|Year : 2017 | Volume
| Issue : 1 | Page : 14-18
Application of novel computing technologies regarding gait analysis, such as CatWalk XT, in spinal cord regeneration, in the fields of experimental neurosurgery and neurophysiology
Wojciech Slusarczyk, Sławomir Gumularz, Radosław Zachara, Marek Hamm, Hanna Cholewa, Dorota Chlebosz, Katarzyna Duda, Maria Kornaś, Arkadiusz Liśkiewicz, Jan Wiaderkiewicz, Wiesław Marcol, Piotr Morawski, Adam Właszczuk, Joanna Lewin-Kowalik
Department of Physiology, School of Medicine in Katowice, Medical University of Silesia, Katowice, Poland
|Date of Web Publication||29-May-2017|
Department of Physiology Medical University of Silesia ul., Medyków 18 40-752, Katowice, Ligota
Source of Support: None, Conflict of Interest: None
Purpose: Our project focuses on assessing the possibility of spinal cord regeneration after its injury. Trying to find an effective therapy is possible nowadays because of modern pharmacology and molecular biology achievements. Materials and Methods: The object of research is male Wistar C rats. To obtain a selective and repeatable spinal tissue damage, we use the pressure impactor. Then, rats are administered with various types of therapeutic substances, which are considered to have a neuroprotective function. Afterward, the effect of our experiment is precisely measured by CatWalk XT device which measures numerous gait parameters. Results: It was found that several parameters significantly varied between therapeutic groups, in particular, MaxContactArea, PrintLength, PrintWidth, PrintArea, SwingSpeed, and StrideLength. Conclusions: CatWalk testing is a simple yet robust tool gait analysis in rats after spinal cord injury.
Keywords: CatWalk, Neuroregeneration, spinal cord
|How to cite this article:|
Slusarczyk W, Gumularz S, Zachara R, Hamm M, Cholewa H, Chlebosz D, Duda K, Kornaś M, Liśkiewicz A, Wiaderkiewicz J, Marcol W, Morawski P, Właszczuk A, Lewin-Kowalik J. Application of novel computing technologies regarding gait analysis, such as CatWalk XT, in spinal cord regeneration, in the fields of experimental neurosurgery and neurophysiology. J Spinal Stud Surg 2017;1:14-8
|How to cite this URL:|
Slusarczyk W, Gumularz S, Zachara R, Hamm M, Cholewa H, Chlebosz D, Duda K, Kornaś M, Liśkiewicz A, Wiaderkiewicz J, Marcol W, Morawski P, Właszczuk A, Lewin-Kowalik J. Application of novel computing technologies regarding gait analysis, such as CatWalk XT, in spinal cord regeneration, in the fields of experimental neurosurgery and neurophysiology. J Spinal Stud Surg [serial online] 2017 [cited 2022 Jan 29];1:14-8. Available from: https://www.jsss-journal.com/text.asp?2017/1/1/14/207209
| Introduction|| |
The authors, cooperating with the Silesian Technical University and the Institute for Research KOMAG, constructed a device called the “pressure impactor (PI).” PI is an appliance which is capable of making a repeatable, isolated spinal cord injury (SCI), mainly within the white matter. PI can perform a spinal injury through a small opening in the dorsolateral part of the vertebra. It allows us to avoid an extensive opening of the bone, called laminectomy. This procedure significantly reduces the possibility of postoperative complications, including the risk of destabilizing the spinal column. Laminectomy procedure can affect the results of the experiment because it is a way of treating the posttraumatic edema of the spinal cord. The new method of producing shock waves using compressed air does not require opening of the dural sac – the injury is caused by the shock wave created by compressed air. The energy is penetrating into the spinal cord through dura mater, so it reflects conditions causing similar injuries in humans [Figure 1]. The PI and the method of SCI have been patented (patent number NN-380468).
|Figure 1: Action of the impactor – schematic drawing. Spinal cord is stabilized in a frame, pressure impactor is damaging the spinal cord with a shock wave.|
Click here to view
Creating a suitable model for spinal cord damage is only the beginning of the whole process. The aim of our experiments is to test the regeneration processes after SCI. Choosing the appropriate neuroprotective and healing therapy and selection of appropriate tests to assess the process of regeneration are the biggest problems. In the case of SCI, the most important indicator of experimental therapy's effectiveness will be the return of the motor function. The authors in previous studies of SCI used the whole panel of functional tests, often using personally designed devices such as recording in a transparent tunnel, “footprint test,” tensometric tunnel, and esthesiometer connected to a tensometer. The crucial disadvantage of these tests was to perform multiple activities that require much experience, time, the skill of interpretation, and manual processing of data. They were also inconvenient for tested animals. Therefore, it was important to find a suitable, simple, and advanced system for assessing the return of motor function in animals after SCI. CatWalk XT 9.0 (Noldus Information Technology, Wageningen, The Netherlands) is a device which can complete all of these tasks.
| Materials and Methods|| |
All of the procedures were approved by the Local Animal Ethical Committee.
