Name: sagittal plane kinematic gait analysis in c57bl/6 mice subjected to mog35-55 induced experimental autoimmune encephalomyelitis
Uploaded: 2017-11-04T13:00:00
Description: Watch this Scientific Journal Video about Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis at JoVE.com

Wij willen erkennen Sid Chedrawe voor zijn technische bijstand met filmen. Dit werk werd gesteund door de financiering van het MS Society of Canada (EGID 2983).

dit protocol is volgens de Canadese Raad betreffende richtsnoeren Animal Care en werd goedgekeurd door het Comité van Dalhousie University op laboratoriumdieren.
 1. bouwen reflecterende Markeringen: <ol><li>met behulp van een hand-held perforator, het gewenste aantal kleine cirkels van een vel reflecterende papier punch. Elk dier moet 5 markeringen voor een enkele opname; twee grote en drie kleine markeringen.</li>
	<li>Met fijne schaar, een rechte snede uitbreiding van de omtrek naar het...

<ol>
	<li>Giladi, N., Horak, F. B., Hausdorff, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classification+of+gait+disturbances:+distinguishing+between+continuous+and+episodic+changes.">Classification of gait disturbances: distinguishing between continuous and episodic changes.</a> Mov Disord. 28 (11), 1469-1473 (2013).</li><li>Kiehn, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decoding+the+organization+of+spinal+circuits+that+control+locomotion.">Decoding the organization of spinal circuits that control locomotion.</a> Nat Rev Neurosci. 17 (4), 224-238 (2016).</li><li>Akay, T., Tourtellotte, W. G., Arber, S., Jessell, T. 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M., 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=Basso+Mouse+Scale+for+locomotion+detects+differences+in+recovery+after+spinal+cord+injury+in+five+common+mouse+strains.">Basso Mouse Scale for locomotion detects differences in recovery after spinal cord injury in five common mouse strains.</a> J Neurotrauma. 23 (5), 635-659 (2006).</li><li>Tatem, K. S., 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=Behavioral+and+locomotor+measurements+using+an+open+field+activity+monitoring+system+for+skeletal+muscle+diseases.">Behavioral and locomotor measurements using an open field activity monitoring system for skeletal muscle diseases.</a> J Vis Exp. (91), e51785 (2014).</li><li>Hetze, S., Romer, C., Teufelhart, C., Meisel, A., Engel, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gait+analysis+as+a+method+for+assessing+neurological+outcome+in+a+mouse+model+of+stroke.">Gait analysis as a method for assessing neurological outcome in a mouse model of stroke.</a> J Neurosci Methods. 206 (1), 7-14 (2012).</li><li>Preisig, D. F., 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=High-speed+video+gait+analysis+reveals+early+and+characteristic+locomotor+phenotypes+in+mouse+models+of+neurodegenerative+movement+disorders.">High-speed video gait analysis reveals early and characteristic locomotor phenotypes in mouse models of neurodegenerative movement disorders.</a> Behav Brain Res. 311, 340-353 (2016).</li><li>Leblond, H., L'Esperance, M., Orsal, D., Rossignol, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treadmill+locomotion+in+the+intact+and+spinal+mouse.">Treadmill locomotion in the intact and spinal mouse.</a> J Neurosci. 23 (36), 11411-11419 (2003).</li><li>Ueno, M., Yamashita, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinematic+analyses+reveal+impaired+locomotion+following+injury+of+the+motor+cortex+in+mice.">Kinematic analyses reveal impaired locomotion following injury of the motor cortex in mice.</a> Exp Neurol. 230 (2), 280-290 (2011).</li><li>Zorner, B., 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=Profiling+locomotor+recovery:+comprehensive+quantification+of+impairments+after+CNS+damage+in+rodents.">Profiling locomotor recovery: comprehensive quantification of impairments after CNS damage in rodents.</a> Nat Methods. 7 (9), 701-708 (2010).</li><li>Pearson, K. G., Acharya, H., Fouad, 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+new+electrode+configuration+for+recording+electromyographic+activity+in+behaving+mice.">A new electrode configuration for recording electromyographic activity in behaving mice.</a> J Neurosci Methods. 148 (1), 36-42 (2005).</li><li>Balkaya, M., Krober, J. M., Rex, A., Endres, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessing+post-stroke+behavior+in+mouse+models+of+focal+ischemia.">Assessing post-stroke behavior in mouse models of focal ischemia.</a> J Cereb Blood Flow Metab. 33 (3), 330-338 (2013).</li><li>Akay, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+measurement+of+muscle+denervation+and+locomotor+behavior+in+individual+wild-type+and+ALS+model+mice.">Long-term measurement of muscle denervation and locomotor behavior in individual wild-type and ALS model mice.</a> J Neurophysiol. 111 (3), 694-703 (2014).</li><li>Taylor, T. N., Greene, J. G., Miller, G. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+phenotyping+of+mouse+models+of+Parkinson's+disease.">Behavioral phenotyping of mouse models of Parkinson's disease.</a> Behav Brain Res. 211 (1), 1-10 (2010).</li><li>Chen, K., 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=Differential+Histopathological+and+Behavioral+Outcomes+Eight+Weeks+after+Rat+Spinal+Cord+Injury+by+Contusion,+Dislocation,+and+Distraction+Mechanisms.">Differential Histopathological and Behavioral Outcomes Eight Weeks after Rat Spinal Cord Injury by Contusion, Dislocation, and Distraction Mechanisms.</a> J Neurotrauma. 33 (18), 1667-1684 (2016).</li><li>de Bruin, N. M., 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=Multiple+rodent+models+and+behavioral+measures+reveal+unexpected+responses+to+FTY720+and+DMF+in+experimental+autoimmune+encephalomyelitis.">Multiple rodent models and behavioral measures reveal unexpected responses to FTY720 and DMF in experimental autoimmune encephalomyelitis.