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1.
J Pathol ; 251(2): 123-134, 2020 06.
Artículo en Inglés | MEDLINE | ID: mdl-32166747

RESUMEN

Spontaneously regressing infantile haemangiomas and aggressive angiosarcomas are vascular tumours with excessive angiogenesis. When analysing haemangiomas and angiosarcomas immunohistochemically with respect to their chaperone profiles we found that angiosarcomas have significantly elevated protein levels of binding immunoglobulin protein (BIP) and PERK with concomitant attenuated IRE1α levels, whereas haemangioma tissue exhibits the same pattern as embryonal skin tissue. We show that BiP is essential for the maintenance of VEGFR2 protein, which is expressed in the endothelium of both tumour types. When studying the effects of BiP, the IRE1α/Xbp1 -, and PERK/ATF4-signalling pathways on the migration and tube-forming potential of endothelial cells, we show that downregulation of BiP, as well as inhibition of the kinase activity of IRE1α, inhibit in vitro angiogenesis. Downregulation of PERK (PKR-like kinase; PKR = protein kinase R) levels promotes Xbp1 splicing in endoplasmic reticulum (ER)-stressed cells, indicating that in angiosarcoma the elevated PERK levels might result in high levels of unspliced Xbp1, which have been reported to promote cell proliferation and increase tumour malignancy. The data presented in this study revealed that in addition to BiP or PERK, the kinase domains of IRE1α and Xbp1 could be potential targets for the development of novel therapeutic approaches for treating angiosarcomas and to control tumour angiogenesis. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.


Asunto(s)
Endorribonucleasas/metabolismo , Células Endoteliales/enzimología , Proteínas de Choque Térmico/metabolismo , Hemangioma/enzimología , Hemangiosarcoma/enzimología , Neovascularización Patológica , Proteínas Serina-Treonina Quinasas/metabolismo , eIF-2 Quinasa/metabolismo , Movimiento Celular , Proliferación Celular , Células Cultivadas , Chaperón BiP del Retículo Endoplásmico , Endorribonucleasas/genética , Células Endoteliales/patología , Regulación Enzimológica de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Proteínas de Choque Térmico/genética , Hemangioma/genética , Hemangioma/patología , Hemangiosarcoma/genética , Hemangiosarcoma/patología , Células Endoteliales de la Vena Umbilical Humana/enzimología , Humanos , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Proteína 1 de Unión a la X-Box/genética , Proteína 1 de Unión a la X-Box/metabolismo , eIF-2 Quinasa/genética
2.
J Exp Clin Cancer Res ; 34: 42, 2015 May 13.
Artículo en Inglés | MEDLINE | ID: mdl-25967108

RESUMEN

BACKGROUND: For the successful therapeutic use of inhibitors of the vascular endothelial growth factor receptor (VEGFR) pathway detailed knowledge of the mechanisms leading to tumor progression is indispensable. The main goal of this study was to determine the relevance of the VEGFR-2 activating pathway for colon carcinoma (CC) metastasis. The initial event is ligand-induced receptor activation through tyrosine autophosphorylation. METHODS: VEGFR-2, its ligands VEGF-C and VEGF-D and the phosphorylated (activated) receptor forms pVEGFR-2(Tyr1175) and pVEGFR-2(Tyr1214) were investigated immunohistochemically in different tumor compartments (intratumoral (zone 1) - invasive front (zone 2) - extratumoral soft tissue (zone 3)) and various cell types (tumor cells, inflammatory cells, macro- and microvasculature) in 84 non-metastatic, lymphogenous-metastatic and haematogenous-metastatic CC. RESULTS: VEGF-D produced by tumor cells has an autocrine affinity for its receptor VEGFR-2. In tumor budding regions VEGF-D-induced receptor activation by autophosphorylation at Tyr1214 seems to be a possible initial event of the VEGFR-2-mediated signaling pathway, but without effect on metastatic behaviour. In inflammatory cells of almost all CC VEGFR-2 phosphorylation at Tyr 1175 and Tyr 1214 was detectable without accompanying receptor expression, suggesting receptor activation without cell surface expression. Peritumoral inflammatory cells also expressed paracrine acting VEGF-C. The autocrine VEGF-D/VEGFR-2 signaling axis and receptor autophosphorylation at Tyr1214 appear to be main events for capillaries in all three tumor zones and for small vessels in zone 1 and 2. Independent of the metastatic status a large number of cases with capillary immunopositivity in the angiogenically active invasive front was documented, especially for VEGF-D, VEGFR-2 and pVEGFR-2(Tyr1214). VEGFR-2 positive extratumoral capillaries were significantly more common in distant metastatic CC. In all tumor compartments the investigated biomolecules were also detected in different frequencies in the macrovasculature, which is responsible for sufficient tumor vascularization. In addition, vascular paracrine-acting VEGF-C production was widely detected, but without zone and vessel-type dependence. CONCLUSIONS: The VEGFR-2 activating pathway is closely involved in tumor cell-associated, vessel-mediated and immuno-inflammatory processes in colon carcinoma and appears to contribute to tumor survival and growth as well as maintenance of the infiltrative phenotype rather than to promote metastasis.


