Investigation of the effect of inflammation on platelet-mediated hypoperfusion in MCAO model and treatment with ADAMTS13.

Abstract

Despite major advances in stroke research only a limited number of stroke patients can benefit from current treatments. Cerebral ischaemia result from sudden loss of blood supply to the brain, which causes brain dysfunction and death. Pre-existing inflammation is an important contributing factor in the no-reflow phenomenon, preventing the brain from restoring perfusion and worsen stroke outcomes. This thesis investigates the effect of systemic inflammation on cerebral blood flow (CBF) using Laser speckle contrast imaging (LSCI). We aim to explore potential mechanisms that may contribute to inflammation-induced impairments in brain reperfusion and tested a novel therapeutic intervention to improve CBF perfusion and ischemic outcomes. First, we sought to establish a suitable in vivo model to investigate the role of interleukin-1β (IL-1β) as a systemic inflammatory agent using LSCI. CBF was measured at 20 min and 4h after reperfusion in C57BL6 mice, given systemic (intraperitoneal) IL-1β or vehicle before 20 minute and 30 min MCAO. Systemic interleukin-1 caused a severe reduction in CBF followed by significant reperfusion defect and increased blood brain barrier (BBB) breakdown compared with vehicle as early as 4h after stroke in the 30 min but not the 20 min MCAO occlusion duration. Restriction in CBF was observed alongside hyper-coagulation of the platelet in the vasculature of mice receiving interleukin-1. In this thesis, we used a novel ADAMTS13 variant Ala1144Val 7 (hereafter referred to as constitutively active (ca) ADAMTS13) in order to evaluate it is effect as a potential treatment of ischemic stroke. ADAMTS13 function is to cleave von Willebrand factor (VWF) therefore preventing and dissolving microvascular platelet aggregation. We tested it is activity on transient MCAO with systemic inflammation and ischaemia/reperfusion injury at a delayed time point of 4h reperfusion. The primary measure of efficacy was the restoration of CBF to the MCA region which was evaluated using LSCI. We demonstrated a significant restoration of CBF and reduced lesion volume and BBB breakdown in animals treated with caADAMTS13 at 6 h reperfusion. Moreover, caADAMTS13 induced a significant dissolution of platelet aggregates and a reduction within the stroked hemisphere of MCAO mice. Our results support evidence that systemic inflammation contribute to the no-reflow phenomenon as well as excessive ischaemic damage. Furthermore, we identified the presence of platelets aggregates at acute time of 4 h following MCAO reperfusion, suggesting their contribution to the early CBF deficit. Our results also introduce a novel caADAMTS13 variant as a potential viable therapeutic option for the treatment of ischaemic stroke.

Date of Award	1 Aug 2022
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Ingo Schiessl (Supervisor) & Stuart Allan (Supervisor)