Name: ovine lumbar intervertebral disc degeneration model utilizing a lateral retroperitoneal drill bit injury
Uploaded: 2017-05-25T15:00:00
Description: Watch this Scientific Journal Video about Ovine Lumbar Intervertebral Disc Degeneration Model Utilizing a Lateral Retroperitoneal Drill Bit Injury at JoVE.com

Dr. Chris Daly is the recipient of the Foundation for Surgery Richard Jepson Research Scholarship. The authors would like to thank Dr. Anne Gibbon, Dr. Dong Zhang and the staff of Monash Animal Services, Monash University for their assistance with animal surgery and care.

The protocol detailed in this manuscript follows the animal care guidelines of Monash University Animal Ethics. Animal ethics approval for this protocol has been granted by Monash University Animal Ethics. Ethics approval number: MMCA/2014/55
1. Sheep Preparation
NOTE: Ewes aged two to four years were used.
<ol>
	<li>Fast sheep for 18 h prior to anesthesia. Provide animals with access to water until 6-12 h prior to operation19.</li>
	<li>Se...

<ol>
	<li>Hoy, D., March, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+global+burden+of+low+back+pain:+estimates+from+the+Global+Burden+of+Disease+2010+study.">The global burden of low back pain: estimates from the Global Burden of Disease 2010 study.</a> Ann Rheum Dis. 73 (6), 968-974 (2014).</li><li>Luoma, K., Riihim&#228;ki, H., Luukkonen, R., Raininko, R., Viikari-Juntura, E., Lammine, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Low+back+pain+in+relation+to+lumbar+disc+degeneration.">Low back pain in relation to lumbar disc degeneration.</a> Spine. 25 (4), 487-492 (2000).</li><li>Daly, C., Ghosh, P., Jenkin, G., Oehme, D., Goldschlager, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Review+of+Animal+Models+of+Intervertebral+Disc+Degeneration:+Pathophysiology,+Regeneration,+and+Translation+to+the+Clinic.">A Review of Animal Models of Intervertebral Disc Degeneration: Pathophysiology, Regeneration, and Translation to the Clinic.</a> BioMed Res Int. 2016 (3), 5952165 (2016).</li><li>Sahlman, J., Inkinen, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Premature+vertebral+endplate+ossification+and+mild+disc+degeneration+in+mice+after+inactivation+of+one+allele+belonging+to+the+Col2a1+gene+for+Type+II+collagen.">Premature vertebral endplate ossification and mild disc degeneration in mice after inactivation of one allele belonging to the Col2a1 gene for Type II collagen.</a> Spine. 26 (23), 2558-2565 (2001).</li><li>Melrose, J., Taylor, T., Ghosh, P., Holbert, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intervertebral+disc+reconstitution+after+chemonucleolysis+with+chymopapain+is+dependent+on+dosage:+An+experimental+study+in+beagle+dogs.">Intervertebral disc reconstitution after chemonucleolysis with chymopapain is dependent on dosage: An experimental study in beagle dogs.</a> Spine. 21 (1), (1996).</li><li>Oehme, D., Goldschlager, T., Shimon, S., Wu, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiological,+Morphological,+Histological+and+Biochemical+Changes+of+Lumbar+Discs+in+an+Animal+Model+of+Disc+Degeneration+Suitable+for+Evaluating+the+potential+regenerative+capacity+of+novel+biological+agents.">Radiological, Morphological, Histological and Biochemical Changes of Lumbar Discs in an Animal Model of Disc Degeneration Suitable for Evaluating the potential regenerative capacity of novel biological agents.</a> J Tiss Sci Eng. , (2015).</li><li>Platenberg, R. C., Hubbard, G. B., Ehler, W. J., Hixson, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spontaneous+disc+degeneration+in+the+baboon+model:+magnetic+resonance+imaging+and+histopathologic+correlation.">Spontaneous disc degeneration in the baboon model: magnetic resonance imaging and histopathologic correlation.</a> J Med Primatol. 30 (5), 268-272 (2001).</li><li>Iatridis, J. C., Mente, P. L., Stokes, I. A. F., Aronsson, D. D., Alini, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Compression-Induced+Changes+in+Intervertebral+Disc+Properties+in+a+Rat+Tail+Model.">Compression-Induced Changes in Intervertebral Disc Properties in a Rat Tail Model.</a> Spine. 24 (10), 996 (1999).</li><li>Silberberg, R., Aufdermaur, M., Adler, J. 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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Minimally+invasive+surgery:+lateral+approach+interbody+fusion:+results+and+review.">Minimally invasive surgery: lateral approach interbody fusion: results and review.</a> Spine. 35 (Suppl 26), S302-S311 (2010).</li><li>Zhang, Y., Drapeau, S., An, H. S., Markova, D., Lenart, B. A., Anderson, D. 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J., Matyas, J. R., Duncan, N. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cytomorphology+of+notochordal+and+chondrocytic+cells+from+the+nucleus+pulposus:+a+species+comparison.">Cytomorphology of notochordal and chondrocytic cells from the nucleus pulposus: a species comparison.</a> J Anat. 205 (5), 357-362 (2004).</li><li>Hoogendoorn, R. J., Helder, M. N., Smit, T. H., Wuisman, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Notochordal+cells+in+mature+caprine+intervertebral+discs.">Notochordal cells in mature caprine intervertebral discs.</a> Eur Cells Mater. 10 (Suppl 3), (2005).</li><li>Pohlmeyer, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Zur vergleichenden Anatomie von Damtier, Schaf und Ziege: Osteologie und postnatale Osteogenese. , (1985).</li><li>Pfirrmann, C. 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This minimally invasive lateral access approach is efficacious and safe with no post-operative herniae, abdominal wound dehiscence or infection observed in this series. Use of the drill bit intervertebral disc injury model with a depth stop provides a reproducible method of inducing a consistent intervertebral disc injury of known dimension (i.e. a 3.5 mm diameter x 12 mm depth injury in this study). In our experience, this method produces a less severe degree of disc degeneration than that observed in conventio...