Asunto(s)
Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Anciano , Anciano de 80 o más Años , Neoplasias del Colon/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Modelos Biológicos , Estadificación de Neoplasias , Neovascularización Patológica/metabolismo , Transducción de Señal , Factor C de Crecimiento Endotelial Vascular/genética , Factor C de Crecimiento Endotelial Vascular/metabolismo , Factor D de Crecimiento Endotelial Vascular/genética , Factor D de Crecimiento Endotelial Vascular/metabolismo
3.
BMC Cancer ; 15: 104, 2015 Mar 07.
Artículo en Inglés | MEDLINE | ID: mdl-25880726

RESUMEN

BACKGROUND: Detailed knowledge of the essential pro-angiogenic biomolecules, the vascular endothelial growth factor (VEGF) family and its receptors, in the characteristically heterogeneous tumor tissue is a pre-requisite for an effective personalized target therapy. The effects of VEGF receptors after ligand binding are mediated through receptor tyrosine autophosphorylation. We determined the relevance of the VEGFR-1 activating pathway for colon cancer (CC) metastasis. METHODS: The expression profiles of VEGFR-1, phosphorylated (activated) VEGFR-1 (pVEGFR-1(Tyr1048), pVEGFR-1(Tyr1213) and pVEGFR-1(Tyr1333)) and the VEGFR-1 ligands (VEGF, PlGF and VEGF-B) were investigated using immunohistochemistry in different tumor compartments (intratumoral - invasive front - extratumoral), cell types (tumor cells - macro- (large and small vessels) and the microvasculature (capillaries) - inflammatory cells) in human sporadic non-metastatic, lymphogenous metastatic and haematogenous metastatic CC. RESULTS: VEGF and PlGF produced by tumor cells have an autocrine affinity for their receptor VEGFR-1. Subsequent PlGF-mediated receptor activation by autophosphorylation at Tyr1048 and Tyr1213 is a potential signaling pathway, which in turn seems to protect against distant metastasis and, in regions of tumor budding, additionally against lymph node metastasis. This autocrine link could be supported by possible formation of PlGF-VEGF heterodimers and PlGF-PlGF homodimers, which are known to have anti-metastatic properties. In contrast, in order to enhance their potential for distant metastasis tumor cells produce paracrine-acting VEGF-B. VEGFR-1 activation in tumor-associated macrovasculature but not capillaries appears to affect metastatic ability. Paracrine-mediated receptor autophosphorylation at Tyr1048 and Tyr1213 in small vessels located intratumorally and along the invasive front appears to be inversely correlated with metastasis, especially distant metastasis. Additionally, macrovessels are able to produce VEGFR-1 ligands, which influence the metastatic potential. Paracrine-acting VEGF-B production by intratumorally located small vessels and autocrine-acting PlGF production by extratumorally located small vessels seem to be associated with the non-metastatic phenotype. In contrast, VEGF-B-expressing extratumoral large and small vessels correlate with distant metastasis. Lymphocyte-associated VEGFR-1 expression in the invasive front without accompanying autophosphorylation could prevent against distant metastasis possibly by acting as a decoy and scavenger receptor. CONCLUSION: VEGFR-1 and its ligands participate in vascular, tumor cell-mediated and immuno-inflammatory processes in a complex biomolecule-dependent and tumor zone-specific manner and hence could influence metastatic behavior in CC.


Asunto(s)
Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Anciano , Anciano de 80 o más Años , Neoplasias del Colon/genética , Expresión Génica , Humanos , Inmunohistoquímica , Ligandos , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Especificidad de Órganos/genética , Fosforilación , Multimerización de Proteína , Transducción de Señal , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genética , Factores de Crecimiento Endotelial Vascular/química , Factores de Crecimiento Endotelial Vascular/metabolismo
4.
BMC Cancer ; 15: 14, 2015 Jan 21.
Artículo en Inglés | MEDLINE | ID: mdl-25603809

RESUMEN

BACKGROUND: Metastatic dissemination can exist before a pathologically and clinically detectable manifestation. The structural heterogeneity of colon cancer (CC) in histological sections with respect to the morphology of tumor aggressiveness and composition of the tumor microenvironment raises the question of whether the microscopical tumor architecture enables a discrimination of groups with different metastatic potential. This would result in an assessment of the prognosis and provision of an ancillary tool for the therapeutic management after surgery, beside the estimation of the local tumor extent. METHODS: In order to identify predictive biomarkers for metastasis of locally advanced CC, which can easily be integrated into the pathologist's daily routine diagnostic activity, we determined tumor budding, peritumoral inflammation, extent of desmoplasia and necrosis, density of macro- and microvascular blood vessels and functional state of lymphatics in the tumor center, invasive margin and tumor-free surrounding tissue in 86 non-metastatic, lymphogenous-metastatic and haematogenous-metastatic, subserosa-invasive CC. RESULTS: Features influencing nodal metastasis in the univariate analysis included high tumor budding (p = 0.004), high large vessel density in the subserosa (p = 0.043), abundant desmoplasia (p = 0.049), non-finger-like desmoplastic pattern (p = 0.051) and absent lymphocellular intratumoral inflammation (p = 0.084). In the multivariate analysis, with the exception of large vessel density, these pathomorphological features were independent risk factors for lymphogenous metastasis (p = 0.023, p = 0.017, p = 0.037, p = 0.012, respectively) with a good discrimination ability (AUC of 0.853). Features associated with distant metastasis in the univariate analysis included high tumor budding (p = 0.002), low intratumoral small vessel density (p = 0.013), absent lymphocellular intratumoral inflammation (p = 0.048) and abundant necrosis (p = 0.073). In the multivariate analysis only tumor budding was an independent predictor for haematogenous metastasis (p = 0.007) with a good discrimination ability (AUC of 0.829). CONCLUSIONS: Thus, mainly tumor budding but also the described structural characteristics of the peritumoral tissue appears to reflect the metastatic potential of locally advanced CC and therefore should be stated in pathological reports.


