Characterization of zebrafish larval inflammatory macrophages

Mathias, Jonathan R.; Dodd, Mary; Walters, Kevin B.; Yoo, Sa Kan; Ranheim, Erik A.; Huttenlocher, Anna

doi:10.1016/j.dci.2009.07.003

Cited by 125 publications

(124 citation statements)

References 45 publications

(83 reference statements)

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“…Monocytes, macrophages and neutrophils are professional phagocytes; however other cells can perform phagocytosis so that the cell requires the detection of the foreign particle through membrane receptors. Cytokines, molecular signals of inflammation, are released by those phagocytes at the inflammation site by injured tissue so as to promote chemotaxis and phagocyte mobilisation (Stuart and Ezekowitz, 2005;Mathias et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Leukocytes respiratory burst activity as indicator of innate immunity of pacu Piaractus mesopotamicus

et al. 2013

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The present study evaluated the assay to quantify the respiratory burst activity of blood leukocytes of pacu as an indicator of the innate immune system, using the reduction of nitroblue tetrazolium (NBT) to formazan as a measure of the production of reactive oxygen species (ROS). In order to assess the accuracy of the assay, fish were challenged by Aeromonas hydrophila and sampled one week after challenge. The A. hydrophila infection increased the leukocyte respiratory burst activity. The protocol showed a reliable and easy assay, appropriate to determine the respiratory burst activity of blood leukocytes of pacu, a neotropical fish, in the present experimental conditions. Keywords: immune methodology, fish immunology, cellular immunity. Atividade respiratória de leucócitos como indicador de imunidade inata de pacu (Piaractus mesopotamicus)Resumo O presente estudo avaliou o ensaio para quantificar a atividade respiratória dos leucócitos do sangue de pacu como um indicador do sistema imune inato, usando a redução do nitroazul tetrazólio (NBT) a formazan como medida da produção de espécies reativas de oxigênio (EROs). Para avaliar a precisão do ensaio, peixes foram desafiados por Aeromonas hydrophila e amostrados uma semana após o desafio. A infecção com A. hydrophila aumentou a atividade respiratória dos leucócitos. O protocolo se mostrou confiável e de fácil aplicação, apropriado para determinar a atividade respiratória de leucócitos do sangue do pacu, peixe neotropical, nas condições experimentais apresentadas.Palavras-chave: metodologia, imunidade de peixe, imunidade mediada por células.

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Section: Introductionmentioning

confidence: 99%

Leukocytes respiratory burst activity as indicator of innate immunity of pacu Piaractus mesopotamicus

et al. 2013

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“…Neutrophils and macrophages in zebrafish larvae can be visualized by immunolabeling with anti-L-plastin antibody, whereas the expression of csf1r (colony-stimulating factor-1 receptor) and the lack of mpx and lyz expression in macrophages can be used to distinguish them from neutrophils. 23,26 The elongated morphology of the cells expressing these markers further points to a macrophage identity. 23 Both the neutrophil and the macrophage populations are involved in the innate immune response toward infection and injury.…”

mentioning

confidence: 99%

“…23 Both the neutrophil and the macrophage populations are involved in the innate immune response toward infection and injury. [23][24][25][26] In order to understand the fate and interaction of nanomaterials with the immune cells, further studies are needed.…”

mentioning

confidence: 99%

“…Nanostructures that are coated with the polymer polyethylene glycol (PEG) have shown to be more resistant to uptake by phagocytic cells. 22 Zebrafish transgenic lines that express green or red fluorescent proteins (DsRED2) under a neutrophil-specific promoter, such as the myeloperoxidase (mpx) promoter 23,24 or the lysozyme C (lyz) promoter, 25 provide useful in vivo models for real-time imaging and genetic analyses of inflammatory responses. Neutrophils and macrophages in zebrafish larvae can be visualized by immunolabeling with anti-L-plastin antibody, whereas the expression of csf1r (colony-stimulating factor-1 receptor) and the lack of mpx and lyz expression in macrophages can be used to distinguish them from neutrophils.…”

mentioning

confidence: 99%

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Mesoporous silica nanoparticles as a compound delivery system in zebrafish embryos

