Advances in imaging of new targets for pharmacological intervention in stroke: Real-time tracking of T-cells in the ischaemic brain

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 × 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 m 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.