Name: subcutaneous neurotrophin 4 infusion using osmotic pumps or direct muscular injection enhances aging rat laryngeal muscles
Uploaded: 2017-06-13T15:00:00
Description: Watch this Scientific Journal Video about Subcutaneous Neurotrophin 4 Infusion Using Osmotic Pumps or Direct Muscular Injection Enhances Aging Rat Laryngeal Muscles at JoVE.com

Dit werk werd ondersteund door subsidies van het Nationaal Instituut voor Doofheid en Andere Communicatie Stoornissen (R21DC010806 naar CAM en JCS en R01DC011285 naar CAM).

Gebruik mannelijke Fischer 344-Brown Noorse ratten op 6 en 30 maanden oud voor dit protocol. Ratten werden verkregen uit het National Institute of Aging knaagdierkolonie. Wij hebben ratten gebruikt voor deze studie omdat de structuur van de larynx van de rat gelijkwaardig is aan die van de mens, functioneel voor de bescherming van de luchtwegen en soortenspecifieke vocalisaties. Deze studie is uitgevoerd in overeenstemming met het PHS-beleid inzake humane zorg en gebruik van laboratoriumdieren, De NIH Gids voor de zorg ...

<ol>
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Larynxale spieren zijn kwetsbaar voor de ongunstige effecten van veroudering. Vorige studies hebben aangetoond veranderingen in veroudering larynxale spieren die veranderingen in vezelgrootte, totaal aantal vezels, regeneratief vermogen, NMJ grootte en hoeveelheid veranderingen omvatten, naast variaties in contractiele functie en myosine isoform verschuivingen 4 , 11 , 27 , 30 , <su...

