RESUMEN
The control of cell growth is regulated through coordinated responses to growth factors and cell-extracellular matrix (ECM) interactions. Integrins, the major family of cell-ECM receptors, are vital to these coordinated responses. Although much is known of the role of integrins in growth promotion, specific examples of integrin-mediated cell growth inhibition are few. On the basis of our findings that the integrin beta8 subunit is expressed in airway epithelial cells and is absent in lung cancers, we investigated the role and mechanism of the integrin alphavbeta8 in mediating growth inhibition. When introduced into either a lung or colon carcinoma cell line, beta8 inhibited cell growth without inducing apoptosis. Ligation of alphavbeta8 also induced cell rounding, inhibited focal contact formation, and initiated an inhibitory signaling pathway as demonstrated by increased expression of the cyclin-dependent kinase inhibitor p21Cip1. The cytoplasmic domain of beta8 was capable of both growth inhibition and causing cell shape changes as shown by the use of a chimeric integrin construct consisting of the beta8-cytoplasmic domain coupled to the beta6-extracellular domain. Finally, when tested in vivo, beta8 potently inhibited tumor growth in nude mice. Together, these results implicate alphavbeta8 as a novel growth-regulatory molecule of epithelial cells.
Asunto(s)
División Celular , Células Epiteliales/citología , Integrinas/fisiología , Actinas/metabolismo , Animales , Apoptosis , Western Blotting , Adhesión Celular , Separación Celular , Neoplasias del Colon/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/metabolismo , Citoplasma/metabolismo , Relación Dosis-Respuesta a Droga , Matriz Extracelular/metabolismo , Citometría de Flujo , Humanos , Inmunohistoquímica , Neoplasias Pulmonares/metabolismo , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Pruebas de Precipitina , Estructura Terciaria de Proteína , Retroviridae/genética , Transducción Genética , Células Tumorales Cultivadas , Vinculina/metabolismoRESUMEN
Integrins are a large family of cell adhesion receptors mediating cell-extracellular matrix (ECM) interactions and are widely distributed in tissues. The beta8 integrin subunit mRNA has been shown to be expressed at higher levels in the central nervous system (CNS) than in other organs [M. Moyle, M.A. Napier, J.W. McLean, Cloning and expression of a divergent integrin subunit beta8, J. Biol. Chem. 266 (29) (1991) 19650-19658] but its cellular and subcellular localization in the CNS are unknown. In this report, we demonstrate that beta8 pairs exclusively with the alphav subunit in the CNS to form the alphavbeta8 heterodimer. Immunohistochemical analysis of the distribution of beta8 in adult mouse and rat brains revealed that the protein is expressed in several regions of the hippocampal formation and in the molecular layer and glomeruli of the granular cell layer of the cerebellum. Punctate and diffuse immunolabeling was observed occasionally surrounding neuronal pericarya and extensively throughout dendritic fields suggesting both pre- and post-synaptic localization and/or expression in non-neuronal cells. By immunoelectron microscopy, beta8 immunoreactivity was detected in dendritic spines where it was often localized at post-synaptic densities, occasionally in axon terminals and in glial processes. Association of beta8 with synaptic membranes was further supported by its enrichment in synaptosomal preparations as detected by immunoblotting. These results demonstrate that alphavbeta8 is present in mature synapses and therefore may play a role in synaptic function.
