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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

Presented here is a protocol to induce diffuse traumatic brain injury using a lateral fluid percussion device followed by the collection of the caecum content for gut microbiome analysis.

Abstract

Increasing evidence shows that the microbiota-gut-brain axis plays an important role in the pathogenesis of brain diseases. Several studies also demonstrate that traumatic brain injuries cause changes to the gut microbiota. However, mechanisms underlying the bidirectional regulation of the brain-gut axis remain unknown. Currently, few models exist for studying the changes in gut microbiota after traumatic brain injury. Therefore, the presented study combines protocols for inducing traumatic brain injury using a lateral fluid percussion device and analysis of caecum samples following injury for investigating alterations in the gut microbiome. Alterations of the gut microbiota composition after traumatic brain injury are determined using 16S-rDNA sequencing. This protocol provides an effective method for studying the relationships between enteric microorganisms and traumatic brain injury.

Introduction

Traumatic brain injury (TBI) is a global public health problem and the leading cause of death and disability in young adults1,2. TBI causes many deaths every year, and survivors experience a variety of physical, psychiatric, emotional, and cognitive disabilities. Therefore, TBI is a heavy burden to a patient's family and societal resources. TBI involves both the primary brain injury that occurs at the time of trauma and any secondary brain injuries that develop hours to months following initial injury. Secondary brain injury is mediated by several biochemical cascades, which are not only detrimental to the bra....

Protocol

All procedures performed were approved by the Experimental Animal Ethics Committee of Zhejiang University. All instruments and materials used in surgery are sterile. The TBI proceudre takes about 20 minutes.

1. Animal care

  1. Use 5- to 6-week-old male C57BL/6J mice (20-25 g of weight) in this experiment.
  2. Maintain mice on a 12 h/12 h light/dark cycle, and make sure they receive food and water ad libitum. Provide the same amounts of food and water to both the sham and TBI.......

Representative Results

Establishment of TBI is shown in Figure 1. After anesthesia and disinfection, the scalp was incised sagittally (Figure 1A). A craniotomy (3 mm in diameter) was trephined into the skull over the right parietal cortex with an electric drill, the dura was kept intact (Figure 1B,C). A plastic injury cannula was placed over the bone window and cemented to the skull using dental acrylic (<.......

Discussion

Presented here is a simple and efficient protocol to determine changes in cecal microbiota after TBI in mice. Induction of brain injury and collection of caecum content samples are critical parts of the protocol.

Despite researchers having studied the changes of gut microbiota following TBI, the brain injury used in these studies were CCI-8 and weight drop/impact-induced models9. However, the CCI model mostly replicates brain contusion, and the w.......

Acknowledgements

The authors have nothing to disclose.

....

Materials

NameCompanyCatalog NumberComments
DNA isolation kit QIAGEN51604For fast purification of genomic DNA from stool samples
Gene analysis serviceGENEWIZGene analyse service
Heating padShanghai SAFE Biotech Co.TR-200heating pad
InjectorThe First Affiliated Hospital, School of Medicine, Zhejiang Universityinjector
LFPI deviceVirginia
Commonwealth University
FP302LFPI device
Micro cranial drillRWD Life Science78061Micro cranial drill
Povidone IodineThe First Affiliated Hospital, School of Medicine, Zhejiang UniversityPovidone Iodine

References

  1. Cheng, P., et al. . Trends in traumatic brain injury mortality in China, 2006-2013: A population-based longitudinal study. 14, e1002332 (2017).
  2. Maas, A. I. R., et al. Traumatic brain injury: integrated approaches to improve prevention, clinical care, and ....

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Traumatic Brain InjuryCaecum MicrobiotaGut brain AxisLateral Fluid Percussion InjuryMouse ModelMicrobiome Analysis

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