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1.
bioRxiv ; 2023 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-38077064

RESUMO

Neural organoids derived from human induced pluripotent stem cells (iPSCs) provide a model to study the earliest stages of human brain development, including neurogenesis, neural differentiation, and synaptogenesis. However, neural organoids lack supportive tissues and some non-neural cell types that are key regulators of brain development. Neural organoids have instead been co-cultured with non-neural structures and cell types to promote their maturation and model interactions with neuronal cells. One structure that does not form de novo with neural organoids is the meninges, a tri-layered structure that surrounds the CNS and secretes key signaling molecules required for mammalian brain development. Most studies of meninges-brain signaling have been performed in mice or using two-dimensional (2D) cultures of human cells, the latter not recapitulating the architecture and cellular diversity of the tissue. To overcome this, we developed a co-culture system of neural organoids generated from human iPSCs fused with fetal leptomeninges from mice with fluorescently labeled meninges (Col1a1-GFP). These proof-of-concept studies test the stability of the different cell types in the leptomeninges (fibroblast and macrophage) and the fused brain organoid (progenitor and neuron), as well as the interface between the organoid and meningeal tissue. We test the longevity of the fusion pieces after 30 days and 60 days in culture, describe best practices for preparing the meninges sample prior to fusion, and examine the feasibility of single or multiple meninges pieces fused to a single organoid. We discuss potential uses of the current version of the LMNO fusion model and opportunities to improve the system.

2.
Development ; 150(19)2023 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-37756588

RESUMO

Perivascular fibroblasts (PVFs) are a fibroblast-like cell type that reside on large-diameter blood vessels in the adult meninges and central nervous system (CNS). PVFs contribute to fibrosis following injury but their homeostatic functions are not defined. PVFs were previously shown to be absent from most brain regions at birth and are only detected postnatally within the cerebral cortex. However, the origin, timing and cellular mechanisms of PVF development are not known. We used Col1a1-GFP and Col1a2-CreERT2 transgenic mice to track PVF development postnatally. Using lineage tracing and in vivo imaging we show that brain PVFs originate from the meninges and are first seen on parenchymal cerebrovasculature at postnatal day (P) 5. After P5, PVF coverage of the cerebrovasculature expands via local cell proliferation and migration from the meninges. Finally, we show that PVFs and perivascular macrophages develop concurrently. These findings provide the first complete timeline for PVF development in the brain, enabling future work into how PVF development is coordinated with cell types and structures in and around the perivascular spaces to support normal CNS vascular function.

3.
bioRxiv ; 2023 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-36993587

RESUMO

Perivascular fibroblasts (PVFs) are a fibroblast-like cell type that reside on large-diameter blood vessels in the adult meninges and central nervous system (CNS). PVFs drive fibrosis following injury but their homeostatic functions are not well detailed. In mice, PVFs were previously shown to be absent from most brain regions at birth and are only detected postnatally within the cerebral cortex. However, the origin, timing, and cellular mechanisms of PVF development are not known. We used Col1a1-GFP and Col1a2-CreERT transgenic mice to track PVF developmental timing and progression in postnatal mice. Using a combination of lineage tracing and in vivo imaging we show that brain PVFs originate from the meninges and are first seen on parenchymal cerebrovasculature at postnatal day (P)5. After P5, PVF coverage of the cerebrovasculature rapidly expands via mechanisms of local cell proliferation and migration from the meninges, reaching adult levels at P14. Finally, we show that PVFs and perivascular macrophages (PVMs) develop concurrently along postnatal cerebral blood vessels, where the location and depth of PVMs and PVFs highly correlate. These findings provide the first complete timeline for PVF development in the brain, enabling future work into how PVF development is coordinated with cell types and structures in and around the perivascular spaces to support normal CNS vascular function. Summary: Brain perivascular fibroblasts migrate from their origin in the meninges and proliferate locally to fully cover penetrating vessels during postnatal mouse development.

