JoVE Logo
Faculty Resource Center

Sign In

Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Neuroscience

A Neuronal Apoptosis Model induced by Spinal Cord Compression in Rat

Published: June 29th, 2021

DOI:

10.3791/62604

1Longhua Hospital, Shanghai University of Traditional Chinese Medicine, 2Spine Research Institute, Shanghai University of Traditional Chinese Medicine, 3Shanghai University of Traditional Chinese Medicine
* These authors contributed equally

Here, we present a protocol to generate a rat spinal cord compression model, assess its behavioral score, and observe the compressed spinal cord region. The behavioral assessments showed decreased monitor motor disability. Hematoxylin and eosin staining and immunostaining revealed considerable neuronal apoptosis in the compressed region of the spinal cord.

As a severe progressive degenerative disease, cervical spondylotic myelopathy (CSM) has a poor prognosis and is associated with physical pain, stiffness, motor or sensory dysfunction, and a high risk of spinal cord injury and acroparalysis. Thus, therapeutic strategies that promote efficient spinal cord regeneration in this chronic and progressive disease are urgently needed. Effective and reproducible animal spinal cord compression models are required to understand the complex biological mechanism underlying CSM. Most spinal cord injury models reflect acute and structural destructive conditions, whereas animal models of CSM present a chronic compression in the spinal cord. This paper presents a protocol to generate a rat spinal cord compression model, which was further evaluated by assessing the behavioral score and observing the compressed spinal cord region. The behavioral assessments showed decreased monitor motor disability, including joint movements, stepping ability, coordination, trunk stability, and limb muscle strength. Hematoxylin and eosin (H&E) staining and immunostaining revealed considerable neuronal apoptosis in the compressed region of the spinal cord.

As a common progressive degenerative disease, CSM accounts for 5-10% of all cervical spondylosis1. If patients suffering from CSM ignore their symptoms and fail to treat them in a timely and effective manner, this could lead to severe complications, such as spinal cord injury and limb paralysis, which would deteriorate with aging, posing a substantial economic and mental burden to patients and their families2,3. The pathogenesis of CSM is complex, involving static and dynamic factors, the hypoxia-ischemia theory, endothelial cell injury, the blood spinal cord barrier destruction theory,....

Log in or to access full content. Learn more about your institution’s access to JoVE content here

The following procedure was performed with approval from the Institutional Animal Care and Use Committee (IACUC), Shanghai University of Traditional Chinese Medicine. All survival surgeries were performed under sterile conditions as outlined by the NIH guidelines. Pain and risk of infections were managed with appropriate analgesics and antibiotics to ensure a successful outcome. This surgical procedure is optimized for Sprague-Dawley (SD) outbred male rats at 12 weeks of age and 400 g weight.

1........

Log in or to access full content. Learn more about your institution’s access to JoVE content here

Spinal cord compressive injury may lead to neuromuscular disability in limbs
As the hydrogel piece expands gradually, it persistently compresses the spinal cord region for a prolonged period, which simulates the forelimb disabilities induced by cervical spinal cord diseases8,10. In the current model, considerable ipsilateral forepaw contracture was observed in most of the rats (9/10) in the model group (Figure 2A

Log in or to access full content. Learn more about your institution’s access to JoVE content here

The goal of this surgical procedure was to generate reproducible, prolonged, neural apoptosis in the rat spinal cord. A key advantage of this model is that the expandable hydrogel implants provide a prolonged compression on the spinal cord, thereby leading to a progressive neural apoptotic response (Figure 2C), which is consistent with the pathological process of CSM. In the current study, the mortality from spinal cord injury was extremely low (~2 in 50), whereas the repeatability of t.......

Log in or to access full content. Learn more about your institution’s access to JoVE content here

This study was supported by the National Key R&D Program of China (2018YFC1704300), National Natural Science Foundation of China (81930116, 81804115, 81873317, and 81704096), Shanghai Sailing Program (18YF1423800), Natural science Foundation of Shanghai (20ZR1473400). This project was also supported by the Shanghai University of Traditional Chinese Medicine (2019LK057).

....

Log in or to access full content. Learn more about your institution’s access to JoVE content here

Name Company Catalog Number Comments
Antibiotic ointment Prevent wound infection
Buprenorphine-SR Pain relief
Isoflurane Veteasy Anesthesia
Inhalant anesthesia equipment Anesthesia
Micro ophthalmic forceps Mingren medical equipment Length: 11 cm, Head diameter: 0.3 mm Clip the muscle
Ophthalmic forceps Shanghai Medical Devices (Group) Co., Ltd. Surgical Instruments Factory JD1050 Clip the skin
Ophthalmic scissors (10 cm) Shanghai Medical Devices (Group) Co., Ltd. Surgical Instruments Factory Y00030 Skin incision
SD male rats Shanghai SLAC Laboratory Animal Co., Ltd SCXK2018-0004 Animal model
Sterile surgical blades (22#) Shanghai Pudong Jinhuan Medical Products Co., Ltd. 35T0707 Muscle incision
Small animal trimmer Hair removal
Veet hair removal cream RECKITT BENCKISER (India) Ltd Hair removal
Venus shears Mingren medical equipment Length: 12.5 cm Muscle incision

