方法文章

A Preclinical Controlled Cortical Impact Model for Traumatic Hemorrhage Contusion and Neuroinflammation

DOI：

10.3791/61393

⸱

2020年6月10日

 ,  , 
1Department of Neurosurgery, Cheng Hsin General Hospital, 2Graduate Institute of Physiology, College of Medicine, National Taiwan University

* These authors contributed equally

本文内容

摘要

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Intraparenchymal hemorrhage and neuroinflammation accompanied by cerebral contusion can trigger severe secondary brain injury. This protocol details a mouse controlled cortical impact (CCI) model allowing researchers to study hemorrhage contusion and post-traumatic immune responses and explore potential therapeutics.

摘要

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Cerebral contusion is a severe medical problem affecting millions of people worldwide each year. There is an urgent need to understand the pathophysiological mechanism and to develop effective therapeutic strategy for this devastating neurological disorder. Intraparenchymal hemorrhage and post-traumatic inflammatory response induced by initial physical impact can aggravate microglia/macrophage activation and neuroinflammation which subsequently worsen brain pathology. We provide here a controlled cortical impact (CCI) protocol that can reproduce experimental cortical contusion in mice by using a pneumatic impactor system to deliver mechanical force with controllable magnitude and velocity onto the dural surface. This preclinical model allows researchers to induce moderately severe focal cerebral contusion in mice and to investigate a wide range of post-traumatic pathological progressions including hemorrhage contusion, microglia/macrophage activation, iron toxicity, axonal injury, as well as short-term and long-term neurobehavioral deficits. The present protocol can be useful for exploring the long-term effects of and potential interventions for cerebral contusion.

引言

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Cerebral contusion is a form of traumatic brain injury that ranks high among the deadliest health issues in modern society1. It is primarily caused by accidental events such as traffic accident that results in external forces applying mechanical energy to the head. Traumatic brain injury affects an approximate of 3.5 million people and accounts for 30% of all acute injury-related deaths in the US each year2. Patients who survive cerebral contusion oftentimes suffer from long-term consequences including focal motor weakness, sensory dysfunction, and mental illness1.

The ....

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方案

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All procedures described in this protocol were conducted under the approval of the Institutional Animal Care and Use Committee at Cheng Hsin General Hospital and National Taiwan University College of Medicine. Eight- to ten-week-old male C57BL/6 wild type mice were used in this protocol.

1. Anesthesia induction

  1. Anesthetize the mouse with ~4% isoflurane mixed with room air at ~0.2 L/min in an induction chamber connected to the isoflurane vaporizer.
  2. Ensure the respiratory pattern is smooth. Check the depth of anesthesia by confirming a lack of toe-pinch reflex in the animal.

2. Pre-surgic....

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结果

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Illustration of stereotactic placement and craniotomy procedure.

The CCI model is known for its stability and reproducibility in producing injury ranging from mild to severe18. Proper stereotactic technique and craniotomy procedure are major determinants in producing stable and reproducible CCI-induced brain injury (Figure 1A,B). An ideal craniotomy procedure would cause minimal histological injury in the s.......

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讨论

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The CCI protocol produces highly reproducible mechanical injury to the brain for cerebral contusion research. The following steps are crucial for generating consistent brain injury in animals using this CCI protocol.

First, the mouse head should be stably mounted on the stereotaxic frame and the anatomical landmarks Bregma and Lambda always in the same horizontal plane. Unsteady or unlevel head placement oftentimes result in varied injury levels between animals. To ensure the animal head is sa.......

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披露

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The authors have nothing to disclose.

致谢

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We thank Danye Jiang for editing the manuscript and insightful input. We thank Jhih Syuan Lin for assisting in manuscript preparation. This work was supported by the Ministry of Science and Technology of Taiwan (MOST 107-2320-B-002-063-MY2) to C.F.C.

....

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甲酚紫乙酸Sigma-AldrichC5042
DAB 染色试剂盒载体SK-4105
山羊抗兔 IgG 二抗，Alexa Fluor 488InvitrogenA11034
山羊抗大鼠 IgG 二抗，Alexa Fluor 594InvitrogenA11007
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参考文献

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  1. Maas, A. I. R., et al. Traumatic brain injury: integrated approaches to improve prevention, clinical care, and research. The Lancet Neurology. 16 (12), 987-1048 (2017).
  2. Taylor, C. A., Bell, J. M., Breiding, M. J., Xu, L.

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