Comprehensive Evaluation of Peripheral Nerve Regeneration in the Acute Healing Phase Using Tissue Clearing and Optical Microscopy in a Rodent Model

Autor: Joanna H. Ng, Conor L. Evans, Yookyung Jung, Mark A. Randolph, Cameron P. Keating, Prabhu Senthil-Kumar, Jie Zhao, Jonathan M. Winograd
Rok vydání: 2014
Předmět:
Male
Nervous system
Pathology
Wallerian degeneration
Critical Care and Emergency Medicine
Neurology
Immunofluorescence
lcsh:Medicine
Nervous System
Nervous System Procedures
Peripheral Nerve Injuries
Neurobiology of Disease and Regeneration
Medicine and Health Sciences
lcsh:Science
Trauma Medicine
Mammals
Microscopy
Confocal
Multidisciplinary
Tissue clearing
medicine.diagnostic_test
Animal Models
Sciatic Nerve
medicine.anatomical_structure
Vertebrates
Peripheral nerve injury
Sciatic nerve
Anatomy
Rats
Transgenic
Plastic Surgery and Reconstructive Techniques
Research Article
medicine.medical_specialty
Immunology
Neurosurgery
Surgical and Invasive Medical Procedures
Research and Analysis Methods
Rodents
Model Organisms
Neurorehabilitation and Trauma
medicine
Animals
Immunoassays
Wound Healing
business.industry
lcsh:R
Organisms
Biology and Life Sciences
medicine.disease
Rats
Nerve Regeneration
Immunologic Techniques
Clinical Immunology
lcsh:Q
Wound healing
business
Neuroscience
Zdroj: PLoS ONE, Vol 9, Iss 4, p e94054 (2014)
PLoS ONE
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0094054
Popis: Peripheral nerve injury (PNI), a common injury in both the civilian and military arenas, is usually associated with high healthcare costs and with patients enduring slow recovery times, diminished quality of life, and potential long-term disability. Patients with PNI typically undergo complex interventions but the factors that govern optimal response are not fully characterized. A fundamental understanding of the cellular and tissue-level events in the immediate postoperative period is essential for improving treatment and optimizing repair. Here, we demonstrate a comprehensive imaging approach to evaluate peripheral nerve axonal regeneration in a rodent PNI model using a tissue clearing method to improve depth penetration while preserving neural architecture. Sciatic nerve transaction and end-to-end repair were performed in both wild type and thy-1 GFP rats. The nerves were harvested at time points after repair before undergoing whole mount immunofluorescence staining and tissue clearing. By increasing the optic depth penetration, tissue clearing allowed the visualization and evaluation of Wallerian degeneration and nerve regrowth throughout entire sciatic nerves with subcellular resolution. The tissue clearing protocol did not affect immunofluorescence labeling and no observable decrease in the fluorescence signal was observed. Large-area, high-resolution tissue volumes could be quantified to provide structural and connectivity information not available from current gold-standard approaches for evaluating axonal regeneration following PNI. The results are suggestive of observed behavioral recovery in vivo after neurorrhaphy, providing a method of evaluating axonal regeneration following repair that can serve as an adjunct to current standard outcomes measurements. This study demonstrates that tissue clearing following whole mount immunofluorescence staining enables the complete visualization and quantitative evaluation of axons throughout nerves in a PNI model. The methods developed in this study could advance PNI research allowing both researchers and clinicians to further understand the individual events of axonal degeneration and regeneration on a multifaceted level.
Databáze: OpenAIRE
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