In Vivo Near-Infrared Imaging of Fibrin Deposition in Thromboembolic Stroke in Mice

imaging of activated factor XIII (FXIIIa), an important mediator of thrombosis or fibrinolytic resistance. The present study was to investigate the fibrin deposition in a thromboembolic stroke mice model by FXIIIa–targeted near-infrared fluorescence (NIRF) imaging., which were correlated with histology after animal euthanasia. NIRF images and lesion volume.Non-invasive detection of fibrin deposition in ischemic mouse brain using NIRF imaging is feasible and this technique may provide an in vivo experimental tool in studying the role of fibrin in stroke

MyD88-TLR4-dependent choroid plexus activation precedes perilesional inflammation and secondary brain edema in a mouse model of intracerebral hemorrhage

Background: The functional neurological outcome of patients with intracerebral hemorrhage (ICH) strongly relates to the degree of secondary brain injury (ICH-SBI) evolving within days after the initial bleeding. Different mechanisms including the incitement of inflammatory pathways, dysfunction of the blood–brain barrier (BBB), activation of resident microglia, and an influx of blood-borne immune cells, have been hypothesized to contribute to ICH-SBI. Yet, the spatiotemporal interplay of specific inflammatory processes within different brain compartments has not been sufficiently characterized, limiting potential therapeutic interventions to prevent and treat ICH-SBI. Methods: We used a whole-blood injection model in mice, to systematically characterized the spatial and temporal dynamics of inflammatory processes after ICH using 7-Tesla magnetic resonance imaging (MRI), spatial RNA sequencing (spRNAseq), functional BBB assessment, and immunofluorescence average-intensity-mapping. Results: We identified a pronounced early response of the choroid plexus (CP) peaking at 12–24 h that was characterized by inflammatory cytokine expression, epithelial and endothelial expression of leukocyte adhesion molecules, and the accumulation of leukocytes. In contrast, we observed a delayed secondary reaction pattern at the injection site (striatum) peaking at 96 h, defined by gene expression corresponding to perilesional leukocyte infiltration and correlating to the delayed signal alteration seen on MRI. Pathway analysis revealed a dependence of the early inflammatory reaction in the CP on toll-like receptor 4 (TLR4) signaling via myeloid differentiation factor 88 (MyD88). TLR4 and MyD88 knockout mice corroborated this observation, lacking the early upregulation of adhesion molecules and leukocyte infiltration within the CP 24 h after whole-blood injection. Conclusions: We report a biphasic brain reaction pattern after ICH with a MyD88-TLR4-dependent early inflammatory response of the CP, preceding inflammation, edema and leukocyte infiltration at the lesion site. Pharmacological targeting of the early CP activation might harbor the potential to modulate the development of ICH-SBI

Post-ischaemic silencing of p66Shc reduces ischaemia/reperfusion brain injury and its expression correlates to clinical outcome in stroke

In light of the limited repertoire of therapeutical options available for the treatment of ischaemic stroke, the identification of novel potential targets is vital; in this respect, the present study demonstrates that the adaptor protein p66Shc holds this potential as an adjunct therapy to thrombolysis. Post-ischaemic silencing of p66Shc protein yielded beneficial effects in a mouse model of I/R brain injury underlying an interesting translational perspective for this target protein. Further, in proof-of-principle clinical experiments using PBMs, we demonstrate that p66Shc gene expression is transiently increased and that its levels correlate to short-term outcome in ischaemic stroke patients. Although these latter experiments are not directly relevant to the experiments performed in mice and in human endothelial cells, they provide novel important information about p66Shc regulation in stroke patients and set the basis for further investigations aimed at assessing the potential for p66Shc to become a novel therapeutic target as an adjunct of thrombolysis for the management of acute ischaemic strok

Extracting Vascular Networks under Physiological Constraints via Integer Programming

Abstract. We introduce an integer programming-based approach to vessel net-work extraction that enforces global physiological constraints on the vessel struc-ture and learn this prior from a high-resolution reference network. The method accounts for both image evidence and geometric relationships between vessels by formulating and solving an integer programming problem. Starting from an over-connected network, it is pruning vessel stumps and spurious connections by evaluating bifurcation angle and connectivity of the graph. We utilize a high-resolution micro computed tomography (µCT) dataset of a cerebrovascular corro-sion cast to obtain a reference network, perform experiments on micro magnetic resonance angiography (µMRA) images of mouse brains and discuss properties of the networks obtained under different tracking and pruning approaches.

