The “no-reflow” phenomenon revisited: Delayed microvascular occlusions as a novel therapeutic target for ischemic stroke

• Applicant: Igor Khalin, Ph.D.
• Subject Area: Experimental Models for the Understanding of Nervous System Diseases ; Clinical Neurology; Neurosurgery and Neuroradiology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 457586042

Ischemic stroke caused by the thrombotic or embolic occlusion of a large cerebral artery is one of the most frequent causes of death and disability worldwide. Recanalization of the occluded vessel with plasminogen activator or/and mechanical thrombectomy has recently been established as standard of care. However, even after successful recanalization less than 50% of patients show clinical improvements. This situation is believed to occur, among others, due to a fateful lack of reperfusion of the parenchymal microcirculation, a process termed “no-reflow” phenomenon more than three decades ago. Since then it was assumed that capillaries get plugged during ischemia and reperfusion is not able to fully restore microvascular flow. In a set of preliminary experiments using novel ultra-bright nanoparticles and in vivo multi-photon microscopy of the cerebral vasculature in mice, we demonstrate that capillary flow is readily restored immediately after recanalization, but is heavily impaired by the delayed formation of microvascular occlusions (dMiVOs) within the first few hours after restoration of large artery flow. Hence, the time course of dMiVO formation suggests that microcirculatory failure after ischemic stroke may be a treatable entity, provided the undelaying mechanisms are known.The aim of the current project is therefore to examine the mechanisms of dMiVO formation on the cellular and molecular level. For this purpose, we developed a novel class of nanoparticles able to visualize and potentially treat dMiVOs in vivo. The specific aims of the current application are: (1) Determine the effect of ischemia duration (10-180 min) on dMiVO formation and fully characterize the spatio-temporal pattern of dMiVO formation following reperfusion (2-72 hours). At the same time, we will evaluate whether the highly fluorescent nanoparticles used to visualize dMiVOs can be used as drug carriers by monitoring their decomposition in vivo by Förster-resonance energy transfer (FRET); (2) Determine the vascular mechanisms associated with dMiVO formation, i.e. blood-brain barrier permeability and damage of endothelial cells and pericytes; (3) Identify the parenchymal events associated with dMiVO formation, i.e. reactivity of astrocytes and survival of neurons; (4) Characterize inflammatory processes associated with dMiVO formation, i.e. the role of microglia in dMiVO formation and modulation of BBB integrity. These aims will be achieved by using a whole range of state-of-the-art in vivo imaging tools (multi-photon microscopy, transgenic mice, viral-based cell labelling, correlation electron microscopy, small animal magnetic resonance imaging).The overall goal of the current proposal is to decipher the mechanisms of dMiVO formation and to evaluate the therapeutic potential of this process.

• DFG Programme: Research Grants
• Co-Investigator: Professor Dr. Nikolaus Plesnila