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1.
Oncol Rep ; 42(4): 1475-1486, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31364740

RESUMO

Gallbladder cancer (GBC) is a lethal aggressive malignant neoplasm of the biliary tract. Potential prognostic markers and therapeutic targets for this disease are urgently required. Cancer­associated fibroblasts (CAFs) play a key role in tumorigenesis and the development of cancer. Nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1) expression has been reported to be involved in tumorigenesis and useful for tumor prognosis. However, NOX1 expression in the stroma of GBCs, particularly gallbladder cancer­associated fibroblasts (GCAFs), and its prognostic significance in GBC patients remains unclear. In the present study, NOX1 expression in the stroma of human gallbladder lesions in vivo was investigated, as well as in GCAFs and co­cultures of GBC­SD+GCAFs in vitro, and their correlation with clinicopathological parameters and the prognosis of GBC patients were evaluated. The results revealed that NOX1 expression was significantly upregulated in the stroma of GBCs compared with precancerous and benign lesions of the gallbladder; NOX1 expression was localized to gallbladder stromal fibroblasts expressing α­smooth muscle actin and fibroblast secreted protein­1. Furthermore, these observations were confirmed by the fact that NOX1 expression was upregulated in GCAFs as determined by Affymetrix gene profile chip analysis and reverse transcription­quantitative PCR. In addition, overexpression was observed in formed spheroids of GBC­SD+GCAF co­cultures by immunohistochemistry and western blotting in vitro. Thus, it was verified that NOX1 expression was upregulated in GCAFs. Furthermore, upregulated stromal NOX1 expression was correlated with aggressive characteristics such as differentiation degree (P=0.042), venous invasion (P=0.041), resection methods (P=0.002), and a lower survival rate (P=0.025, log­rank test) of patients with GBC. Stromal NOX1 expression (P=0.047) was an independent prognostic factor for the overall survival rate of patients with GBC. GBC patients with upregulated NOX1 expression in GCAFs had a poorer prognosis. These results revealed that stromal NOX1 may be a novel biomarker and/or target, and may contribute to the discovery of new tumor markers and potential targeted therapeutics for human GBCs.


Assuntos
Fibroblastos Associados a Câncer/enzimologia , Neoplasias da Vesícula Biliar/enzimologia , NADPH Oxidase 1/biossíntese , Actinas/biossíntese , Idoso , Proteínas de Ligação ao Cálcio/biossíntese , Fibroblastos Associados a Câncer/patologia , Feminino , Neoplasias da Vesícula Biliar/patologia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Prognóstico , Regulação para Cima
2.
Oncol Lett ; 17(3): 3055-3065, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30867734

RESUMO

Adequate blood supply is essential for tumor survival, growth and metastasis. The tumor microenvironment (TME) is dynamic and complex, comprising cancer cells, cancer-associated stromal cells and their extracellular products. The TME serves an important role in tumor progression. Cancer-associated fibroblasts (CAFs) are the principal component of stromal cells within the TME, and contribute to tumor neo-angiogenesis by altering the proteome and degradome. The present paper reviews previous studies of the molecular signaling pathways by which CAFs promote tumor neo-angiogenesis and highlights therapeutic response targets. Also discussed are potential strategies for antitumor neo-angiogenesis to improve tumor treatment efficacy.

3.
J Invest Surg ; 32(8): 689-696, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29693474

RESUMO

Objective: The objectives of this study were to examine the clinical profile of critically ill patients with septic acute kidney injury (AKI) and to investigate clinical characteristics associated with the outcome of patients. Methods: Data from 582 critically ill patients were collected and retrospectively reviewed. Patients were divided into two groups: without AKI development and with AKI development. Baseline characteristics, laboratory, and other clinical data were compared between these two groups, and correlations between the characteristics and AKI development were examined. Patients with AKI development were further divided into two groups according to the survival outcome, and variables associated with the outcome were determined. Results: AKI was developed in 54.12% (n = 315) of patients, and these patients had blood pressure, SOFA score, APACHE II score, GCS, and various blood chemistry and hematology characteristics significantly different from the patients without AKI. Demographic characteristics (e.g. age and weight) were comparable between the two groups of patients. Among the 315 patients with AKI, 136 of them died during the study period. Multivariate logistic regression analysis revealed that the outcome of patients was associated with lung infection, coagulation system dysfunction, staphylococcus aureus infection, and use of various treatments (epinephrine, norepinephrine, and the use of mechanical ventilation) after AKI development. Conclusion: AKI occurred in approximately half of the critically ill patients admitted to ICU. The site and type of infections, as well as the use of vasopressor agents, were associated with the outcome.