The experiment was conducted on adult male Wistar C rat, weight 300 ± 50 g. The SCI was performed at the thoracic (Th) 11 vertebra using the PI. After intraperitoneal anesthesia with chloral hydrate (420 mg/kg), animals were placed on a heated platform and attached to a stabilizing mechanism (frame stabilizing the animal's head and spine-vertebrae Th10 and Th12).
After the skin incision, the dorsal and lateral surfaces of vertebras from Th10 to Th12 were dissected. Using a drill, a spinous process of the vertebra Th11 was removed. Later, controlling the operating field with a microscope, an opening on the arc of Th11 was made (hole Ø 3 mm).
Next, the PI was installed in the operating field. Compressed air caused a short-term pressure on the spinal cord, without causing damage to the dural sac and blood vessels. Parameters of the shock wave (time of impulse - 0.1 s; pressure – 200 kPa) have been selected carefully to induce the selective damage of the spinal cord half with transient limb paresis [Figure 2] and [Figure 3].
|Figure 2: Intraoperative photograph. Pressure impactor with spine-stabilizing frame and operating microscope. Vertebrae thoracic 10–12 are exposed.|
Click here to view
|Figure 3: Intraoperative photograph (operating microscope). The tip of the pressure impactor is touching the dura mater at the level of thoracic 11.|
Click here to view
After the injury, neuroprotective therapy was applied. The progress of the regeneration of the spinal cord and recovery of motor functions was assessed using CatWalk (CW) gait analysis; moreover, histological and immunohistochemical examination of spinal cord sections taken 6 weeks after the injury was performed. Before the surgery, all rats were subjected to a preliminary assessment using CW, to determine the gait parameters. Functional studies using CW were regularly performed at day 4, 7, 14, 28, and 42 postinjury. The CW examination consists of letting the animals walk (preferably three times) through a tunnel with transparent floor, which is lit up. The device uses a light-emitting diode, which provides light inside the glass floor of the tunnel [Figure 4].
The emitted light is then reflected everywhere, except the places where the animal's paws touch the floor. In these spots, the light refracts. A high-speed camera, located beneath the glass floor, records the floodlit areas. The images are automatically sent to the computer and analyzed. If the animal applies more pressure in a particular area, the image is subsequently brighter – it allows noticing even subtle differences between parameters responding to different paws [Figure 5].
If the animal presses more on a particular area, obtained image is proportionally brighter – showing even subtle differences between parameters of the individual paw. In the case of issues during the gait analysis (e.g., if the animal stops during the walk, it turns back), the computer automatically rejects the run and asks for a next one. A computer program automatically identifies particular rat's paw (dividing paws into front/hind and right/left), remembers, and analyzes individual fingers of each paw during the walk. Analysis of the walk was achieved with the following detection parameters: camera gain = 16.01 and intensity threshold = 0.12. Four groups of rats, each consisting of five animals, were assessed:
- K – Control – Injury without therapy
- NAP – Neuroprotective therapy with NAP after injury
- Sch – Therapy with Schwann cells after injury
- MG – Therapy with activated microglial cell after injury.
| Results|| |
The authors obtained numerous statistically significant results characterizing gait impairment and return of kinesthetic function during the treatment. They will be the subject of a separate article.
During the functional tests of movement using CW, we achieved the following parameters independent of the presence/number of footprints and their classification:
- Run duration (s): The time of whole registered run
- Average speed (cm/s): The average speed of run
- Maximum variation (%): The maximal registered variation of speed.
The above-mentioned parameters are calculated based on time of whole registered run and are independent of selected and classified footprints.
