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BS70 Development of an in vitro model of ischemic stroke and reperfusion injury
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BS70 Development of an in vitro model of ischemic stroke and reperfusion injury
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BS70 Development of an in vitro model of ischemic stroke and reperfusion injury
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Journal Article
BS70 Development of an in vitro model of ischemic stroke and reperfusion injury
2023
Overview
IntroductionIschaemic stroke, and subsequent reperfusion injury is a multicellular process, with limited treatment options. The successful translation of novel therapies is currently hampered by the lack of existing cellular models that reflect human disease and consider the multicellular nature of ischaemic stroke. In this project we developed a simple, cost effective and adaptable model of the human neurovascular unit, that is compatible with flow to enable the pathological basis of ischaemic stroke and neuroinflammation to be investigated. The model also offers a novel non-animal-based platform to identify and test novel therapeutic strategies for the treatment and prevention of ischaemic stroke and reperfusion injury.AimsTo characterise the role of activated platelets and inflammatory mediators on cells of the neurovascular unit to assess their involvement in ischemic reperfusion injury.MethodMonocultures of human microglia (HMC-3), astrocyte-like glioma (U87) and microvascular endothelial cells (hCMECs/D3) were treated with activated platelet releasate, and cell activation, proliferation, migration, and survival were monitored. Human washed platelet releasate was prepared by centrifugation.Quantitative PCR (qPCR) in the presence of SYBR Green I was performed using CFX PCR system.High resolution 3D imagingHMC-3 (Red), U87 (green) and HMEC/D3 (Blue) were incubated with cell tracker in serum free media for 30 minutes. HMC-3 and U87 were seeded within a 1.5 mg.ml collagen rat tail matrix in a microfluidic chamber. HMEC/D3 cell line was seeded on top of the collagen matrix within the fluid chamber. 3D Z stack images taken with Stellaris 5 Confocal microscope.Results/ConclusionTreatment of the different neurovascular cell types with hypoxia, TNFα and platelet releasate were also found to alter expression of key mediators of neuroinflammation (IL-1B, IL-6) and thrombosis (vWF and CD39) alongside targets of interest for the research team (LXRα, LXRβ, Pim-1, Pim-2, Pim-3 and SIRT1, P2Y2), identifying potential roles for these mediators in the regulation of neuroinflammation following ischaemic stroke and reperfusion injury.Conflict of InterestAbstract from work I did in previous PDRA. I have approval to submit
Publisher
BMJ Publishing Group Ltd and British Cardiovascular Society,BMJ Publishing Group LTD
Subject
