. Methods The anti-proliferation activity of PPEE was tested on 143B, MG-63, U-2 OS, and hFOB1.19 cells using MTT assay. The pro-apoptotic and cell cycle arrest effects of PPEE were confirmed by Hoechst 33342 staining and flow cytometry. The anti-vasculogenic mimicry effects of PPEE were investigated by Matrigel culture assays. The related proteins expression was evaluated by Western blottig. Results PPEE evidently suppressed cell proliferation of 143B, MG-63, and U-2 OS cells with IC50 values of 10-60 μg/mL, but showed little cytotoxicity against normal osteoblastic cell. PPEE induced G2/M phase arrest associated with elevated phosphorylation of CDK1, Cdc25C, Chk2 and down-regulation of cyclin B1 expression. It also promoted apoptosis in 143B cells by up-regulating the expression of cleaved Caspase-3, Caspase-8, and Caspase-9, and Bax/Bcl-2 ratio. Additionally, PPEE inhibited 143B cells vasculogenic mimicry formation at non-cytotoxic concentrations through decreasing the expression of FAK, Mig-7, MMP-2, and MMP-9. Finally, four weeks’ daily oral administration of PPEE exhibited potent antitumor and anti-vasculogenic mimicry activity in 143B xenograft model with low toxicity. Conclusion These findings demonstrated that PPEE possesses anti-osteosarcoma and anti-vasculogenic mimicry activity in vitro and in vivo, and its underlying mechanisms may be related to inducing apoptosis, blocking cell cycle, and destroying vasculogenic mimicry formation of osteosarcoma cells. Therefore, PPEE is a potential candidate for osteosarcoma treatment.
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@#Objective: To investigate the effects of ginsenoside Rg3 on the formation of vasculogenic mimicry (VM) in gastric cancer cell line SGC7901 and its molecular mechanism. Methods: MTT assay was used to detect the effect of different concentrations of Rg3 on the proliferation of SGC7901 cells. SGC7901 cells were grouped as follows: BML-284 group, XAV-939 group, Rg3 group, Rg3+ BML-284 group and blank group. Transwell chamber assay was used to detect cell invasion and migration; the formation of VM was observed by tube formation assay; the secretion of MMP-9 and MMP2 was detected by ELISA; the mRNA expressions of GSK-3β and Wnt2B were detected by qPCR; the expression of β-Catenin protein in cells was analyzed by WB; and nuclear entry of β-Catenin was examined by Immunofluorescence. Results: Ginsenoside Rg3 inhibited the proliferation of SGC7901 cells in a time- and concentrationdependent manner; compared with the blank group, 40 mg/L Rg3 significantly inhibited the invasion and migration of SGC7901 cells (both P<0.05) and VM formation (P<0.05); in the meanwhile, the expressions of intracellular GSK-3β, Wnt2B mRNA and β-catenin protein, as well as the nuclear entry of β-catenin were significantly inhibited (all P<0.05). The invasion, migration and VM formation of SGC7901 cells in Rg3+BML-284 group were not significantly different from those in the blank group (all P>0.05). Conclusion: Rg3 can inhibit cell invasion, migration and VM formation in SGC7901 cells by inhibiting the activation of Wnt/β-Catenin pathway.
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The growth of solid tumors is dependent on new blood vessels generated from existed ones,that is tumor angiogenesis.Much material consumption is required in the invasion and metastasis of malignant tumors,to meet the own needs of oxygen and nutrients and remove the metabolic wastes from them,tumor cells will generate new vasculature through different ways.Vasculogenic mimicry is a kind of angiogenesis dominated by tumor cells,it is one style of vascular pattern that is different from traditional endothelial cells pattern.Vasculogenic mimicry plays an important role in the invasion and metastasis of tumors.This article will review the relationship between vasculogenic mimicry and tumor invasion and metastasis,tumor angiogenesis,and related signaling pathways.