The following parameters are analyzed as a group, comparing the difference between particular paws or pair of paws of the animal. In the majority, results are averaged of all registered footprints of particular paw:
- Stand (s): Time of paw's contact with floor. Stand index: Rate describing speed of detaching paw from the floor
- MaxContactArea (%): The moment when the animal starts pushing off the paw, defined as the percent of time that passed from the moment of contact with the floor
- Max contact max intensity (0–255): The maximal intensity of footprint during the maximal contact area of paw with the floor – it is given the value of footprint that had the maximal intensity of all prints from particular run
- Max contact mean intensity (0–255): The mean intensity during the maximal contact of paw with the floor – the average of all registered footprints for particular paw
- PrintArea (cm 2): The area of footprint registered during the maximal contact of paw with the floor
- Max intensity (%): The moment of step in which the maximal intensity of footprint was registered
- Max intensity mean (0–255): The maximal registered intensity of footprint during the contact of paw with the floor – the average of all classified paws
- Min intensity mean (0–255): The minimal registered intensity of footprint during the contact of paw with the floor – the average of all classified paws
- Mean intensity mean (0–255): The averaged registered intensity of footprint during the contact of paw with the floor – the average of all classified paws
- Mean intensity of the 15 most intense pixels (0–255): Mean intensity of 15 most intensive imprints
- Swing (s): Time of having the paw lifted up between steps
- SwingSpeed (cm/s): Speed of transporting the lifted up paw
- StrideLength (cm): Length of one step. Distance between consecutive imprints of the same paw
- Step cycle (s): Sum of step and swing. It presents the time that passes between consecutive imprints of the same paw
- Duty cycle (%): Stand/(stand + swing) × 100%. Percentage that shows how much of the step is in contact with the surface
- Single stance (s): Time in which only one of the front or hind pair of paws is in contact with the surface
- Initial dual stance (s): Moment in which for the first time both of the front or hind paws are at the same time in contact with the surface
- Terminal dual stance (s): Moment in which for the second time both of the front or hind paws are at the same time in contact with the surface
- Step sequence (1-2-3-4): Order of putting down the paws. 1 = LH, 2 = LF, 3 = RH, 4 = RF (L – left, R – right, H – hind, F – front)
- Regularity index (%): Percentage of correct step sequences in regard to all step sequences
- Base of support (cm): Distance between RF and LF or RH and LH
- Cadence (step/s): Number of steps per se cond
- Print positions (cm): Distance between the hind paw and previously put down front paw at the right or left side of the animal. A positive value means that the hind paw is put behind the front one
- Phase dispersions (−50%–>75%): Shows ratio of putting down paws in the step cycle, which can let us evaluate the level of coordination between the paws in question
- Couplings (0–100%): Shows the same ratio as phase dispersions, but the evaluated paw is always put down after the one which serves as a baseline
- Support formula: Amount of paws being in contact with the surface between consecutive imprints of the left hind paw
- Footfall formula: Shows information regarding which paws are in contact with the surface in the following moment
- Standing on %: Percentage of time in which the animal uses the following combination of paws.
Gait analysis using CW has let us obtain statistically significant results in groups, where standard gait observation and neurological examination showed no obvious differences. We have found statistically significant differences in the following parameters: MaxContactArea, PrintLength, PrintWidth, PrintArea, SwingSpeed, and StrideLength. It confirms what other researchers have already proved – CW being a practical tool, useful to distinguish even subtle gait dysfunctions.
| Discussion|| |
In the past years, we have noted a significant progress in research regarding central and peripheral nervous system (CNS and PNS) regeneration. To evaluate changes occurring during experiments, standardized methods should be used, to enable comparing results between different research teams. Gait analysis is crucial in research involving CNS and PNS. It is extremely important due to the huge number of patients hopefully awaiting treatment of their severe disability – being unable to move on their own. In the opinion of the authors, based on own experience and literature, the CW examination enables to perform a wide analysis of gait. Its importance has been so far recognized in neurodegenerative disorders, for example, in the animal model of Parkinson's disease,, cerebral hemorrhages, polyneuropathies, peripheral nerve injuries,,,,, SCIs ,,, neuropathic pain , as well as osteoarthritis, and inflammatory joint diseases., Literature offers a variety of publications describing how to use CW in the most effective way., CW, a system for quantitative gait analysis, allows us to notice even subtle neurological changes.
| Conclusions|| |
CW testing may successfully replace other, more complicated methods of gait analysis. It allows to evaluate static and dynamic parameters at the same time. CW examination may be performed by a person with minimal experience. For the animal, it lasts only around a dozen seconds, and in this short time, it enables a multifunctional gait analysis. To assess motor functions, the majority of authors use two tests, which are CW and open field test (Basso, Beattie, and Bresnahan), as they are complementary.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Marcol W, Slusarczyk W, Gzik M, Larysz-Brysz M, Bobrowski M, Grynkiewicz-Bylina B, et al.
