RESUMEN
Renal cell carcinoma (RCC) is characterized by organ-specific metastases. The chemokine stromal derived factor-1 (SDF-1/CXCL12) and its receptor CXCR4 have been suggested to regulate organ-specific metastasis in various other cancers. On this basis, we hypothesized that the biological axis of CXCL12 via interaction with its receptor, CXCR4, is a major mechanism for RCC metastasis. We demonstrated that CXCR4 was significantly expressed on circulating cytokeratin+ RCC cells from patients with known metastatic RCC. We detected up-regulation of CXCR4 mRNA and protein levels on a human RCC cell line by either knockdown of the von Hippel-Lindau (VHL) tumor suppressor protein, or incubating the cells under hypoxic conditions. The enhanced CXCR4 expression was mediated through the interaction of the Hypoxia Inducible Factor-1alpha (HIF-1alpha) with the promoter region of the CXCR4 gene. Furthermore, the expression of CXCR4 on human RCC directly correlated with their metastatic ability in vivo in both heterotopic and orthotopic SCID mouse models of human RCC. Neutralization of CXCL12 in SCID mice abrogated metastasis of RCC to target organs expressing high levels of CXCL12; without altering tumor cell proliferation, apoptosis, or tumor-associated angiogenesis. Therefore, our data suggest that the CXCL12/CXCR4 biological axis plays an important role in regulating the organ-specific metastasis of RCC.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Carcinoma de Células Renales/secundario , Quimiocinas CXC/fisiología , Neoplasias Renales/patología , Receptores CXCR4/metabolismo , Animales , Biomarcadores de Tumor/genética , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/metabolismo , Hipoxia de la Célula , Línea Celular Tumoral , Quimiocina CXCL12 , Quimiocinas CXC/antagonistas & inhibidores , Quimiocinas CXC/farmacología , Quimiotaxis/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Queratinas/análisis , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Ratones , Ratones SCID , Regiones Promotoras Genéticas , Interferencia de ARN , Receptores CXCR4/genética , Activación Transcripcional , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/antagonistas & inhibidores , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genéticaRESUMEN
BACKGROUND: Peripheral blood monocytes and monocyte-derived macrophages are key regulatory components in many chronic inflammatory pathologies of the vasculature including the formation of atherosclerotic lesions. However, the molecular and biochemical events underlying monocyte maturation are not fully understood. METHODS: We have used freshly isolated human monocytes and the model human monocyte cell line, THP-1, to investigate changes in the expression of a panel of monocyte and macrophage markers during monocyte differentiation. We have examined these changes by RT-PCR and FACS analysis. Furthermore, we cloned the CCR2 promoter and analyzed specific changes in transcriptional activation of CCR2 during monocyte maturation. RESULTS: The CC chemokine receptor 2 (CCR2) is rapidly downregulated as monocytes move down the macrophage differentiation pathway while other related chemokine receptors are not. Using a variety of biochemical and transcriptional analyses in the human THP-1 monocyte model system, we show that both monocytes and THP-1 cells express high levels of CCR2, whereas THP-1 derived macrophages fail to express detectable CCR2 mRNA or protein. We further demonstrate that multiple signaling pathways activated by IFN-gamma and M-CSF, or by protein kinase C and cytoplasmic calcium can mediate the downregulation of CCR2 but not CCR1. CONCLUSION: During monocyte-to-macrophage differentiation CCR2, but not CCR1, is downregulated and this regulation occurs at the level of transcription through upstream 5' regulatory elements.
RESUMEN
An increase in fat mass associated with obesity results from recruitment and differentiation of adipocyte progenitor cells. The precise origin of these cells is unknown, although accumulating evidence suggests that circulating stem cells can differentiate into cells of mesenchymal lineage. It is currently unclear whether a progenitor adipocyte population exists in circulation. One potential candidate is the fibrocyte, which may represent a common progenitor cell for several mesenchymal lineages. We demonstrate that these circulating progenitors become adipocytes when cultured under adipogenic conditions, with intracellular lipids accumulation and up-regulation of proteins specific for adipocyte differentiation, including leptin, PPARgamma, and FABP4. cDNA microarray analysis revealed gene clusters that were differentially regulated during adipogenesis of fibrocytes, which were similar to visceral and subcutaneous adipose tissue preadipocyte-to-adipocyte differentiation. Moreover, these progenitors engrafted and formed human adipose tissue following injection into SCID mice. Although fibrocytes express an array of chemokine receptors, we observed an up-regulation of CCR2 expression following fibrocytes differentiation into adipocytes, which was associated with increased chemotactic response to CCL2. This paradigm supports the notion that elevated CCL2 levels in visceral adipose tissue associated with Metabolic Syndrome is a chemotactic niche, whereby fibrocytes can home to and differentiate into adipocytes to perpetuate its tissue formation.