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Cholangiocarcinoma is a primary malignant tumor derived from the epithelium of the intrahepatic and extrahepatic bile ducts, and vasculogenic mimicry induced by hypoxia and lack of nutrition is a key factor for malignant proliferation, recurrence, and metastasis of cholangiocarcinoma. Previous studies have shown that autophagy maintains cell nutrition metabolism under the condition of a lack of nutrition, and preliminary experiments have confirmed that autophagy was associated with VM in cholangiocarcinoma and there was high expression of PAT4 in cholangiocarcinoma cells; on the basis of these studies, it is pointed out that in cholangiocarcinoma, protective autophagy regulates VM formation by maintaining intracellular metabolic balance and cellular homeostasis. As a nutrient sensor in tumor microenvironment, PAT4 mediates protective autophagy via the PI3K-Akt-mTORC1 signaling pathway to regulate VM formation. Histological, cellular molecular, and in vivo experiments have confirmed that autophagy regulates VM formation by maintaining cell metabolism, stem cell features, and extracellular matrix remodeling, which helps to investigate the signal mechanism for PAT4 mediating autophagy to regulate VM. It is suggested that autophagy is the source of energy and nutrition in cholangiocarcinoma cells under the condition of a lack of nutrition, and PAT4 is the trigger point for autophagy in regulating VM formation. These findings provide new thoughts for the metabolism of cholangiocarcinoma cells and lay a theoretical foundation for antiangiogenic drugs combined with autophagy inhibitors in the treatment of highly aggressive tumors.
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OBJECTIVE@#To isolate tumor stem-like cells from human epithelial ovarian cancer SKOV3 cells and explore their role in the formation of vascularization mimicry (VM).@*METHODS@#SKOV3 cells were passaged to the 7th generation by suspension culture in serum-free medium, and the percentages of CD133- and CD117-positive cells in the 1st, 3rd, 5th and 7th generations were analyzed using flow cytometry. The proliferative activity of the cells sorted from the 7th generation SKOV3 cells was assessed with colony formation assay. A three-dimensional cell culture model was established to compare the ability of VM formation between the sorted cells and the parental SKOV3 cells. The expression levels of matrix metalloproteinases-2 (MMP-2) and MMP-9 in the two groups were detected using real-time PCR and Western blotting.@*RESULTS@#Some SKOV3 cells formed typical cell spheres with suspension growth in serum-free medium and were passaged to the 7th generation. Flow cytometry revealed that the percentage of CD133-positive cells increased with cell passaging. The cloning efficiency of the sorted cells was significantly higher than that of the parental SKOV3 cells (50.33% 5.33%, < 0.001). The VM formation ability of the sorted cells was stronger than that of the parental SKOV3 cells in the three-dimensional cell culture system. RT-PCR and Western blotting showed that the expression levels of MMP-2 and MMP-9 were significantly higher in the 7th passage cells than in the parental cells ( < 0.05).@*CONCLUSIONS@#The sorted cells from SKOV3 cells cultured in serum-free medium exhibit biological properties of tumor stem cells with strong VM formation ability, suggesting their role in VM formation.
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Female , Humans , Carcinoma, Ovarian Epithelial , Pathology , Cell Line, Tumor , Cell Movement , Matrix Metalloproteinase 2 , Metabolism , Matrix Metalloproteinase 9 , Metabolism , Neoplastic Stem Cells , Cell Biology , Neovascularization, Pathologic , Pathology , Ovarian Neoplasms , PathologyABSTRACT
OBJECTIVE@#To investigate the inhibitory effects of silencing migration-inducing gene-7 (Mig-7) on vasculogenic mimicry formation, migration and invasion of human glioma cells and whether MEK/ERK signaling pathway mediates these effects.@*METHODS@#Human glioma U251 cells were infected by lentiviral vectors carrying a small interfering RNA targeting Mig-7 gene (sh-Mig-) or a negative control shRNA (sh-NC), and real-time quantitative PCR was used to detect the expression level of Mig- mRNA in the cells. Three-dimensional culture and Transwell chamber invasion assay were used to observe the effect of Mig- gene silencing on vasculogenic mimicry formation and invasion ability of the U251 cells. Western blotting was performed to detect the changes in the protein expression levels of MEK/ERK in the infected cells.@*RESULTS@#We successfully obtained a U251 cell line with stable low expression of Mig- gene using RNA interference technique. Compared with the cells infected with sh-NC lentivirus and the non- infected cells, U251 cells infected with the lentiviral vector carrying sh-Mig- showed significantly decreased expression level of Mig- ( < 0.01) with obviously lowered vasculogenic mimicry formation and invasion abilities ( < 0.05). Mig- silencing also significantly lowered the expressions of MEK and ERK proteins in U251 cells ( < 0.05).@*CONCLUSIONS@#Silencing of Mig-7 gene inhibits vasculogenic mimicry formation and invasion of U251 cells possibly by suppressing MEK/ERK signaling, suggesting the important role of Mig-7 gene in vasculogenic mimicry formation and invasion of human glioma cells.