4.
Oncotarget ; 8(30): 49897-49914, 2017 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-28611292

RESUMO

Human gallbladder cancer (GBC) is a lethal aggressive malignant neoplasm. Identification of potential molecular biomarkers and development of targeted therapeutics for GBC patients is very necessary. In this study, we firstly investigated the correlation between ring finger protein 125 (RNF125) expression and the metastasis and prognosis of GBC, and the underlying molecular mechanism. RNF125 expression in a cohort of GBC tissues was examined; its correlation with clinicopathological and prognostic factors of GBC patients was analyzed. Moreover, the metastasis-related difference expressed genes in highly and lowly aggressive GBC cell lines were identified; and the influence of RNF125 knockdown on the metastatic phenotypes and characteristic EMT markers in highly aggressive GBC NOZ cells was detected. Furthermore, the underlying molecular mechanism of RNF125 effect was explored. The results showed that RNF125 was highly expressed in GBC tissues and related with aggressive characteristics such as Nevin stage (P = 0.041) etc. and unfavorable prognosis of GBC patients (P = 0.023, log-rank test). And, RNF125 was proved to a positive metastasis-related gene in vitro. RNF125 knockdown inhibited the invasion and migration, enhanced the adhesion, upregulated E-cadherin and ß-catenin expression, and downregulated vimentin and N-cadherin expression (all P < 0.001) of NOZ cells in vitro. RNF125 promoting effect on GBC tumor progression was identified to relate with the activation of TGF-ß1-SMAD3-ID1 signaling pathway. These findings firstly confirm that high RNF125 expression is related with aggressive characteristics and unfavorable prognosis of GBC patients; RNF125 promotes the invasion and metastasis of human GBCs via activating the TGF-ß1-SMAD3-ID1 signaling pathway.


Assuntos
Neoplasias da Vesícula Biliar/metabolismo , Neoplasias da Vesícula Biliar/mortalidade , Proteína 1 Inibidora de Diferenciação/metabolismo , Transdução de Sinais , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais , Adesão Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Biologia Computacional/métodos , Transição Epitelial-Mesenquimal/genética , Feminino , Neoplasias da Vesícula Biliar/patologia , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , Gradação de Tumores , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Carga Tumoral , Ubiquitina-Proteína Ligases/genética
5.
Cancer Cell Int ; 17: 20, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28194091

RESUMO

BACKGROUND: Human gallbladder cancer (GBC) is an aggressive malignant neoplasm with a poor prognosis. The development of ideal tools for example tumor cell lines for investigating biological behavior, metastatic mechanism and potential treatment in GBCs is essential. In present study, we established and characterized a GBC cell line derived from primary tumor. METHODS: Primary culture method was used to establish this cell line from a primary GBC. Light and electron microscopes, flow cytometry, chromosome analysis, heterotransplantation and immunohistochemistry were used to characterize the epidemic tumor characteristics and phenotypes of this cell line. RESULTS: A novel GBC cell line, named TJ-GBC2, was successfully established from primary GBC. This cell line had characteristic epithelial tumor morphology and phenotypes in consistent with primary GBC, such as polygon and irregular cell shape, increased CA19-9 and AFP levels, and positive expression of CK7, CK8, CK19 and E-cadherin with negative vimentin. Moreover, about 25% of the cells were in the S-G2/M phase; abnormity in structure and number of chromosome with a peak number of 90-105 and 80% hypertetraploid were observed. Furthermore, this cell line had higher invasion and highest migration abilities compared to other GBC cell lines; and metastatic-related marker MMP9 and nm23 were positively expressed. CONCLUSIONS: A novel highly aggressive GBC cell line TJ-GBC2 was successfully established from primary GBC. TJ-GBC2 cell line may be efficient tool for further investigating the biological behaviors, metastatic mechanism and potential targeted therapy of human GBC.

6.
BMC Cancer ; 15: 527, 2015 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-26187792

RESUMO

BACKGROUND: Tumor lymphangiogenesis plays an important role in promoting growth and metastasis of tumors, but no antilymphangiogenic agent is used clinically. Based on the effect of norcantharidin (NCTD) on lymphangiogenesis of human lymphatic endothelial cells (LECs), we firstly investigated the antilymphangiogenic activity of NCTD as a tumor lymphangiogenic inhibitor for human colonic adenocarcinomas (HCACs). METHODS: In vivo and in vitro experiments to determine the effects of NCTD on tumor growth and lymphangiogenesis of the in-situ colonic xenografts in nude mice, and lymphatic tube formation of the three-dimensional (3-D) of the co-culture system of HCAC HT-29 cells and LECs were done. Proliferation, apoptosis, migration, invasion, Ki-67, Bcl-2 and cell cycle of LECs and the co-culture system in vitro were respectively determined. Streparidin-peroxidase staining, SABC, western blotting and RT-PCR were respectively used to examine the expression of LYVE-1, D2-40, CK20 (including their LMVD), and VEGF-A, VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 in vitro and in vivo. RESULTS: NCTD inhibited tumor growth and lymphangiogenesis of the in-situ colonic xenografts in vivo, and these observations were confirmed by facts that lymphatic tube formation, proliferation, apoptosis, migration, invasion, S-phase cell cycle, and Ki-67 and Bcl-2 expression in vitro, and LYVE-1, D2-40, CK20 expression and their LMVD in vitro and in vivo were inhibited and affected. Furthermore, the expression of VEGF-A, VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 at protein/mRNA levels in the process of lymphatic tube formation in vitro and tumor lymphangiogenesis in vivo was downregulated; NCTD in combination with mF4-31C1 or Sorafenib enhanced these effects. CONCLUSIONS: NCTD inhibits tumor growth and lymphangiogenesis of HCACs through "multi-points priming" mechanisms i.e. affecting related malignant phenotypes, inhibiting Ki-67 and Bcl-2 expression, inducing S-phase cell cycle arrest, and directly or indirectly downregulating VEGF-A,-C,-D/VEGFR-2,-3 signaling pathways. The present finding strongly suggests that NCTD could serve as a potential antilymphangiogenic agent for tumor lymphangiogenesis and is of importance to explore NCTD is used for antitumor metastatic comprehensive therapy for HCACs.