Sharif¹,

Porta²,

Meijer³

et al. 2012

IJN

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Silica nanoparticles can be efficiently employed as carriers for therapeutic drugs in vitro. Here, we use zebrafish embryos as a model organism to see whether mesoporous silica nanoparticles (MSNPs) can be incorporated to deliver compounds in vivo. We injected 35-40 nL (10 mg/mL) of custom-synthesized, fluorescently-tagged 200 nm MSNPs into the left flank, behind the yolk sac extension, of 2-day-old zebrafish embryos. We tracked the distribution and translocation of the MSNPs using confocal laser scanning microscopy. Some of the particles remained localized at the injection site, whereas others entered the bloodstream and were carried around the body. Embryo development and survival were not significantly affected by MSNP injection. Acridine orange staining revealed that MSNP injections did not induce significant cell death. We also studied cellular immune responses by means of lysC::DsRED2 transgenic embryos. MSNP-injected embryos showed infiltration of the injection site with neutrophils, similar to controls injected with buffer only. In the same embryos, counterstaining with L-plastin antibody for leukocytes revealed the same amount of cellular infiltration of the injection site in embryos injected with MSNPs or with buffer only. Next, we used MSNPs to deliver two recombinant cytokines (macrophage colony-stimulating factor and receptor for necrosis factor ligand) to zebrafish embryos. These proteins are known to activate cells involved in bone remodeling, and this can be detected with the marker tartrate-resistant acid phosphatase. Coinjection of these proteins loaded onto MSNPs produced a significant increase in the number of tartrate-resistant acid phosphatase-positive cells after 2-3 days of injection. Our results show that MSNPs can be used to deliver bioactive compounds into zebrafish larvae without producing higher mortality or gross evidence of teratogenicity.

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“…The first laboratory exercise, which was performed during laboratory sessions 1 and 2, required students to identify the cell type expressing enhanced green fluorescent protein (EGFP) in a transgenic zebrafish line by comparing forward [25][26][27][28][29][30].…”

Section: Laboratory Exercise 1: Identification Of Transgenic Zebrafismentioning

confidence: 99%

Immunological tools: Engaging students in the use and analysis of flow cytometry and enzyme‐linked immunosorbent assay (ELISA)

Ott

Carson

2014

Biochem Molecular Bio Educ

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Flow cytometry and enzyme-linked immunosorbent assay (ELISA) are commonly used techniques associated with clinical and research applications within the immunology and medical fields. The use of these techniques is becoming increasingly valuable in many life science and engineering disciplines as well. Herein, we report the development and evaluation of a novel half-semester course that focused on introducing undergraduate and graduate students to advance conceptual and technical skills associated with flow cytometry and ELISA, with emphasis on applications, experimental design, and data analysis. This course was offered in the North Carolina State University Biotechnology Program over three semesters and consisted of weekly lectures and laboratories. Students performed and/or analyzed flow cytometry and ELISA in three separate laboratory exercises:(1) identification of transgenic zebrafish hematopoietic cells, (2) analysis of transfection efficiency, and (3) analysis of cytokine production upon lipopolysaccharide stimulation. Student learning outcomes were achieved as demonstrated by multiple means of assessment, including three laboratory reports, a data analysis laboratory practicum, and a cumulative final exam. Further, anonymous student selfassessment revealed increased student confidence in the knowledge and skill sets defined in the learning outcomes.

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Characterization of zebrafish larval inflammatory macrophages

Cited by 125 publications

References 45 publications

Leukocytes respiratory burst activity as indicator of innate immunity of pacu Piaractus mesopotamicus

Leukocytes respiratory burst activity as indicator of innate immunity of pacu Piaractus mesopotamicus

Mesoporous silica nanoparticles as a compound delivery system in zebrafish embryos

Immunological tools: Engaging students in the use and analysis of flow cytometry and enzyme‐linked immunosorbent assay (ELISA)

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