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Humans , Cell Line, Tumor , Cell Movement , Cell Proliferation , Gene Silencing , Glioma , Genetics , Pathology , Neoplasm Proteins , Metabolism , RNA, Small Interfering , Signal TransductionABSTRACT
Objective To evaluate the effect of luteolin on the growth,migration and vasculogenic mimicry formation of a melanoma cell line B16.Methods In vitro cultured B16 melanoma cells were divided into 4 groups:low-,middle-and high-dose luteolin groups treated with 2.5,5,10 μmol/L luteolin respectively,and control group treated with 0.1％ dimethyl sulfoxide (DMSO).Scratch assay,Transwell invasion assay and vascular channel formation assay were performed to assess the migration,invasion of and vascular channel formation by melanoma cells.A model of subcutaneous transplanted B 16 melanoma was established in 12 C57 mice,which were randomly and equally divided into 4 groups:control group gavaged with ultrapure water,low-,middle-and high-dose luteolin groups gavaged with 10,20,40 mg/kg luteolin respectively every day.The above treatment for the tumor-bearing mice lasted till day 28,and then these mice were sacrificed.Meanwhile,the lung and tumor tissues of the mice were excised,and the growth,metastasis and vasculogenic mimicry of transplanted melanoma were observed.Immunofluorescence and immunohistochemical studies were performed to evaluate the effects of luteolin on the expression of vascular endothelial cadherin (VE-cadherin),vascular endothelial growth factor receptor 1 (VEGFR1),VEGFR2,matrix metalloproteinase-2 (MMP-2) and MMP-9 in the transplanted melanoma.Means were compared among several groups by using one-way analysis of variation or rank sum test.Results In vitro study showed that the relative scratch width at 48 hours significantly differed among the control group,low-,middle-and high-dose luteolin groups (0.47 ± 0.04,0.64 ± 0.04,0.73 ± 0.03,0.84 ± 0.04 respectively;F =34.51,P ＜ 0.001),and the migration ability of B16 cells was significantly lower in the low-,middle-and high-dose luteolin groups than in the control group (all P ＜ 0.05).At 24 hours,there were significant differences in the number of cells crossing the Transwell membrane among the control group,low-,middle-and high-dose luteolin groups (281.00 ± 8.79,169.00 ± 15.35,92.00 ± 14.79 and 57.00 ± 13.72 respectively;F =275.30,P ＜ 0.001),and the invasive ability was significantly lower in the low-,middle-and high-dose luteolin groups than in the control group (P ＜ 0.01).Meanwhile,the number of formed vascular channels also differed among the above 4 groups (20.00 ± 2.77,11.00 ± 1.28,7.00 ± 1.86 and 2.00 ± 1.32 respectively;F =48.61,P ＜ 0.001),and the number of vascular channels was significantly lower in the low-,middle-and high-dose luteolin groups than in the control group (all P ＜ 0.01).In vivo study showed that the tumor size significantly differed among the control group,low-,middle-and high-dose luteolin groups (5.10 ± 1.72,4.02 ± 2.13,2.98 ± 0.92,1.49 ± 1.13 cm3 respectively;F =28.76,P ＜ 0.001),and was significantly lower in the low-,middle-and high-dose luteolin groups than in the control group (t =3.86,7.11 and 13.06 respectively,all P ＜ 0.01).CD31-PAS double staining showed that the number of vasculogenic mimicry was significantly higher in the control group than in the low-,middle-and high-dose luteolin groups (all P ＜ 0.01).In vivo and in vitro studies both showed that the expression of vasculogenic mimicry-related markers in the cells or mouse tumor tissues was significantly lower in the high-dose luteolin group than in the control group (P ＜ 0.05).Conclusion Luteolin can effectively inhibit the growth,metastasis and vasculogenic mimicry formation of melanoma.