Assuntos
Inibidores da Angiogênese/administração & dosagem , Antineoplásicos/administração & dosagem , Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Neoplasias do Colo/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Linfangiogênese/efeitos dos fármacos , Metástase Linfática/prevenção & controle , Inibidores da Angiogênese/farmacologia , Animais , Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células Endoteliais/citologia , Células HT29 , Humanos , Técnicas In Vitro , Camundongos , Camundongos Nus , Receptores de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Int J Oncol ; 46(2): 627-40, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25405519

RESUMO

Vasculogenic mimicry (VM) is a tumor microcirculation pattern in highly aggressive gallbladder cancers (GBCs). We recently reported the anti­VM activity of norcantharidin (NCTD) in highly aggressive GBC­SD cells and xenografts. In this study, we further investigated that NCTD enhanced tissue inhibitor of matrix metalloproteinase­2 (TIMP­2) anti­VM activity for GBCs and the underlying mechanisms. In vivo and in vitro experiments were performed to determine the effects of NCTD in combination with TIMP­2 on tumor growth, host survival, VM formation, hemodynamic of GBC­SD xenografts, and VM­like networks and malignant phenotypes of GBC­SD cells. Expression of matrix metalloproteinase (MMP)­2 and membrane type 1­MMP (MT1­MMP) among human GBCs, GBC­SD cells and xenografts were determined, respectively. The results showed that expression of MMP­2 and MT1­MMP in human GBCs, GBC­SD cells and xenografts was significantly related to VM in GBCs; a shorter survival time of VM­positive patients with high expression of MMP­2 or MT1­MMP compared to that of the patients with low expression. After treatment with NCTD+TIMP­2, tumor growth, VM formation, VM hemodynamic of the xenografts in vivo were significantly inhibited as compared to control, NCTD or TIMP­2 group, with a prolonged survival time of the xenograft mice (log­rank test, P=0.0115); and these observations were confirmed by VM­like networks by 3­D matrices and showed that proliferation, apoptosis, invasion, migration of GBC­SD cells in vitro were markedly affected. Furthermore, expression of MMP­2 and MT1­MMP in VM formation of the xenografts in vivo and GBC­SD cells in vitro was downregulated as compared to control, NCTD or TIMP­2 group. Thus, we concluded that NCTD enhances TIMP­2 antitumor and anti­VM activities in GBCs through downregulating MMP­2 and MT1­MMP.


Assuntos
Neoplasias da Vesícula Biliar/genética , Metaloproteinase 14 da Matriz/biossíntese , Metaloproteinase 2 da Matriz/biossíntese , Neovascularização Patológica/genética , Inibidor Tecidual de Metaloproteinase-2/biossíntese , Animais , Apoptose/efeitos dos fármacos , Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias da Vesícula Biliar/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
8.
PLoS One ; 9(5): e96982, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24811250

RESUMO

Vasculogenic mimicry (VM) is a newly-defined tumor microcirculation pattern in highly aggressive malignant tumors. We recently reported tumor growth and VM formation of gallbladder cancers through the contribution of the ephrin type a receptor 2 (EphA2)/focal adhesion kinase (FAK)/Paxillin signaling pathways. In this study, we further investigated the anti-VM activity of norcantharidin (NCTD) as a VM inhibitor for gallbladder cancers and the underlying mechanisms. In vivo and in vitro experiments to determine the effects of NCTD on tumor growth, host survival, VM formation of GBC-SD nude mouse xenografts, and vasculogenic-like networks, malignant phenotypes i.e., proliferation, apoptosis, invasion and migration of GBC-SD cells. Expression of VM signaling-related markers EphA2, FAK and Paxillin in vivo and in vitro were examined by immunofluorescence, western blotting and real-time polymerase chain reaction (RT-PCR), respectively. The results showed that after treatment with NCTD, GBC-SD cells were unable to form VM structures when injecting into nude mouse, growth of the xenograft was inhibited and these observations were confirmed by facts that VM formation by three-dimensional (3-D) matrix, proliferation, apoptosis, invasion, migration of GBC-SD cells were affected; and survival time of the xenograft mice was prolonged. Furthermore, expression of EphA2, FAK and Paxillin proteins/mRNAs of the xenografts was downregulated. Thus, we concluded that NCTD has potential anti-VM activity against human gallbladder cancers; one of the underlying mechanisms may be via blocking the EphA2/FAK/Paxillin signaling pathway.


Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Neoplasias da Vesícula Biliar/irrigação sanguínea , Neoplasias da Vesícula Biliar/patologia , Microcirculação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Antineoplásicos/uso terapêutico , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Efrina-A2/metabolismo , Feminino , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Neoplasias da Vesícula Biliar/tratamento farmacológico , Humanos , Masculino , Camundongos , Paxilina/metabolismo , Análise de Sobrevida , Ensaios Antitumorais Modelo de Xenoenxerto
9.
BMC Cancer ; 14: 193, 2014 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-24628713

RESUMO

BACKGROUND: Vasculogenic mimicry (VM) is a novel tumor blood supply in some highly aggressive malignant tumors. Recently, we reported VM existed in gallbladder carcinomas (GBCs) and the formation of the special passage through the activation of the PI3K/MMPs/Ln-5γ2 signaling pathway. GBC is a highly aggressive malignant tumor with disappointing treatments and a poor prognosis. Norcantharidin (NCTD) has shown to have multiple antitumor activities against GBCs, etc; however the exact mechanism is not thoroughly elucidated. In this study, we firstly investigated the anti-VM activity of NCTD as a VM inhibitor for GBCs and its underlying mechanisms. METHODS: In vitro and in vivo experiments to determine the effects of NCTD on proliferation, invasion, migration, VM formation, hemodynamic and tumor growth of GBC-SD cells and xenografts were respectively done by proliferation, invasion, migration assays, H&E staining and CD31-PAS double stainings, optic/electron microscopy, tumor assay, and dynamic micro-MRA. Further, immunohistochemistry, immunofluorescence, Western blotting and RT-PCR were respectively used to examine expression of VM signaling-related markers PI3-K, MMP-2, MT1-MMP and Ln-5γ2 in GBC-SD cells and xenografts in vitro and in vivo. RESULTS: After treatment with NCTD, proliferation, invasion, migration of GBC-SD cells were inhibited; GBC-SD cells and xenografts were unable to form VM-like structures; tumor center-VM region of the xenografts exhibited a decreased signal in intensity; then cell or xenograft growth was inhibited. Whereas all of untreated GBC-SD cells and xenografts formed VM-like structures with the same conditions; the xenograft center-VM region exhibited a gradually increased signal; and facilitated cell or xenograft growth. Furthermore, expression of MMP-2 and MT1-MMP products from sections/supernates of 3-D matrices and the xenografts, and expression of PI3-K, MMP-2, MM1-MMP and Ln-5γ2 proteins/mRNAs of the xenografts were all decreased in NCTD or TIMP-2 group; (all P < 0.01, vs. control group); NCTD down-regulated expression of these VM signaling-related markers in vitro and in vivo. CONCLUSIONS: NCTD inhibited tumor growth and VM of human GBCs in vitro and in vivo by suppression of the PI3-K/MMPs/Ln-5γ2 signaling pathway. It is firstly concluded that NCTD may be a potential anti-VM agent for human GBCs.


Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Neoplasias da Vesícula Biliar/patologia , Neovascularização Patológica/patologia , Transdução de Sinais/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Neoplasias da Vesícula Biliar/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Metaloproteinases da Matriz/genética , Metaloproteinases da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias Experimentais , Neovascularização Patológica/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Int J Oncol ; 42(6): 2103-15, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23588386

