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In Vivo Non-Invasive Tracking of Macrophage Recruitment to Experimental Stroke.

Selt M, Tennstaedt A, Beyrau A, Nelles M, Schneider G, Löwik C, Hoehn M - PLoS ONE (2016)

Bottom Line: However, the underlying molecular mechanisms needed for proper regulation still remain to be elucidated.The labelling procedure of the isolated cells did not significantly influence cell characteristics and resulted in detection of as few as 500 labelled cells in vivo.Hypointensity in MRI of the graft appeared unaltered in spatial location.

View Article: PubMed Central - PubMed

Affiliation: In-vivo-NMR Laboratory, Max Planck Institute for Metabolism Research, Cologne, Germany.

ABSTRACT
Brain-infiltrating monocyte-derived macrophages are one of the key players in the local immune response after stroke. It is now widely accepted that the inflammatory response is not an exclusively destructive process. However, the underlying molecular mechanisms needed for proper regulation still remain to be elucidated. Here, we propose an in vitro labelling strategy for multimodal in vivo observation of macrophage dynamics distinguished from brain-residing microglia response. Prior to intracerebral transplantation into the striatum of recipient mice or systemic administration, monocytes and macrophages, isolated from luciferase-expressing mice, were labelled with superparamagnetic iron oxide particles. Temporo-spatial localization was monitored by magnetic resonance imaging, whereas survival of grafted cells was investigated using bioluminescence imaging. The labelling procedure of the isolated cells did not significantly influence cell characteristics and resulted in detection of as few as 500 labelled cells in vivo. Two weeks after stereotactic transplantation, the luciferase signal was sustained traceable, with approximately 18% of the original luciferase signal detectable for monocytes and about 30% for macrophages. Hypointensity in MRI of the graft appeared unaltered in spatial location. In a therapeutically relevant approach, systemic cell administration after stroke resulted in accumulation mostly in thoracic regions, as could be visualized with BLI. For detection of homing to ischemic brain tissue more cells need to be administered. Nevertheless, during parallel MRI sessions recruitment of i.v. injected cells to the lesion site could be detected by day 2 post stroke as scattered hypointense signal voids. With further increase in sensitivity, our multi-facetted labelling strategy will provide the basis for in vivo tracking and fate specification of tissue-infiltrating macrophages and their distinct role in stroke-related neuro-inflammation.

No MeSH data available.


Related in: MedlinePlus

Representative dot plots and histograms of flow cytometric investigations of the effect of SPIO labelling on cell functionality.(A) Unlabelled control (upper row) and Nanomag labelled luc+ monocytes (middle row) were stained against monocytic markers Ly6c and CD115. Upon iron particle labelling, monocytes reduced their expression levels for CD115, which is clearly visible in the depicted histogram for CD115 marker (lower row). (B) Luc+ MΦ without and with SPIO-labelling were tested for their expression levels of CD68, CD80, CD86, CD206 and F4/80. The monocytic markers Ly6c and CD115 served as negative controls. For both, labelled and unlabelled MΦ, F4/80-positive cells were also positive for CD68, CD80, CD86 and CD206. No change in expression levels of Ly6c could be detected upon particle uptake, whereas the CD115+ population of control cells minimized due to contrast agent incorporation. n = 2 for both cell types.
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pone.0156626.g004: Representative dot plots and histograms of flow cytometric investigations of the effect of SPIO labelling on cell functionality.(A) Unlabelled control (upper row) and Nanomag labelled luc+ monocytes (middle row) were stained against monocytic markers Ly6c and CD115. Upon iron particle labelling, monocytes reduced their expression levels for CD115, which is clearly visible in the depicted histogram for CD115 marker (lower row). (B) Luc+ MΦ without and with SPIO-labelling were tested for their expression levels of CD68, CD80, CD86, CD206 and F4/80. The monocytic markers Ly6c and CD115 served as negative controls. For both, labelled and unlabelled MΦ, F4/80-positive cells were also positive for CD68, CD80, CD86 and CD206. No change in expression levels of Ly6c could be detected upon particle uptake, whereas the CD115+ population of control cells minimized due to contrast agent incorporation. n = 2 for both cell types.

Mentions: Influence of SPIO-labelling on functionality of luc+ monocytes and differentiated luc+ MΦ was analysed via flow cytometric measurements. Labelled (168 μg Fe/ml) as well as unlabelled control monocytes were stained for the monocytic markers CD115 and Ly6c (Fig 4A). 86.6% of the unlabelled control cells were double positive for both tested markers. Only a small population (10.7%) did not express CD115. This situation changed upon contrast agent uptake. Only 13.5% of the cells were double positive, but 84.5% reduced their CD115-expression and only expressed Ly6c.