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Objective:To examine the expression of IQGAP1 in hepatocellular carcinoma and its effect on vasculogenic mimicry(VM). Methods:Immunohistochemical staining was performed to investigate the expression of IQGAP1.CD31/PAS double staining was per-formed to detect VM and analyze the correlation of IQGAP1 and VM.HepG2 cells were transfected with an IQGAP1 overexpression plasmid to induce exogenous expression of IQGAP1,and an IQGAP1 knockdown plasmid was transfected into SMMC7721 cells to re-duce IQGAP1 levels.The expression of cancer stem cell markers CD133,CD44,Sox2,and ALDH1 was analyzed by Western blot and compared with that in the control.Cellular functional experiments were used to determine the role of IQGAP1 in promoting cancer cells' ability of invasiveness and migration, proliferation, and VM formation. Results: Immunohistochemical analysis revealed that IQGAP1 was mainly located in the cell membrane and/or cytoplasm,and the staining intensity was correlated with tumor grade,me-tastasis,and VM(P<0.05).Cells transfected with the overexpression plasmid showed enhanced CD133,CD44,Sox2,and ALDH1 levels due to the increase in IQGAP1 and exhibited increased invasion ability,proliferation,and VM formation.In the SMMC7721 cells trans-fected with the knockdown plasmid,CD133,CD44,Sox2,and ALDH1 levels were decreased and motility was inhibited.Conclusions:IQGAP1 supports malignant behavior in hepatocellular carcinoma and may promote VM by increasing stemness.
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<p><b>Objective</b>To investigate the effect of Qilan Capsules (QLC) on the expressions of the related proteins HIF-1α, VEGF-α, EphA2 and MMP-1 in the formation of vasculogenic mimicry (VM) in prostate cancer.</p><p><b>METHODS</b>Prostate cancer PC-3 cells were cultured, transfected with siRNA, and divided into eight groups, blank control, HIF-1α siRNA, VEGF-α siRNA, EphA2 siRNA, QLC intervention, QLC + HIF-1α siRNA, QLC + VEGF-α siRNA, and QLC + EphA2 siRNA. The expressions of the HIF-1α, VEGF-α and EphA2 proteins in the pathway of VEGF were determined by Western blot.</p><p><b>RESULTS</b>Compared with the blank control group, the expression of HIF-1α was evidently decreased in the HIF-lα siRNA and QLC + HIF-lα siRNA groups (0.624 7 ± 0.042 8 vs 0.032 8 ± 0.002 5 and 0.036 8 ± 0.018 1, P < 0.05), so were that of VEGF-α in the VEGF-α siRNA and QLC + VEGF-α siRNA groups (0.068 9 ± 0.005 1 vs 0.016 9 ± 0.000 7 and 0.010 9 ± 0.000 8, P < 0.05), that of EphA2 in the EphA2 siRNA and QLC + EphA2 siRNA groups though with no statistically significant difference (0.1684 ± 0.0126 vs 0.134 5 ± 0.028 6 and 0.165 4 ± 0.039 8, P > 0.05), and that of MMP-1 in the HIF-lα siRNA, VEGF-α siRNA and EphA2 siRNA groups (1.696 1 ± 0.152 7 vs 0.435 9 ± 0.036 9, 0.198 7 ± 0.009 0 and 0.0218 ± 0.000 7, P < 0.05).</p><p><b>CONCLUSIONS</b>Qilan Capsules can suppress VM formation in prostate cancer by inhibiting the expressions of HIF-1α, VEGF-α and MMP-1, which plays a role in the clinical treatment of prostate cancer by checking the growth and development of the blood supply system in the tumor tissue.</p>
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Humans , Male , Capsules , Drugs, Chinese Herbal , Pharmacology , Hypoxia-Inducible Factor 1, alpha Subunit , Metabolism , Matrix Metalloproteinase 1 , Metabolism , Molecular Mimicry , Prostatic Neoplasms , Metabolism , RNA, Small Interfering , Metabolism , Receptor, EphA2 , Metabolism , Transfection , Vascular Endothelial Growth Factor A , MetabolismABSTRACT
