RESUMEN
Microscopic lasers represent a promising tool for the development of cutting-edge photonic devices thanks to their ability to enhance light-matter interaction at the microscale. In this work, we realize liquid microlasers with tunable emission by exploiting the self-formation of three-dimensional liquid droplets into a polymeric matrix driven by viscoelastic dewetting. We design a flexible device to be used as a smart photonic label which is detachable and reusable on various types of substrates such as paper or fabric. The innovative lasing emission mechanism proposed here is based on whispering gallery mode emission coupled to random lasing, the latter prompted by the inclusion of dielectric compounds into the active gain medium. The wide possibility of modulating the emission wavelength of the microlasers by acting on different parameters, such as the cavity size, type and volume fraction of the dielectrics, and gain medium, offers a multitude of spectroscopic encoding schemes for the realization of photonic barcodes and labels to be employed in anticounterfeiting applications and multiplexed bioassays.
RESUMEN
Ultrasound (US) induced transient membrane permeabilisation has emerged as a hugely promising tool for the delivery of exogenous vectors through the cytoplasmic membrane, paving the way to the design of novel anticancer strategies by targeting functional nanomaterials to specific biological sites. An essential step towards this end is the detailed recognition of suitably marked nanoparticles in sonoporated cells and the investigation of the potential related biological effects. By taking advantage of Synchrotron Radiation Fourier Transform Infrared micro-spectroscopy (SR-microFTIR) in providing highly sensitive analysis at the single cell level, we studied the internalisation of a nanoprobe within fibroblasts (NIH-3T3) promoted by low-intensity US. To this aim we employed 20 nm gold nanoparticles conjugated with the IR marker 4-aminothiophenol. The significant Surface Enhanced Infrared Absorption provided by the nanoprobes, with an absorbance increase up to two orders of magnitude, allowed us to efficiently recognise their inclusion within cells. Notably, the selective and stable SR-microFTIR detection from single cells that have internalised the nanoprobe exhibited clear changes in both shape and intensity of the spectral profile, highlighting the occurrence of biological effects. Flow cytometry, immunofluorescence and murine cytokinesis-block micronucleus assays confirmed the presence of slight but significant cytotoxic and genotoxic events associated with the US-nanoprobe combined treatments. Our results can provide novel hints towards US and nanomedicine combined strategies for cell spectral imaging as well as drug delivery-based therapies.
Asunto(s)
Fibroblastos/metabolismo , Oro/química , Rayos Infrarrojos , Nanopartículas del Metal/química , Análisis de la Célula Individual , Sincrotrones , Ultrasonografía , Animales , Supervivencia Celular , Ratones , Micronúcleo Germinal/metabolismo , Células 3T3 NIH , Espectroscopía Infrarroja por Transformada de Fourier , Propiedades de SuperficieRESUMEN
Multiple myeloma (MM) is characterized by the impaired osteogenic differentiation of human mesenchymal stromal cells (hMSCs). Canonical Wnt signaling is critical for the regulation of bone formation, however, recent evidence suggests that the non-canonical Wnt agonist Wnt5a stimulates human osteoblastogenesis through its co-receptor Ror2. The effects of MM cells on non-canonical Wnt signaling and the effect of the activation of this pathway on MM-induced osteoblast exhaustion are not known and were investigated in this study. We found that the osteogenic differentiation of bone marrow hMSCs toward osteoprogenitor cells (PreOB) significantly increased Ror2 expression, and that MM cells inhibit Ror2 expression by PreOB in co-culture by inhibiting the non-canonical Wnt5a signaling. The activation of the non-canonical Wnt pathway in hMSCs by means of Wnt5a treatment and the overexpression of Wnt5 or Ror2 by lentiviral vectors increased the osteogenic differentiation of hMSCs and blunted the inhibitory effect of MM in co-culture. Consistently, Wnt5a inhibition by specific small interfering RNA reduced the hMSC expression of osteogenic markers. Our findings demonstrate that the Wnt5a/Ror2 pathway is involved in the pathophysiology of MM-induced bone disease and that the activation of the non-canonical Wnt5a/Ror2 pathway in hMSCs increases osteogenic differentiation and may counterbalance the inhibitory effect of MM cells.