Asunto(s)
Vasos Sanguíneos/patología , Carcinoma/patología , Neoplasias del Colon/patología , Inflamación/patología , Actinas/metabolismo , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor , Carcinoma/genética , Neoplasias del Colon/genética , Femenino , Humanos , Inflamación/genética , Metástasis Linfática/patología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Pronóstico
5.
Histol Histopathol ; 28(5): 647-53, 2013 05.
Artículo en Inglés | MEDLINE | ID: mdl-23417498

RESUMEN

The biological behavior of VEGF-B, a ligand of the receptor VEGFR-1, is still enigmatic. Despite its high sequence homology to the better known angiogenetic factor VEGF-A, the function of VEGF-B has remained elusive. Especially, its role in tumor biology has thus far not been defined. In the present study we address the question of whether VEGF-B could play a role in the metastatic process of colorectal carcinomas (CRC). Using immunohistochemistry we investigated its expression in the tumor tissue of 91 non-metastatic, lymphogenous-metastatic and haematogenous-metastastic CRC. Independently of metastatic status, VEGF-B was expressed in endothelial as well as in tumor cells. 81% of the CRC showed a positive, partly focal, partly disseminated endothelial expression in intratumoral vessels and the vascular fraction throughout the invasive tumor margin. Almost all of the VEGF-B positive vessels were of blood vascular origin. Many of these were thick-walled blood vessels with atherosclerotic changes characterizing preexistent but not angiogenetic vasculature. Thus it appears that VEGF-B might be an important ligand to ensure the blood supply for tumor survival. 46% of the CRC presented an additional tumoral VEGF-B expression which significantly correlated with haematogenous metastasis (p=0.006). These morphological results provide evidence for a probable pathobiological significance of VEGF-B in the tumor progression of CRC, especially in the case of haematogenous metastasis. It appears that VEGF-B might be an important ligand in the signalling between the tumor and preexisting blood vessels to ensure a functional blood supply for tumor survival.


Asunto(s)
Adenocarcinoma/metabolismo , Biomarcadores de Tumor/metabolismo , Neoplasias Colorrectales/metabolismo , Progresión de la Enfermedad , Factor B de Crecimiento Endotelial Vascular/metabolismo , Adenocarcinoma/irrigación sanguínea , Adenocarcinoma/patología , Vasos Sanguíneos/metabolismo , Vasos Sanguíneos/patología , Neoplasias Colorrectales/irrigación sanguínea , Neoplasias Colorrectales/patología , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Humanos , Inmunohistoquímica , Metástasis de la Neoplasia/fisiopatología , Neovascularización Patológica/fisiopatología , Estudios Retrospectivos , Transducción de Señal/fisiología
6.
Virchows Arch ; 455(2): 187-90, 2009 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-19562369

RESUMEN

We present a case of a histiocytic sarcoma incidentally detected in peripheral lung tissue resected for a spontaneous pneumothorax. Furthermore, we discuss the practical approach to pulmonary Langerhans cell histiocytosis, the main differential diagnosis of this lesion in the lung, based on morphological and immunohistochemical features. A 23-year-old male patient presented with recurrent pneumothoraces. The pulmonary tissue showed a single round granuloma-like lesion measuring 4 mm in diameter in close neighbourhood to a bronchial wall. The granuloma consisted of histiocytic cells with enlarged pale nuclei, plasma cells, lymphocytes and scanty eosinophilic granulocytes giving the impression of a granuloma of pulmonary Langerhans cell histiocytosis on haematoxylin and eosin (H&E) stains. Immunohistochemically, the histiocytic cells were negative for CD1a and S-100. They were positive for CD68, HLA-DR, CD14, CD4, CD11c, CD45LCA and lysozyme. MIB1 (Ki67) showed a nuclear staining of approximately 10% of the histiocytic cells. In summary, these findings were in keeping with a histiocytic sarcoma, a rare haematopoetic neoplasm. By demonstrating this particular case, we emphasise the importance of proving the diagnosis of pulmonary Langerhans cell histiocytosis by means of immunohistochemistry. In case of a negative CD1a reaction in a histiocytic lesion, further immunohistochemical studies have to be performed in order not to misdiagnose a malignant haematopoetic lesion.