Asunto(s)
Adipocitos/citología , Tejido Adiposo/citología , Tejido Adiposo/patología , Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Animales , Diferenciación Celular , Linaje de la Célula , Quimiocina CCL2/metabolismo , Quimiocinas/metabolismo , ADN Complementario/metabolismo , Regulación de la Expresión Génica , Genoma , Humanos , Lípidos/química , Mesodermo , Síndrome Metabólico/metabolismo , Ratones , Ratones SCID , Modelos Biológicos , Familia de Multigenes , Análisis de Secuencia por Matrices de Oligonucleótidos , Programas Informáticos , Células Madre/citología , Factor de Crecimiento Transformador beta/metabolismo , Regulación hacia ArribaRESUMEN
The regulation of the transcription factor nuclear factor-kappaB (NF-kappaB) during B-cell development was examined using cells isolated from the bone marrow of transgenic mice expressing a kappaB luciferase reporter gene. The results indicate that the highest level of NF-kappaB activity is present in cells expressing the pre-B-cell receptor. Furthermore, cross-linking of Igbeta on CD43(+) pre-B cells is able to activate NF-kappaB in recombination-activating gene 1-deficient mice, preceding their further differentiation into CD43(-) pre-B cells. Expression of a dominant negative form of IkappaBalpha using a transgenic approach or by retroviral infection leads to a reduction in the number of CD43(+) pre-B cells. These data therefore indicate that activation of NF-kappaB in CD43(+) pre-B cells, as a result of signaling by the pre-B-cell receptor, facilitates the continued development of large, CD43(+) pre-B cells into small CD43(-) pre-B cells.
Asunto(s)
Linfocitos B/inmunología , Células de la Médula Ósea/inmunología , Glicoproteínas de Membrana/inmunología , FN-kappa B/inmunología , Animales , Anticuerpos , Linfocitos B/metabolismo , Células de la Médula Ósea/metabolismo , Diferenciación Celular/inmunología , Regulación de la Expresión Génica/inmunología , Técnicas de Transferencia de Gen , Genes Reporteros , Proteínas Fluorescentes Verdes , Antígenos Comunes de Leucocito/análisis , Luciferasas , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , FN-kappa B/metabolismo , Receptores de Células Precursoras de Linfocitos B , Proteínas Proto-Oncogénicas c-rel/análisis , Receptores de Antígenos de Linfocitos B , Retroviridae/genéticaRESUMEN
Non-small cell lung cancer (NSCLC) expresses a particularly aggressive metastatic phenotype, and patients with this disease have a poor prognosis. CXC chemokine receptor 4 (CXCR4) is a cell surface receptor that has been shown to mediate the metastasis of many solid tumors including lung, breast, kidney, and prostate. In addition, overexpression of the epidermal growth factor receptor (EGFR) is associated with the majority of NSCLC and has been implicated in the process of malignant transformation by promoting cell proliferation, cell survival, and motility. Here we show for the first time that activation of the EGFR by EGF increases CXCR4 expression and the migratory capacity of NSCLC cells. Furthermore, many solid tumors are associated with low oxygen tension, and when NSCLC cells were cultured with EGF under hypoxic conditions, CXCR4 expression was dramatically enhanced. A molecular analysis of these events indicated that augmented CXCR4 expression was regulated by the phosphatidylinositol 3-kinase/PTEN/AKT/mammalian target of rapamycin signal transduction pathway, activation of hypoxia inducible factor (HIF) 1alpha, and ultimately HIF-1-dependent transcription of the CXCR4 gene. Thus, a combination of low oxygen tension and overexpression of EGFR within the primary tumor of NSCLC may provide the microenvironmental signals necessary to upregulate CXCR4 expression and promote metastasis.