Lower back pain is the leading cause of disability worldwide1. Lumbar intervertebral disc degeneration associated discogenic pain is considered a significant contributor to lower back pain2. There is an increasing demand for reliable animal models of intervertebral disc disease for broadening the understanding of the degenerative process and for the investigation of potential therapies.
Numerous animal models of intervertebral disc degeneration exist3. Animals models used in the investigation of degenerative disc disease range in size from mice4, to larger mammals such as dogs5, sheep6, and non-human primates7. Methods used to induce intervertebral disc degeneration can be broadly classified into the categories of mechanical (e.g. intervertebral disc compression8 or surgical injury6), chemical (e.g. chemical nucleolysis5) or, less commonly, spontaneous degeneration (e.g. the sand rat9).
Given the complexity of human intervertebral disc degeneration, a perfect animal model does not exist. However, important considerations in choosing an appropriate animal model to mimic this condition closely have been identified3. Such considerations include the absence of notochordal cells (primitive cells with possible progenitor cell function10 absent from the adult nucleus pulposus in humans, sheep, goats and chondrodystrophic dogs but present in most mammals), similarities in animal and intervertebral disc size relative to humans, comparable biomechanical forces to the clinical condition, mechanistic and ethical considerations3.
Non-human primates meet many of the above criteria. Baboon and macaque models of spontaneous intervertebral disc degeneration have been described11,12,13. Both species spend large amounts of time in erect or semi-erect postures &#8212; a distinct advantage relative to other animal models. However, ethical and practical consideration (e.g. expense, housing, delayed onset of spontaneous degeneration) restrict their use in many institutions.
The ovine spine is an established model of intervertebral disc degeneration, with advantages including cellular, biomechanical and anatomical similarities to the human spine10,14,15. Despite the quadrupedal stature of sheep the ovine lumbar intervertebral disc is exposed to similar stresses to the human disc14. The ovine model is also more widely accepted, from an ethical perspective, than non-human primate models. Varied methods have been described to initiate the degenerative process, many of which require direct access to the intervertebral disc. Due to the termination of the spinal cord in the sacral region and ossification of the posterior longitudinal ligament in the ovine lumbar spine, posterior approaches to the intervertebral disc are technically challenging and less commonly used in the sheep16. The traditional access routes to the sheep lumbar spine, i.e. via anterior or anterolateral approaches, require large abdominal incisions, are fraught with risks of hernia, and damage to internal viscera and neurovascular structures16. The use of a relatively small lateral incision away from dependent abdominal areas may decrease such risks17.
We present an ovine model of degenerative lumbar intervertebral disc disease using drill bit injury performed through a minimally invasive lateral approach, and inspired by the work of Zhang et. al18. The goal of this protocol is to enable a reliable lumbar disc injury model that is readily reproducible, produces a consistent injury, and is safe and well tolerated. This approach is well-suited to investigators seeking to induce a milder degree of lumbar intervertebral disc degeneration than that observed with traditional surgical annulotomy (unpublished data) for the investigation of either intervertebral disc degeneration or regenerative therapies. These findings will be described in a forthcoming publication.