Asunto(s)
Sarcoma Histiocítico/diagnóstico , Histiocitosis de Células de Langerhans/diagnóstico , Neoplasias Pulmonares/diagnóstico , Neumotórax/diagnóstico , Diagnóstico Diferencial , Sarcoma Histiocítico/complicaciones , Sarcoma Histiocítico/patología , Histiocitosis de Células de Langerhans/patología , Humanos , Neoplasias Pulmonares/complicaciones , Neoplasias Pulmonares/patología , Masculino , Neumotórax/etiología , Neumotórax/patología , Adulto Joven
7.
Oncol Rep ; 21(4): 933-9, 2009 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-19287991

RESUMEN

Colorectal carcinoma growth and progression is dependent on the vasculature of the tumor microenvironment. Tumor-derived endothelial cells differ functionally from their normal counterpart. For this reason we isolated microvascular endothelial cells from human colon cancer tissue (HCTEC) and compared them with endothelial cells from normal colonic tissue (HCMEC) of the same donor. Since hypoxia is a universal hallmark of carcinomas, we examined its effects on HCTEC of five patients in comparison with the corresponding HCMEC, with respect to the secretion of the soluble form of the two important vascular endothelial growth factor (VEGF) receptors, VEGFR-1 and -2. After dissociation by dispase/collagenase of central non-necrotic tumor areas obtained from colon carcinomas, HCTEC were isolated using CD31-coated magnetic beads and cultivated as monolayers. Subsequent characterization studies demonstrated the endothelial phenotype, including VEGFR-1 and -2 mRNA and protein expression as well as E-selectin expression, up-regulated after LPS, TNFalpha and IL-1beta stimulation. sVEGFR expression analyses were performed using ELISA. In comparison with HCMEC markedly lower sVEGFR-1 protein concentrations were found in HCTEC. These low sVEGFR-1 levels remain unchanged under hypoxia. In contrast, sVEGFR-2 was significantly decreased in both HCMEC and HCTEC under hypoxic conditions (p

Asunto(s)
Hipoxia de la Célula , Colon/irrigación sanguínea , Neoplasias del Colon/irrigación sanguínea , Células Endoteliales/química , Receptor 1 de Factores de Crecimiento Endotelial Vascular/análisis , Receptor 2 de Factores de Crecimiento Endotelial Vascular/análisis , Separación Celular , Células Cultivadas , Colon/química , Colon/citología , Neoplasias del Colon/química , Ensayo de Inmunoadsorción Enzimática , Humanos , Microcirculación
8.
Int J Mol Med ; 23(1): 49-55, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-19082506

RESUMEN

The functionality of large-vessel endothelial cells, such as human umbilical vein endothelial cells (HUVEC), may differ significantly from that in the microvasculature. We established a method for the isolation of human colonic microvascular endothelial cells (HCMEC). Since colonic diseases are often accompanied by hypoxia we examined its effects on HCMEC of five individuals in comparison with HUVEC, with respect to the secretion of the soluble form of the two important vascular endothelial growth factor (VEGF) receptors, VEGFR-1 and 2. After dissociation by dispase/collagenase of mucosal and submucosal tissue obtained from normal adult colon, HCMEC were isolated using CD31-coated magnetic beads and cultivated as monolayers. Subsequent characterization studies demonstrated the endothelial phenotype, including VEGFR-1 and 2 mRNA and protein expression. sVEGFR expression analyses were performed using ELISA. Under hypoxic conditions significantly enhanced levels of sVEGFR-1 on HUVEC were observed (p<0.001), while in HCMEC there was a markedly variable reaction to hypoxia, with cases of enhanced, unchanged and reduced expression. sVEGFR-2 was significantly decreased in HCMEC under hypoxia (p<0.001). In contrast, the responses of sVEGFR-2 levels to hypoxia in HUVEC were variable, that is, either unchanged or up-regulated. The different secretion profiles of sVEGFR-1 and 2 between HUVEC and HCMEC under normoxia and hypoxia underline the importance of using a functionally adequate and relevant microvasculature for in vitro studies of colonic diseases. The homogeneously reduced sVEGFR-2 levels in hypoxic HCMEC provide evidence for a novel microvascular endothelium-specific biomarker in hypoxia-response processes.


Asunto(s)
Hipoxia de la Célula , Colon/citología , Células Endoteliales/citología , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Células Cultivadas , Colon/irrigación sanguínea , Células Endoteliales/metabolismo , Regulación de la Expresión Génica , Humanos , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética
9.
BMC Cancer ; 8: 320, 2008 Nov 04.
Artículo en Inglés | MEDLINE | ID: mdl-18983642

