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Intravital imaging of a massive lymphocyte response in the cortical dura of mice after peripheral infection by trypanosomes.

Coles JA, Myburgh E, Ritchie R, Hamilton A, Rodgers J, Mottram JC, Barrett MP, Brewer JM - PLoS Negl Trop Dis (2015)

Bottom Line: Making use of optical signals from the skull bone, blood vessels and dural cells, we conclude that up to 40 dpi, the extravascular trypanosomes were essentially confined to the dura, as were the great majority of the T cells.Inhibition of T cell activation by intraperitoneal injection of abatacept reduced the numbers of meningeal T cells at 12 dpi and their mean speed fell from 11.64 ± 0.34 μm/min (mean ± SEM) to 5.2 ± 1.2 μm/min (p = 0.007).The population and motility of the trypanosomes tended to decline after about 30 dpi.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Molecular Parasitology, University of Glasgow, Glasgow, United Kingdom; Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom.

ABSTRACT
Peripheral infection by Trypanosoma brucei, the protozoan responsible for sleeping sickness, activates lymphocytes, and, at later stages, causes meningoencephalitis. We have videoed the cortical meninges and superficial parenchyma of C56BL/6 reporter mice infected with T.b.brucei. By use of a two-photon microscope to image through the thinned skull, the integrity of the tissues was maintained. We observed a 47-fold increase in CD2+ T cells in the meninges by 12 days post infection (dpi). CD11c+ dendritic cells also increased, and extravascular trypanosomes, made visible either by expression of a fluorescent protein, or by intravenous injection of furamidine, appeared. The likelihood that invasion will spread from the meninges to the parenchyma will depend strongly on whether the trypanosomes are below the arachnoid membrane, or above it, in the dura. Making use of optical signals from the skull bone, blood vessels and dural cells, we conclude that up to 40 dpi, the extravascular trypanosomes were essentially confined to the dura, as were the great majority of the T cells. Inhibition of T cell activation by intraperitoneal injection of abatacept reduced the numbers of meningeal T cells at 12 dpi and their mean speed fell from 11.64 ± 0.34 μm/min (mean ± SEM) to 5.2 ± 1.2 μm/min (p = 0.007). The T cells occasionally made contact lasting tens of minutes with dendritic cells, indicative of antigen presentation. The population and motility of the trypanosomes tended to decline after about 30 dpi. We suggest that the lymphocyte infiltration of the meninges may later contribute to encephalitis, but have no evidence that the dural trypanosomes invade the parenchyma.

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Arrival of trypanosomes in the dura mater.A. In dural vessels, trypanosomes were observed to move rapidly (red streaks, S2 Video) while leukocytes (dark, excluding plasma marker) could be arrested (e.g., at arrow). 26 dpi. B. Approximate tracks of trypanosomes within the focal depth of an XY scan, acquired over 12 s. A few excursions away from restricted localities are seen (e.g., arrow). Time is color-coded from blue to white (scale). 13 dpi. C. At another site in the dura of the same mouse1000 frames were acquired over 121 sec. Arrow indicates a movement out of the generally restricted volume. D-F. Frames from S14 Video showing extravasation of a trypanosome. The blood marker (dextran-fluorescein, 70 kDa) appears yellow-green, the trypanosomes express mCherry. In (E) blood plasma leaks from the vessel (arrow). 4 sec later, a trypanosome appears outside the vessel (F, arrow). Excitation wavelength 1050 nm. Exceptionally, D-F, are from a CD1 mouse infected with T.b.brucei Lister S427, at 3 dpi.
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pntd.0003714.g010: Arrival of trypanosomes in the dura mater.A. In dural vessels, trypanosomes were observed to move rapidly (red streaks, S2 Video) while leukocytes (dark, excluding plasma marker) could be arrested (e.g., at arrow). 26 dpi. B. Approximate tracks of trypanosomes within the focal depth of an XY scan, acquired over 12 s. A few excursions away from restricted localities are seen (e.g., arrow). Time is color-coded from blue to white (scale). 13 dpi. C. At another site in the dura of the same mouse1000 frames were acquired over 121 sec. Arrow indicates a movement out of the generally restricted volume. D-F. Frames from S14 Video showing extravasation of a trypanosome. The blood marker (dextran-fluorescein, 70 kDa) appears yellow-green, the trypanosomes express mCherry. In (E) blood plasma leaks from the vessel (arrow). 4 sec later, a trypanosome appears outside the vessel (F, arrow). Excitation wavelength 1050 nm. Exceptionally, D-F, are from a CD1 mouse infected with T.b.brucei Lister S427, at 3 dpi.

Mentions: Previous authors have suggested that trypanosomes arrive in the brain parenchyma either by diapedesis [12, 17] or by transport in CSF from the choroid plexus [57, 58], so we first looked for evidence of similar processes in the dura. Diapedesis of trypanosomes would require interaction lasting at least many seconds with the vascular endothelium, but in 24 C57BL/6 mice (and a further 12 CD-1 mice included in [20]), we never observed a trypanosome slowed or arrested on vascular endothelium (Fig 10A and S2 Video). It is therefore unlikely that trypanosomes extravasated from dural vessels by a diapedesis similar to that of leukocytes.