4.
Dev Cell ; 58(8): 635-644.e4, 2023 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-36996816

RESUMO

The arachnoid barrier, a component of the blood-cerebrospinal fluid barrier (B-CSFB) in the meninges, is composed of epithelial-like, tight-junction-expressing cells. Unlike other central nervous system (CNS) barriers, its' developmental mechanisms and timing are largely unknown. Here, we show that mouse arachnoid barrier cell specification requires the repression of Wnt-ß-catenin signaling and that constitutively active ß-catenin can prevent its formation. We also show that the arachnoid barrier is functional prenatally and, in its absence, a small molecular weight tracer and the bacterium group B Streptococcus can cross into the CNS following peripheral injection. Acquisition of barrier properties prenatally coincides with the junctional localization of Claudin 11, and increased E-cadherin and maturation continues after birth, where postnatal expansion is marked by proliferation and re-organization of junctional domains. This work identifies fundamental mechanisms that drive arachnoid barrier formation, highlights arachnoid barrier fetal functions, and provides novel tools for future studies on CNS barrier development.


Assuntos
Meninges , beta Catenina , Camundongos , Animais , Aracnoide-Máter , Barreira Hematoencefálica , Sistema Nervoso Central , Junções Íntimas
5.
Res Sq ; 2023 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-38168409

RESUMO

Neural organoids derived from human induced pluripotent stem cells (iPSCs) provide a model to study the earliest stages of human brain development, including neurogenesis, neural differentiation, and synaptogenesis. However, neural organoids lack supportive tissues and some non-neural cell types that are key regulators of brain development. Neural organoids have instead been co-cultured with non-neural structures and cell types to promote their maturation and model interactions with neuronal cells. One structure that does not form de novo with neural organoids is the meninges, a tri-layered structure that surrounds the CNS and secretes key signaling molecules required for mammalian brain development. Most studies of meninges-brain signaling have been performed in mice or using two-dimensional (2D) cultures of human cells, the latter not recapitulating the architecture and cellular diversity of the tissue. To overcome this, we developed a co-culture system of neural organoids generated from human iPSCs fused with fetal leptomeninges from mice with fluorescently labeled meninges (Col1a1-GFP). These proof-of-concept studies test the stability of the different cell types in the leptomeninges (fibroblast and macrophage) and the fused brain organoid (progenitor and neuron), as well as the interface between the organoid and meningeal tissue. We test the longevity of the fusion pieces after 30 days and 60 days in culture, describe best practices for preparing the meninges sample prior to fusion, and examine the feasibility of single or multiple meninges pieces fused to a single organoid. We discuss potential uses of the current version of the LMNO fusion model and opportunities to improve the system.

6.
Nutrients ; 14(11)2022 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-35684062

RESUMO

The incidence of several diet and lifestyle-related diseases, previously seen only in adults, is increasing in prevalence in young people. The Black population, and particularly Black males, are at high risk of developing lifestyle-related diseases. Adolescence and young adulthood are considered a transitional period with increasing independence and responsibility, along with the development of lifelong lifestyle habits. This systematic review aimed to establish which methods and approaches to nutritional education interventions are the most effective in improving the nutritional/dietary intake in healthy young Black males. Eligibility criteria were designed using PICOS and included controlled trials of nutrition education interventions designed to improve dietary intake in healthy young Black or mixed-race males aged 14-21 years old. Medline, Cinahl and Scopus were searched in April 2021, resulting in 20,375 records being screened, and subsequently 72 full-text articles were reviewed. Risk of bias was assessed using the ROBINS-I tool. One study met the eligibility criteria. Results are presented in a narrative format as meta-analysis was not possible. This systematic review revealed a lack of evidence on the effectiveness of nutritional education interventions in this high-risk population. Limitations are noted and recommendations have been made.


Assuntos
Ingestão de Alimentos , Terapia Nutricional , Adolescente , Adulto , Dieta , Educação em Saúde , Humanos , Estilo de Vida , Masculino , Terapia Nutricional/métodos , Adulto Jovem
7.
Behav Sci (Basel) ; 12(6)2022 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-35735394

RESUMO

Service industry workers experience challenging labor conditions in the United States, including pay below the minimum wage, expected emotional labor, and harassment. Additionally, in part because they work long shifts in high stress environments in restaurants and bars, many build and form personal workplace relationships (PWRs). In 2021, we interviewed 38 service industry workers and managers during the COVID-19 pandemic where we examined occupational challenges they faced in the state of Texas, USA. Through our interpretive research, this essay showcases our inductive findings on how service industry workers and managers utilize communication to create and sustain PWRs. We identified how some PWRs are sustained through a unique form of occupational identification that cultivates a "service industry family", which we term familial personal workplace relationships (familial PWRs). This extends past organizational communication scholarship on family to consider occupational identification. Furthermore, our research reveals that while PWRs may build communities through care and support, they also perpetuate organizational violence, like sexual harassment and bullying.

