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Visualisation of chicken macrophages using transgenic reporter genes: insights into the development of the avian macrophage lineage.

Balic A, Garcia-Morales C, Vervelde L, Gilhooley H, Sherman A, Garceau V, Gutowska MW, Burt DW, Kaiser P, Hume DA, Sang HM - Development (2014)

Bottom Line: The Fms intronic regulatory element (FIRE) within CSF1R is shown to be highly conserved in amniotes and absolutely required for myeloid-restricted expression of fluorescent reporter genes.The cell lineage specificity of reporter gene expression was confirmed by demonstration of coincident expression with the endogenous CSF1R protein.In transgenic birds, expression of the reporter gene provided a defined marker for macrophage-lineage cells, identifying the earliest stages in the yolk sac, throughout embryonic development and in all adult tissues.

View Article: PubMed Central - PubMed

Affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Sciences, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK adam.balic@roslin.ed.ac.uk helen.sang@roslin.ed.ac.uk.

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Macrophages associated with the embryonic vasculature are highly motile and phagocytic, and undergo local division. Time-lapse imaging of region above the vitelline artery near the embryo proper. The aorta of CSF1R-eGFP embryos was injected with Texas Red-labelled zymosan 1 h prior to the beginning of imaging. Most zymosan particles adhered to the blood vessel walls (yellow arrows). eGFP+ macrophages are highly motile. Between 100 and 125 min from the start of filming, a zymosan particle (yellow arrow) becomes associated with a macrophage; this macrophage re-enters the circulation, removing the zymosan particle by 150 min. At 0 min, a zymosan particle is contained within a macrophage (white arrow); from 0-75 min this macrophage is both motile and exhibits changes in morphology. At 100 min, this macrophage (white arrow) no longer exhibits movement and does not extend any cellular processes. A similar macrophage without a phagocytised zymosan particle (blue arrow) exhibits identical behaviour. At 100-150 min, both undergo division (white and blue arrows), and daughter cells resume active patrolling of the vasculature. Scale bar: 50 µm.
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DEV105593F4: Macrophages associated with the embryonic vasculature are highly motile and phagocytic, and undergo local division. Time-lapse imaging of region above the vitelline artery near the embryo proper. The aorta of CSF1R-eGFP embryos was injected with Texas Red-labelled zymosan 1 h prior to the beginning of imaging. Most zymosan particles adhered to the blood vessel walls (yellow arrows). eGFP+ macrophages are highly motile. Between 100 and 125 min from the start of filming, a zymosan particle (yellow arrow) becomes associated with a macrophage; this macrophage re-enters the circulation, removing the zymosan particle by 150 min. At 0 min, a zymosan particle is contained within a macrophage (white arrow); from 0-75 min this macrophage is both motile and exhibits changes in morphology. At 100 min, this macrophage (white arrow) no longer exhibits movement and does not extend any cellular processes. A similar macrophage without a phagocytised zymosan particle (blue arrow) exhibits identical behaviour. At 100-150 min, both undergo division (white and blue arrows), and daughter cells resume active patrolling of the vasculature. Scale bar: 50 µm.

Mentions: Vitelline vasculature macrophages imaged in HH17 MacGreen embryos were highly motile and were observed both within blood vessels and in a perivascular locations but not integrated into the blood vessel walls, as described by Al-Roubaie et al. (2012). Within blood vessels, macrophages were observed crawling on the blood vessel walls, both as isolated cells and as cell clusters (supplementary material Movie 2). This crawling behaviour is reminiscent of ‘patrolling’ behaviour reported for a subset of blood monocytes in mice that respond to microbial infection (Auffray et al., 2007). A well-established model for studying the interactions of microbes with phagocytes is the recognition and phagocytosis of microbial-derived zymosan particles (Underhill, 2003). We determined the capacity of patrolling macrophages within the vitelline blood vessels to recognise and phagocytose zymosan particles by injection of Texas Red-labelled zymosan particles into the dorsal aorta of HH17 MacGreen embryos. These particles were observed throughout the embryonic and extra-embryonic vasculature where they stuck to the blood vessel walls. Patrolling macrophages moved towards and engulfed zymosan particles, and then either continued to crawl along the vessel walls or entered the circulation (Fig. 4; supplementary material Movie 3). Cell division of patrolling embryonic macrophages associated with the vasculature was frequently observed and macrophages containing zymosan particles were also divided. This process involved the cessation of patrolling behaviour, retraction of cellular processes and rounding of cells before cell division. After cell division, both daughter cells resumed a ramified morphology and patrolling behaviour (Fig. 4; supplementary material Movie 3), indicating that mature yolk sac-derived macrophages are a self-renewing population.Fig. 4.