RESUMEN

BACKGROUND: Hypoxia-inducible factor 1 alpha (HIF-1alpha) is involved in processes promoting carcinogenesis of many tumors. However, its role in the development of colorectal cancer is unknown. To investigate the significance of HIF-1alpha during colorectal carcinogenesis and progression we examined its expression in precursor lesions constituting the conventional and serrated pathways, as well as in non-metastatic and metastatic adenocarcinomas. METHODS: Immunohistochemistry and Western blot is used to analyse HIF-1alpha expression in normal colonic mucosa, hyperplastic polyps (HPP), sessile serrated adenomas (SSA), low-grade (TA-LGD) and high-grade (TA-HGD) traditional adenomas as well as in non-metastatic and metastatic colorectal adenocarcinomas. Eight colorectal carcinoma cell lines are tested for their HIF-1alpha inducibility after lipopolysaccharide (LPS) stimulation using western blot and immunocytochemistry. RESULTS: In normal mucosa, HPP and TA-LGD HIF-1alpha was not expressed. In contast, perinuclear protein accumulation and nuclear expression of HIF-1alpha were shown in half of the examined SSA and TA-HGD. In all investigated colorectal carcinomas a significant nuclear HIF-1alpha overexpression compared to the premalignant lesions was observed but a significant correlation with the metastatic status was not found. Nuclear HIF-1alpha expression was strongly accumulated in perinecrotic regions. In these cases HIF-1alpha activation was seen in viable cohesive tumor epithelia surrounding necrosis and in dissociated tumor cells, which subsequently die. Enhanced distribution of HIF-1alpha was also seen in periinflammatory regions. In additional in vitro studies, treatment of diverse colorectal carcinoma cell lines with the potent pro-inflammatory factor lipopolysaccharide (LPS) led to HIF-1alpha expression and nuclear translocation. CONCLUSION: We conclude that HIF-1alpha expression occurs in early stages of colorectal carcinogenesis and achieves a maximum in the invasive stage independent of the metastatic status. Perinecrotic activation of HIF-1alpha in invasive tumors underlines a dual role of HIF-1alpha by regulating both pro-survival and pro-death processes. HIF-1alpha up-regulation in response to LPS-mediated stimulation and periinflammatory expression in invasive carcinomas suggest its involvement in inflammatory events. These patterns of HIF-1alpha inducibility could contribute indirectly to the acquisition of a metastatic phenotype.


Asunto(s)
Transformación Celular Neoplásica/metabolismo , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Subunidad alfa del Factor 1 Inducible por Hipoxia/biosíntesis , Lesiones Precancerosas/metabolismo , Línea Celular Tumoral , Transformación Celular Neoplásica/patología , Pólipos del Colon/metabolismo , Pólipos del Colon/patología , Progresión de la Enfermedad , Humanos , Inmunohistoquímica , Lipopolisacáridos/farmacología , Metástasis de la Neoplasia , Lesiones Precancerosas/patología
10.
Int J Oncol ; 32(3): 585-92, 2008 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-18292935

RESUMEN

Adaptation to hypoxia, a universal hallmark of carcinomas, is a critical step for tumor cell survival and growth. One of the principal regulators of hypoxia-responsive pathways is the transcription factor hypoxia-inducible factor-1 alpha (HIF-1 alpha). Currently, it is known that tumoral production of members of the vascular endothelial growth factor (VEGF)-family (VEGFs) may promote tumor growth and progression by acting on carcinoma cells that express the cognate receptors (VEGFRs). However, the influence of hypoxia in the formation of such a tumoral VEGF/VEGFR loop is not completely understood. In the present study we examined the potential existence of a HIF-1 alpha/VEGF/VEGFR autocrine loop on commonly occurring carcinomas. The experiments were performed on five colorectal carcinoma cell lines, one breast (MCF7) and one lung (A549) adenocarcinoma cell line under normoxic and oxygen stress conditions using HIF-1 alpha-EIA, VEGFs-ELISA as well as RT-PCR and immunofluorescence for VEGFRs. HIF-1 alpha overexpression was found already after 2 h of exposure to hypoxia in all above mentioned cell lines, thus documenting that activation of the transcription factor HIF-1 alpha is an early cellular event. Under hypoxic conditions a significant upregulation and activation of HIF-1 alpha accompanied by an increased production of VEGF in MCF7 and A549 was observed. The well-differentiated colorectal carcinoma cell lines were 'hypoxia-resistant' showing unchanged levels of HIF-1 alpha and VEGF under hypoxia. None of the cell lines used in this study expressed the VEGF receptors VEGFR-1 and VEGFR-2 under normoxia and hypoxia. Additionally, all colorectal carcinoma cell lines were negative for VEGFR-3 transcripts in both conditions. However, VEGFR-3 mRNA and protein were expressed and under hypoxia overexpressed in MCF7 and A549. Hypoxic cultures of both cell lines secreted in elevated levels the VEGFR-3 ligand VEGF-C but not VEGF-D. Our findings suggest that under hypoxic conditions an autocrine loop between VEGF-C/VEGFR-3 and HIF-1 alpha is possible in breast carcinoma and lung carcinoma but not in colorectal carcinoma cell lines.