Objective To detect the presence of vasculogenic mimicry (VM) in endometrial carcinoma (EC) tissue specimens,and to investigate the expression and relationship between VM and vascular endothelial growth factor (VEGF) in EC,and discuss the relationship between the two and the survival of patients after surgery.Methods The expression of VM in EC tissue,atypical hyperplasia and normal endometrial tissue was detected by CD34-PAS double staining.The expression of VEGF protein was detected by immunohistochemical SP method,and the relationship between the clinicopathological factors and the survival of EC patients was analyzed.Results The positive expression rates of VM and VEGF were 39.59％ and 68.02％ in EC tissues,4.71％ and 21.18％ in atypical hyperplasia endometrium respectively,1.59％ and 4.76％ in normal endometrium respectively,the expression of the two in the EC group were statistically significant compared with the other groups(x2 =60.668,P ＜ 0.05;x2 =98.976,P ＜ 0.05).Both V M and VEGF protein were associated with surgical-pathological stage and histological differentiation (P ＜ 0.05),positive correlation was found between them (r =0.293,P =0.000).Kaplan-Meier univariate analysis of the prognosis of patients with EC showed that the 5-year overall survival rates of VM-positive and VEGF-positive patients were 66.67％ and 79.t0％ respectively,which were lower than 95.80％ and 95.24％ in VM negative and VEGF negative patients(P all ＜ 0.05).Cox multivariate analysis of prognostic factors in EC patients showed that surgical-pathological stage,histological differentiation,lymph node metastasis,VM positive and overexpression of VEGF were significant factors influencing EC prognosis.Conclusion VM was present in EC tissue specimens.Combined detection of VM and VEGF plays an important role in the invasion,metastasis and prognosis of EC patients,both of which may serve as a reference for the diagnosis and evaluation of EC prognosis.
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<p><b>OBJECTIVE</b>To observe effects of on hepatocarcinoma (HCC) cell vasculogenic mimicry (VM) and explore the molecular mechanism by which inhibits HCC metastasis and invasion.</p><p><b>METHODS</b>Forty male SD rats were randomly divided into 4 groups for gastric lavage of normal saline or high, moderate or low doses of (twice daily) for 4 consecutive days. The sera were collected from the rats for treatment of cultured human HCC HepG2 cells. VM formation in the cells was detected using an image acquisition and analysis system 24 h after incubation of the cells with the sera and with the RhoA/ROCK inhibitor Y-27632(P). The expression levels of RhoA and ROCK1 in the cells were detected using Western blotting, and the contents of VE-cadherin and PI3K in the culture supernatant were determined using ELISA.</p><p><b>RESULTS</b>Treatment with the sera from -treated rats significantly inhibited formation of VM in HepG2 cells, and the diameters of VM formed were significantly greater than those in the positive control group ( < 0.01). Y-27632 completely inhibited the formation of VM in HepG2 cells ( < 0.01). Treatments with and Y-27632 both inhibited the expression of RhoA and ROCK1 ( < 0.05) and significantly lowered the contents of VE-cadherin and PI3K in the culture supernatant ( < 0.05).</p><p><b>CONCLUSIONS</b> can inhibit the formation of VM in HCC cells possibly by inhibiting the RhoA/ROCK pathways and the expressions of VE-cadherin and PI3K.</p>