Asunto(s)
Encéfalo/metabolismo , Dendritas/química , Cadenas beta de Integrinas , Integrinas/análisis , Neuroglía/química , Terminales Presinápticos/química , Secuencia de Aminoácidos , Animales , Anticuerpos/análisis , Encéfalo/ultraestructura , Humanos , Inmunohistoquímica , Integrinas/inmunología , Ratones , Ratones Endogámicos , Microscopía Inmunoelectrónica , Datos de Secuencia Molecular , Plasticidad Neuronal/fisiología , Ratas , Ratas Sprague-Dawley , Células Tumorales CultivadasRESUMEN
In this study we identified tenascin-C (TN-C) and one of its integrin receptors, alpha(v)beta6, in oral squamous-cell carcinoma (SCC) specimens. Neither TN-C nor alpha(v)beta6 are expressed in normal oral mucosa. We also studied 2 human oral squamous-cell carcinoma cell lines: the highly invasive HSC-3 cells, and the poorly invasive SCC-25 cells. We determined that adhesion of these cells to TN-C involves both alpha2 and alpha(v) integrins. Migration on TN-C by oral SCC cells required fibroblast-conditioned medium and did not occur in its absence. This migration was blocked by anti-alpha2 and anti-alpha(v) antibodies and was partially inhibited by antibodies to hepatocyte growth factor, epidermal growth factor and transforming growth factor-beta1. When seeded on TN-C, the poorly invasive SCC-25 cells formed alpha(v)beta6-positive focal contacts; the HSC-3 cells did not. HSC-3, SCC-25 and PTF cells secrete TN-C into the culture medium, as determined by Western blot. However, when HSC-3 cells were inoculated into the floor of the mouth of nude mice, only murine TN-C could be identified in the reactive stroma adjacent to the resulting tumor nests, demonstrating that in vivo, HSC-3 cells do not secrete TN-C. Our results demonstrate that alpha(v)beta6 and tenascin-C are neo-expressed in oral squamous-cell carcinoma, and that the tumor stromal environment is influential in oral SCC behavior.
Asunto(s)
Antígenos de Neoplasias , Carcinoma de Células Escamosas/metabolismo , Neoplasias de la Boca/metabolismo , Células del Estroma/metabolismo , Tenascina/metabolismo , Animales , Carcinoma de Células Escamosas/patología , Adhesión Celular , Movimiento Celular , Medios de Cultivo Condicionados , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Integrinas/metabolismo , Ratones , Neoplasias de la Boca/patología , Células del Estroma/patología , Células Tumorales CultivadasRESUMEN
Myelination of the peripheral nervous system (PNS) requires the migration of Schwann cells during both development and regeneration. We have characterized the expression pattern of Schwann cell integrins and analyzed their role in migration on different ECM substrates known to be present within the PNS. We found that Schwann cells in cell culture express four beta1 integrins, alpha1 beta1, alpha2 beta1, alpha6 beta1, and another unidentified beta1 integrin, as well as two alpha v integrins, alpha v beta3 and alpha v beta8. Using the Varani migration assay, we found that laminin-1, laminin-2 (merosin), and fibronectin increased Schwann cell migration, while vitronectin and collagen did not increase migration compared to an uncoated plastic substrate. Schwann cell migration on laminin-1 and laminin-2 (merosin) was blocked by antibodies against beta1 integrins, but not affected by RGD peptides or antibodies against beta3 integrins. In contrast, migration on fibronectin was unaffected by antibodies against beta1 and beta3 integrins but was blocked by RGD peptides. This in vitro study shows that there is a division of labor of Schwann cell integrins in the regulation of migration on peripheral nerve ECM components; beta1 integrins mediate migration on laminin-1 and laminin-2 (merosin), while alpha v integrins mediate migration on fibronectin. Taken together, these results suggest that multiple interactions between Schwann cell integrins and ECM within the PNS will contribute to Schwann cell migration during myelination of the PNS.
Asunto(s)
Movimiento Celular/fisiología , Matriz Extracelular/metabolismo , Integrinas/biosíntesis , Sistema Nervioso Periférico/fisiología , Células de Schwann/citología , Animales , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Fibronectinas/farmacología , Laminina/farmacología , Ligandos , Ratones , Ratas , Células de Schwann/metabolismo , Vitronectina/farmacologíaRESUMEN
Thermophilic bacteria resembling Thermus aquaticus were isolated from hot water taken from domestic and commercial hot-water tanks. Cold water from the same locations never yielded thermophilic bacteria, suggesting that the bacteria were growing in the tanks. In contrast to the T. aquaticus isolates from hot springs, the present isolates were rarely pigmented. In general, the hotter sources more frequently yielded bacteria.