8.
Neurophotonics ; 9(2): 021911, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35402637

RESUMO

Significance: Fibroblasts are found associated with blood vessels in various locations across the central nervous system (CNS): in the meninges, the choroid plexus, and in the parenchyma within perivascular spaces. CNS fibroblasts have been characterized using transcriptional profiling and a Col1a1-GFP mouse line used to identify CNS fibroblasts in vivo; however, we still know very little regarding their functions and identity. Aim: Current methods for visualizing CNS fibroblasts are lacking and, in particular, prevent adequate assessment of fibroblast-vessel interactions. We aimed to develop new ways to visualize CNS fibroblasts in greater detail. Approach: Here, we describe methods for whole mount visualization of meningeal and choroid plexus fibroblasts, and CUBIC optical tissue clearing methods for visualization of parenchymal vessel-associated fibroblasts. Results: We show that these methods can be used for visualization of vessel-fibroblast interactions in these CNS structures and provide significant improvement over traditional sectioning and staining methods. In addition, we can combine these techniques with immunohistochemistry methods for labeling different cell types in the meninges and blood vasculature as well as EdU-based cell proliferation assays. Conclusions: We expect these methods will advance studies of CNS fibroblast development and functions in homeostasis, injury, and disease.

9.
Nat Rev Neurosci ; 23(1): 23-34, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34671105

RESUMO

Recent transcriptomic, histological and functional studies have begun to shine light on the fibroblasts present in the meninges, choroid plexus and perivascular spaces of the brain and spinal cord. Although the origins and functions of CNS fibroblasts are still being described, it is clear that they represent a distinct cell population, or populations, that have likely been confused with other cell types on the basis of the expression of overlapping cellular markers. Recent work has revealed that fibroblasts play crucial roles in fibrotic scar formation in the CNS after injury and inflammation, which have also been attributed to other perivascular cell types such as pericytes and vascular smooth muscle cells. In this Review, we describe the current knowledge of the location and identity of CNS perivascular cell types, with a particular focus on CNS fibroblasts, including their origin, subtypes, roles in health and disease, and future areas for study.


Assuntos
Doenças do Sistema Nervoso Central/fisiopatologia , Sistema Nervoso Central/lesões , Sistema Nervoso Central/fisiologia , Fibroblastos/fisiologia , Animais , Sistema Nervoso Central/citologia , Humanos
10.
Front Cell Neurosci ; 15: 761506, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34690706

RESUMO

[This corrects the article DOI: 10.3389/fncel.2021.703944.].

11.
Trends Neurosci ; 44(11): 849-851, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34452754

RESUMO

The cerebral cortex requires a dense, highly organized network of vasculature that ensures high-volume and continuous oxygen delivery to metabolically active neuronal circuits. In a recent paper, Coelho-Santos et al. used in vivo two-photon microscopy to reveal how this precise network is constructed during a short window of mouse postnatal development.


Assuntos
Capilares , Pericitos , Animais , Encéfalo/irrigação sanguínea , Córtex Cerebral , Humanos , Camundongos , Vênulas
12.
Front Cell Neurosci ; 15: 703944, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34276313