<table border="0" cellpadding="0" cellspacing="0" width="947">
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		<tr height="42">
			<td height="42" style="height:42px;width:296px;">Medetomidine Hydrochloride (10 mL Injection)</td>
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			<td height="42" style="height:42px;width:296px;">Chlorhexidine 5%w/v</td>
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			<td height="42" style="height:42px;width:296px;">Atipamezole (Antisedan 0.06 mg/kg - 0.08 mg/kg)</td>
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			<td style="width:119px;">PFIANT10</td>
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			<td height="21" style="height:21px;width:296px;">Drill Bit Stop 4 mm</td>
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			<td height="21" style="height:21px;width:296px;">3-0 absorbable synthetic braided sutures</td>
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ovine lumbar intervertebral disc degeneration model utilizing a lateral retroperitoneal drill bit injury

Intervertebral disc degeneration is a significant contributor to the development of back pain and the leading cause of disability worldwide. Numerous animal models of intervertebral disc degeneration have been developed. The ideal animal model should closely mimic the human intervertebral disc with regard to morphology, biomechanical properties and the absence of notochordal cells. The sheep lumbar intervertebral disc model fulfils these criteria. We present an ovine model of intervertebral disc degeneration utilizing a drill bit injury through a lateral retroperitoneal approach. The lateral approach significantly reduces the incision and potential morbidity associated with the traditional anterior approach to the ovine spine. Utilization of a drill-bit method of injury affords the ability to produce a consistent and reproducible injury, of precise dimensions, that initiates a consistent degree of intervertebral disc degeneration. The focal nature of the annular and nucleus pulposus defect more closely mimics the clinical condition of focal intervertebral disc herniation. Sheep recover rapidly following this procedure and are typically mobile and eating within the hour. Intervertebral disc degeneration ensues and sheep undergo necropsy and subsequent analysis at periods from eight weeks. We believe that the drill bit injury model of intervertebral disc degeneration offers advantages over more conventional annular injury models.

Pre-operatively, sheep underwent baseline 3T magnetic resonance imaging (MRI) for assessment of underlying intervertebral disc morphology and degeneration. Sheep underwent additional intra-operative lateral radiography for confirmation of intervertebral disc level and calculation of disc height index. A pre-operative sagittal plane slice from 3T MRI and an intra-operative radiograph are demonstrated in Figure 1.
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Watch this Scientific Journal Video about Ovine Lumbar Intervertebral Disc Degeneration Model Utilizing a Lateral Retroperitoneal Drill Bit Injury at JoVE.com

Ovine Lumbar Intervertebral Disc Degeneration Model Utilizing a Lateral Retroperitoneal Drill Bit Injury

Intervertebral disc degeneration is a significant contributor to back pain and a leading cause of disability worldwide. Numerous animal models of intervertebral disc degeneration exist. We demonstrate an ovine model of intervertebral disc degeneration, utilizing a drill bit, which achieves a consistent disc injury and reproducible level of disc degeneration.

Intervertebral disc degeneration is a significant contributor to the development of back pain and the leading cause of disability worldwide. Numerous ...