Larynxale spieren trekken snel en consistent samen en zijn kwetsbaar voor de nadelige effecten van veroudering. Deze constante activiteit wordt geacht bij te dragen aan spraakproblemen of dysfagie waargenomen bij personen ouder dan 65 jaar 1 , 2 , 3 , 4 , 5 , 6 , 7 . Verscheidene moleculaire en pathofysiologische mechanismen dragen bij aan deze leeftijdgerelateerde disfunctie. Deze mechanismen kunnen remodellering van larynxmucosa, spiervezelatrofie of -verlies omvatten, gebrek aan regeneratie of atrofie van spiervezels, waardoor de vocale vouwen worden gebogen en de glottisluiting 8 , 9 , 10 , 11 niet mogelijk is . Er is geen bewezen medische therapie op dit moment dat kan comDeze leeftijdsgebonden veranderingen in deze spieren verbeteren of voorkomen. Modulatie van de effectiviteit van neuromusculaire transmissie kan de neuromotorische prestaties sterk beïnvloeden. De familie van neurotrophins omvat zenuwgroeifactor (NGF), hersenverwante zenuwgroeifactor (BDNF), neurotrophine 3 (NTF3) en NTF4 12 , 13 . Neurotrofinen hebben aangetoond dat ze synaptische werkzaamheid 1 , 4 moduleren. Hepatocytgroeifactor, transformerende groeifactor bèta en fibroblastgroeifactor, is onlangs bij mensen gebruikt voor de behandeling van vocal fold scarring 15-17. NTF4 regelt ook de effectiviteit van de NMJ; Muizen die NTF4 ontbreken tonen gedemonteerde NMJ&#39;s 11 , 18 , 19 . Deze studies leiden tot veelbelovende effecten van behandelaarsT van veroudering van larynxale spierstoornissen en denervering met groeifactoren. Directe injectie therapeutica aan de weefsels van de vocale vouwen zijn niet bij elkaar in de mens. Bijvoorbeeld worden lokale injecties van botulinum toxine momenteel gebruikt als een effectieve behandeling voor neurologische bewegingsstoornissen die de spieren in de larinks beïnvloeden, zoals spasmodische dysfonie en bilaterale herhalende larynxale zenuwverlamming 20 , 21 . Hyaluronzuurhydrogel is een ander injecteerbaar, dat gebruikt wordt om vocale vouwverschrikking en glottale insufficiëntie 22 , 23 te behandelen. Injectie laryngoplastie kan worden gebruikt om een ​​verscheidenheid aan communicatie stoornissen te behandelen 24 . Deze directe injectiemethoden houden belofte om de vocale functie te verbeteren en te slikken in verouderende populaties.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Neurotrophin 4</td>
			<td>Pepro Tech</td>
			<td>450-04</td>
			<td>200ng in 50&#956;l</td>
		</tr>
		<tr>
			<td>Alzet Osmotic Pump</td>
			<td>DURECT Corporation</td>
			<td>2001D</td>
			<td></td>
		</tr>
		<tr>
			<td>30&#176; endoscope</td>
			<td>Stoltz</td>
			<td>61029D</td>
			<td></td>
		</tr>
		<tr>
			<td>50mm 30 gauge 100-&#956;l syringe</td>
			<td>Hamilton</td>
			<td>84850 and 201812</td>
			<td></td>
		</tr>
		<tr>
			<td>saline (sodium chloride solution)</td>
			<td>Sigma-Aldrich</td>
			<td>S8776</td>
			<td></td>
		</tr>
		<tr>
			<td>ketamine hydrochloride</td>
			<td>Henry Schein</td>
			<td>56344</td>
			<td></td>
		</tr>
		<tr>
			<td>xylazine hydrochloride</td>
			<td>Henry Schein</td>
			<td>33198</td>
			<td></td>
		</tr>
		<tr>
			<td>25 G 5/8 needle</td>
			<td>Becton-Dickinson</td>
			<td>305901</td>
			<td></td>
		</tr>
		<tr>
			<td>1 ml syringe</td>
			<td>Becton-Dickinson</td>
			<td>309659</td>
			<td></td>
		</tr>
		<tr>
			<td>ophthalmic ointment</td>
			<td>Henry Schein</td>
			<td>8897</td>
			<td></td>
		</tr>
		<tr>
			<td>clippers</td>
			<td>Oster</td>
			<td>44-018</td>
			<td></td>
		</tr>
		<tr>
			<td>ethanol</td>
			<td>Decon</td>
			<td>2716</td>
			<td></td>
		</tr>
		<tr>
			<td>iodine (Betadine)</td>
			<td>Purdue Pharma L.P.</td>
			<td>606404</td>
			<td></td>
		</tr>
		<tr>
			<td>heating pad</td>
			<td>Sunbeam</td>
			<td>731-5</td>
			<td></td>
		</tr>
		<tr>
			<td>5-0 nylon suture thread</td>
			<td>AD Surgical</td>
			<td>PMN-518R6</td>
			<td></td>
		</tr>
		<tr>
			<td>crile hemostat</td>
			<td>Fine Science Tools</td>
			<td>13005-14</td>
			<td></td>
		</tr>
		<tr>
			<td>delicate suture tying forceps</td>
			<td>Fine Science Tools</td>
			<td>11063-07</td>
			<td></td>
		</tr>
		<tr>
			<td>meloxicam</td>
			<td>Henry Schein</td>
			<td>49756</td>
			<td></td>
		</tr>
		<tr>
			<td>carprofen</td>
			<td>Merritt Veterinary Supplies</td>
			<td>148700</td>
			<td></td>
		</tr>
		<tr>
			<td>antibiotic ointment</td>
			<td>Henry Schein</td>
			<td>57110</td>
			<td></td>
		</tr>
		<tr>
			<td>acepromizine Aceproject</td>
			<td>Henry Schein</td>
			<td>3845</td>
			<td></td>
		</tr>
		<tr>
			<td>isoflurane Isothesia</td>
			<td>Henry Schein</td>
			<td>50033</td>
			<td></td>
		</tr>
		<tr>
			<td>induction box (anesthetizing box)</td>
			<td>Harvard Apparatus</td>
			<td>50-0116</td>
			<td></td>
		</tr>
		<tr>
			<td>oxygen compressed tank</td>
			<td>Scott Gross</td>
			<td>UN1072</td>
			<td></td>
		</tr>
		<tr>
			<td>plexiglas platform</td>
			<td>Small Parts Inc (Amazon)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>rubber tipped forceps</td>
			<td>Fine science tools rubber</td>
			<td>11075-00</td>
			<td></td>
		</tr>
		<tr>
			<td>liquid rubber for forceps above</td>
			<td>Lowe&#39;s</td>
			<td>42518</td>
			<td></td>
		</tr>
		<tr>
			<td>plastic spectula (BD syringe cut to length)</td>
			<td>Becton-Dickinson</td>
			<td>309659</td>
			<td></td>
		</tr>
		<tr>
			<td>halogen light source rhino-laryngeal stroboscope</td>
			<td>Kay-Pentax</td>
			<td>RLS 9100 B</td>
			<td></td>
		</tr>
		<tr>
			<td>video recorder</td>
			<td>Kay-Pentax</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>sucrose</td>
			<td>Sigma-Aldrich</td>
			<td>S0389-500G</td>
			<td></td>
		</tr>
		<tr>
			<td>phosphate buffered saline</td>
			<td>Sigma-Aldrich</td>
			<td>P4417-100TAB</td>
			<td></td>
		</tr>
		<tr>
			<td>cryostat Mictotom HM525</td>
			<td>Thermo Scientific</td>
			<td>HM 525</td>
			<td></td>
		</tr>
		<tr>
			<td>Gill 1 hematoxylin</td>
			<td>VWR</td>
			<td>10143-142</td>
			<td></td>
		</tr>
		<tr>
			<td>Shandon eosin-Y alcoholic</td>
			<td>Thermo Fisher Scientific</td>
			<td>6766007</td>
			<td></td>
		</tr>
		<tr>
			<td>anti-sodium channel Nav1.5 antibody produced in rabbit</td>
			<td>Sigma-Aldrich</td>
			<td>S0819</td>
			<td></td>
		</tr>
		<tr>
			<td>Texas red-X phalloidin</td>
			<td>Sigma-Aldrich</td>
			<td>T7471</td>
			<td></td>
		</tr>
		<tr>
			<td>alpha- bungarotoxin alexa fluor 488 conjugate</td>
			<td>Thermo Fisher Scientific</td>
			<td>B-13422</td>
			<td></td>
		</tr>
		<tr>
			<td>Small animal anaesthesia machine</td>
			<td>Smiths Medical</td>
			<td>CDS 9000</td>
			<td></td>
		</tr>
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subcutaneous neurotrophin 4 infusion using osmotic pumps or direct muscular injection enhances aging rat laryngeal muscles

