RESUMEN
Macrophages play an integral role in initiating innate immune defences and regulating inflammation. They are also involved in maintaining homeostasis and the resolution of inflammation, by promoting tissue repair and wound healing. There is evidence that like neutrophils, macrophages can release extracellular traps following exposure to a range of pathogenic and pro-inflammatory stimuli. Extracellular traps are released by a specialised cell death pathway termed 'ETosis', and consist of a backbone of DNA and histones decorated with a range of other proteins. The composition of extracellular trap proteins can be influenced by both the cell type and the local environment in which the traps are released. In many cases, these proteins have an antimicrobial role and assist with pathogen killing. Therefore, the release of extracellular traps serves as a means to both immobilise and destroy invading pathogens. In addition to their protective role, extracellular traps are also implicated in disease pathology. The release of neutrophil extracellular traps (NETs) is causally linked to the development of wide range of human diseases. However, whether macrophage extracellular traps (METs) play a similar role in disease pathology is less well established. Moreover, macrophages are also involved in the clearance of extracellular traps, which could assist in the resolution of tissue damage associated with the presence of extracellular traps. In this review, we will provide an overview of the pathways responsible for macrophage extracellular trap release, and discuss the role of these structures in innate immunity and disease pathology and possible therapeutic strategies.
Asunto(s)
Trampas Extracelulares , Trampas Extracelulares/metabolismo , Humanos , Inmunidad Innata , Inflamación/metabolismo , Macrófagos , Neutrófilos/metabolismoRESUMEN
We show how a bird's-eye view of genomic structure can be obtained at â¼1-kb resolution from long (â¼2 Mb) DNA molecules extracted from whole chromosomes in a nanofluidic laboratory-on-a-chip. We use an improved single-molecule denaturation mapping approach to detect repetitive elements and known as well as unique structural variation. Following its mapping, a molecule of interest was rescued from the chip; amplified and localized to a chromosome by FISH; and interrogated down to 1-bp resolution with a commercial sequencer, thereby reconciling haplotype-phased chromosome substructure with sequence.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos , ADN , Genoma Humano , Técnicas Analíticas Microfluídicas , Mapeo Cromosómico/instrumentación , Mapeo Cromosómico/métodos , Cromosomas Humanos/química , Cromosomas Humanos/genética , ADN/química , ADN/genética , Humanos , Hibridación Fluorescente in Situ/instrumentación , Hibridación Fluorescente in Situ/métodos , Masculino , Técnicas Analíticas Microfluídicas/instrumentación , Técnicas Analíticas Microfluídicas/métodosRESUMEN
Mitotic chromosome structure is pivotal to cell division but difficult to observe in fine detail using conventional methods. DNA catenation has been implicated in both sister chromatid cohesion and chromosome condensation, but has never been observed directly. We have used a lab-on-a-chip microfluidic device and fluorescence microscopy, coupled with a simple image analysis pipeline, to digest chromosomal proteins and examine the structure of the remaining DNA, which maintains the canonical 'X' shape. By directly staining DNA, we observe that DNA catenation between sister chromatids (separated by fluid flow) is composed of distinct fibres of DNA concentrated at the centromeres. Disrupting the catenation of the chromosomes with Topoisomerase IIα significantly alters overall chromosome shape, suggesting that DNA catenation must be simultaneously maintained for correct chromosome condensation, and destroyed to complete sister chromatid disjunction. In addition to demonstrating the value of microfluidics as a tool for examining chromosome structure, these results lend support to certain models of DNA catenation organization and regulation: in particular, we conclude from our observation of centromere-concentrated catenation that spindle forces could play a driving role in decatenation and that Topoisomerase IIα is differentially regulated at the centromeres, perhaps in conjunction with cohesin.
Asunto(s)
Cromosomas Humanos/ultraestructura , ADN Encadenado/ultraestructura , Metafase/genética , Cromátides/ultraestructura , Humanos , Técnicas Analíticas Microfluídicas , Microscopía FluorescenteRESUMEN
We describe the structure and operation of a micro/nanofluidic device in which individual metaphase chromosomes can be isolated and processed without being displaced during exchange of reagents. The change in chromosome morphology as a result of introducing protease into the device was observed by time-lapse imaging; pressure-driven flow was then used to shunt the chromosomal DNA package into a nanoslit. A long linear DNA strand (>1.3 Mbp) was seen to stretch out from the DNA package and along the length of the nanoslit. Delivery of DNA in its native metaphase chromosome package as well as the microfluidic environment prevented DNA from shearing and will be important for preparing ultra-long lengths of DNA for nanofluidic analysis.