The overall goal of this Lateral Retroperitoneal approach, Drill Bit Intervertebral Disc Injury method is to create a safe, consistent, and reproducible Ovine model of degenerative intervertebral discs. This Ovine Lumbar Intervertebral Disc Degeneration model can help answer key questions in the field of translational spinal research by providing another method for investigating the path or physiology of disc degeneration and the potential investigation of regenerative therapies. The main advantage of this approach is that it allows for consistent surgical injury through a safe, surgical approach in an appropriate animal model.After anesthetizing and intubating a sheep and preparing the operating suite according to the text protocol, place the animal on the operating table in the right lateral position. Using electronic clippers, shave the region defined superiorly by the lower ribs, inferiorly by the Iliac bone, medially by the Contralateral Lumbar Transverse Processes and approximately 10 centimeters lateral to the Ipsilateral Lumbar Transverse Processes. Palpate the Iliac crest, lumbar transverse processes, and cross a vertebral angle for the landmarks for this surgical incision site.Then, use a sterile pen to mark these landmarks. Next, with chlorhexidine and alcohol iodine antiseptic wash, disinfect the lateral abdomen. Then, place a sterile fenestrated square drape over the surgical site and a large sterile square drape on the overhead table followed by the sterile instruments.Under surgical loupe magnification and a headlight, inject local anesthetic, and then use a 22 scalpel blade attached to a four scalpel handle to make a longitudinal incision parallel to and one centimeter anterior to one to two lumbar transverse processes above and below the intervertebral disc levels of interest. Using monopolar diathermy, divide the underlying subcutaneous tissue and the lateral aspect of the abdominal wall musculature. Directing the dissection towards the tips of the lumbar transverse processes, above and below the intervertebral discs of interest.Next, divide the thoracolumbar fascia longitudinally at it's attachment to the transverse processes. Visualize and preserve the underlying quadratus lumborum, soleus muscles and the traversing neurovascular bundles. Using diathermy, maintain hemostasis through the procedure.Sweep the fingers between the plane of the peritoneum and posterior abdominal wall musculature at the exposed intervertebral disc levels to perform digital blunt dissection. Now, using diva retractor, retract the quadratus lumborum and soleus muscles posterior laterally to further expose the intervertebral discs. Then, palpate for the concave intervertebral bodies and the convex intervertebral discs.Position the retractors immediately over the discs and take care to ensure lumbar vessels are not damaged. Then, using surgical loupe magnification with the headlight, identify the lumbar vessels which are located approximately one centimeter caudal to the inferior end plate. After performing an intraoperative lateral x-ray to confirm the disc level, depending on the disc levels desired, expose the intervertebral discs by separating the surrounding structures.For levels L three, four, and above, use digital blunt dissection to sweep aside the muscular attachments over the discs. For levels L four, five, and below, use bipolar diathermian scissors to sharply divide the thicker tendonus muscular attachments over the disc. Define the drill bit injury point by observing the left lateral and interior extremities at the intervertebral disc.The disc level should be confirmed with lateral x-ray prior to drill bit injury. Fit a brad point drill bit into the power drill. Ensure the diameter of the drill bit is slightly less than the intervertebral disc height.For example, 3.5 millimeters for lumbar intervertebral discs in 60 to 70 kilogram sheep. Apply a drill bit stop to provide an unprotected drill bit length of approximately half the diameter of the lumbar intervertebral disc. For example, 12 millimeters for lumbar intervertebral discs in 60 to 70 kilogram sheep.Next, apply the drill bit to the entry point and direct it in the trajectory slightly cranial to the center of the intervertebral disc, which will minimize the risk of end plate injury. With the drill on low power, slowly advance the drill bit into the intervertebral disc for one second. If excessive resistance is encountered, adjust the trajectory in a slight cranial or caudal fashion.Once hemostasis is achieved, use Ringer's solution to irrigate the wound. Then, perform layered closure, preferably using 2-0 absorbable synthetic braided sutures to the lateral abdominal wall tissues and continuous 3-0 undyed absorbable synthetic subcuticular suture to the skin. Shown here is a pre-operative sagittal plane slice from a 3T MRI of ovine lumbar one half to the lumbosacral intervertebral disc with no evidence of significant pre-operative intervertebral disc degeneration.This intraoperative lateral radiograph confirms the intervertebral disc level and enables calculation of the disc height index. The gross morphological appearance presented here demonstrates the drill bit injury tract penetrating the annulus fibrosus and extending into the nucleus pulposus. The injury is also evident in this 9.4T MRI.While attempting this approach, it's important to direct the drill bit cranially and advance the drill bit slowly to minimize the risk of end plate injury. After watching this video, you should have a good understanding of how to produce drill bit injury in the ovine model through a lateral retroperitoneal approach. Don't forget that working with scalpels and cautery can be hazardous.Personal protective equipment and appropriate safety precautions must be followed at all times. It is important to maintain surgical sterility.