</a> Behav Brain Res. 300, 160-174 (2016).</li><li>Steinman, L., Zamvil, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+to+successfully+apply+animal+studies+in+experimental+allergic+encephalomyelitis+to+research+on+multiple+sclerosis.">How to successfully apply animal studies in experimental allergic encephalomyelitis to research on multiple sclerosis.</a> Ann Neurol. 60 (1), 12-21 (2006).</li><li>Emerson, M. R., Gallagher, R. J., Marquis, J. G., LeVine, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhancing+the+ability+of+experimental+autoimmune+encephalomyelitis+to+serve+as+a+more+rigorous+model+of+multiple+sclerosis+through+refinement+of+the+experimental+design.">Enhancing the ability of experimental autoimmune encephalomyelitis to serve as a more rigorous model of multiple sclerosis through refinement of the experimental design.</a> Comp Med. 59 (2), 112-128 (2009).</li><li>Bittner, S., Afzali, A. M., Wiendl, H., Meuth, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myelin+oligodendrocyte+glycoprotein+(MOG35-55)+induced+experimental+autoimmune+encephalomyelitis+(EAE)+in+C57BL/6+mice.">Myelin oligodendrocyte glycoprotein (MOG35-55) induced experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice.</a> J Vis Exp. (86), (2014).</li><li>Beeton, C., Garcia, A., Chandy, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+and+clinical+scoring+of+chronic-relapsing+experimental+autoimmune+encephalomyelitis.">Induction and clinical scoring of chronic-relapsing experimental autoimmune encephalomyelitis.</a> J Vis Exp. (5), e224 (2007).</li><li>Barthelmes, J., 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=Induction+of+Experimental+Autoimmune+Encephalomyelitis+in+Mice+and+Evaluation+of+the+Disease-dependent+Distribution+of+Immune+Cells+in+Various+Tissues.">Induction of Experimental Autoimmune Encephalomyelitis in Mice and Evaluation of the Disease-dependent Distribution of Immune Cells in Various Tissues.</a> J Vis Exp. (111), (2016).</li><li>Shaw, M. K., Zhao, X. Q., Tse, H. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Overcoming+unresponsiveness+in+experimental+autoimmune+encephalomyelitis+(EAE)+resistant+mouse+strains+by+adoptive+transfer+and+antigenic+challenge.">Overcoming unresponsiveness in experimental autoimmune encephalomyelitis (EAE) resistant mouse strains by adoptive transfer and antigenic challenge.</a> J Vis Exp. (62), e3778 (2012).</li><li>Stromnes, I. M., Goverman, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Passive+induction+of+experimental+allergic+encephalomyelitis.">Passive induction of experimental allergic encephalomyelitis.</a> Nat Protoc. 1 (4), 1952-1960 (2006).</li><li>Stromnes, I. M., Goverman, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Active+induction+of+experimental+allergic+encephalomyelitis.">Active induction of experimental allergic encephalomyelitis.</a> Nat Protoc. 1 (4), 1810-1819 (2006).</li><li>Fiander, M. D., Stifani, N., Nichols, M., Akay, T., Robertson, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinematic+gait+parameters+are+highly+sensitive+measures+of+motor+deficits+and+spinal+cord+injury+in+mice+subjected+to+experimental+autoimmune+encephalomyelitis.">Kinematic gait parameters are highly sensitive measures of motor deficits and spinal cord injury in mice subjected to experimental autoimmune encephalomyelitis.</a> Behav Brain Res. 317, 95-108 (2017).</li><li>Jones, M. V., 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=Behavioral+and+pathological+outcomes+in+MOG+35-55+experimental+autoimmune+encephalomyelitis.">Behavioral and pathological outcomes in MOG 35-55 experimental autoimmune encephalomyelitis.</a> J Neuroimmunol. 199 (1-2), 83-93 (2008).</li><li>van den Berg, R., Laman, J. D., van Meurs, M., Hintzen, R. Q., Hoogenraad, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rotarod+motor+performance+and+advanced+spinal+cord+lesion+image+analysis+refine+assessment+of+neurodegeneration+in+experimental+autoimmune+encephalomyelitis.">Rotarod motor performance and advanced spinal cord lesion image analysis refine assessment of neurodegeneration in experimental autoimmune encephalomyelitis.</a> J Neurosci Methods. 262, 66-76 (2016).</li><li>Sasaki, M., Lankford, K. L., Brown, R. J., Ruddle, N. H., Kocsis, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Focal+experimental+autoimmune+encephalomyelitis+in+the+Lewis+rat+induced+by+immunization+with+myelin+oligodendrocyte+glycoprotein+and+intraspinal+injection+of+vascular+endothelial+growth+factor.">Focal experimental autoimmune encephalomyelitis in the Lewis rat induced by immunization with myelin oligodendrocyte glycoprotein and intraspinal injection of vascular endothelial growth factor.</a> Glia. 58 (13), 1523-1531 (2010).</li><li>Merkler, D., Ernsting, T., Kerschensteiner, M., Bruck, W., Stadelmann, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+focal+EAE+model+of+cortical+demyelination:+multiple+sclerosis-like+lesions+with+rapid+resolution+of+inflammation+and+extensive+remyelination.">A new focal EAE model of cortical demyelination: multiple sclerosis-like lesions with rapid resolution of inflammation and extensive remyelination.</a> Brain. 129 (Pt 8), 1972-1983 (2006).</li><li>Franco-Pons, N., Torrente, M., Colomina, M. 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Bij muizen met EAE, twee veelgebruikte methoden voor het meten van motorische tekorten zijn klinische scoren en latentie vallen van een rotarod27,28. Deze technieken hebben verschillende beperkingen. Hoewel handig en gebruikte, wordt klinische scoren beperkt door de opbrengst alleen ordinaal niveau gegevens, wat betekent dat de omvang van de verschillen tussen klinische scores zijn niet bekend. Klinische scoren lijdt ook niet in staat om nauwkeurige informatie ov...