Air gun impactor – A novel model of graded white matter spinal cord injury in rodents. J Reconstr Microsurg 2012;28:561-8.
Westin JE, Janssen ML, Sager TN, Temel Y. Automated gait analysis in bilateral parkinsonian rats and the role of L-DOPA therapy. Behav Brain Res 2012;226:519-28.
Zhou M, Zhang W, Chang J, Wang J, Zheng W, Yang Y, et al.
Gait analysis in three different 6-hydroxydopamine rat models of Parkinson's disease. Neurosci Lett 2015;584:184-9.
Liu Y, Ao LJ, Lu G, Leong E, Liu Q, Wang XH, et al.
Quantitative gait analysis of long-term locomotion deficits in classical unilateral striatal intracerebral hemorrhage rat model. Behav Brain Res 2013;257:166-77.
Huehnchen P, Boehmerle W, Endres M. Assessment of paclitaxel induced sensory polyneuropathy with “Catwalk” automated gait analysis in mice. PLoS One 2013;8:e76772.
Liskiewicz A, Wlaszczuk A, Gendosz D, Larysz-Brysz M, Kapustka B, Laczynski M, et al.
Sciatic nerve regeneration in rats subjected to ketogenic diet. Nutr Neurosci 2016;19:116-24.
Murakami K, Kuniyoshi K, Iwakura N, Matsuura Y, Suzuki T, Takahashi K, et al.
Vein wrapping for chronic nerve constriction injury in a rat model: Study showing increases in VEGF and HGF production and prevention of pain-associated behaviors and nerve damage. J Bone Joint Surg Am 2014;96:859-67.
Simões GF, Benitez SU, Oliveira AL. Granulocyte colony-stimulating factor (G-CSF) positive effects on muscle fiber degeneration and gait recovery after nerve lesion in MDX mice. Brain Behav 2014;4:738-53.
Bozkurt A, Deumens R, Scheffel J, O'Dey DM, Weis J, Joosten EA, et al.
CatWalk gait analysis in assessment of functional recovery after sciatic nerve injury. J Neurosci Methods 2008;173:91-8.
Deumens R, Marinangeli C, Bozkurt A, Brook GA. Assessing motor outcome and functional recovery following nerve injury. Methods Mol Biol 2014;1162:179-88.
Kanno H, Pressman Y, Moody A, Berg R, Muir EM, Rogers JH, et al.
Combination of engineered Schwann cell grafts to secrete neurotrophin and chondroitinase promotes axonal regeneration and locomotion after spinal cord injury. J Neurosci 2014;34:1838-55.
Gensel JC, Tovar CA, Hamers FP, Deibert RJ, Beattie MS, Bresnahan JC. Behavioral and histological characterization of unilateral cervical spinal cord contusion injury in rats. J Neurotrauma 2006;23:36-54.
Garcia-Ovejero D, González S, Paniagua-Torija B, Lima A, Molina-Holgado E, De Nicola AF, et al.
Progesterone reduces secondary damage, preserves white matter, and improves locomotor outcome after spinal cord contusion. J Neurotrauma 2014;31:857-71.
Chiang CY, Sheu ML, Cheng FC, Chen CJ, Su HL, Sheehan J, et al.
Comprehensive analysis of neurobehavior associated with histomorphological alterations in a chronic constrictive nerve injury model through use of the CatWalk XT system. J Neurosurg 2014;120:250-62.
Parvathy SS, Masocha W. Gait analysis of C57BL/6 mice with complete Freund's adjuvant-induced arthritis using the CatWalk system. BMC Musculoskelet Disord 2013;14:14.
Ishikawa G, Koya Y, Tanaka H, Nagakura Y. Long-term analgesic effect of a single dose of anti-NGF antibody on pain during motion without notable suppression of joint edema and lesion in a rat model of osteoarthritis. Osteoarthritis Cartilage 2015;23:925-32.
Chen YJ, Cheng FC, Sheu ML, Su HL, Chen CJ, Sheehan J, et al.
Detection of subtle neurological alterations by the Catwalk XT gait analysis system. J Neuroeng Rehabil 2014;11:62.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]