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Objective To observe the effects of curcumin (Cur) on vasculogenic mimicry (VM) and PI3K pathway in choroid melanoma cell line OCM-1 in vitro 3-dimentional culture.Methods Three-dimensional culture of OCM-1 cells was performed for the observation of VM formation and the influence of concentration gradient (0μmol · L-1,10 μmol · L-1,20 μmol · L-1,30 μmol · L-1,40 μmol · L-1) Cur on VM,and immunohis to chemistry staining was used to detect the expression of PI3K and EphA2 in OCM-1 cell line.Results Cur could markedly inhibit the formation of VM in choroid melanoma cell line OCM-1 in vitro 3-dimensional culture with a dose-dependent manner.Cur suppressed EphA2 and PI3K expression in a concentration-dependent manner,and the difference between groups with 0 μmol · L-1 Cur group was significant statistically (all P =0.000),but there was no significant difference between 30 μmol · L-1and40 μmol · L-1 group (EphA2,P=0.300;PI3K,P=0.188).Conclusion Cur can significantly inhibit VM formation in choroid melanoma cell line OCM-1,of which the mechanism may be related to the downregulation expression of EphA2 and PI3K,thus inhibiting PI3K signaling pathway.
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Objective To observe vasculogenic mimicry (VM) of human choroidal melanoma cell line OCM-1 cultured in vitro and the expression of PI3K and EphA2 protein,as well as to explore the possible mechanisms.Methods OCM-1 cells were cultured in vitro and stained with periodic acid Schiff (PAS) on days 7,which aimed to observe the formation of PAS-positive cyclic structures,that is,VM formation.Then immunohistochemical staining was performed to detect PI3K and EphA2 on day 1,4,7 and the results were observed.Ressults On day 4 of 3-demintional culture,most of OCM-1 cells were polygonal and the cytoplasm was abundant;the nuclei were round and the nucleoli were visible.A small part of the tumor cells were long spindle.It was found that several long spindle cells were connected to each other to form hemicyclic structure.After 7 days,a large number of tumor cells became long spindle,growing along the collagen scaffold,and long protrusions appeared,forming a ring structure.PAS staining showed that the tumor cells were mostly arranged in a row,and tumor cells imitated the formation of body blood vessels,resulting in cell band and pipeline-like cell layers,with one layer of extracellular matrix (PAS-positive substance) making up the ring structure.Moreover,the expression levels of PI3K and EphA2 on day 4 and 7 were significantly higher than those on day 1 (all P ＜ 0.05),and their expression levels on day 7 were higher than those on day 4 (all P ＜ 0.05).Conclusion EphA2 and PI3K may play an important role in the VM formation in 3-dimentional culture of human choroidal melanoma cell line OCM-1.
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Objective To explore the effect of fibroblast growth factor receptor(FGFR)inhibitor BGJ398 on the vasculogenic mimicry(VM)formation of glioma cells. Methods The phosphor-FGFR(pFGFR)was de-tected by Western blot,the expressions of MMP2 and MMP14 were detected by Western blot and immunocytochem-istry;the VM formation of U87MG and U251MG was tested by tube formation assay;subcutaneously implanted tu-mor model in nude mouse was established and tumor sections were CD34/PAS double-stained to detect the forma-tion of VM in vivo. Results Western blot showed that pFGFR in the experimental groups decreased significantly (P < 0.05);western blot and immunohistochemical staining showed that the expression of MMP2 and MMP14 in the experimental groups decreased significantly compared to the control group. In the tube formation assay ,the tube formation of U87MG and U251MG cells were restrained. In the subcutaneously implanted tumor model ,the VM number of the experimental group(13.85 ± 3.96)was significantly lower than that in the control group(26.40 ± 5.06,P < 0.05). Conclusions In vivo and in vitro experiments confirmed that BGJ398 can inhibit the activa-tion of FGFR,and inhibit the VM formation of glioma cells. These indicate FGFR signaling pathway is involved in the formation of VM.