Gait is een serie van repetitieve bewegingen van de ledematen gebruikt om de motoriek. Gait bestaat uit stap cycli, die zijn verdeeld in twee fasen: de fase van de houding, die is wanneer de voet naar achteren op de grond beweegt voor het voortbewegen van het lichaam naar voren; en de fase van de schommel, waar de voet uit de grond en bewegende vooruit is. Gait storingen zijn waarmerk kenmerken van vele neurodegeneratieve aandoeningen, zoals dwarslaesie (SCI), multiple sclerose (MS), Amyotrofische laterale sclerose (ALS), de ziekte van Parkinson (PD) en lijn; preklinische knaagdier modellen van deze aandoeningen recapituleren vaak hun respectieve gait waardeverminderingen1. De fundamentele controlemechanismen van voortbewegen in muizen zijn intensief bestudeerde2,3. Daarnaast zijn er Muismodellen van vele menselijke neurologische aandoeningen4. Ganganalyse in muizen is dan ook een aantrekkelijke aanpak voor het meten van meerdere aspecten van motorische tekorten die anatomische correlaten hebben gekend. De studie van gait in muismodellen kan inzicht verwerven in de neuropathologische grondslagen van motorische tekorten in neurodegeneratieve aandoeningen, en de identificatie van potentiële behandelingen inschakelen.
Sommige technieken die zijn gebruikt voor het meten van gait in knaagdieren omvatten visuele inspectie (bijvoorbeeld, de Basso muis schaal5 en open veld test6) en analyseren van de gang van de ventrale vliegtuig7. Meer recentelijk, methoden om te meten sagittale vlak kinematica van stuk bewegingen hebben opgedaan populariteit omdat ze meer informatie over de uitvoering van de beweging bieden en bijgevolg gevoeliger voor subtiele veranderingen in gang8, zijn 9 , 10 , 11. kinematische technieken ontwikkeld om te bestuderen stuk beweging in het sagittale vlak tijdens het lopen op een loopband9,12 zijn uitvoerig bestudeerd in het kader van SCI ALS, traumatische corticale letsels, beroerte, en De ziekte van Huntington8,9,10,11,13,14,15,16. Deze technieken hebben daarentegen beperkt gebruik in de studie van motorische tekorten voor Muismodellen van multiple sclerose17gezien.
Experimentele autoimmune encefalomyelitis (EAE) is de meest gebruikte muismodel van MS18. De twee belangrijkste methoden van inducerende EAE is via actieve of passieve inoculatie. In actieve EAE, zijn muizen geïmmuniseerd met myeline antigenen, het veroorzaken van autoreactieve T cellen gemedieerde neuroinflammation en demyelinisatie in het ruggemerg en de kleine hersenen. Passieve EAE, aan de andere kant, is veroorzaakt door overdracht van autoreactieve T cellen van een muis met actieve EAE naar een naïeve muis19. Zoals elders beschreven, het verloop van de ziekte en de Neuropathologie worden beïnvloed door het centrale zenuwstelsel (CNS) antigeen en muis stam20,21,22,23,24 ,25. Controle muizen worden EAE experimenten geïnjecteerd met compleet Freund van adjuvans (CFA) zonder de myeline-antigeen. EAE wordt gekenmerkt door de oplopende verlamming die begint met staart zwakte en kan potentieel betrekking hebben op de voorpoten, wat resulteert in ataxie en verlamming20. We hebben onlangs gekenmerkt gait veranderingen in C57Bl/6 muizen onderworpen aan myeline Oligodendrocyt glycoproteïne 35-55 (MOG35-55)-geïnduceerde EAE. Deze studies hebben aangetoond ganganalyse te zijn superieur dan klassieke gedragsanalyse omdat afwijkingen van de normale enkel verkeer zijn sterk gecorreleerd met de mate van verlies van de witte stof in de lumbale ruggemerg van EAE muizen26. De sterkte van de correlatie tussen witte massa verlies en twee andere maatregelen op de traditionele gedrags (klinische scoren en rotarod) was daarentegen veel zwakkere26.
We beschrijven hier het gebruik van kinematische ganganalyse te detecteren verkeer tekorten in het sagittale vlak van EAE muizen lopen op een loopband. Vijf reflecterende markeringen werden geplaatst op een stuk te identificeren van verkeer van de heup-, knie- en enkel gewrichten in high-speed video-opnamen. De software van de analyse van de beweging werd gebruikt om kinematische gegevens over gezamenlijke excursies te extraheren. Het nut van deze technieken te kwantificeren verkeer tekorten voor het model35-55 MOG van EAE worden besproken. Deze technieken zijn ook van toepassing op de studie van gait tekorten in andere Muismodellen van neurodegeneratieve aandoeningen.