Asunto(s)
Células de la Médula Ósea/citología , Diferenciación Celular , Mieloma Múltiple/patología , Osteoblastos/citología , Osteogénesis , Proteínas Proto-Oncogénicas/metabolismo , Receptores Huérfanos Similares al Receptor Tirosina Quinasa/metabolismo , Células Madre/citología , Proteínas Wnt/metabolismo , Animales , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Western Blotting , Células de la Médula Ósea/metabolismo , Estudios de Casos y Controles , Proliferación Celular , Técnicas de Cocultivo , Citometría de Flujo , Perfilación de la Expresión Génica , Humanos , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Ratones , Mieloma Múltiple/genética , Mieloma Múltiple/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Osteoblastos/metabolismo , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/genética , ARN Mensajero/genética , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Huérfanos Similares al Receptor Tirosina Quinasa/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Células Madre/metabolismo , Proteínas Wnt/antagonistas & inhibidores , Proteínas Wnt/genética , Proteína Wnt-5aRESUMEN
VP2 is the outermost Bluetongue virus (BTV) antigenic protein, forming triskelion motifs on the virion surface. Although VP2 has been expressed successfully through many systems, its paracrine expression as a soluble form by mammalian cells represents a difficult task. In the present paper two fragments of VP2 have been expressed successfully into the medium of transiently transfected mammalian cells through a fusion peptides strategy. The crude conditioned medium containing the secreted peptide could be employed for immunodiagnostic assay development or vaccine purposes.
Asunto(s)
Virus de la Lengua Azul/genética , Proteínas de la Cápside/genética , Proteínas de la Cápside/metabolismo , Línea Celular , Medios de Cultivo/química , Humanos , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismoRESUMEN
BoHV-4 replication cycle is dependent on the S-phase of the cell-cycle at the stage of viral DNA synthesis. Because p21 is a rate-limiting regulator of the G1/S-phase transition and up-regulated by DNA-damaging agents, in this study p21 expression in BoHV-4 infected cells was investigated. The p21 promoter was found to be highly activated in a dose- and time-dependent manner following BoHV-4 infection only in cells which are permissive for BoHV-4 replication. Thus p21 expression reports on BoHV-4 replication and could represent a host cell defensive response to infection-associated cellular damage.
Asunto(s)
Enfermedades de los Bovinos/virología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/biosíntesis , Infecciones por Herpesviridae/veterinaria , Herpesvirus Bovino 4/fisiología , Infecciones Tumorales por Virus/veterinaria , Replicación Viral , Animales , Biomarcadores , Bovinos , Enfermedades de los Bovinos/metabolismo , Línea Celular , Genes Reporteros , Infecciones por Herpesviridae/metabolismo , Infecciones por Herpesviridae/virología , Interacciones Huésped-Patógeno , Ratones , Regiones Promotoras Genéticas , Factores de Tiempo , Infecciones Tumorales por Virus/metabolismo , Infecciones Tumorales por Virus/virologíaRESUMEN
A bovine herpesvirus 4 was isolated from the milk cell fraction of a healthy cow and his full genome cloned as a bacterial artificial chromosome. So cloned viral genome was used as a vector platform to deliver in vitro and in vivo an optimized secreted chimeric peptide obtained by the fusion of the bovine viral diarrhoea virus glycoprotein E2 ectodomain with the bovine herpesvirus 1 glycoprotein D ectodomain. Recombinant virus infected cells robustly expressed and secreted the chimeric peptide into the culture medium and inoculated animals with the recombinant virus successfully responded toward antigens, gE2 and gD. Thus, this work has implications for the development of safe and effective polyvalent vaccines.