RESUMO

The meninges are the fibrous covering of the central nervous system (CNS) which contain vastly heterogeneous cell types within its three layers (dura, arachnoid, and pia). The dural compartment of the meninges, closest to the skull, is predominantly composed of fibroblasts, but also includes fenestrated blood vasculature, an elaborate lymphatic system, as well as immune cells which are distinct from the CNS. Segregating the outer and inner meningeal compartments is the epithelial-like arachnoid barrier cells, connected by tight and adherens junctions, which regulate the movement of pathogens, molecules, and cells into and out of the cerebral spinal fluid (CSF) and brain parenchyma. Most proximate to the brain is the collagen and basement membrane-rich pia matter that abuts the glial limitans and has recently be shown to have regional heterogeneity within the developing mouse brain. While the meninges were historically seen as a purely structural support for the CNS and protection from trauma, the emerging view of the meninges is as an essential interface between the CNS and the periphery, critical to brain development, required for brain homeostasis, and involved in a variety of diseases. In this review, we will summarize what is known regarding the development, specification, and maturation of the meninges during homeostatic conditions and discuss the rapidly emerging evidence that specific meningeal cell compartments play differential and important roles in the pathophysiology of a myriad of diseases including: multiple sclerosis, dementia, stroke, viral/bacterial meningitis, traumatic brain injury, and cancer. We will conclude with a list of major questions and mechanisms that remain unknown, the study of which represent new, future directions for the field of meninges biology.

13.
Dev Cell ; 54(1): 43-59.e4, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32634398

RESUMO

The meninges are a multilayered structure composed of fibroblasts, blood and lymphatic vessels, and immune cells. Meningeal fibroblasts secrete a variety of factors that control CNS development, yet strikingly little is known about their heterogeneity or development. Using single-cell sequencing, we report distinct transcriptional signatures for fibroblasts in the embryonic dura, arachnoid, and pia. We define new markers for meningeal layers and show conservation in human meninges. We find that embryonic meningeal fibroblasts are transcriptionally distinct between brain regions and identify a regionally localized pial subpopulation marked by the expression of µ-crystallin. Developmental analysis reveals a progressive, ventral-to-dorsal maturation of telencephalic meninges. Our studies have generated an unparalleled view of meningeal fibroblasts, providing molecular profiles of embryonic meningeal fibroblasts by layer and yielding insights into the mechanisms of meninges development and function.


Assuntos
Encéfalo/metabolismo , Fibroblastos/metabolismo , Meninges/metabolismo , Transcriptoma , Animais , Encéfalo/citologia , Encéfalo/embriologia , Cristalinas/genética , Cristalinas/metabolismo , Humanos , Meninges/citologia , Meninges/embriologia , Camundongos , Camundongos Endogâmicos C57BL , RNA-Seq , Análise de Célula Única
14.
Front Plant Sci ; 10: 1757, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-32161600

RESUMO

Most of our crops are grown in monoculture with single genotypes grown over wide acreage. An alternative approach, where segregating populations are used as crops, is an exciting possibility, but outcomes of natural selection upon this type of crop are not well understood. We tracked allelic frequency changes in evolving composite cross populations of wheat grown over 10 generations under organic and conventional farming. At three generations, each population was genotyped with 19 SSR and 8 SNP markers. The latter were diagnostic for major functional genes. Gene diversity was constant at SSR markers but decreased over time for SNP markers. Population differentiation between the four locations could not be detected, suggesting that organic vs. non-organic crop management did not drive allele frequency changes. However, we did see changes for genes controlling plant height and phenology in all populations independently and consistently. We interpret these changes as the result of a consistent natural selection towards wild-type. Independent selection for alleles that are associated with plant height suggests that competition for light was central, resulting in the predominance of stronger intraspecific competitors, and highlighting a potential trade-off between individual and population performance.

15.
Appl Soil Ecol ; 120: 35-43, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29104370

RESUMO

Rhizobia play important roles in agriculture owing to their ability to fix nitrogen through a symbiosis with legumes. The specificity of rhizobia-legume associations means that underused legume species may depend on seed inoculation with their rhizobial partners. For black medic (Medicago lupulina) and lucerne (Medicago sativa) little is known about the natural prevalence of their rhizobial partner Ensifer meliloti in UK soils, so that the need for inoculating them is unclear. We analysed the site-dependence of rhizobial seed inoculation effects on the subsequent ability of rhizobial communities to form symbioses with four legume species (Medicago lupulina, M. sativa, Trifolium repens and T. pratense). At ten organic farms across the UK, a species-diverse legume based mixture (LBM) which included these four species was grown. The LBM seed was inoculated with a mix of commercial inocula specific for clover and lucerne. At each site, soil from the LBM treatment was compared to the soil sampled prior to the sowing of the LBM (the control). From each site and each of the two treatments, a suspension of soils was applied to seedlings of the four legume species and grown in axenic conditions for six weeks. Root nodules were counted and their rhizobia isolated. PCR and sequencing of a fragment of the gyrB gene from rhizobial isolates allowed identification of strains. The number of nodules on each of the four legume species was significantly increased when inoculated with soil from the LBM treatment compared to the control. Both the proportion of plants forming nodules and the number of nodules formed varied significantly by site, with sites significantly affecting the Medicago species but not the Trifolium species. These differences in nodulation were broadly reflected in plant biomass where site and treatment interacted; at some sites there was a significant advantage from inoculation with the commercial inoculum but not at others. In particular, this study has demonstrated the commercial merit of inoculation of lucerne with compatible rhizobia.