<table border="0" cellpadding="0" cellspacing="0" width="893">
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">Camera</td>
			<td style="width:167px;">Nikon</td>
			<td style="width:344px;">Nikon D750</td>
			<td style="width:167px;">Used to film the video</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Reflective tape</td>
			<td>B&#38;L Engineering</td>
			<td>MKR-Tape-2</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Fine scissors</td>
			<td>Fine Science Tools</td>
			<td>15023-10</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Forceps</td>
			<td>Fine Science Tools</td>
			<td>11252-20</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Glue gun</td>
			<td>Craftsmart</td>
			<td>E231647</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">scalpel handle #4</td>
			<td>Roboz</td>
			<td>R5-9884</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Scalpel Blade No.10</td>
			<td>Feather</td>
			<td>2020-12</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">C57BL/6 mice</td>
			<td>Charles River Laboratories</td>
			<td style="width:344px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Anesthetic machine</td>
			<td>EZ Anesthesia</td>
			<td>EZ-AF9000 Auto Flow System</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Recirculating water heating blanket</td>
			<td>Androit</td>
			<td>HTP-1500</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">topical eye lubricant</td>
			<td>Refresh</td>
			<td>DIN00210889</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Shaver</td>
			<td>Oster</td>
			<td>78997-010</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">High speed camera</td>
			<td>Fastec</td>
			<td>Fastec IL3-100</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">High power light</td>
			<td>Smith Victor Corporation</td>
			<td>Model 700 SG (600 Watt quartz light, 120 Volts)</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Light Stand</td>
			<td>Promaster</td>
			<td>LS1</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Treadmill</td>
			<td colspan="2">Custom built at the Zoological Institute, University of Cologne</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Microsoft Excel 2016</td>
			<td>Microsoft</td>
			<td>Version 2016</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">KinemaJ</td>
			<td>Nicolas Stifani</td>
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sagittal plane kinematic gait analysis in c57bl/6 mice subjected to mog35-55 induced experimental autoimmune encephalomyelitis