Asunto(s)
Carcinoma/genética , Hipoxia de la Célula/genética , Neoplasias Colorrectales/genética , Factor C de Crecimiento Endotelial Vascular/genética , Receptor 3 de Factores de Crecimiento Endotelial Vascular/genética , Comunicación Autocrina/genética , Comunicación Autocrina/fisiología , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Carcinoma/metabolismo , Línea Celular Tumoral , Neoplasias Colorrectales/metabolismo , Regulación Neoplásica de la Expresión Génica , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Regulación hacia Arriba , Factor C de Crecimiento Endotelial Vascular/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Receptor 3 de Factores de Crecimiento Endotelial Vascular/metabolismo
11.
Int J Mol Med ; 21(1): 19-31, 2008 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18097612

RESUMEN

Recently, we identified htid-1, the human counterpart of the Drosophila tumor suppressor gene lethal(2)tumorous imaginal discs [l(2)tid], as a direct molecular ligand of the adenomatous polyposis coli (APC) tumor suppressor. The gene encodes three cytosolic (Tid50, Tid48 and Tid46) and three mitochondrial (Tid43, Tid40 and Tid38) proteins. In the colorectal epithelium the cytosolic forms hTid50/hTid48 interact under physiological conditions with the N-terminal region of APC. This complex which associates with additional proteins such as Hsp70, Hsc70, Actin, Dvl and Axin defines a novel physiological state of APC unrelated to beta-catenin degradation. Here we show that the expression of the genes htid-1 and APC was altered in colorectal tumors. These changes concerned both the localization and the expression level of all three htid-1 splice variants and of APC. Furthermore, we showed that the protein products of the two tumor suppressors co-localized in the basal and apical region of normal colon epithelia and that loss of differentiation capacity of colorectal cancers correlated with a shift in their expression patterns from compartmentalized to diffuse cytoplasmic. These findings support our hypothesis that the building of the multi-component complex mentioned above is associated with the maintenance of the polarity of cells and tissues. In addition, we provide evidence that colon cancer progression correlates with up-regulation of htid-1 and its ligand Hsp70. Since the Tid proteins are members of the DnaJ-like protein family, an essential component of the Hsp70/Hsc70 chaperone machinery, our findings describe a novel, causal link between the function of chaperone machines, APC-mediated Wg/Wnt signaling and tumor development.


Asunto(s)
Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Proteínas del Choque Térmico HSP40/genética , Proteínas del Choque Térmico HSP40/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Proteína de la Poliposis Adenomatosa del Colon/genética , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Anticuerpos Antineoplásicos , Diferenciación Celular , Cartilla de ADN , Progresión de la Enfermedad , Proteínas HSP70 de Choque Térmico/metabolismo , Humanos , Mucosa Intestinal/patología , Empalme del ARN , ARN Neoplásico/genética , ARN Neoplásico/metabolismo
12.
Anticancer Res ; 27(5A): 3245-50, 2007.
Artículo en Inglés | MEDLINE | ID: mdl-17970067

RESUMEN

BACKGROUND: Vascular endothelial growth factor (VEGF), originally identified as an endothelium-specific factor, can also bind to malignant cells, a mechanism by which a tumor could regulate its own progression. The biological effects of VEGF are mediated by three receptors (VEGFRs), VEGFR-1, VEGFR-2 and VEGFR-3. This study aimed at defining the expression of VEGFs in colorectal cancer (CRC) epithelia and their relationship to the metastatic status. MATERIALS AND METHODS: Using immunohistochemistry, the levels of tumoral immunoreactivity for VEGFs in 105 nonmetastatic, lymphogenously-metastatic and haematogenously-metastatic CRC specimens were assessed. Statistical analysis was performed using Fisher's exact probability test. RESULTS: VEGFR-1 immunoreactivity was positive in only 50% of the cases. However lack of expression of VEGFR-1 was significantly associated with lymphogenous and haematogenous metastases. VEGFR-2 and VEGFR-3 were expressed in all investigated specimens to varying degrees. Low levels of VEGFR-2 were significantly associated with distant metastases. No significant changes were detected in VEGFR-3 expression. CONCLUSION: Epithelial expression of VEGFR-1 and VEGFR-2 appear to have a protective effect against tumor aggressiveness in CRC.


Asunto(s)
Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Receptores de Factores de Crecimiento Endotelial Vascular/biosíntesis , Células Epiteliales/metabolismo , Células Epiteliales/patología , Humanos , Inmunohistoquímica , Metástasis de la Neoplasia
13.
Int J Oncol ; 31(2): 269-75, 2007 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-17611682

RESUMEN

The transcription factor NFkappaB regulates the expression of several tumor-related molecules associated with tumor progression and metastasis. However, the precise mechanisms by which its activation mediates these processes in diverse tumors are unknown. In this study we determined the expression of NFkappaB in various colorectal carcinoma cell lines, in a series of 90 non-metastatic and metastatic colorectal tumors and in an in vitro 3D-spheroid model of HT-29 cells simulating morphological hallmark of these adenocarcinomas, namely neoplastic glandular nests around a necrotic center. We show that the inactive cytoplasmic NFkappaB form is evidently up-regulated in the tumor epithelium, especially in the metastatic cases, as compared to normal tissue. We found that in situ nuclear NFkappaB staining is characteristic for cells that are still viable but dissociated from the surrounding cohesive tumor tissue and destined to die. Evidence for a possible association between NFkappaB expression and loss of cell adhesion mediated by E-cadherin function has been provided in vivo and in vitro using the HT-29 3D-spheroid model. In both cases, we found a strong correlation between activation of NFkappaB and loss of E-cadherin expression. Considering the fact that cancer cell necrosis plays a crucial role in metastasis, NFkappaB activation mediated by loss of E-cadherin may represent an essential, even initial event in this process. Furthermore, we present in vitro data implicating LPS, the endotoxin of gram-negative bacteria, in the triggering of NFkappaB up-regulation. Thus, release of bacterial endotoxin may essentially contribute to the progression of colon cancer in vivo.