16.
J Exp Bot ; 68(8): 2055-2063, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-27927999

RESUMO

Climate change can threaten the reproductive success of plants, both directly, through physiological damage during increasingly extreme weather events, and indirectly, through disruption of plant-pollinator interactions. To explore how plant-pollinator interactions are modified by extreme weather, we exposed faba bean (Vicia faba) plants to elevated temperature for 5 d during flowering, simulating a heatwave. We then moved the plants to flight cages with either bumblebees or no pollinators, or to two field sites, where plants were enclosed in mesh bags or pollinated by wild insect communities. We used a morphological marker to quantify pollen movement between experimental plants. There was a substantial increase in the level of outcrossing by insect pollinators following heat stress. Proportion outcrossed seed increased from 17 % at control temperature, to 33 % following heat stress in the flight cages, and from 31 % to 80 % at one field site, but not at the other (33 % to 32 %). Abiotic stress can dramatically shift the relative contributions of cross- and self-pollination to reproduction in an insect pollinated plant. The resulting increases in gene flow have broad implications for genetic diversity and functioning of ecosystems, and may increase resilience by accelerating the selection of more stress-tolerant genotypes.


Assuntos
Mudança Climática , Produtos Agrícolas/fisiologia , Temperatura Alta , Polinização/fisiologia , Reprodução/fisiologia , Vicia faba/fisiologia , Animais , Insetos
17.
PLoS One ; 9(10): e111225, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25356771

RESUMO

A variety of foods have been implicated in symptoms of patients with Irritable Bowel Syndrome (IBS) but wheat products are most frequently cited by patients as a trigger. Our aim was to investigate the effects of breads, which were fermented for different lengths of time, on the colonic microbiota using in vitro batch culture experiments. A set of in vitro anaerobic culture systems were run over a period of 24 h using faeces from 3 different IBS donors (Rome Criteria-mainly constipated) and 3 healthy donors. Changes in gut microbiota during a time course were identified by fluorescence in situ hybridisation (FISH), whilst the small-molecular weight metabolomic profile was determined by NMR analysis. Gas production was separately investigated in non pH-controlled, 36 h batch culture experiments. Numbers of bifidobacteria were higher in healthy subjects compared to IBS donors. In addition, the healthy donors showed a significant increase in bifidobacteria (P<0.005) after 8 h of fermentation of a bread produced using a sourdough process (type C) compared to breads produced with commercial yeasted dough (type B) and no time fermentation (Chorleywood Breadmaking process) (type A). A significant decrease of δ-Proteobacteria and most Gemmatimonadetes species was observed after 24 h fermentation of type C bread in both IBS and healthy donors. In general, IBS donors showed higher rates of gas production compared to healthy donors. Rates of gas production for type A and conventional long fermentation (type B) breads were almost identical in IBS and healthy donors. Sourdough bread produced significantly lower cumulative gas after 15 h fermentation as compared to type A and B breads in IBS donors but not in the healthy controls. In conclusion, breads fermented by the traditional long fermentation and sourdough are less likely to lead to IBS symptoms compared to bread made using the Chorleywood Breadmaking Process.