ovine-lumbar-intervertebral-disc-degeneration-model-utilizing-lateral

Research

JoVE Journal

Medicine

Renal Ischaemia Reperfusion Injury: A Mouse Model of Injury and Regeneration

Renal ischaemia reperfusion injury (IRI) is a common cause of acute kidney injury (AKI) in patients and occlusion of renal blood flow is unavoidable during renal transplantation. Experimental models that accurately and reproducibly recapitulate renal IRI are crucial in dissecting the pathophysiology of AKI and the development of novel therapeutic agents. Presented here is a mouse model of renal IRI that results in reproducible AKI. This is achieved by a midline laparotomy approach for the surgery with one incision allowing both a right nephrectomy that provides control tissue and clamping of the left renal pedicle to induce ischaemia of the left kidney. By careful monitoring of the clamp position and body temperature during the period of ischaemia this model achieves reproducible functional and structural injury. Mice sacrificed 24 hr&#160;following surgery demonstrate loss of renal function with elevation of the serum or plasma creatinine level as well as structural kidney damage with acute tubular necrosis evident. Renal function improves and the acute tissue injury resolves during the course of 7 days following renal IRI such that this model may be used to study renal regeneration. This model of renal IRI has been utilized to study the molecular and cellular pathophysiology of AKI as well as analysis of the subsequent renal regeneration.

The mouse model of renal ischaemia reperfusion injury described here comprises of a right nephrectomy that provides control tissue and clamping of the left renal pedicle to induce ischaemia that results in acute kidney injury. This model uses a midline laparotomy approach with all steps performed via one incision.

Renal ischaemia reperfusion injury (IRI) is a common cause of acute kidney injury (AKI) in patients and occlusion of renal blood flow is unavoidable ...

Utilizing Murine Inducible Telomerase Alleles in the Studies of Tissue Degeneration/Regeneration and Cancer

Telomere dysfunction-induced loss of genome integrity and its associated DNA damage signaling and checkpoint responses are well-established drivers that cause tissue degeneration during ageing. Cancer, with incidence rates greatly increasing with age, is characterized by short telomere lengths and high telomerase activity. To study the roles of telomere dysfunction and telomerase reactivation in ageing and cancer, the protocol shows how to generate two murine inducible telomerase knock-in alleles 4-Hydroxytamoxifen (4-OHT)-inducible TERT-Estrogen Receptor (mTERT-ER) and Lox-Stopper-LoxTERT (LSL-mTERT). The protocol describes the procedures to induce telomere dysfunction and reactivate telomerase activity in mTERT-ER and LSL-mTERT mice in vivo. The representative data show that reactivation of telomerase activity can ameliorate the tissue degenerative phenotypes induced by telomere dysfunction. In order to determine the impact of telomerase reactivation on tumorigenesis, we generated prostate tumor model G4 PB-Cre4 PtenL/L p53L/L LSL-mTERTL/L and thymic T-cell lymphoma model G4 Atm-/- mTERTER/ER. The representative data show that telomerase reactivation in the backdrop of genomic instability induced by telomere dysfunction can greatly enhance tumorigenesis. The protocol also describes the procedures used to isolate neural stem cells (NSCs) from mTERT-ER and LSL-mTERT mice and reactivate telomerase activity in NSCs in vitro. The representative data show that reactivation of telomerase can enhance the self-renewal capability and neurogenesis in vitro. Finally, the protocol describes the procedures for performing telomere FISH (Fluorescence In Situ Hybridization) on both mouse FFPE (Formalin Fixed and Paraffin Embedded) brain tissues and metaphase chromosomes of cultured cells.

Telomere and telomerase play essential roles in ageing and tumorigenesis. The goal of this protocol is to show how to generate two murine inducible telomerase knock-in alleles and how to utilize them in the studies of tissue degeneration/regeneration and cancer.

Telomere dysfunction-induced loss of genome integrity and its associated DNA damage signaling and checkpoint responses are well-established drivers that ...

A Repetitive Concussive Head Injury Model in Mice

Despite the concussion/ mild traumatic brain injury (mTBI) being the most frequent occurrence of traumatic brain injury, there is still a lack of knowledge on the injury and its effects. To develop a better understanding of concussions, animals are often used because they provide a controlled, rigorous, and efficient model. Studies have adapted traditional animal models to perform mTBI to stimulate mild injury severity by changing the injury parameters. These models have been used because they can produce morphologically similar brain injuries to the clinical condition and provide a spectrum of injury severities. However, they are limited in their ability to present the identical features of injuries in patients. Using a traditional impact system, a repetitive concussive injury (rCHI) model can induce mild to moderate human-like concussion. The injury degree can be determined by measuring the period of loss of consciousness (LOC) with a sign of a transient termination of breathing. The rCHI model is beneficial to use for its accuracy and simplicity in determining mTBI effects and potential treatments.

Concussion presents the most common type of traumatic brain injury. Therefore, a repetitive concussive animal model, which replicates the important features of an injury in patients, may provide a means to study concussion in a rigorous, controlled, and efficient manner.