RESUMO

Vasculogenic mimicry (VM) is a new tumor blood supply in some highly aggressive malignant tumors. We previously reported VM in human gallbladder carcinomas, 3-D matrices in vitro and nude mouse xenografts in vivo of highly aggressive GBC-SD cells and its clinical significance. In this study, we further studied the underlying mechanisms of VM in gallbladder carcinomas via the 3-D matrix in vitro, the nude mouse xenografts in vivo of GBC-SD or SGC-996 cells, immunohistochemistry (H&E staining and CD31-PAS double staining), electron microscopy, expression of MMP-2, MT1-MMP, PI3K, Ln-5γ2, EphA2, FAK and Paxillin-P proteins/mRNAs determined by SABC, ELISA, immunofluorescence, western blotting and qRT-PCR, respectively. It was shown that all of untreated highly aggressive GBC-SD cells and xenografts formed vasculogenic-like structures within 2 weeks of seeding and injecting, and facilitated the growth of tumor cells or xenografts; whereas poorly aggressive SGC-996 cells or GBC-SD cells treated by TIMP-2 were unable to form the vasculogenic-like structures with the same conditions; and tumor xenograft growth was inhibited. Expression of MMP-2, MT1-MMP proteins/mRNAs from sections and supernates of 3-D matrix in vitro, expression of PI3K, MMP-2, MT1-MMP, Ln-5γ2, EphA2, FAK and Paxillin-P proteins/mRNAs from sections of xenografts in vivo in untreated GBC-SD group was upregulated significantly (all P<0.001); however, expression of these VM signal-related proteins/mRNAs in the SGC-996 group and GBC-SD treated by the TIMP-2 group was significantly downregulated (all P<0.001). Thus, we identified for the first time that highly aggressive GBC-SD cells formed VM in vitro and in vivo through the upregulation of PI3K/MMPs/Ln-5γ2 and/or EphA2/FAK/Paxillin signaling. PI3K/MMPs/Ln-5γ2 and EphA2/FAK/Paxillin as key signaling pathways in a coordinated manner contributed to tumor growth and VM of gallbladder carcinomas and provided novel targets that could be potentially exploited for therapeutic intervention of human gallbladder carcinomas.


Assuntos
Quinase 1 de Adesão Focal/metabolismo , Neoplasias da Vesícula Biliar/irrigação sanguínea , Neoplasias da Vesícula Biliar/patologia , Metaloendopeptidases/metabolismo , Neovascularização Patológica/metabolismo , Paxilina/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Receptor EphA2/metabolismo , Animais , Feminino , Quinase 1 de Adesão Focal/genética , Neoplasias da Vesícula Biliar/metabolismo , Humanos , Laminina/genética , Laminina/metabolismo , Masculino , Camundongos , Camundongos Nus , Paxilina/genética , Fosfatidilinositol 3-Quinases/genética , Receptor EphA2/genética , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Asian Pac J Cancer Prev ; 13(6): 2823-8, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22938466

RESUMO

OBJECTIVE: To explore a new model of in-situ xenograft lymphangiogenesis of human colonic adenocarcinomas in nude mice. METHOD: On the basis of establishing subcutaneous xenograft lymphangiogenesis model of human colonic adenocarcinoms, in-situ xenografts were established through the in situ growth of the HT-29 human colonic adenocarcinoma cell line in nude mice. The numbers of lymphangiogenic microvessels, the expression of lymphatic endothelial cell markers lymphatic vessel endothelial hyaloronic acid receptor-1 (LYVE-1), D2-40 and the lymphatic endothelial growth factors vascular endothelial growth factor-C (VEGF-C), -D (VEGF-D) and receptor-3 (VEGFR-3) were compared by immunohistochemical staining, Western bolt and quantitative RT-PCR in xenograft in-situ models. RESULTS: Some microlymphatics with thin walls, large and irregular or collapsed cavities and increased LMVD, with strong positive of LYVE-1, D2-40 in immunohistochemistry, were observed, identical with the morphological characteristics of lymphatic vessels and capillaries. Expression of LYVE-1 and D2-40 proteins and mRNAs were significantly higher in xenografts in-situ than in the negative control group (both P<0.01). Moreover, the expression of VEGF-C, VEGF-D and VEGFR-3 proteins and mRNAs were significantly higher in xenografts in-situ (both P<0.01), in conformity with the signal regulation of the VEGF-C,-D/VEGFR-3 axis of tumor lymphangiogenesis. CONCLUSIONS: In-situ xenografts of a human colonic adenocarcinoma cell line demonstrate tumor lymphangiogenesis. This novel in-situ animal model should be useful for further studying mechanisms of lymph node metastasis, drug intervention and anti-metastasis therapy in colorectal cancer.


Assuntos
Adenocarcinoma/fisiopatologia , Neoplasias do Colo/fisiopatologia , Linfangiogênese , Vasos Linfáticos/metabolismo , Animais , Linhagem Celular Tumoral , Glicoproteínas/metabolismo , Humanos , Proteínas de Membrana Transportadoras , Camundongos , Camundongos Nus , Transplante de Neoplasias , Transplante Heterólogo , Fator C de Crescimento do Endotélio Vascular/metabolismo , Fator D de Crescimento do Endotélio Vascular/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismo
12.
Int J Oncol ; 41(5): 1762-72, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22922710