The new cardioprotector Monohydroxyethylrutoside protects against doxorubicin-induced inflammatory effects in vitro

The new cardioprotector Monohydroxyethylrutoside protects against doxorubicin-induced inflammatory effects in vitro. Abou El Hassan MA, Verheul HM, Jorna AS, Schalkwijk C, van Bezu J, van der Vijgh WJ, Bast A. Department of Medical Oncology, Free University Medical Center, PO Box 7057, 1007 MB Amsterdam, The Netherlands. [email protected] Besides its cardiotoxic effect, doxorubicin also elicits inflammatory effects in vivo. 7-Monohydroxyethylrutoside (monoHER) has recently been used as a protector against doxorubicin-induced cardiotoxicity in vivo. It is not known yet whether monoHER can also protect against doxorubicin-induced inflammatory effects. The aim of the present study was (1) to illustrate the inflammatory effects of doxorubicin in vitro and (2) to evaluate a possibly protective effect of monoHER. In order to demonstrate the inflammatory effects of doxorubicin and the possible protection of monoHER, proliferating human umbilical cord vascular endothelial cells (HUVECs) were incubated with different concentrations of doxorubicin ranging from 12.5 to 600 nM with(out) 200 micro M monoHER. Resting (confluent) HUVECs were incubated with (0.5-25 micro M) doxorubicin with(out) monoHER (0.2-1.2 mM) and the viability of endothelial cells and their propensity to adhere to neutrophils were measured 24 h after treatment. The localisation of adhered neutrophils was determined with immunofluorescence microscopy. To further characterise the mechanism of doxorubicin-induced neutrophil adhesion, the expression of the HUVECs surface adhesion molecules was determined after doxorubicin treatment. Doxorubicin decreased the viability and proliferation capacity of HUVECs in a concentration-dependent manner. The proliferating HUVECs were much more sensitive to doxorubicin (IC(50)=60.0+/-20.8 nM) than resting cells (LC(50)=4.0+/-0.3 micro M). Doxorubicin also increased the adhesion of neutrophils reaching a plateau value at a doxorubicin concentration of > or =0.4 micro M (P=0.0113). The induced neutrophil adhesion was accompanied by overexpression of VCAM and E-selectin but not ICAM. Although monoHER did not reverse the effect of doxorubicin on the proliferation of endothelial cells, it significantly protected resting HUVECs against the cytotoxic effect of doxorubicin (< or =25 micro M, P<0.0015). In addition, monoHER completely protected against the stimulatory effect of doxorubicin on neutrophil adhesion, and inhibited the doxorubin-induced expression of VCAM and E-selectin on the surface of treated HUVECs. This study illustrates that monoHER, which protects against doxorubicin's cardiotoxic effect, can also protect against doxorubicin-induced inflammatory effects. These data prompt further investigation about the possible link between doxorubicin-induced inflammatory effects and its cardiotoxicity in viv

Purinergic Receptor Stimulation Reduces Cytotoxic Edema and Brain Infarcts in Mouse Induced by Photothrombosis by Energizing Glial Mitochondria

Treatments to improve the neurological outcome of edema and cerebral ischemic stroke are severely limited. Here, we present the first in vivo single cell images of cortical mouse astrocytes documenting the impact of single vessel photothrombosis on cytotoxic edema and cerebral infarcts. The volume of astrocytes expressing green fluorescent protein (GFP) increased by over 600% within 3 hours of ischemia. The subsequent growth of cerebral infarcts was easily followed as the loss of GFP fluorescence as astrocytes lysed. Cytotoxic edema and the magnitude of ischemic lesions were significantly reduced by treatment with the purinergic ligand 2-methylthioladenosine 5′ diphosphate (2-MeSADP), an agonist with high specificity for the purinergic receptor type 1 isoform (P2Y1R). At 24 hours, cytotoxic edema in astrocytes was still apparent at the penumbra and preceded the cell lysis that defined the infarct. Delayed 2MeSADP treatment, 24 hours after the initial thrombosis, also significantly reduced cytotoxic edema and the continued growth of the brain infarction. Pharmacological and genetic evidence are presented indicating that 2MeSADP protection is mediated by enhanced astrocyte mitochondrial metabolism via increased inositol trisphosphate (IP3)-dependent Ca2+ release. We suggest that mitochondria play a critical role in astrocyte energy metabolism in the penumbra of ischemic lesions, where low ATP levels are widely accepted to be responsible for cytotoxic edema. Enhancement of this energy source could have similar protective benefits for a wide range of brain injuries

Mouse models of neurodegenerative disease: preclinical imaging and neurovascular component.

Neurodegenerative diseases represent great challenges for basic science and clinical medicine because of their prevalence, pathologies, lack of mechanism-based treatments, and impacts on individuals. Translational research might contribute to the study of neurodegenerative diseases. The mouse has become a key model for studying disease mechanisms that might recapitulate in part some aspects of the corresponding human diseases. Neurode- generative disorders are very complicated and multifacto- rial. This has to be taken in account when testing drugs. Most of the drugs screening in mice are very di cult to be interpretated and often useless. Mouse models could be condiderated a ‘pathway models’, rather than as models for the whole complicated construct that makes a human disease. Non-invasive in vivo imaging in mice has gained increasing interest in preclinical research in the last years thanks to the availability of high-resolution single-photon emission computed tomography (SPECT), positron emission tomography (PET), high eld Magnetic resonance, Optical Imaging scanners and of highly speci c contrast agents. Behavioral test are useful tool to characterize di erent ani- mal models of neurodegenerative pathology. Furthermore, many authors have observed vascular pathological features associated to the di erent neurodegenerative disorders. Aim of this review is to focus on the di erent existing animal models of neurodegenerative disorders, describe behavioral tests and preclinical imaging techniques used for diagnose and describe the vascular pathological features associated to these diseases