Despite the concussion/ mild traumatic brain injury (mTBI) being the most frequent occurrence of traumatic brain injury, there is still a lack of ...

Transvaginal Mesh Insertion in the Ovine Model

This protocol describes mesh insertion into the rectovaginal septum in sheep using a single vaginal incision technique, with and without the trocar-guided insertion of anchoring arms. Parous sheep underwent the dissection of the rectovaginal septum, followed by the insertion of an implant with or without four anchoring arms, both designed to fit the ovine anatomy. The anchoring arms were put in place using a trocar and an "outside-in" technique. The cranial arms were passed through the obturator, gracilis, and adductor magnus muscles. The caudal arms were fixed near the sacrotuberous ligament, through the coccygeus muscles. This technique allows for the mimicking of surgical procedures performed in women suffering from pelvic organ prolapse. The anatomical spaces and elements are easily identified. The most critical part of the procedure is the insertion of the cranial trocar, which can easily penetrate the peritoneal cavity or the surrounding pelvic organs. This can be avoided by a more extensive retroperitoneal dissection and by guiding the trocar more laterally. This approach is designed only for experimental testing of novel implants in large animal models, as trocar-guided insertion is currently not used clinically.

This protocol describes mesh implantation in the ovine rectovaginal septum using a single vaginal incision technique, with and without the trocar-guided insertion of anchoring arms.

This protocol describes mesh insertion into the rectovaginal septum in sheep using a single vaginal incision technique, with and without the trocar-guided ...

A Mouse Model of Single and Repetitive Mild Traumatic Brain Injury

Mild Traumatic Brain Injury (mTBI) can result in the acute loss of brain function, including a period of confusion, a loss of consciousness (LOC), focal neurological deficits and even amnesia. Athletes participating in contact sports are at high risk of exposure to large number of mTBIs. In terms of the level of injury in a sporting athlete, a mTBI is defined as a mild injury that does not cause gross pathological changes, but does cause short-term neurological deficits that are spontaneously resolved. Despite previous attempts to model mTBI in mice and rats, many have reported gross adverse effects including skull fractures, intracerebral bleeding, axonal injury and neuronal cell death. Herein, we describe our highly reproducible animal model of mTBI that reproduces clinically relevant symptoms. This model uses a custom made pneumatic impactor device to deliver a closed-head trauma. This impact is made under precise velocity and deformation parameters, creating a reliable and reproducible model to examine the mechanisms that contribute to effects of single or repetitive concussive mTBI.

Athletes absorb several hundred mild traumatic brain injuries (mTBI)/concussions every year; however, the consequence of these on the brain is poorly understood. Therefore, an animal model of single and repetitive mTBI that consistently replicates clinically relevant symptoms provides the means to advance the study of mTBI and concussion.

Mild Traumatic Brain Injury (mTBI) can result in the acute loss of brain function, including a period of confusion, a loss of consciousness (LOC), focal ...

Spinal Cord Lateral Hemisection and Asymmetric Behavioral Assessments in Adult Rats

Incomplete spinal cord injury (SCI) often leads to impairments of sensorimotor functions and is clinically the most frequent type of SCI. Human Brown-S&#233;quard syndrome&#160;is a common type of incomplete SCI caused by a lesion to one half of the spinal cord which results in paralysis and loss of&#160;proprioception&#160;on the same (or ipsilesional) side as the injury, and loss of pain and temperature sensation on the opposite (or contralesional) side. Adequate methodologies for producing a spinal cord lateral hemisection (HX) and assessing neurological impairments are essential to establish a reliable animal model of Brown-S&#233;quard syndrome. Although lateral hemisection model plays a pivotal role in basic and translational research, standardized protocols for creating such a hemisection and assessing unilateralized function are lacking. The goal of this study is to describe step-by-step procedures to produce a rat spinal lateral HX at the 9th thoracic (T9) vertebral level. We, then, describe a combined behavior scale for HX (CBS-HX) that provides a simple and sensitive assessment of asymmetric neurological performance for unilateral SCI. The CBS-HX, ranging from 0 to 18, is composed of 4 individual assessments which include unilateral hindlimb stepping (UHS), coupling, contact placing, and grid walking. For CBS-HX, the ipsilateral and contralateral hindlimbs are assessed separately. We found that, after a T9 HX, the ipsilateral hindlimb showed impaired behavior function whereas the contralateral hindlimb showed substantial recovery. The CBS-HX effectively discriminated behavioral functions between ipsilateral and contralateral hindlimbs and detected temporal progression of recovery of the ipsilateral hindlimb. The CBS-HX components can be analyzed separately or in combination with other measures when needed. Although we only provided visual descriptions of the surgical procedures and behavioral assessments of a thoracic HX, the principle may be applied to other incomplete SCIs and at other levels of the injury.

