scholarly journals Optical Tissue Clearing Enables Rapid, Precise and Comprehensive Assessment of Three-Dimensional Morphology in Experimental Nerve Regeneration Research

2021 ◽  
Author(s):  
Simeon C. Daeschler ◽  
Jennifer Zhang ◽  
Tessa Gordon ◽  
Gregory H. Borschel
Keyword(s):  
Nerve Regeneration ◽  
Nerve Repair ◽  
Three Dimensional ◽  
Retrograde Labelling ◽  
Tissue Clearing ◽  
Processing Times ◽  
Morphological Studies ◽  
Target Organs ◽  
Subcellular Resolution ◽  
Rapid Processing

AbstractMorphological analyses are key outcome assessments for nerve regeneration studies but are historically limited to tissue sections. Novel optical tissue clearing techniques enabling three-dimensional imaging of entire organs at a subcellular resolution have revolutionized morphological studies of the brain. To extend their applicability to experimental nerve repair studies we adapted these techniques to nerves and their motor and sensory targets in rats. The solvent-based protocols rendered harvested peripheral nerves and their target organs transparent within 24 h while preserving tissue architecture and fluorescence. Optical clearing is compatible with conventional laboratory techniques, including retrograde labelling studies, and computational image segmentation, providing fast and precise cell quantitation. Further, optically cleared organs enable three-dimensional morphometry at an unprecedented scale including dermatome-wide innervation studies, tracing of intramuscular nerve branches and mapping of neurovascular networks. Given their wide-ranging applicability, rapid processing times and low costs, tissue clearing techniques are likely to be a key technology for next-generation nerve repair studies.

Tissue clearing‐based method for unobstructed three‐dimensional imaging of mouse penis with subcellular resolution

2020 ◽  
Vol 13 (7) ◽  
Author(s):  
Paweł Matryba ◽  
Artur Wolny ◽  
Monika Pawłowska ◽  
Anna Sosnowska ◽  
Zuzanna Rydzyńska ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Three Dimensional Imaging ◽  
Tissue Clearing ◽  
Subcellular Resolution

scholarly journals Peripheral Nerve Regeneration and Muscle Reinnervation

2020 ◽  
Vol 21 (22) ◽  
pp. 8652 ◽  
Author(s):  
Tessa Gordon
Keyword(s):  
Peripheral Nerve ◽  
Nerve Regeneration ◽  
Nerve Repair ◽  
Peripheral Nerve Regeneration ◽  
Low Frequency ◽  
Nerve Fibers ◽  
Target Organs ◽  
Neuronal Soma ◽  
Original Target ◽  
Surgical Manipulations

Injured peripheral nerves but not central nerves have the capacity to regenerate and reinnervate their target organs. After the two most severe peripheral nerve injuries of six types, crush and transection injuries, nerve fibers distal to the injury site undergo Wallerian degeneration. The denervated Schwann cells (SCs) proliferate, elongate and line the endoneurial tubes to guide and support regenerating axons. The axons emerge from the stump of the viable nerve attached to the neuronal soma. The SCs downregulate myelin-associated genes and concurrently, upregulate growth-associated genes that include neurotrophic factors as do the injured neurons. However, the gene expression is transient and progressively fails to support axon regeneration within the SC-containing endoneurial tubes. Moreover, despite some preference of regenerating motor and sensory axons to “find” their appropriate pathways, the axons fail to enter their original endoneurial tubes and to reinnervate original target organs, obstacles to functional recovery that confront nerve surgeons. Several surgical manipulations in clinical use, including nerve and tendon transfers, the potential for brief low-frequency electrical stimulation proximal to nerve repair, and local FK506 application to accelerate axon outgrowth, are encouraging as is the continuing research to elucidate the molecular basis of nerve regeneration.

scholarly journals A rat study of the use of end-to-side peripheral nerve repair as a “babysitting” technique to reduce the deleterious effect of chronic denervation

2019 ◽  
Vol 131 (2) ◽  
pp. 622-632 ◽  
Author(s):  
Olawale A. R. Sulaiman ◽  
Tessa Gordon
Keyword(s):  
Nerve Regeneration ◽  
Functional Recovery ◽  
Deleterious Effect ◽  
Nerve Repair ◽  
Nerve Fibers ◽  
Nerve Stump ◽  
Muscle Weight ◽  
Target Organs ◽  
Chronic Denervation ◽  
Distal Nerve