Kinematische ganganalyse in het sagittale vlak is vaak gebruikt voor het karakteriseren van motorische tekorten bij multiple sclerose (MS). We beschrijven de toepassing van deze technieken te identificeren gait tekorten in een muismodel van MS, bekend als experimentele autoimmune encefalomyelitis (EAE). Verlamming en motor tekorten in muizen onderworpen aan EAE meestal geschat aan de hand van een klinische scoren schaal. Deze schaal geeft echter alleen ordinale dat de gegevens weinig informatie over de precieze aard van de motorische tekorten verschaft. EAE ziekte ernst is ook beoordeeld door rotarod prestaties, waarmee een maatregel van algemene motorische coördinatie. Kinematische ganganalyse van de hind-limb in het sagittale vlak genereert daarentegen zeer nauwkeurige informatie over hoe beweging geschaad is. Voor het uitvoeren van deze procedure, zijn reflecterende markeringen geplaatst op een hind-limb gezamenlijke beweging detecteren terwijl een muis is lopen op een loopband. De software van de analyse van de beweging wordt gebruikt voor het meten van de beweging van de markers tijdens het wandelen. Kinematische gait parameters zijn vervolgens afgeleid van de resulterende gegevens. Laten we zien hoe deze gait parameters te kwantificeren verminderde bewegingen van de gewrichten van het heup-, knie- en enkel in EAE kunnen worden gebruikt. Deze technieken kunnen worden gebruikt om beter te begrijpen van de ziekte mechanismen en identificeren van potentiële behandelingen voor MS en andere neurodegeneratieve aandoeningen die afbreuk doen aan de mobiliteit.

Figuur 1 is een schematische voorstelling van de gebruikte voor kinematische ganganalyse procedure. Eerste, reflecterende markeringen zijn gemaakt en geplaatst op een muis op 5 anatomische punten. Gait is vervolgens opgenomen, terwijl de muis is lopen op een loopband. De software van de analyse van de beweging wordt gebruikt voor het uitpakken kinematische gegevens voor een latere anayse.
Figuur 2A -C vertegenwoordige...

Watch this Scientific Journal Video about Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis at JoVE.com

Sagittal Plane Kinematic Gait Analysis in C57BL/6 Mice Subjected to MOG35-55 Induced Experimental Autoimmune Encephalomyelitis

Sagittale vlak kinematische ganganalyse in C57BL/6 muizen onderworpen aan MOG35-55 geïnduceerde experimentele Autoimmune encefalomyelitis

Kinematische ganganalyse in het sagittale vlak levert zeer nauwkeurige informatie over hoe verkeer wordt uitgevoerd. We beschrijven de toepassing van deze technieken te identificeren gait tekorten voor muizen onderworpen aan auto-immuun-gemedieerde demyelinisatie. Deze methoden kunnen ook worden gebruikt om te karakteriseren gait tekorten voor andere Muismodellen met verminderde motoriek.