RESUMO

Lymph node metastasis of tumors is a crucial early step in the metastatic process. Tumor lymphangiogenesis plays an important role in promoting tumor metastasis to regional lymph nodes. Norcantharidin (NCTD) has been reported to possess potent anti-angiogenesis and antitumor properties in several cell lines and xenograft tumor models. However, its role in tumor-associated lymphangiogenesis and lymphatic metastasis remains unclear. Here, we investigated the effect of NCTD on proliferation, apoptosis, migration, invasion and the lymphatic tube formation, lymphangiogenesis, of human lymphatic endothelial cells (HLECs) in vitro by MTT, proliferation assay, Hoechst staining and flow cytometry, scraping line method, Matrigel invasion assay, inverted or fluorescence microscope and transmission electron microscope. Moreover, the underlying mechanisms, such as VEGF-C, VEGF-D, VEGFR-3 at protein and mRNA levels in lymphangiogenesis using 3-dimensional (3-D) culture of HLECs were measured by immunohistochemistry, western blotting and real-time polymerase chain reaction (RT-PCR). It was shown that NCTD inhibited proliferation, migration, invasion and lymphatic tube formation (forming-lymphatic and/or formed-lymphatic) of HLECs, induced HLEC apoptosis (all P<0.01) significantly, in a dose- and time-dependent manner (IC50 6.8 µg/ml); and downregulated the expression of VEGF-C, VEGF-D and VEGFR-3 at protein or/and mRNA levels (P<0.01) in HLEC lymphatic tube formation. Thus, we identified for the first time that NCTD inhibited HLEC lymphangiogenesis by simultaneously blocking VEGF-C and VEGF-D/VEGFR-3 in vitro. The present findings may be of importance to explore the therapeutical target or strategy of NCTD for tumor lymphangiogenesis and lymphatic metastasis.


Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Endotélio Linfático/efeitos dos fármacos , Linfangiogênese/efeitos dos fármacos , Fator C de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator D de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Endotélio Linfático/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Fator C de Crescimento do Endotélio Vascular/genética , Fator C de Crescimento do Endotélio Vascular/metabolismo , Fator D de Crescimento do Endotélio Vascular/genética , Fator D de Crescimento do Endotélio Vascular/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismo
13.
Oncol Rep ; 27(6): 1990-2002, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22470047

RESUMO

As a novel mode of tumor neovascularization, vasculogenic mimicry (VM) has been reported to increase tumor-related mortality in many different solid tumors. In the present study, two established human gallbladder carcinoma (GBC) cell lines (highly aggressive GBC-SD and poorly aggressive SGC-996) cultured on a three-dimensional matrix were assessed for the ability of VM channel formation under normoxic or hypoxic conditions. In addition, the relationship between HIF-1α gene expression and VM channel formation of GBC cells in vitro was measured using the small interfering RNA (siRNA) technique, western blotting and real-time reverse transcription (RT)-PCR analysis. Furthermore, H&E and CD31/periodic acid-Schiff (PAS) staining were used to observe VM in GBC tissue samples. Additionally, all seventy-one specimens with VM and non-VM were stained for hypoxia inducible factor-1 α (HIF-1α) and its correlation with clinicopathological features and prognosis was analyzed simultaneously. We found that hypoxia could induce more VM channel formation and elevated HIF-1α expression in highly aggressive GBC-SD cells. HIF-1α siRNA efficiently knocked down HIF-1α expression and GBC VM networks under either normoxic or hypoxic conditions. VM was present in human primary GBC and overexpression of HIF-1α was significantly correlated with depth of invasion and perineural involvement in the non-VM group. Moreover, VM and HIF-1α were independent factors for the overall survival of GBC patients and correlated with decreased survival. In conclusion, VM was present in human GBC. As a critical mediator in VM formation, high expression of HIF-1α was associated with VM and tumor progression in GBC patients.


Assuntos
Neoplasias da Vesícula Biliar/metabolismo , Neoplasias da Vesícula Biliar/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neovascularização Patológica , Adulto , Idoso , Idoso de 80 Anos ou mais , Hipóxia Celular , Linhagem Celular Tumoral , Feminino , Neoplasias da Vesícula Biliar/irrigação sanguínea , Neoplasias da Vesícula Biliar/mortalidade , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Pessoa de Meia-Idade , Prognóstico , Interferência de RNA , RNA Interferente Pequeno
14.
Int J Oncol ; 40(5): 1501-14, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22200632

RESUMO

Our objective was to explore the antiangiogenic activity of norcantharidin (NCTD) as an angiogenic inhibitor for gallbladder cancers. In vitro and in vivo experiments to determine the effects of NCTD on HUVECs, chicken CAM capillaries and gallbladder cancer xenograft angiogenesis in nude mice were respectively done. The MTT method was used to assay the cytotoxicity of NCTD on HUVECs. Immunofluorescence was used to evaluate HUVEC apoptosis. The scraping line method, matrigel invasion assay and tube formation assay were used to detect the migration, invasion and tube formation of HUVECs. A digital camera was used to observe chicken CAM capillaries. Experiments with NCTD in a xenograft model were used to observe the effect of NCTD on xenograft growth and survival of mice with xenografts. CD34 immunohistochemistry, flow cytometry and micro-MRA were used, respectively, to determine MVD, cell apoptosis and hemodynamic analysis of the xenografts. Immunohistochemistry and RT-PCR were used, respectively, to detect the expression of VEGF, Ang-2, TSP, TIMP-2 proteins/mRNAs of the xenografts. The xenograft MVD associated with tumor volume, the PCNA/apoptosis ratio and related-protein expression was evaluated simultaneously. We found that NCTD effectively inhibited the proliferation, migration, invasion and capillary-like tube formation of HUVECs in vitro; it reduced angiogenesis and directly destroyed the formed CAM capillaries in vivo. In the experiments in mice, NCTD not only inhibited significantly xenograft proliferation and growth, prolonged survival time of mice with xenografts, decreased the xenograft MVD and vascular perfusion, but also, similarly to ES, decreased significantly the expression of VEGF or Ang-2 protein/mRNA, increased the expression of TSP or TIMP-2 protein/mRNA. Moreover, the xenograft MVD was positively related with tumor volume, PCNA/apoptosis ratio, and VEGF or Ang-2 expression, respectively (all P<0.05), but negatively correlated with TSP or TIMP-2 expression (both P<0.05). These data showed that NCTD could serve as a potential antiangiogenic agent for gallbladder cancers.


