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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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Related in: MedlinePlus

As infection progressed, trypanosomes in the dura made shorter displacements.Automated tracking suggests that at 11 dpi (A) extravascular trypanosomes make longer displacements compared to 32 dpi (B). Scale bar applies to both images. C. Maximum excursions measured on 12s videos. Each symbol is the mean of the maximum excursions for all trypanosomes imaged one image field. Pooled results from 3 mice 11–14 dpi and 2 mice 36 and 39 dpi. D. Another frame from the mouse of (A) showing trypanosomes and T cells in approximately the same plane, but apparently not interacting (see also S9 Video). All scale bars are 20 μm.
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pntd.0003714.g011: As infection progressed, trypanosomes in the dura made shorter displacements.Automated tracking suggests that at 11 dpi (A) extravascular trypanosomes make longer displacements compared to 32 dpi (B). Scale bar applies to both images. C. Maximum excursions measured on 12s videos. Each symbol is the mean of the maximum excursions for all trypanosomes imaged one image field. Pooled results from 3 mice 11–14 dpi and 2 mice 36 and 39 dpi. D. Another frame from the mouse of (A) showing trypanosomes and T cells in approximately the same plane, but apparently not interacting (see also S9 Video). All scale bars are 20 μm.

Mentions: The number of extravascular trypanosomes in the dura appeared to peak after 30 dpi and then fall to zero (Fig 3D). This means that either trypanosomes moved out of the dura, or were destroyed within the dura, at a rate that could exceed the rate of entry. Automated tracking of trypanosomes suggested they displaced less at later infection times (Fig 11A and 11B). To check this, without relying on automated tracking of the rapidly moving trypanosomes, we visually followed their positions on the videos and noted the co-ordinates of the two extreme positions visited by each trypanosome during the standard imaging period of 12 s. This analysis confirmed that the excursions were fewer and shorter at later stages of infection (the mean values being 10.8 μm, SD = 7.7 μm at 36–39 dpi compared to 29.2 μm ± 13.9 μm at 11–14 dpi, p < 0.0001; Fig 11C). This progressive reduction in movement, and the decline in the dural population after 30 dpi (Fig 3D), suggest that the dura becomes an unfavorable environment for trypanosomes.


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)

As infection progressed, trypanosomes in the dura made shorter displacements.Automated tracking suggests that at 11 dpi (A) extravascular trypanosomes make longer displacements compared to 32 dpi (B). Scale bar applies to both images. C. Maximum excursions measured on 12s videos. Each symbol is the mean of the maximum excursions for all trypanosomes imaged one image field. Pooled results from 3 mice 11–14 dpi and 2 mice 36 and 39 dpi. D. Another frame from the mouse of (A) showing trypanosomes and T cells in approximately the same plane, but apparently not interacting (see also S9 Video). All scale bars are 20 μm.
© Copyright Policy
Related In: Results  -  Collection

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

pntd.0003714.g011: As infection progressed, trypanosomes in the dura made shorter displacements.Automated tracking suggests that at 11 dpi (A) extravascular trypanosomes make longer displacements compared to 32 dpi (B). Scale bar applies to both images. C. Maximum excursions measured on 12s videos. Each symbol is the mean of the maximum excursions for all trypanosomes imaged one image field. Pooled results from 3 mice 11–14 dpi and 2 mice 36 and 39 dpi. D. Another frame from the mouse of (A) showing trypanosomes and T cells in approximately the same plane, but apparently not interacting (see also S9 Video). All scale bars are 20 μm.
Mentions: The number of extravascular trypanosomes in the dura appeared to peak after 30 dpi and then fall to zero (Fig 3D). This means that either trypanosomes moved out of the dura, or were destroyed within the dura, at a rate that could exceed the rate of entry. Automated tracking of trypanosomes suggested they displaced less at later infection times (Fig 11A and 11B). To check this, without relying on automated tracking of the rapidly moving trypanosomes, we visually followed their positions on the videos and noted the co-ordinates of the two extreme positions visited by each trypanosome during the standard imaging period of 12 s. This analysis confirmed that the excursions were fewer and shorter at later stages of infection (the mean values being 10.8 μm, SD = 7.7 μm at 36–39 dpi compared to 29.2 μm ± 13.9 μm at 11–14 dpi, p < 0.0001; Fig 11C). This progressive reduction in movement, and the decline in the dural population after 30 dpi (Fig 3D), suggest that the dura becomes an unfavorable environment for trypanosomes.

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