Asunto(s)
Cadherinas/biosíntesis , Carcinoma/metabolismo , Neoplasias Colorrectales/metabolismo , Regulación Neoplásica de la Expresión Génica , FN-kappa B/metabolismo , Adenocarcinoma , Adhesión Celular , Línea Celular Tumoral , Núcleo Celular/metabolismo , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Progresión de la Enfermedad , Activación Enzimática , Humanos , Necrosis , Neoplasias/metabolismo
14.
Int J Mol Med ; 20(1): 21-9, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17549384

RESUMEN

Signaling mediating colorectal cancer (CRC) progression is incompletely understood. Previously, we identified lipopolysaccharide (LPS), an endotoxin of ubiquitously existing colonic bacteria, as a pivotal stimulus increasing the metastatic potential of human CRC. Since the ubiquitous colonic bacteria release large amounts of LPS this observation could be of enormous relevance for the progression of CRC. In this study we present data contributing to the elucidation of its mode of action. Since both receptors CD14 and TLR4 act as LPS mediators, we determined their expression in various CRC cell lines and in 115 non-metastatic, lymphogenous-metastatic and haematogenous-metastatic CRC specimens. Here we showed that CD14 was not expressed in normal colon epithelium, in non-metastatic and metastatic CRC. Furthermore, we showed that diverse CRC cell lines did not express CD14 under normal conditions and after LPS stimulation. Thus, CD14 can be ruled out as a mediator of LPS-induced signaling related to CRC progression. In contrast, we found that normal colon epithelium and CRC cell lines were positive for TLR4. Furthermore, both lymphogenous and haematogenous metastatic cases showed either loss of expression or strong downregulation of TLR4 as compared to normal tissue and to non-metastatic tumors. We found that LPS stimulation resulted in significant TLR4 upregulation in cells expressing lower constitutive TLR4 levels such as CaCo2, whereas no significant response to LPS was observed in cells characterized by relatively high amounts of constitutive TLR4. Our data suggest that TLR4 expression may be associated with mechanisms preventing CRC progression.


Asunto(s)
Carcinoma/patología , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Lipopolisacáridos/metabolismo , Receptor Toll-Like 4/metabolismo , Células CACO-2 , Carcinoma/cirugía , Línea Celular Tumoral , Neoplasias Colorrectales/cirugía , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Inmunohistoquímica , Receptores de Lipopolisacáridos/metabolismo , Lipopolisacáridos/farmacología , Metástasis Linfática/patología , Metástasis de la Neoplasia/patología , Estadificación de Neoplasias , Estudios Retrospectivos , Receptor Toll-Like 4/genética
15.
Oncol Rep ; 16(1): 109-13, 2006 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-16786131

RESUMEN

Despite many pathophysiological analyses, the process of tumor dissemination of hepatocellular carcinoma (HCC) remains vague. In diverse tumor entities, expression of the chemokine receptor, CCR7, has been linked to tumor dissemination and poor prognosis. Therefore, we evaluated, whether CCR7 exerts similar effects in human HCC. CCR7 expression analysis was performed in vitro on human hepatoma cell lines (Huh7, Hep3B, wt HepG2, p53 dominant negative transfected HepG2). In addition, CCR7 expression was evaluated in 39 patients with hepatocellular cancer and correlated with both, tumor and patients characteristics. Human hepatocellular carcinoma samples and hepatoma cell lines displayed variable intensities of CCR7 expression. In patients, CCR7 expression was significantly associated with progressed local tumors (P = 0.02) and lymphatic metastasis (P = 0.02). Strong expression of CCR7 promotes intrahepatic and lymphatic HCC dissemination.


Asunto(s)
Carcinoma Hepatocelular/genética , Regulación Neoplásica de la Expresión Génica , Predisposición Genética a la Enfermedad , Neoplasias Hepáticas/genética , Receptores de Quimiocina/biosíntesis , Anciano , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Femenino , Genes Dominantes , Humanos , Hígado/metabolismo , Neoplasias Hepáticas/metabolismo , Metástasis Linfática , Masculino , Persona de Mediana Edad , Pronóstico , Receptores CCR7
16.
J Magn Reson Imaging ; 22(1): 101-8, 2005 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-15971183

