Elsevier

Neurobiology of Disease

Volume 74, February 2015, Pages 305-313
Neurobiology of Disease

Cerebral collateral flow defines topography and evolution of molecular penumbra in experimental ischemic stroke

https://doi.org/10.1016/j.nbd.2014.11.019Get rights and content

Highlights

  • Molecular penumbra has heterogeneous topography.

  • The extension of molecular penumbra correlated with ischemic core.

  • Collateral flow is associated with reduced areas of ischemic penumbra and core.

  • Collateral flow is associated with increased areas of intact tissue.

Abstract

Intracranial collaterals are dynamically recruited after arterial occlusion and are emerging as a strong determinant of tissue outcome in both human and experimental ischemic stroke. The relationship between collateral flow and ischemic penumbra remains largely unexplored in pre-clinical studies. The aim of the present study was to investigate the pattern of collateral flow with regard to penumbral tissue after transient middle cerebral artery (MCA) occlusion in rats. MCA was transiently occluded (90 min) by intraluminal filament in adult male Wistar rats (n = 25). Intracranial collateral flow was studied in terms of perfusion deficit and biosignal fluctuation analyses using multi-site laser Doppler monitoring. Molecular penumbra was defined by topographical mapping and quantitative signal analysis of Heat Shock Protein 70 kDa (HSP70) immunohistochemistry. Functional deficit and infarct volume were assessed 24 h after ischemia induction. The results show that functional performance of intracranial collaterals during MCA occlusion inversely correlated with HSP70 immunoreactive areas in both the cortex and the striatum, as well as with infarct size and functional deficit. Intracranial collateral flow was associated with reduced areas of both molecular penumbra and ischemic core and increased areas of intact tissue in rats subjected to MCA occlusion followed by reperfusion. Our findings prompt the development of collateral therapeutics to provide tissue-saving strategies in the hyper-acute phase of ischemic stroke prior to recanalization therapy.

Introduction

Intracranial collateral circulation represents a multiple-level subsidiary vascular network which is dynamically recruited after occlusion of cerebral arteries to provide a source of residual blood flow (Liebeskind, 2003).

In both humans and rodents, a significant supply of collateral flow after middle cerebral artery (MCA) occlusion is provided by the circle of Willis through the anterior cerebral artery (ACA) and the leptomeningeal anastomoses between the cortical branches of ACA and MCA. However, a significant degree of inter-individual variability exists in the functional performance of intracranial collaterals under ischemic conditions in humans (Qureshi et al., 2008, Liu et al., 2011) and rodents (Armitage et al., 2010, Riva et al., 2012).

Cerebral collateral flow is emerging as a powerful determinant of functional and tissue outcome in unselected ischemic stroke patients (Maas et al., 2009, Menon et al., 2011) and in stroke patients treated with intravenous rtPA (Brunner et al., 2012, Miteff et al., 2009) or endovascular recanalization (Bang et al., 2011, Bang et al., 2008).

For these reasons, an in-depth understanding of the physiology, adaptive response and modulation of intracranial collateral circulation is of foremost importance for acute stroke pathophysiology and therapy.

Ischemic penumbra was originally defined as “tissue at risk”, which has been made functionally silent and metabolically metastable by ischemic injury but still has the potential for full recovery if reperfusion is timely achieved (Branston et al., 1974, Hossmann, 1994, Baron, 1999). The concept of ischemic penumbra is gradually evolving, after pre-clinical and clinical studies showed a heterogeneous and variable pattern of perfusion deficit, molecular response, topographical distribution and evolution of the penumbra in relation to the ischemic core (del Zoppo et al., 2011, Manning et al., 2014).

HSP70 is the major inducible heat shock protein (Sharp et al., 2000), whose expression after focal cerebral ischemia reflects an endogenous cell response to injury. The neuronal expression of HSP70 is considered as a molecularly defined penumbra, where injured neurons have activated endogenous pathways for protein renaturation and protection against further ischemia.

In the present study, we investigated the relationship between intracranial collateral flow during transient MCA occlusion and the development of molecular penumbra and ischemic infarct after 24 h. The hemodynamic monitoring of central and peripheral MCA territories was analyzed in terms of perfusion deficit and biosignal fluctuation. Hemodynamic data were correlated with the corresponding areas of molecular penumbra, infarct volume and functional deficit.

Section snippets

Study design

The experimental protocol was approved by the Committee on Animal Care of the University of Milano Bicocca, in accordance with the national guidelines on the use of laboratory animals (D.L. 116/1992) and the European Union Directive for animal experiments (2010/63/EU), under project license from the Italian Ministry of Health.

A group of consecutive animals undergoing successful MCA occlusion (see below) were used to explore the effect of cerebral collateral flow on molecular penumbra. Cerebral

Multi-site cerebral perfusion monitoring during MCA occlusion

Hemodynamic monitoring with two LD probes was performed during transient MCA occlusion, as shown in Fig. 1. The position of the probes was decided according to the rat cerebral vascular territories and verified by gelatin-ink perfusion experiments (Fig. 1A). Probe 1 (bregma − 1 mm, 5 mm from midline) was positioned within the central MCA territory, while Probe 2 (bregma + 2 mm, 2 mm from midline) was positioned in the borderzone territory between the cortical branches of ACA and MCA to monitor

Discussion

The functional performance of intracranial collaterals is crucially involved in the pathophysiology of acute ischemic stroke (Liebeskind, 2012). Functionally active cerebral collaterals have been associated with reduced penumbra loss (Jung et al., 2013) and better outcomes (Cortijo et al., 2014) in acute stroke patients, even with delayed reperfusion.

Therapeutic modulation of cerebral collateral flow is being proposed (Shuaib et al., 2011) and may be used for widening the therapeutic window for

Conclusions

The degree of cerebral collateral perfusion inversely correlated with both ischemic core and molecular penumbra during transient proximal MCA occlusion. Our findings prompt the development of collateral therapeutics to provide tissue-saving strategies in the hyper-acute phase of ischemic stroke prior to recanalization therapy.

Conflict of interest

The authors declare no conflict of interest.

The following are the supplementary data related to this article.

Acknowledgments

We thank Dr. Virginia Rodriguez Menendez for the technical assistance with immunohistochemistry and Miss Elena Pirovano for the figure preparation. This study was supported by the Italian Ministry of University and Research (MIUR, FAR12-01-138-2002100) and Université Lyon 1. These funding sources were not involved in the study design, collection, analysis and interpretation of data, writing of the report or decision to submit the present article for publication.

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