Visualisation of chicken macrophages using transgenic reporter genes: insights into the development of the avian macrophage lineage.

Balic A, Garcia-Morales C, Vervelde L, Gilhooley H, Sherman A, Garceau V, Gutowska MW, Burt DW, Kaiser P, Hume DA, Sang HM - Development (2014)

Macrophages associated with the embryonic vasculature are highly motile and phagocytic, and undergo local division. Time-lapse imaging of region above the vitelline artery near the embryo proper. The aorta of CSF1R-eGFP embryos was injected with Texas Red-labelled zymosan 1 h prior to the beginning of imaging. Most zymosan particles adhered to the blood vessel walls (yellow arrows). eGFP+ macrophages are highly motile. Between 100 and 125 min from the start of filming, a zymosan particle (yellow arrow) becomes associated with a macrophage; this macrophage re-enters the circulation, removing the zymosan particle by 150 min. At 0 min, a zymosan particle is contained within a macrophage (white arrow); from 0-75 min this macrophage is both motile and exhibits changes in morphology. At 100 min, this macrophage (white arrow) no longer exhibits movement and does not extend any cellular processes. A similar macrophage without a phagocytised zymosan particle (blue arrow) exhibits identical behaviour. At 100-150 min, both undergo division (white and blue arrows), and daughter cells resume active patrolling of the vasculature. Scale bar: 50 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4197536&req=5

DEV105593F4: Macrophages associated with the embryonic vasculature are highly motile and phagocytic, and undergo local division. Time-lapse imaging of region above the vitelline artery near the embryo proper. The aorta of CSF1R-eGFP embryos was injected with Texas Red-labelled zymosan 1 h prior to the beginning of imaging. Most zymosan particles adhered to the blood vessel walls (yellow arrows). eGFP+ macrophages are highly motile. Between 100 and 125 min from the start of filming, a zymosan particle (yellow arrow) becomes associated with a macrophage; this macrophage re-enters the circulation, removing the zymosan particle by 150 min. At 0 min, a zymosan particle is contained within a macrophage (white arrow); from 0-75 min this macrophage is both motile and exhibits changes in morphology. At 100 min, this macrophage (white arrow) no longer exhibits movement and does not extend any cellular processes. A similar macrophage without a phagocytised zymosan particle (blue arrow) exhibits identical behaviour. At 100-150 min, both undergo division (white and blue arrows), and daughter cells resume active patrolling of the vasculature. Scale bar: 50 µm.
Mentions: Vitelline vasculature macrophages imaged in HH17 MacGreen embryos were highly motile and were observed both within blood vessels and in a perivascular locations but not integrated into the blood vessel walls, as described by Al-Roubaie et al. (2012). Within blood vessels, macrophages were observed crawling on the blood vessel walls, both as isolated cells and as cell clusters (supplementary material Movie 2). This crawling behaviour is reminiscent of ‘patrolling’ behaviour reported for a subset of blood monocytes in mice that respond to microbial infection (Auffray et al., 2007). A well-established model for studying the interactions of microbes with phagocytes is the recognition and phagocytosis of microbial-derived zymosan particles (Underhill, 2003). We determined the capacity of patrolling macrophages within the vitelline blood vessels to recognise and phagocytose zymosan particles by injection of Texas Red-labelled zymosan particles into the dorsal aorta of HH17 MacGreen embryos. These particles were observed throughout the embryonic and extra-embryonic vasculature where they stuck to the blood vessel walls. Patrolling macrophages moved towards and engulfed zymosan particles, and then either continued to crawl along the vessel walls or entered the circulation (Fig. 4; supplementary material Movie 3). Cell division of patrolling embryonic macrophages associated with the vasculature was frequently observed and macrophages containing zymosan particles were also divided. This process involved the cessation of patrolling behaviour, retraction of cellular processes and rounding of cells before cell division. After cell division, both daughter cells resumed a ramified morphology and patrolling behaviour (Fig. 4; supplementary material Movie 3), indicating that mature yolk sac-derived macrophages are a self-renewing population.Fig. 4.

Bottom Line: The Fms intronic regulatory element (FIRE) within CSF1R is shown to be highly conserved in amniotes and absolutely required for myeloid-restricted expression of fluorescent reporter genes.The cell lineage specificity of reporter gene expression was confirmed by demonstration of coincident expression with the endogenous CSF1R protein.In transgenic birds, expression of the reporter gene provided a defined marker for macrophage-lineage cells, identifying the earliest stages in the yolk sac, throughout embryonic development and in all adult tissues.

View Article: PubMed Central - PubMed

Affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Sciences, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK adam.balic@roslin.ed.ac.uk helen.sang@roslin.ed.ac.uk.

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Related in: MedlinePlus