OBJECTIVEFunctional recovery is disappointing after surgical repair of nerves that are injured far from their target organs and/or after delayed repair. In the former case, a nerve transfer that transects a distal nerve fascicle to innervate denervated targets is one strategy to promote nerve regeneration and functional recovery. An alternate strategy tested in this study is to perform an end-to-side neurorrhaphy to “babysit” (protect) the denervated distal nerve stump at the time of nerve repair and reduce the deleterious effect of chronic denervation on nerve regeneration.METHODSIn the hindlimbs of Sprague-Dawley rats, the common peroneal (CP) nerve was transected unilaterally and the distal CP nerve stump inserted through a perineurial window into the intact tibial (TIB) nerve, i.e., CP-TIB end-to-side neurorrhaphy. In the first experiment, TIB nerve motoneurons that had regenerated and/or sprouted axons into the CP nerve within 3 months were stimulated to elicit contractions, and thereafter, identified with retrograde dyes for counting. In the second experiment, the intact TIB nerve was transected and cross-sutured to a 3-month chronically denervated distal CP nerve stump that had either been “protected” by ingrown TIB nerves after CP-TIB neurorrhaphy or remained chronically denervated. Thereafter, the number of retrogradely labeled TIB nerve motoneurons that had regenerated their nerves within 3 months were counted and reinnervated tibialis anterior (TA) muscles weighed.RESULTSA mean (± SE) of 231 ± 83 TIB nerve motoneurons grew into the end-to-side CP distal nerve stump with corresponding ankle flexion; 32% regenerated their axons and 24% sprouted axons from the intact TIB nerve, eliciting ankle flexor-extensor co-contraction. In the second experiment, after a 3-month period of TIB nerve regeneration, significantly more TIB motoneurons regenerated their axons into “protected” than “unprotected” CP distal nerve stumps within 3 months (mean 332 ± 43.6 vs 235 ± 39.3 motoneurons) with corresponding and significantly higher numbers of regenerated nerve fibers, resulting in significantly better recovery of reinnervated TA muscle weight.CONCLUSIONSThese experiments in rats demonstrated that delayed nerve repair is more effective when the deleterious effects of chronic denervation of the distal nerve stump are reduced by protecting the nerve stump with ingrowing nerve fibers across an end-to-side insertion of the distal nerve stump into a neighboring intact nerve. Such an end-to-side neurorrhaphy may be invaluable as a means of preventing the atrophy of distal nerve stumps and target organs after chronic denervation, which allows for effective reinnervation of the protected distal nerve stumps and target organs over distance and time.

Tissue clearing techniques for three‐dimensional optical imaging of intact human prostate and correlations with multi‐parametric MRI

The Prostate ◽  
2021 ◽  
Author(s):  
Stefano Cipollari ◽  
Neema Jamshidi ◽  
Liutao Du ◽  
Kyunghyun Sung ◽  
Danshan Huang ◽  
...  
Keyword(s):  
Optical Imaging ◽  
Three Dimensional ◽  
Human Prostate ◽  
Tissue Clearing

scholarly journals IL-17F depletion accelerates chitosan conduit guided peripheral nerve regeneration

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Feixiang Chen ◽  
Weihuang Liu ◽  
Qiang Zhang ◽  
Ping Wu ◽  
Ao Xiao ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Sciatic Nerve ◽  
Peripheral Nerve ◽  
Nerve Regeneration ◽  
Knockout Mice ◽  
Inflammatory Markers ◽  
Nerve Repair ◽  
Peripheral Nerve Regeneration ◽  
Wild Type ◽  
Anti Inflammatory

AbstractPeripheral nerve injury is a serious health problem and repairing long nerve deficits remains a clinical challenge nowadays. Nerve guidance conduit (NGC) serves as the most promising alternative therapy strategy to autografts but its repairing efficiency needs improvement. In this study, we investigated whether modulating the immune microenvironment by Interleukin-17F (IL-17F) could promote NGC mediated peripheral nerve repair. Chitosan conduits were used to bridge sciatic nerve defect in IL-17F knockout mice and wild-type mice with autografts as controls. Our data revealed that IL-17F knockout mice had improved functional recovery and axonal regeneration of sciatic nerve bridged by chitosan conduits comparing to the wild-type mice. Notably, IL-17F knockout mice had enhanced anti-inflammatory macrophages in the NGC repairing microenvironment. In vitro data revealed that IL-17F knockout peritoneal and bone marrow derived macrophages had increased anti-inflammatory markers after treatment with the extracts from chitosan conduits, while higher pro-inflammatory markers were detected in the Raw264.7 macrophage cell line, wild-type peritoneal and bone marrow derived macrophages after the same treatment. The biased anti-inflammatory phenotype of macrophages by IL-17F knockout probably contributed to the improved chitosan conduit guided sciatic nerve regeneration. Additionally, IL-17F could enhance pro-inflammatory factors production in Raw264.7 cells and wild-type peritoneal macrophages. Altogether, IL-17F may partially mediate chitosan conduit induced pro-inflammatory polarization of macrophages during nerve repair. These results not only revealed a role of IL-17F in macrophage function, but also provided a unique and promising target, IL-17F, to modulate the microenvironment and enhance the peripheral nerve regeneration.