Assuntos
Inibidores da Angiogênese/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Membrana Corioalantoide/irrigação sanguínea , Neoplasias da Vesícula Biliar/tratamento farmacológico , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Neovascularização Patológica/prevenção & controle , Neovascularização Fisiológica/efeitos dos fármacos , Angiopoietina-2/genética , Angiopoietina-2/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Embrião de Galinha , Relação Dose-Resposta a Droga , Feminino , Citometria de Fluxo , Neoplasias da Vesícula Biliar/irrigação sanguínea , Neoplasias da Vesícula Biliar/genética , Neoplasias da Vesícula Biliar/metabolismo , Neoplasias da Vesícula Biliar/patologia , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Imuno-Histoquímica , Angiografia por Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Imagem de Perfusão/métodos , Reação em Cadeia da Polimerase , Ratos , Fluxo Sanguíneo Regional , Trombospondinas/genética , Trombospondinas/metabolismo , Fatores de Tempo , Inibidor Tecidual de Metaloproteinase-2/genética , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Carga Tumoral/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Oncol Rep ; 26(5): 1133-42, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21750868

RESUMO

Astrocyte elevated gene-1 (AEG-1), as an HIV-1 or TNF-α-inducible transcript, is correlated with various aspects of tumor malignancy. However, the status of AEG-1 expression and its clinical significance in human gallbladder carcinoma (GBC) remains unknown. In the present study, we investigated AEG-1 expression in two GBC cell lines (GBC-SD and SGC-996) and GBC tissues by immunohistochemical, Western blot and real-time PCR analysis. We found that AEG-1 was highly expressed in GBC samples (63.4%, 26 of 41) compared with normal gallbladder mucosa (p=0.0003) and highly invasive GBC-SD cell lines at both the protein (p=0.0043) and mRNA levels (p=0.0001), and strongly correlated with differentiation degree (p=0.006), Nevin stage (p=0.0344), Ki-67 expression (p=0.0024) and liver infiltration (p=0.0332) in these patients. Multivariate analysis indicated that AEG-1 overexpression was an independent prognostic marker for GBC patients. Moreover, patients with high AEG-1 levels had shorter survival time (p=0.008). Our results suggest that the AEG-1 protein is a valuable marker of GBC progression and could be a potential therapeutic target.


Assuntos
Biomarcadores Tumorais/biossíntese , Moléculas de Adesão Celular/biossíntese , Neoplasias da Vesícula Biliar/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Western Blotting , Moléculas de Adesão Celular/genética , Feminino , Neoplasias da Vesícula Biliar/genética , Neoplasias da Vesícula Biliar/patologia , Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Regulação para Cima
16.
World J Gastroenterol ; 17(4): 540-2, 2011 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-21274386

RESUMO

Granulocytic sarcoma (GS) is an extramedullary tumor mass consisting of immature myeloid cells. Isolated pancreatic granulocyte sarcoma is extremely rare. We report a very unusual pancreatic granulocytic sarcoma in a patient without acute myeloid leukemia. The patient presented with acute epigastric pain because of splenic infarction due to a mass consisting of myeloblasts in the pancreatic tail. The patients underwent splenectomy and distal pancreatectomy. Pathology and immunohistochemistry suggested a GS. Despite local surgery, an isolated tumor recurred 2 mo after operation and the patient died 3 mo after removal of the tumor. Only 7 reported cases of pancreatic GS were identified in the literature and the mass was located in the pancreatic head. This is the first report of GS in the pancreatic tail with splenic infarction.


