British journal of pharmacology · 2009
Advances in imaging of new targets for pharmacological intervention in stroke: real-time tracking of T-cells in the ischaemic brain.
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Ortolano F, Maffia P, Dever G, Rodolico G, Millington O R, De Simoni M G, Brewer J M, Bushell T J, Garside P, Carswell H V
Аннотация
BACKGROUND AND PURPOSE: T-cells may play a role in the evolution of ischaemic damage and repair, but the ability to image these cells in the living brain after a stroke has been limited. We aim to extend the technique of real-time in situ brain imaging of T-cells, previously shown in models of immunological diseases, to models of experimental stroke.
EXPERIMENTAL APPROACH: Male C57BL6 mice (6-8 weeks) (n= 3) received a total of 2-5 x 10(6) carboxyfluorescein diacetate succinimidyl ester (CFSE)-labelled lymphocytes from donor C57BL6 mice via i.v. injection by adoptive transfer. Twenty-four hours later, recipient mice underwent permanent left distal middle cerebral artery occlusion (MCAO) by electrocoagulation or by sham surgery under isoflurane anaesthesia. Female hCD2-green fluorescent protein (GFP) transgenic mice that exhibit GFP-labelled T-cells underwent MCAO. At 24 or 48 h post-MCAO, a sagittal brain slice (1500 microm thick) containing cortical branches of the occluded middle cerebral artery (MCA) was dissected and used for multiphoton laser scanning microscopy (MPLSM).
KEY RESULTS: Our results provide direct observations for the first time of dynamic T-cell behaviour in living brain tissue in real time and herein proved the feasibility of MPLSM for ex vivo live imaging of immune response after experimental stroke.
CONCLUSIONS AND IMPLICATIONS: It is hoped that these advances in the imaging of immune cells will provide information that can be harnessed to a therapeutic advantage.
EXPERIMENTAL APPROACH: Male C57BL6 mice (6-8 weeks) (n= 3) received a total of 2-5 x 10(6) carboxyfluorescein diacetate succinimidyl ester (CFSE)-labelled lymphocytes from donor C57BL6 mice via i.v. injection by adoptive transfer. Twenty-four hours later, recipient mice underwent permanent left distal middle cerebral artery occlusion (MCAO) by electrocoagulation or by sham surgery under isoflurane anaesthesia. Female hCD2-green fluorescent protein (GFP) transgenic mice that exhibit GFP-labelled T-cells underwent MCAO. At 24 or 48 h post-MCAO, a sagittal brain slice (1500 microm thick) containing cortical branches of the occluded middle cerebral artery (MCA) was dissected and used for multiphoton laser scanning microscopy (MPLSM).
KEY RESULTS: Our results provide direct observations for the first time of dynamic T-cell behaviour in living brain tissue in real time and herein proved the feasibility of MPLSM for ex vivo live imaging of immune response after experimental stroke.
CONCLUSIONS AND IMPLICATIONS: It is hoped that these advances in the imaging of immune cells will provide information that can be harnessed to a therapeutic advantage.
Предметные рубрики MeSH
Adoptive TransferAnimalsBrainDisease Models, AnimalFeasibility StudiesFemaleFluoresceinsFluorescent DyesGreen Fluorescent ProteinsInfarction, Middle Cerebral ArteryMaleMiceMice, Inbred C57BLMice, TransgenicMicroscopy, Fluorescence, MultiphotonMolecular ImagingMolecular Probe TechniquesSuccinimidesT-LymphocytesTime Factors