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Hipoxia , Neoplasias Pulmonares/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Receptores CXCR4/metabolismo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Western Blotting , Línea Celular Tumoral , Proliferación Celular , Separación Celular , Supervivencia Celular , Quimiocina CXCL12 , Quimiocinas CXC/metabolismo , Quimiotaxis , Relación Dosis-Respuesta a Droga , Citometría de Flujo , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia , Metástasis de la Neoplasia , Oxígeno/metabolismo , Fosfohidrolasa PTEN , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-akt , ARN Mensajero/metabolismo , Transducción de Señal , Sirolimus/farmacología , Serina-Treonina Quinasas TOR , Transcripción Genética , Activación Transcripcional , Transfección , Regulación hacia ArribaRESUMEN
Previous reports have identified a circulating pool of CD45(+) collagen I(+) CXCR4(+) (CD45(+)Col I(+)CXCR4(+)) cells, termed fibrocytes, that traffic to areas of fibrosis. No studies have demonstrated that these cells actually contribute to fibrosis, however. Pulmonary fibrosis was originally thought to be mediated solely by resident lung fibroblasts. Here we show that a population of human CD45(+)Col I(+)CXCR4(+) circulating fibrocytes migrates in response to CXCL12 and traffics to the lungs in a murine model of bleomycin-induced pulmonary fibrosis. Next, we demonstrated that murine CD45(+)Col I(+)CXCR4(+) fibrocytes also traffic to the lungs in response to a bleomycin challenge. Maximal intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) fibrocytes directly correlated with increased collagen deposition in the lungs. Treatment of bleomycin-exposed animals with specific neutralizing anti-CXCL12 Ab's inhibited intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) circulating fibrocytes and attenuated lung fibrosis. Thus, our results demonstrate, we believe for the first time, that circulating fibrocytes contribute to the pathogenesis of pulmonary fibrosis.
Asunto(s)
Quimiocinas CXC/metabolismo , Fibroblastos/metabolismo , Lesión Pulmonar , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/fisiopatología , Actinas/metabolismo , Animales , Bleomicina/toxicidad , Quimiocina CXCL12 , Quimiotaxis/inmunología , Colágeno Tipo I/metabolismo , Modelos Animales de Enfermedad , Fibroblastos/química , Fibroblastos/inmunología , Humanos , Cinética , Antígenos Comunes de Leucocito/inmunología , Ratones , Ratones SCID , Músculo Liso/metabolismo , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/genética , Fibrosis Pulmonar/patología , Receptores CXCR4/metabolismo , Receptores de Complemento 3b/inmunologíaRESUMEN
The CXC chemokine family are unique cytokines known for their ability to have dual functions in the regulation of angiogenesis related to the following: (1) the presence or absence of the structural/functional motif (Glutamic acid-Leucine-Arginine; 'ELR' motif) that immediately precedes the first cysteine amino acid residue in the primary structure of these chemokines; (2) interferon-inducible gene expression; and (3) specific receptor interaction on endothelial cells. In this review we will appraise the biology of these angiogenic and angiostatic CXC chemokines, and discuss their disparate angiogenic activity in the context of the pathogenesis of cancer.