Larynxale dysfunctie bij ouderen is een belangrijke oorzaak van handicap, van stoornissen aan dysfagie en verlies van luchtweg beschermende reflexen. Er zijn weinig, indien aanwezig, therapieën die gericht zijn op leeftijdgerelateerde larynxale spierdisfunctie. Neurotrofinen zijn betrokken bij spierinervation en differentiatie van neuromusculaire kruispunten (NMJ&#39;s). Het is van mening dat neurotrofinen neuromusculaire transmissie versterken door de vrijlating van neurotransmitteren te verhogen. De neuromusculaire kruispunten (NMJ&#39;s) worden kleiner en minder overvloedig in verouderende larynxale spieren van de rat, met bewijs van functionele denervation. We hebben de effecten van NTF4 onderzocht voor toekomstig klinisch gebruik als therapeutisch voor het verbeteren van de functie bij het verouderen van menselijke larynxspieren. Hier geven we het gedetailleerde protocol voor systemische toepassing en directe injectie van NTF4 om het vermogen van veroudering van de rat larynxale spier te onderzoeken in verband met NTF4 applicatie. Bij deze methode kregen ratten ofwel systematisch via o NTF4Smotische pomp of door directe injectie door de vocale plooien. Larynxale spieren werden vervolgens gedissecteerd en gebruikt voor histologisch onderzoek naar morfologie en leeftijdverwante denervation.

De ratten werden geëuthaniseerd na 2 weken osmotische pompinfectie of 1 week na directe injectie van NTF4. Larynges werden geoogst, geplaatst in cryoprotectant (30% sucrose en 70% fosfaat gebufferde zoutoplossing) en vervolgens serieel gesneden in 10 μm breedten met een cryostat. Veroudering van larynxale spieren wordt beïnvloed door toediening van NTF4 25 . Naast de jonge en oude rat, vergeleken we de geïnjecteerde en niet-geïnjecteerde kant van de thyroaryt...

Watch this Scientific Journal Video about Subcutaneous Neurotrophin 4 Infusion Using Osmotic Pumps or Direct Muscular Injection Enhances Aging Rat Laryngeal Muscles at JoVE.com

Subcutaneous Neurotrophin 4 Infusion Using Osmotic Pumps or Direct Muscular Injection Enhances Aging Rat Laryngeal Muscles

Hier presenteren wij een protocol om het gebruik van neurotrophin 4 (NTF4) systematisch te beschrijven en rechtstreeks te remodellen larynxale spierveroudering.

Subcutane Neurotrofine 4 Infusie Met Osmotische Pompen Of Directe Spierinjectie Verbetert Veroudering Rat Larynx Spieren

Laryngeal dysfunction in the elderly is a major cause of disability, from voice disorders to dysphagia and loss of airway protective reflexes. Few, if ...

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