Assuntos
Pão , Microbioma Gastrointestinal , Síndrome do Intestino Irritável/microbiologia , Adulto , Técnicas de Cultura Celular por Lotes , Ácidos Graxos Voláteis/biossíntese , Fezes/microbiologia , Feminino , Fermentação , Farinha , Gases/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Hibridização in Situ Fluorescente , Cinética , Masculino , Metaboloma , Metabolômica
18.
Cell Microbiol ; 16(4): 519-34, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24152255

RESUMO

Outer membrane vesicles (OMV) are released by many bacteria, and contain immunogenic antigens in addition to harmful inflammatory factors, like lipopolysaccharides. Chemically detoxified OMV have been used in vaccines against Neisseria meningitidis (Nm); however, little is known about their interaction with antigen presenting cells. In this study, we investigated the interaction of Nm OMV with human dendritic cells (DC) to gain further understanding of their biological activity. We engineered a novel serogroup B Nm that is unencapsulated (siaD), expresses pentacylated lipid A (lpxL1), hence conferring reduced toxicity, and expresses an lgtB oligosaccharide structure designed to target OMV to DC via DC-SIGN. We show that the lgtB moiety is critical for internalization of NOMV by DC. Furthermore, the lgtB moiety significantly enhances DC maturation, IL-10 and IL-23 production in the presence of a pentacylated lipid A. While different DC phenotypes were observed for each NOMV, this had little effect on Th1 and Th2 cell differentiation; however, lgtBsignificantly increased Th17 cell expansion in the presence of pentacylated lipid A. We believe that lpxL1/lgtB NOMV should be considered further as a vaccine vector, particularly considering the importance of lgtB in antigen uptake and further human studies on antigen-specific responses should be considered.


Assuntos
Micropartículas Derivadas de Células/metabolismo , Células Dendríticas/imunologia , Lipídeo A/imunologia , Neisseria meningitidis Sorogrupo B/imunologia , Oligossacarídeos/metabolismo , Células Cultivadas , Humanos , Lipídeo A/toxicidade
19.
Funct Plant Biol ; 40(7): 700-707, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32481142

RESUMO

Thermal imaging is a valuable tool for the clarification of gas exchange dynamics between a plant and its environment. The presence of stomata in wheat (Triticum aestivum L.) glumes and awns offers an opportunity to assess the photosynthetic activity of ears up to and during flowering. Knowledge of the spatial and temporal thermodynamics of the wheat ear may provide insight into interactions between floret developmental stage (FDS), temperature depression (TD) and ambient environment, with potential to use these as high-throughput screening tools for breeders. A controlled environment study was conducted using six spring wheat genotypes of the elite recombinant inbred line Seri-Babax. Average ear temperature was recorded using a hand-held infrared camera and gas exchange was measured by enclosing ears in a custom-built cuvette. FDS was monitored and recorded daily throughout the study. Plants were grown in pots and exposed to a combination of two temperature and two water regimes. In the studied wheat lines, TD varied from 0.1°C to 0.6°C according to the level of stress imposed. The results indicated that TD does not occur at FDS F3, the peak of active flowering, but during the stages before pollen release and stigma maturity (F1-F2). These findings suggest that ear temperature during the early stages of anthesis, before pollen release and full extension of the stigma, are likely to be the most relevant for identifying heat stress tolerant genotypes.

20.
Methods Mol Biol ; 799: 217-35, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-21993649

RESUMO

Dendritic cells (DC) play a key role in the development of natural immunity to microbes. The DC form a bridge between the innate and adaptive immune system by providing key instructions particularly to antigen naïve T-cells. The interaction of DC with T lymphocytes involves three signals: (1) antigen processing and presentation in context of MHC Class I and/or II, (2) expression of T cell co-stimulatory molecules, and (3) cytokine production. Studying the interactions of DCs with specific pathogens allows for better understanding of how protective immunity is generated, and may be particularly useful for assessing vaccine components. In this chapter, we describe methods to generate human monocyte-derived DCs and assess their maturation, activation, and function, using interaction with the gram-negative bacterial pathogen Neisseria meningitidis as a model.


Assuntos
Técnicas de Cultura de Células/métodos , Células Dendríticas/citologia , Células Dendríticas/microbiologia , Proteínas de Membrana/metabolismo , Neisseria meningitidis/imunologia , Linfócitos T/imunologia , Citocinas/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Citometria de Fluxo , Humanos , Receptores de Lipopolissacarídeos , Microscopia Confocal , Monócitos/citologia
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