Intravital imaging of a massive lymphocyte response in the cortical dura of mice after peripheral infection by trypanosomes.

Coles JA, Myburgh E, Ritchie R, Hamilton A, Rodgers J, Mottram JC, Barrett MP, Brewer JM - PLoS Negl Trop Dis (2015)

Arrival of trypanosomes in the dura mater.A. In dural vessels, trypanosomes were observed to move rapidly (red streaks, S2 Video) while leukocytes (dark, excluding plasma marker) could be arrested (e.g., at arrow). 26 dpi. B. Approximate tracks of trypanosomes within the focal depth of an XY scan, acquired over 12 s. A few excursions away from restricted localities are seen (e.g., arrow). Time is color-coded from blue to white (scale). 13 dpi. C. At another site in the dura of the same mouse1000 frames were acquired over 121 sec. Arrow indicates a movement out of the generally restricted volume. D-F. Frames from S14 Video showing extravasation of a trypanosome. The blood marker (dextran-fluorescein, 70 kDa) appears yellow-green, the trypanosomes express mCherry. In (E) blood plasma leaks from the vessel (arrow). 4 sec later, a trypanosome appears outside the vessel (F, arrow). Excitation wavelength 1050 nm. Exceptionally, D-F, are from a CD1 mouse infected with T.b.brucei Lister S427, at 3 dpi.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400075&req=5

pntd.0003714.g010: Arrival of trypanosomes in the dura mater.A. In dural vessels, trypanosomes were observed to move rapidly (red streaks, S2 Video) while leukocytes (dark, excluding plasma marker) could be arrested (e.g., at arrow). 26 dpi. B. Approximate tracks of trypanosomes within the focal depth of an XY scan, acquired over 12 s. A few excursions away from restricted localities are seen (e.g., arrow). Time is color-coded from blue to white (scale). 13 dpi. C. At another site in the dura of the same mouse1000 frames were acquired over 121 sec. Arrow indicates a movement out of the generally restricted volume. D-F. Frames from S14 Video showing extravasation of a trypanosome. The blood marker (dextran-fluorescein, 70 kDa) appears yellow-green, the trypanosomes express mCherry. In (E) blood plasma leaks from the vessel (arrow). 4 sec later, a trypanosome appears outside the vessel (F, arrow). Excitation wavelength 1050 nm. Exceptionally, D-F, are from a CD1 mouse infected with T.b.brucei Lister S427, at 3 dpi.
Mentions: Previous authors have suggested that trypanosomes arrive in the brain parenchyma either by diapedesis [12, 17] or by transport in CSF from the choroid plexus [57, 58], so we first looked for evidence of similar processes in the dura. Diapedesis of trypanosomes would require interaction lasting at least many seconds with the vascular endothelium, but in 24 C57BL/6 mice (and a further 12 CD-1 mice included in [20]), we never observed a trypanosome slowed or arrested on vascular endothelium (Fig 10A and S2 Video). It is therefore unlikely that trypanosomes extravasated from dural vessels by a diapedesis similar to that of leukocytes.

Bottom Line: Making use of optical signals from the skull bone, blood vessels and dural cells, we conclude that up to 40 dpi, the extravascular trypanosomes were essentially confined to the dura, as were the great majority of the T cells.Inhibition of T cell activation by intraperitoneal injection of abatacept reduced the numbers of meningeal T cells at 12 dpi and their mean speed fell from 11.64 ± 0.34 μm/min (mean ± SEM) to 5.2 ± 1.2 μm/min (p = 0.007).The population and motility of the trypanosomes tended to decline after about 30 dpi.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Molecular Parasitology, University of Glasgow, Glasgow, United Kingdom; Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, United Kingdom.

ABSTRACT
Peripheral infection by Trypanosoma brucei, the protozoan responsible for sleeping sickness, activates lymphocytes, and, at later stages, causes meningoencephalitis. We have videoed the cortical meninges and superficial parenchyma of C56BL/6 reporter mice infected with T.b.brucei. By use of a two-photon microscope to image through the thinned skull, the integrity of the tissues was maintained. We observed a 47-fold increase in CD2+ T cells in the meninges by 12 days post infection (dpi). CD11c+ dendritic cells also increased, and extravascular trypanosomes, made visible either by expression of a fluorescent protein, or by intravenous injection of furamidine, appeared. The likelihood that invasion will spread from the meninges to the parenchyma will depend strongly on whether the trypanosomes are below the arachnoid membrane, or above it, in the dura. Making use of optical signals from the skull bone, blood vessels and dural cells, we conclude that up to 40 dpi, the extravascular trypanosomes were essentially confined to the dura, as were the great majority of the T cells. Inhibition of T cell activation by intraperitoneal injection of abatacept reduced the numbers of meningeal T cells at 12 dpi and their mean speed fell from 11.64 ± 0.34 μm/min (mean ± SEM) to 5.2 ± 1.2 μm/min (p = 0.007). The T cells occasionally made contact lasting tens of minutes with dendritic cells, indicative of antigen presentation. The population and motility of the trypanosomes tended to decline after about 30 dpi. We suggest that the lymphocyte infiltration of the meninges may later contribute to encephalitis, but have no evidence that the dural trypanosomes invade the parenchyma.

Show MeSH
Related in: MedlinePlus