Kinematic gait analysis in the sagittal plane has frequently been used to characterize motor deficits in multiple sclerosis (MS). We describe the ...

The overall goal of this procedure is to measure functional motor impairments in the experimental autoimmune encephalomyelitis or EAE, mouse model multiple sclerosis using kinematic gait analysis. The application of kinematic gait analysis to mouse-walking behavior has been previously established and described by others. Neurological deficits in an EAE result from neural inflammation and sporadic white matter loss throughout the spinal cord and the cerebellum.Traditionally, motor stability in EAE mice has been assessed using clinical scoring systems, in which mice are assigned a clinical score based on the observer's impression of the severity of motor deficits. Data from clinical scores are ordinal and do not correlate well with the spinal cord histopathology. Kinematic gait analysis has recently been shown to be a better behavioral correlate of white matter loss than clinical scores, in addition to providing an objective description of walking deficits in the EAE mice.This technique involves placing reflective markers on the high limbs of mice and allowing them to walk on a treadmill while being recorded with a high-speed camera. Kinematic parameters are then extracted from the video using motion analysis software. The first step is to make the markers that will be placed on the animal's hind leg.The reflection of light off of these markers allows the coordinates of anatomical points on the leg to be extracted from the videos. To begin, punch the desired number of circles from the sheet of reflective paper. Each mouse requires five markers per recording, two large and three small.Using fine scissors, make a cut extending from the perimeter to the center of the circle. Peel off the paper backing to reveal the adhesive surface using fine forceps. Grip the marker with forceps and using your finger, roll the marker in on itself to form a cone.To make a small marker, make a longer cut and curl the cone more tightly. To make a large marker, make a shorter cut and curl the cone more loosely. Using a hot glue gun, fill the inside of the cone with glue and adhere it to a piece of cardboard.It is important to fully fill the marker with glue to prevent the marker from collapsing when handling during recording. Once the glue is dry, cut the marker off using a scalpel. Ensure that you cut away from your body.The next step is to prepare the animal for recording. To do this, the reflective markers must be adhered to the hind limb of the mouse at the proper anatomical locations. This is conducted under light anesthesia.Place the mouse in an induction chamber and anesthetize with 2.5%isoflurane. Once unconscious, transfer the mouse from the induction chamber to a nose cone or a recirculating water heating pad. Apply a topical lubricant to both eyes.Shave the desired hind limb extending from the ankle to the spine and the bottom of the ribs. In this demonstration, we are recoding movements of the right hind limb, but either or both limbs may be used. Locate the iliac crest by bringing both knees together with your thumb and forefinger and palpating just below the ribs.Mark this location using a permanent marker. Locate the hip by extending the leg and moving it back and forth. Place a marker over the hip joint, which is the point of articulation between the head of the femur and the pelvis.Using a flexible ruler, measure and record the length of the tibia or shank of the mouse. Measure and record the length of the femur or thigh as well. To isolate the fourth toe for marker placement, tape down the rest of the foot.Grasp a small marker with forceps and dip the flat end in glue. Place the small marker on the tip of the fourth toe. Place another small marker on the metatarsophalangeal joint.Place the last small marker on the ankle. Place a large marker on the hip joint directly over the mark on the skin. Place the second large marker on the mark over the iliac crest.Remove the tape from the foot. Place the mouse in a recovery cage and transport immediately to the gait recording room. The next step is recording the mouse walking on a treadmill.This picture is of our treadmill recording setup, showing the light, high-speed camera, and treadmill. Prior to recording the mouse's gait, take a picture of a calibration block with known dimensions on the treadmill. This will allow the pixels in the video to be converted to real measurements.It is critical that the camera angle and position at which the calibration picture is taken remains the same during the recording of walking behavior. Place the mouse in the treadmill. Increase the treadmill's speed gradually to orient the mouse in the correct direction.Slowly accelerate to 20 centimeters per second, which is the ideal running speed for obtaining recordings of steady gait. For accurate analysis, it's best to record eight to 12 step cycles of steady walking behavior. This is an example of consistent walking.The following is the same video shown at half speed. Because mice walk so quickly, it can be useful to view the videos at slower speeds to count step cycles and to better appreciate the gait pattern. This is one example of an EAE mouse walking on the treadmill.This mouse is unable to support its body weight as its pelvis is