In Vivo Non-Invasive Tracking of Macrophage Recruitment to Experimental Stroke.

Selt M, Tennstaedt A, Beyrau A, Nelles M, Schneider G, Löwik C, Hoehn M - PLoS ONE (2016)

Representative dot plots and histograms of flow cytometric investigations of the effect of SPIO labelling on cell functionality.(A) Unlabelled control (upper row) and Nanomag labelled luc+ monocytes (middle row) were stained against monocytic markers Ly6c and CD115. Upon iron particle labelling, monocytes reduced their expression levels for CD115, which is clearly visible in the depicted histogram for CD115 marker (lower row). (B) Luc+ MΦ without and with SPIO-labelling were tested for their expression levels of CD68, CD80, CD86, CD206 and F4/80. The monocytic markers Ly6c and CD115 served as negative controls. For both, labelled and unlabelled MΦ, F4/80-positive cells were also positive for CD68, CD80, CD86 and CD206. No change in expression levels of Ly6c could be detected upon particle uptake, whereas the CD115+ population of control cells minimized due to contrast agent incorporation. n = 2 for both cell types.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4920382&req=5

pone.0156626.g004: Representative dot plots and histograms of flow cytometric investigations of the effect of SPIO labelling on cell functionality.(A) Unlabelled control (upper row) and Nanomag labelled luc+ monocytes (middle row) were stained against monocytic markers Ly6c and CD115. Upon iron particle labelling, monocytes reduced their expression levels for CD115, which is clearly visible in the depicted histogram for CD115 marker (lower row). (B) Luc+ MΦ without and with SPIO-labelling were tested for their expression levels of CD68, CD80, CD86, CD206 and F4/80. The monocytic markers Ly6c and CD115 served as negative controls. For both, labelled and unlabelled MΦ, F4/80-positive cells were also positive for CD68, CD80, CD86 and CD206. No change in expression levels of Ly6c could be detected upon particle uptake, whereas the CD115+ population of control cells minimized due to contrast agent incorporation. n = 2 for both cell types.
Mentions: Influence of SPIO-labelling on functionality of luc+ monocytes and differentiated luc+ MΦ was analysed via flow cytometric measurements. Labelled (168 μg Fe/ml) as well as unlabelled control monocytes were stained for the monocytic markers CD115 and Ly6c (Fig 4A). 86.6% of the unlabelled control cells were double positive for both tested markers. Only a small population (10.7%) did not express CD115. This situation changed upon contrast agent uptake. Only 13.5% of the cells were double positive, but 84.5% reduced their CD115-expression and only expressed Ly6c.

Bottom Line: However, the underlying molecular mechanisms needed for proper regulation still remain to be elucidated.The labelling procedure of the isolated cells did not significantly influence cell characteristics and resulted in detection of as few as 500 labelled cells in vivo.Hypointensity in MRI of the graft appeared unaltered in spatial location.

View Article: PubMed Central - PubMed

Affiliation: In-vivo-NMR Laboratory, Max Planck Institute for Metabolism Research, Cologne, Germany.

ABSTRACT
Brain-infiltrating monocyte-derived macrophages are one of the key players in the local immune response after stroke. It is now widely accepted that the inflammatory response is not an exclusively destructive process. However, the underlying molecular mechanisms needed for proper regulation still remain to be elucidated. Here, we propose an in vitro labelling strategy for multimodal in vivo observation of macrophage dynamics distinguished from brain-residing microglia response. Prior to intracerebral transplantation into the striatum of recipient mice or systemic administration, monocytes and macrophages, isolated from luciferase-expressing mice, were labelled with superparamagnetic iron oxide particles. Temporo-spatial localization was monitored by magnetic resonance imaging, whereas survival of grafted cells was investigated using bioluminescence imaging. The labelling procedure of the isolated cells did not significantly influence cell characteristics and resulted in detection of as few as 500 labelled cells in vivo. Two weeks after stereotactic transplantation, the luciferase signal was sustained traceable, with approximately 18% of the original luciferase signal detectable for monocytes and about 30% for macrophages. Hypointensity in MRI of the graft appeared unaltered in spatial location. In a therapeutically relevant approach, systemic cell administration after stroke resulted in accumulation mostly in thoracic regions, as could be visualized with BLI. For detection of homing to ischemic brain tissue more cells need to be administered. Nevertheless, during parallel MRI sessions recruitment of i.v. injected cells to the lesion site could be detected by day 2 post stroke as scattered hypointense signal voids. With further increase in sensitivity, our multi-facetted labelling strategy will provide the basis for in vivo tracking and fate specification of tissue-infiltrating macrophages and their distinct role in stroke-related neuro-inflammation.

No MeSH data available.


Related in: MedlinePlus