  1. Lebl, D. R., Bono, C. M. Update on the diagnosis and management of cervical spondylotic myelopathy. The Journal of the American Academy of Orthopaedic Surgeons. 23 (11), 648-660 (2015).
  2. Haddas, R., et al. Spine and lower extremity kinematics during gait in patients with cervical spondylotic myelopathy. The Spine Journal. 18 (9), 1645-1652 (2018).
  3. Song, D. W., Wu, Y. D., Tian, D. D. Association of Vdr-Foki and Vdbp-Thr420 Lys polymorphisms with cervical spondylotic myelopathy: A case-control study in the population of China. Journal of Clinical Laboratory Analysis. 33 (2), 22669 (2019).
  4. Kurokawa, R., Murata, H., Ogino, M., Ueki, K., Kim, P. Altered blood flow distribution in the rat spinal cord under chronic compression. Spine. 36 (13), 1006-1009 (2011).
  5. Wen, C. Y., et al. Is Diffusion anisotropy a biomarker for disease severity and surgical prognosis of cervical spondylotic myelopathy. Radiology. 270 (1), 197-204 (2014).
  6. Long, H. Q., Li, G. S., Hu, Y., Wen, C. Y., Xie, W. H. Hif-1A/Vegf signaling pathway may play a dual role in secondary pathogenesis of cervical myelopathy. Medical Hypotheses. 79 (1), 82-84 (2012).
  7. Karadimas, S. K., Erwin, W. M., Ely, C. G., Dettori, J. R., Fehlings, M. G. Pathophysiology and natural history of cervical spondylotic myelopathy. Spine. 38, 21-36 (2013).
  8. Wilson, J. R., et al. State of the art in degenerative cervical myelopathy: an update on current clinical evidence. Neurosurgery. 80, 33-45 (2017).
  9. Baptiste, D. C., Fehlings, M. G. Pathophysiology of cervical myelopathy. The spine Journal. 6, 190-197 (2006).
  10. Wilcox, J. T., et al. Generating level-dependent models of cervical and thoracic spinal cord injury: exploring the interplay of neuroanatomy, physiology, and function. Neurobiology of Disease. 105, 194-212 (2017).
  11. Takano, M., et al. Inflammatory cascades mediate synapse elimination in spinal cord compression. Journal of Neuroinflammation. 11, 40 (2014).
  12. Hu, Y., et al. Somatosensory-evoked potentials as an indicator for the extent of ultrastructural damage of the spinal cord after chronic compressive injuries in a rat model. Clinical Neurophysiology. 122 (7), 1440-1447 (2011).
  13. Yang, T., et al. Inflammation level after decompression surgery for a rat model of chronic severe spinal cord compression and effects on ischemia-reperfusion injury. Neurologia Medico-Chirurgica. 55 (7), 578-586 (2015).
  14. Ijima, Y., et al. Experimental rat model for cervical compressive myelopathy. Neuroreport. 28 (18), 1239-1245 (2017).
  15. Yamamoto, S., Kurokawa, R., Kim, P. Cilostazol, a selective type iii phosphodiesterase inhibitor: prevention of cervical myelopathy in a rat chronic compression model. Journal of Neurosurgery. Spine. 20 (1), 93-101 (2014).
  16. Holly, L. T., et al. Dietary therapy to promote neuroprotection in chronic spinal cord injury. Journal of Neurosurgery. Spine. 17 (2), 134-140 (2012).
  17. Zhao, P., et al. In vivo diffusion tensor imaging of chronic spinal cord compression: a rat model with special attention to the conus medullaris. Acta Radiologica. 57 (12), 1531-1539 (2016).
  18. Kurokawa, R., Nagayama, E., Murata, H., Kim, P. Limaprost alfadex, a prostaglandin E1 derivative, prevents deterioration of forced exercise capability in rats with chronic compression of the spinal cord. Spine. 36 (11), 865-869 (2011).
  19. Lee, J., Satkunendrarajah, K., Fehlings, M. G. Development and characterization of a novel rat model of cervical spondylotic myelopathy: the impact of chronic cord compression on clinical, neuroanatomical, and neurophysiological outcomes. Journal of Neurotrauma. 29 (5), 1012-1027 (2012).
  20. Chen, B., et al. Reactivation of dormant relay pathways in injured spinal cord by Kcc2 manipulations. Cell. 174 (3), 521-535 (2018).
  21. Yu, W. R., Liu, T., Kiehl, T. R., Fehlings, M. G. Human neuropathological and animal model evidence supporting a role for Fas-mediated apoptosis and inflammation in cervical spondylotic myelopathy. Brain. 134, 1277-1292 (2011).
  22. Yu, W. R., et al. Molecular mechanisms of spinal cord dysfunction and cell death in the spinal hyperostotic mouse: implications for the pathophysiology of human cervical spondylotic myelopathy. Neurobiology of Disease. 33 (2), 149-163 (2009).
  23. Iyer, A., Azad, T. D., Tharin, S. Cervical spondylotic myelopathy. Clinical Spine Surgery. 29 (10), 408-414 (2016).

This article has been published

Video Coming Soon

JoVE Logo

Privacy

Terms of Use

Policies

Research

Education

ABOUT JoVE

Copyright © 2024 MyJoVE Corporation. All rights reserved