very low to the ground. Additionally, it is having difficulty lifting its foot off the ground during the swing phase. This is an example of another EAE mouse that has less severe motor deficits.The EAE mouse is walking on its toes with reduced movement at the ankle joint. This gives the mouse an uneven gait. The following are examples of behaviors that will compound analysis.Recordings, including these behaviors, should not be used. Lagging occurs when the mouse stops walking and travels to the back of the treadmill, but then resumes walking. This may occur in any mouse, but will occur more frequently at lower speeds.Rearing occurs when the mouse shifts its weight on its back hind limbs and raises its head and upper body. This behavior is common in anxious mice. The following are examples of poor lighting that will lead to issues with analysis of the markers'position in time and space.If the lighting is too dim, markers may not reflect enough light to be recognized by the computer software. In this video, the toe markers are not easily visible due to insufficient lighting. If the lighting is too bright, objects other than the markers may reflect too much light and be recognized as a marker.In this video, the toe markers appear to be merging with the reflective parts of the treadmill. Data from the reflective markers on mice walking can be used to create stick models of the leg from which kinematic parameters can be extracted. This is a recording of a mouse walking with a stick model of the leg superimposed.Note that in this video, there is a sixth marker on the knee, which is not necessary because the location of the knee can be triangulated from the positions of the hip and ankle joints, and the measured lengths of the femur and tibia. Because markers over the knee joint are often inaccurate due to skin slippage, triangulation is the preferred method. A mouse's step cycle can be divided into two main phases.The stance phase and the swing phase. This stick diagram can be used to qualitatively illustrate the movement of the hind limb in time and space. An example of this could be to assess the movement of the hind limb at different times points throughout the study.This example shows that the hind leg is compressed during the stance phase, indicating that the mouse has difficulty supporting its body weight. This is reflected by increased flexion of the knee and ankle joints. At the later time point, this has partially recovered.This video illustrates the relationship between the walking behavior and joint angles over time. The hip, knee, and ankle angle waveforms can be extracted from each recording. The joint angles can then be averaged over eight to 12 consecutive step cycles, resulting in an average step cycle that can be used for further analysis.This graph represents a knee joint waveform averaging from 10 consecutive step cycles. The data has been normalized, so that the length of the stance and swing phases are 100 frames respectively. The clear background represents the swing phase and the green background represents the stance phase.For this mouse, the walking behavior from week to week was very consistent and the step-cycle waveforms from each week overlapped considerably. However, there can also be substantial variability in gait in healthy mice as seen in this graph. The degree of variability shown here is acceptable and within the range of what might be expected from a mouse.This graph represents the knee step cycle from the EAE mouse recorded on three consecutive weeks. There is a slight change in the shape of the step cycle at week two and a substantial deviation by week three, which the mouse's knee is much more flexed and does not extend during walking. There are numerous parameters that can be measured using this technique.We'll briefly describe three. The average angle is obtained by averaging all angles throughout the normalized step cycle. In this instance, the average angle decreases throughout the study, suggesting that the mice are not extending their knees as much as normal.The range of motion is obtained by subtracting the smallest angle from the largest angle in the normalized step cycle. This parameter can give you insight into joint flexibility, rigidity, or weakness. In this example, the knee range of motion decreases throughout the study indicating that the mice are not able to move their knee normally, possibly due to muscle weakness.Root-Mean-Square difference is a method used to measure deviation of step-cycle waveforms from the baseline recording. This parameter tells you how much deviation there is from the initial recording. Kinematic gait analysis is a valuable technique that can be used to sensitively detect and describe changes in gait.The application of kinematic gait analysis to EAE studies may be a valuable tool in understanding the functional consequences of spinal cord pathology in this model. It may facilitate discovery of novel treatments for multiple sclerosis. Additionally, kinematic gait analysis is not limited to the context of EAE.This technique has previously been used in mouse models with spinal cord injury, amyotrophic lateral sclerosis, Huntington's disease and stroke, and can be applied to other mouse models with neurological disorders, including Parkinson's disease.

Research

JoVE Journal

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