Assuntos
Neoplasias Pancreáticas/diagnóstico , Sarcoma Mieloide/diagnóstico , Evolução Fatal , Feminino , Humanos , Pessoa de Meia-Idade , Pâncreas/patologia , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/cirurgia , Sarcoma Mieloide/patologia , Sarcoma Mieloide/cirurgia
17.
World J Gastrointest Surg ; 2(4): 117-27, 2010 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-21160860

RESUMO

"Vasculogenic mimicry (VM)", is a term that describes the unique ability of highly aggressive tumor cells to express a multipotent, stem cell-like phenotype, and form a pattern of vasculogenic-like networks in three-dimensional culture. As an angiogenesis-independent pathway, VM and/or periodic acid-schiff-positive patterns are associated with poor prognosis in tumor patients. Moreover, VM is resistant to angiogenesis inhibitors. Here, we will review the advances in research on biochemical and molecular signaling pathways of VM in tumors and on potential anti-VM therapy strategy.

18.
Hepatobiliary Pancreat Dis Int ; 9(4): 414-22, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20688607

RESUMO

BACKGROUND: Gallbladder carcinoma, a lethal malignant neoplasm with poor prognosis, has dismal results of surgical resection and chemoradiotherapy. We previously reported that norcantharidin (NCTD) is useful against growth, proliferation, and invasion of human gallbladder carcinoma GBC-SD cells in vitro. In this study, we further studied the inhibitory effect of NCTD on the growth of xenografted tumors of human gallbladder carcinoma in nude mice in vivo and the underlying mechanisms. METHODS: The tumor xenograft model of human gallbladder carcinoma in nude mice in vivo was established with subcutaneous GBC-SD cells. The experimental mice were randomly divided into control, 5-FU, NCTD, and NCTD+5-FU groups which were given different treatments. Tumor growth in terms of size, growth curve, and inhibitory rate was evaluated. Cell cycle, apoptosis, and morphological changes of the xenografted tumors were assessed by flow cytometry and light/electron microscopy. The expression of the cell cycle-related proteins cyclin-D1 and p27 as well as the apoptosis-related proteins Bcl-2, Bax, and survivin were determined by the streptavidin-biotin complex (SABC) method and RT-PCR. RESULTS: NCTD inhibited the growth of the xenografted tumors in a dose- and time-dependent manner. Tumor volume decreased (5.61+/-0.39 vs. 9.78+/-0.61 cm3, P=0.000) with an increased tumor inhibitory rate (42.63% vs. 0%, P=0.012) in the NTCD group compared with the control group. The apoptosis rate increased (15.08+/-1.49% vs. 5.49+/-0.59%, P=0.0001) along with a decreased percentage of cells in S phase (43.47+/-2.83% vs. 69.85+/-1.96%, P=0.0001) in the NTCD group compared with the control group. The morphological changes of apoptosis such as nuclear shrinkage, chromatin aggregation, chromosome condensation, and typical apoptosis bodies in the xenografted tumor cells induced by NCTD were observed by light and electron microscopy. The expression of cyclin-D1, Bcl-2 and survivin proteins/mRNAs decreased significantly, with increased expression of p27 and Bax proteins/mRNAs in the NCTD group compared with the control group. CONCLUSION: NCTD inhibits the growth of xenografted tumors of human gallbladder carcinoma in nude mice by inducing apoptosis and blocking the cell cycle in vivo.


Assuntos
Apoptose/efeitos dos fármacos , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Carcinoma/patologia , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias da Vesícula Biliar/patologia , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Carcinoma/metabolismo , Linhagem Celular Tumoral , Cromatina/efeitos dos fármacos , Cromatina/ultraestrutura , Ciclina D1/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Neoplasias da Vesícula Biliar/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fatores de Tempo , Proteína X Associada a bcl-2/metabolismo
19.
Zhonghua Wei Chang Wai Ke Za Zhi ; 12(5): 511-4, 2009 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-19742347

RESUMO

OBJECTIVE: To examine the expression of lymph vessel endothelial hyaluronan receptor-1 (LYVE-1) in human colorectal carcinoma and to evaluate the relationship of LYVE-1 with lymph mode metastasis and prognosis. METHODS: Colonic cancer samples of 40 cases were collected. The expression of LYVE-1 was determined by RT-PCR and quantified by real-time quantitative PCR. LVD and MVD were detected by immunohistochemistry staining. The relationship of LYVE-1 and LVD with lymph mode metastasis and prognosis were analyzed. All the patients were followed up for at least 3 years. RESULTS: The expression of LYVE-1 and the count of LVD were significantly higher in tumor tissue than those in common colon tissue (P<0.05). In the majority of tumors, the higher count of LVD indicated lymphangiogenesis. The recurrence rates in low LVD group and high LVD group were 46.7% and 60.0% respectively (P<0.05). The survival rates in the above two groups were 90.1% and 56.7% respectively (P<0.05). No significant correlation was found between LYVE-1 and recurrence rate (P>0.05) or overall survival (P>0.05). CONCLUSION: LYVE-1 indicates an increase of lymphangiogenesis in colorectal carcinoma and LVD can be used to evaluate the prognosis for colorectal cancer patients.


Assuntos
Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas de Transporte Vesicular/metabolismo , Adulto , Idoso , Feminino , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Prognóstico , RNA Mensageiro/genética
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