Here we describe surgical procedures to produce a reliable spinal cord lateral hemisection (HX) at the 9th thoracic level in adult rats and neurobehavioral assessments designed for detecting asymmetric deficits after such a unilateral injury.

Incomplete spinal cord injury (SCI) often leads to impairments of sensorimotor functions and is clinically the most frequent type of SCI. Human ...

A Mobile Outside-in Technique of Transforaminal Lumbar Endoscopy for Lumbar Disc Herniations

Percutaneous endoscopic transforaminal lumbar discectomy (PETLD) has now become a standard of care for the management of lumbar disc disease. There are two techniques for the introduction of a working cannula with respect to disc&#8212;outside-in and inside-out. The aim of this prospective study is to describe the technical aspects of a novel mobile outside-in method in dealing with different types of disc prolapse. A total of 184 consecutive patients with unilateral lower limb radiculopathy due to lumbar disc prolapse were operated on with the mobile outside-in technique of PETLD. Their clinical outcomes were evaluated based on the type of disc prolapse they had, a visual analog scale (VAS) leg pain score, the Oswestry Disability Index (ODI), and the Macnab criteria. The completeness of the decompression was documented with a postoperative magnetic resonance imaging. The mean age of the patients was 50 &#177; 16 years and the male/female ratio was 2:1. The mean follow-up was 19 &#177; 6 months. A total of 190 lumbar levels were operated on (L1&#8211;L2: n = 4, L2&#8211;L3: n = 17, L3&#8211;L4: n = 27, L4&#8211;5: n = 123, and L5&#8211;S1: n = 19). Divided into types, the patient distribution was central: n = 14, paracentral: n = 74, foraminal: n = 28, far lateral: n = 13, superior-migrated: n = 8, inferior migrated: n = 38, and high canal compromise: n = 9. The mean operative time was 35 &#177; 12 (25 - 56) min and the mean hospital stay was 1.2 &#177; 0.5 (1&#8211;3) days. The VAS score for leg pain improved from 7.5 &#177; 1 to 1.7 &#177; 0.9. The ODI improved from 70 &#177; 8.3 to 23 &#177; 5. According to the Macnab criteria, 75 patients (40.8%) had excellent results, 104 patients (56.5%) had good results, and 5 patients (2.7%) had fair results. Recurrence (including early and late) was seen in 15 out of the 190 levels that were operated on (7.89%). This article presents a novel outside-in approach that relies on a precise landing within the foramen in a mobile manner and does not solely depend upon the enlargement of the foramen. It is more versatile in application and useful in the management of all types of disc prolapse, even in severe canal compromise and high migration.

Here we present a protocol of a novel outside-in technique of transforaminal endoscopic discectomy for lumbar disc herniations. The technical aspects of the technique, the wide indications of use, and the results of the treatment in 184 patients are described in detail.

Percutaneous endoscopic transforaminal lumbar discectomy (PETLD) has now become a standard of care for the management of lumbar disc disease. There are ...

A Proinflammatory, Degenerative Organ Culture Model to Simulate Early-Stage Intervertebral Disc Disease.

Symptomatic intervertebral disc (IVD) degeneration (IDD) is a major socioeconomic burden and is characterized by inflammation and tissue degradation. Due to the lack of causative therapies, there is an urgent need for innovative experimental organ culture models to study the mechanisms involved in the progression of the disease, find therapeutic targets, and reduce the need for animal models. We here present a novel, three-dimensional organ culture model protocol mimicking the proinflammatory and catabolic microenvironment, which is present during IDD. 
Initially, bovine caudal IVDs were dissected, cleaned, and cultured in the tissue culture medium. Dynamic physiologic or pathologic loading was applied in a custom-made bioreactor for 2 hours per day. IVDs were assigned to a control group (high glucose medium, physiological loading, phosphate-buffered saline injection) and a pathological group (low glucose medium, pathological loading, tumor necrosis factor-alpha injection) for four days. Gene expression analysis from collected nucleus pulposus cells of the IVDs and enzyme-linked immunosorbent assay of the conditioned organ culture media was performed. 
Our data revealed a higher expression of inflammatory markers and reduced disc heights after loading in the pathological group compared to the control group. This protocol is reliable to simulate IVD inflammation and degeneration and can be further expanded to broaden its application scope.