RESUMEN

PURPOSE: To assess the diagnostic accuracy of integrated parallel acquisition technique (iPAT) in local staging of rectal carcinoma in comparison to conventional high-resolution MRI. MATERIALS AND METHODS: A total of 28 patients with a neoplasm of the rectum and 15 control patients underwent MRI of the pelvis. High-resolution images were acquired conventionally and with iPAT using a modified sensitivity encoding (mSENSE). Image quality, signal-to-noise and contrast-to-noise ratios (SNR, CNR), tumor extent, nodal status, and delineation of the circumferential resection margin (CRM) were compared. In 19 patients with a carcinoma, MR findings were correlated with the histopathological diagnosis. Tumor distance to the CRM was matched with resection specimen in 12 cases. RESULTS: The comparison of both MR techniques revealed no clinically relevant differences in tumor staging and delineation of the CRM, though SNR and CNR were significantly lower in mSENSE images. Tumor stage was concordant in 17 of 19 cases compared to histopathology. In four of nine patients with T3 and T4 carcinomas, the histopathological resection margin was < or =2 mm, in five cases MRI predicted a margin of < or =2 mm. CONCLUSION: The application of iPAT in local staging of rectal carcinoma is time-saving and does not degrade diagnostic accuracy. Tumor stage, nodal status, and the CRM can be assessed equally compared to conventional acquisition techniques.


Asunto(s)
Imagen por Resonancia Magnética/métodos , Estadificación de Neoplasias/métodos , Neoplasias del Recto/diagnóstico , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Ganglios Linfáticos/patología , Masculino , Persona de Mediana Edad , Neoplasias del Recto/patología , Sensibilidad y Especificidad
17.
Clin Cancer Res ; 11(5): 1743-50, 2005 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-15755995

RESUMEN

PURPOSE: The expression of chemokine receptors CXCR4 and CCR7 has been associated with tumor dissemination and poor prognosis in a limited number of tumor entities. However, no data are currently available on the impact of chemokine receptor expression on disease progression and prognosis in human colorectal cancer. EXPERIMENTAL DESIGN: The expression of CXCR4 and CCR7 was evaluated in 96 patients with histologically confirmed colorectal cancers and in four colorectal cancer cell lines by immunohistochemical staining. Furthermore, cell migration assays were done with SW480, SW620, and LS174T cancer cells to confirm the effect of the CXCR4 ligand stromal cell-derived factor 1alpha on migration. RESULTS: Human colorectal cancer specimens and cell lines displayed a CXCR4 and CCR7 expression with variable intensities. Interestingly, strong expression of CXCR4, but not of CCR7, was significantly associated with higher Union International Contre Cancer stages 3/4 (P = 0.0017), lymph node metastasis (P = 0.00375), and distant metastasis (P = 0.00003) and further correlated with a reduced 3-year survival rate (P = 0.1). Strong CXCR4 and CCR7 expression positively correlated with the location of the primary tumor in the rectum (P < 0.01). Furthermore, activation of CXCR4-expressing cancer cells by stromal cell-derived factor 1alpha resulted in a significant increase of cell migration (P < 0.014). CONCLUSION: Strong expression of CXCR4 by colorectal cancer cells is significantly associated with lymphatic and distant dissemination in patients with colorectal cancer as well as with cancer cell migration in vitro.


Asunto(s)
Neoplasias Colorrectales/patología , Metástasis de la Neoplasia , Receptores CXCR4/biosíntesis , Receptores CXCR4/fisiología , Receptores de Quimiocina/biosíntesis , Receptores de Quimiocina/fisiología , Movimiento Celular , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Humanos , Inmunohistoquímica , Metástasis Linfática , Masculino , Persona de Mediana Edad , Pronóstico , Receptores CCR7 , Células Tumorales Cultivadas
18.
Anticancer Res ; 22(5): 2641-9, 2002.
Artículo en Inglés | MEDLINE | ID: mdl-12529976

RESUMEN

The mechanisms of tumor metastasis in vitro have been principally investigated using monolayer culture systems. In vivo, however, multicellular clusters of tumor cells rather than individual cells appear to penetrate the microvasculature. Thus, the multicellular tumor spheroid model represents a more suitable tool to study tumor cell-endothelial interactions. Our interest has centered on the role of inflammation on tumor intra- and extravasation. The expression of three endothelial cell adhesion molecules (CAM), ICAM-1, VCAM-1 and E-selectin, on HUVEC after incubation with supernatants of previously LPS-, TNF-alpha- or IL-1 beta-stimulated HT-29 (colon adenocarcinoma) and ST-ML-12 (melanoma) tumor cells growing as monolayers and spheroids, was compared using cell enzyme immunoassay. The results indicated that important differences in the expression of CAM between monolayers and spheroids exist depending on the tumor cell line. The possible significance of LPS for colon carcinomas was underlined by the spheroid-model results. With respect to the influence of intestinal bacteria on the behaviour of colon carcinomas, the spheroid model could be a useful in vitro system for a more realistic simulation of in vivo conditions.


Asunto(s)
Moléculas de Adhesión Celular/biosíntesis , Citocinas/farmacología , Endotelio Vascular/metabolismo , Células HT29/metabolismo , Células HT29/patología , Lipopolisacáridos/farmacología , Melanoma/metabolismo , Melanoma/patología , Selectina E/biosíntesis , Células HT29/efectos de los fármacos , Humanos , Molécula 1 de Adhesión Intercelular/biosíntesis , Interleucina-1/farmacología , Esferoides Celulares , Células Tumorales Cultivadas/efectos de los fármacos , Factor de Necrosis Tumoral alfa/farmacología , Molécula 1 de Adhesión Celular Vascular/biosíntesis
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