Asunto(s)
Quimiocinas CXC/fisiología , Neoplasias/irrigación sanguínea , Neovascularización Patológica , HumanosAsunto(s)
Carcinoma de Pulmón de Células no Pequeñas/secundario , Quimiocinas CXC/metabolismo , Neoplasias Pulmonares/patología , Proteínas de Neoplasias/metabolismo , Receptores de Quimiocina/metabolismo , Animales , Quimiocina CXCL12 , Quimiotaxis , Humanos , Ratones , Ratones SCID , Receptores CCR4RESUMEN
Non-small cell lung cancer (NSCLC) growth, angiogenesis, invasion, and metastases to specific organs is dependent on an orchestrated series of events that include: cellular transformation; establishment of a pro-angiogenic environment; tumour cell proliferation, invasion and entry into the circulation; and tumour cell trafficking and metastatic tumour growth in specific organs based on the concept of Paget's theory of 'seed and soil' related to homing of tumour cells to a specific organ. The events that destine a tumour cell to invade and metastasize to distant organs are analogous to leukocyte trafficking. Chemokines have had an increasingly important role in mediating the homing of leukocytes under both conditions of homeostasis and inflammatory/immunological responses. Recently, the biology of chemokines has extended beyond their role in mediating leukocyte trafficking. Specifically, CXC chemokines have been found to be important in the regulation of angiogenesis, and in promoting tumour cell migration and organ-specific metastases. Data will be presented to highlight the importance of CXC chemokine ligands and receptors in mediating NSCLC tumour-associated angiogenesis, 'immunoangiostasis', and organ specific metastases. These findings may ultimately lead to clinical strategies to attenuate the pathobiology of CXC chemokines in promoting NSCLC tumour growth and metastases.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/irrigación sanguínea , Quimiocinas CXC/fisiología , Neoplasias Pulmonares/irrigación sanguínea , Metástasis de la Neoplasia , Neovascularización Patológica/metabolismo , Animales , Carcinoma de Pulmón de Células no Pequeñas/secundario , Humanos , Neoplasias Pulmonares/patologíaRESUMEN
Non-small cell lung cancer is characterized by a specific metastatic pattern. The mechanism for organ-specific metastasis is poorly understood, although evidence has suggested that the chemokine stromal derived factor-1 (CXCL12) and its cognate receptor CXCR4 may regulate breast cancer metastasis. We hypothesized that the CXCL12-CXCR4 biological axis is important in mediating non-small cell lung cancer metastases. Our results indicate that both non-small cell lung cancer tumor specimens resected from patients and non-small cell lung cancer cell lines express CXCR4, but not CXCL12. Non-small cell lung cancer cell lines undergo chemotaxis in response to CXCL12. CXCL12-CXCR4 activation of non-small cell lung cancer cell lines showed intracellular calcium mobilization and mitogen-activated protein kinase activation with enhanced extracellular signal-related kinase-1/2 phosphorylation without change in either proliferation or apoptosis. Target organs in a murine model that are the preferred destination of human non-small cell lung cancer metastases elaborate higher levels of CXCL12 than does the primary tumor, and suggest the generation of chemotactic gradients. The administration of specific neutralizing anti-CXCL12 antibodies to severe combined immunodeficient mice expressing human non-small cell lung cancer abrogated organ metastases, without affecting primary tumor-derived angiogenesis. These data suggest that the CXCL12-CXCR4 biological axis is involved in regulating the metastasis of non-small cell lung cancer.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/patología , Quimiocinas CXC/análisis , Modelos Animales de Enfermedad , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/patología , Metástasis Linfática/patología , Receptores CXCR4/análisis , Animales , Apoptosis , Western Blotting , Quimiocina CXCL12 , Quimiocinas CXC/antagonistas & inhibidores , Quimiocinas CXC/genética , Ensayo de Inmunoadsorción Enzimática , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Inmunohistoquímica , Ratones , Ratones SCID , ARN Neoplásico/análisis , ARN Neoplásico/genética , Receptores CXCR4/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Quimera por Trasplante , Células Tumorales CultivadasRESUMEN
Mortality related to adult respiratory distress syndrome (ARDS) ranges from 35% to 65%. Lung-protective ventilator strategies can reduce mortality during ARDS. The protective strategies limit tidal volumes and peak pressures while maximizing positive end-expiratory pressure. The efficacy of this approach is due to a reduction of shear-stress of the lung and release of inflammatory mediators. Ventilator-induced lung injury (VILI) is characterized by inflammation. The specific mechanism(s) that recruit leukocytes during VILI have not been elucidated. Because the murine CXC chemokines KC/CXCL1 and MIP-2/CXCL2/3, via CXCR2, are potent neutrophil chemoattractants, we investigated their role in a murine model of VILI. We compared two ventilator strategies in C57BL/6 mice: high peak pressure and high stretch (high peak pressure/stretch) versus low peak pressure/stretch for 6 hours. Lung injury and neutrophil sequestration from the high-peak pressure/stretch group were greater than those from the low-peak pressure/stretch group. In addition, lung expression of KC/CXCL1 and MIP-2/CXCL2/3 paralleled lung injury and neutrophil sequestration. Moreover, in vivo inhibition of CXCR2/CXC chemokine ligand interactions led to a marked reduction in neutrophil sequestration and lung injury. These findings were confirmed using CXCR2(-/-) mice. Together these experiments support the notion that increased expression of KC/CXCL1 and MIP-2/CXCL2/3 and their interaction with CXCR2 are important in the pathogeneses of VILI.