scholarly journals EZ Clear for simple, rapid, and robust mouse whole organ clearing

2022 ◽  
Author(s):  
Chih-Wei Hsu ◽  
Juan Cerda ◽  
Jason M Kirk ◽  
Williamson D. Turner ◽  
Tara L. Rasmussen ◽  
...  
Keyword(s):  
Biomedical Research ◽  
Room Temperature ◽  
Adult Mouse ◽  
Dimensional Space ◽  
Three Dimensional ◽  
Fluorescence Signal ◽  
Tissue Architecture ◽  
Tissue Clearing ◽  
Clearing Method ◽  
Three Dimensional Space

Tissue clearing for whole organ cell profiling has revolutionized biology and imaging for exploration of organs in three-dimensional space without compromising tissue architecture. But complicated, laborious procedures, or expensive equipment, as well as the use of hazardous, organic solvents prevents the widespread adoption of these methods. Here we report a simple and rapid tissue clearing method, EZ Clear, that can clear whole adult mouse organs in 48 hours in just three simple steps. Samples stay at room temperature and remain hydrated throughout the clearing process, preserving endogenous and synthetic fluorescence, without altering sample size. After wholemount clearing and imaging, EZ Cleared samples can be further processed for downstream embedding and cryosectioning followed by standard histology or immunostaining, without loss of endogenous or synthetic fluorescence signal. Overall, the simplicity, speed, and flexibility of EZ Clear make it easy to adopt and apply to diverse approaches in biomedical research.

scholarly journals Recent Findings on the Effects of Pharmacological Agents on the Nerve Regeneration after Peripheral Nerve Injury

2020 ◽  
Vol 18 (11) ◽  
pp. 1154-1163
Author(s):  
Samira Bolandghamat ◽  
Morteza Behnam-Rassouli
Keyword(s):  
Peripheral Nerve ◽  
Nerve Regeneration ◽  
Nerve Injury ◽  
Molecular Mechanisms ◽  
Functional Disability ◽  
Nerve Repair ◽  
Invasive Treatment ◽  
Pharmacological Agents ◽  
Non Invasive ◽  
Repair Techniques

: Peripheral nerve injuries (PNIs) are accompanied with neuropathic pain and functional disability. Despite improvements in surgical repair techniques in recent years, the functional recovery is yet unsatisfied. Indeed a successful nerve repair depends not only on the surgical strategy but also on the cellular and molecular mechanisms involved in traumatic nerve injury. In contrast to all strategies suggested for nerve repair, pharmacotherapy is a cheap, accessible and non-invasive treatment that can be used immediately after nerve injury. This study aimed to review the effects of some pharmacological agents on the nerve regeneration after traumatic PNI evaluated by functional, histological and electrophysiological assessments. In addition, some cellular and molecular mechanisms responsible for their therapeutic actions, restricted to neural tissue, are suggested. These findings can not only help to find better strategies for peripheral nerve repair, but also to identify the neuropathic effects of various medications and their mechanisms of action.

scholarly journals Learning curve in rat dissection for experimental sciatic nerve repair

2019 ◽  
pp. 243-248
Author(s):  
Marin Andrei ◽  
Marin Georgiana Gabriela ◽  
Dobrete Nicoleta Amalia ◽  
Enescu Dan Mircea
Keyword(s):  
Sciatic Nerve ◽  
Learning Curve ◽  
Nerve Regeneration ◽  
Experimental Model ◽  
Rat Model ◽  
Surgical Intervention ◽  
Nerve Repair ◽  
Learning Curves ◽  
Rat Sciatic Nerve ◽  
Nerve Surgery

The baseline for any key research in nerve regeneration is an experimental model and the sciatic nerve in the rat model is the workhorse in this field. Although physically resistant to external traumas, a surgical intervention constitutes a major distress even for a rat. In the following presentation, we will analyse the learning curves for different stages in the rat sciatic nerve surgery as well as possible factors which influence these times.

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