This protocol presents a novel experimental model of proinflammatory, degenerative bovine organ culture to simulate early-stage intervertebral disc degeneration.

Symptomatic intervertebral disc (IVD) degeneration (IDD) is a major socioeconomic burden and is characterized by inflammation and tissue degradation. Due ...

Chronic Ovine Model of Right Ventricular Failure and Functional Tricuspid Regurgitation

The pathophysiology of severe functional tricuspid regurgitation (FTR) associated with right ventricular dysfunction is poorly understood, leading to suboptimal clinical results. We set out to establish a chronic ovine model of FTR and right heart failure to investigate the mechanisms of FTR. Twenty adult male sheep (6-12 months old, 62 &#177; 7 kg) underwent a left thoracotomy and baseline echocardiography. A pulmonary artery band (PAB) was placed and cinched around the main pulmonary artery (PA) to at least double the systolic pulmonary artery pressure (SPAP), inducing right ventricular (RV) pressure overload and signs of RV dilatation. PAB acutely increased the SPAP from 21 &#177; 2 mmHg to 62 &#177; 2 mmHg. The animals were followed for 8 weeks, symptoms of heart failure were treated with diuretics, and surveillance echocardiography was used to assess for pleural and abdominal fluid collection. Three animals died during the follow-up period due to stroke, hemorrhage, and acute heart failure. After 2 months, a median sternotomy and epicardial echocardiography were performed. Of the surviving 17 animals, 3 developed mild tricuspid regurgitation, 3 developed moderate tricuspid regurgitation, and 11 developed severe tricuspid regurgitation. Eight weeks of pulmonary artery banding resulted in a stable chronic ovine model of right ventricular dysfunction and significant FTR. This large animal platform can be used to further investigate the structural and molecular basis of RV failure and functional tricuspid regurgitation.

Right ventricular failure and functional tricuspid regurgitation are associated with left-sided heart disease and pulmonary hypertension, which contribute significantly to morbidity and mortality in patients. Establishing a chronic ovine model to study right ventricular failure and functional tricuspid regurgitation will help in understanding their mechanisms, progression, and possible treatments.

The pathophysiology of severe functional tricuspid regurgitation (FTR) associated with right ventricular dysfunction is poorly understood, leading to ...

Full-Endoscopic Isolation Zone Technique for the Treatment of Lumbar Disc Herniation

Lumbar disc herniation (LDH) is a type of serious sinus or sciatic nerve dysfunction caused by nucleus pulposus protrusion and annulus fibrosus tears. Its clinical symptoms often include severe low back pain, limited lumbar movement, sciatic nerve pain in the lower limbs, and even cauda equina syndrome. The common treatment for LDH is a conservative treatment scheme involving medicine, rest, and physical therapy. However, if the conservative treatment scheme is ineffective, a surgical treatment approach is adopted. Traditional open lumbar surgery has some disadvantages, including the potential for severe surgical trauma, severe blood loss during the operation, instability of the lumbar spine, and loss of the lumbar motor unit. Among the minimally invasive surgical schemes, full-endoscopic spine surgery (FESS) is undoubtedly the most appropriate and has the advantages of minimal trauma, high safety, quick postoperative recovery, and the retention of the stable structure and the motor unit of the lumbar spine. However, simultaneously, incomplete removal of the nucleus pulposus and residual nerve dysfunction after surgery can occur. To avoid these shortcomings, we studied a specific spinal endoscopy technique, the "isolation zone" surgical strategy, which can effectively block the pain from the nerve conduction pathway by completely relieving the nerve compression and nerve dysfunction through the orderly treatment of the protruding nucleus pulposus, the fissure of the annulus fibrosus, the sinus nerve, and the surrounding inflammatory soft tissues.

Here, we present a protocol for the &#34;isolation zone&#34; technique in the treatment of lumbar disc herniation (LDH) under full-endoscopic spine surgery (FESS), including intervertebral foramen formation, targeted catheterization, nucleus pulposus resection, and annulus fibrosus formation, which together completely block the pain from the nerve conduction pathway.

Lumbar disc herniation (LDH) is a type of serious sinus or sciatic nerve dysfunction caused by nucleus pulposus protrusion and annulus fibrosus tears. Its ...