Name: assessing functional metrics of skeletal muscle health in human skeletal muscle microtissues
Uploaded: 2021-02-18T15:00:00
Description: Watch this Scientific Journal Video about Assessing Functional Metrics of Skeletal Muscle Health in Human Skeletal Muscle Microtissues at JoVE.com

Vi vil gerne takke Mohammad Afshar, Haben Abraha, Mohsen Afshar-Bakooshli og Sadegh Davoudi for at have bidraget til opfindelsen af myoTACTIC-kulturplatformen og for at etablere de fremstillings- og analysemetoder, der er beskrevet heri. HL modtaget støtte fra en Naturvidenskab og Engineering Research Council (NSERC) Training Program i Organ-on-a-Chip Engineering and Entrepreneurship Scholarship og en University of Toronto Wildcat graduate stipendier. PMG er Canada Research Chair i Endogenous Repair og modtog støtte til denne undersøgelse fra Ontario Institute for Regenerativ Medicin, Stem Cell Network, og fra Medicine by Design, en Canada First Research Excellence Program. Skemadiagrammer blev oprettet med BioRender.com.

1. PDMS MyoTACTIC plade fabrikation
BEMÆRK: PDMS MyoTACTIC plade fabrikation kræver en PU negativ skimmel, som kan fremstilles som tidligere beskrevet30. Den computerstøttede design (CAD) SolidWorks-fil til MyoTACTIC-pladedesignet er blevet gjort tilgængeligt på GitHub (https://github.com/gilbertlabcode/MyoTACTIC-SolidWork-CAD-file).
<ol><li>Forbered ~ 110 g PDMS polymeropløsning i en engangsplastkop ved et 1:15-forhold mellem monomer og hærdningsmiddel ved hj?...

<ol>
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H., 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=3D+bioprinted+human+skeletal+muscle+constructs+for+muscle+function+restoration.">3D bioprinted human skeletal muscle constructs for muscle function restoration.</a> Scientific Reports. 8 (1), 12307 (2018).</li><li>Takahashi, H., Shimizu, T., Okano, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engineered+human+contractile+myofiber+sheets+as+a+platform+for+studies+of+skeletal+muscle+physiology.">Engineered human contractile myofiber sheets as a platform for studies of skeletal muscle physiology.</a> Scientific Reports. 8 (1), 1-11 (2018).</li><li>Afshar Bakooshli, 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=A+3D+culture+model+of+innervated+human+skeletal+muscle+enables+studies+of+the+adult+neuromuscular+junction.">A 3D culture model of innervated human skeletal muscle enables studies of the adult neuromuscular junction.</a> eLife. 8, 1-29 (2019).</li><li>Madden, L., Juhas, M., Kraus, W. 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Dette manuskript beskriver metoder til at fremstille og analysere en 3D hMMT kulturmodel, der kan anvendes til undersøgelser af grundlæggende muskelbiologi, sygdom modellering, eller for kandidat molekyle test. MyoTACTIC-platformen er omkostningsvenlig, nem at fremstille og kræver et relativt lille antal celler til at producere skeletmuskulaturmikrotiss. hMMTs dannet inden for MyoTACTIC kulturplatformen består af justerede, flerkernede og striated myotubes og reagerer på elektriske stimuli ved at indlede calciumtran...

Skeletmuskulatur er en af de mest rigelige væv i den menneskelige krop og understøtter centrale kropsfunktioner såsom bevægelse, varme homøostase og stofskifte1. Historisk set er dyremodeller og todimensionelle (2D) cellekultursystemer blevet brugt til at studere biologiske processer og sygdomspatogenese samt til test af farmakologiske forbindelser til behandling af skeletmuskulatursygdomme2,3. Mens dyremodeller i høj grad har forbedret vores viden om skeletmuskulatur i sundhed og sygdom, er deres translationelle virkning blevet hæmmet af høje omkostninger, etiske overvejelser og interspecies forskelle2,4. Ved at henvende sig til menneskelige cellebaserede systemer til at studere skeletmuskulatur er 2D-cellekultursystemer gunstige på grund af deres enkelhed. Der er dog en begrænsning. Dette format undlader ofte at generobre de cellecelle- og celle-ekstracellulære matrixinteraktioner, der forekommer naturligt i kroppen5,6. I løbet af de sidste mange år har tredimensionelle (3D) skeletmuskulaturmodeller vist sig som et stærkt alternativ til hele dyremodeller og konventionelle 2D-kultursystemer ved at tillade modellering af fysiologisk og patologisk relevante processer ex vivo7,8. Faktisk har en overflod af undersøgelser rapporteret strategier til at modellere menneskelige skeletmuskulatur i et bioartificialt 3D-kulturformat1. En begrænsning for mange af disse undersøgelser er, at aktiv kraft kvantificeres efter fjernelse af muskelvæv fra kulturplatformene og tilknytningen til en krafttransducer, som er destruktiv og dermed begrænset til at fungere som en endpoint assay9,10,11,12,13,14,15,16,17,18 ,19,20,21. Andre har designet kultursystemer, der giver mulighed for ikke-invasive metoder til måling af aktiv kraft, men ikke alle er modtagelige for molekyletestapplikationer med højt indhold7,8,9,10,14,18,22,23,24,25,26,27,28 ,29.
Denne protokol beskriver en detaljeret metode til fremstilling af humane muskelmikrotiss (hMMTs) i skeletmuskulaturen (Myo) microTissue Array deviCe To Investigate forCe (MyoTACTIC) platform; en 96 brønd plade enhed, der understøtter bulk produktion af 3D skeletmuskulatur microtissues30. MyoTACTIC pladefremstillingsmetoden muliggør generering af en 96 brøndpolylsiloxan (PDMS) kulturplade og alle tilsvarende brøndfunktioner i et enkelt støbetrin, hvorved hver brønd kræver et relativt lille antal celler til mikrotissuedannelse. Microtissues dannet i MyoTACTIC indeholder justeret, striated, og multinukleated myotubes, der er reproducerbare fra godt til brønd af enheden, og ved modning, kan reagere på kemiske og elektriske stimuli in situ30. Heri, teknikken til at fremstille en PDMS MyoTACTIC kultur plade enhed fra en polyurethan (PU) replika, en optimeret metode til at gennemføre udødeliggjorte menneskelige myogene stamfader celler til at fremstille hMMTs, og den funktionelle vurdering af manipuleret hMMT kraft generation og calcium håndtering egenskaber er skitseret og diskuteret.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.9% Saline Solution, Sterile</td>
			<td>House Brand</td>
			<td>1010</td>
			<td>10 mL aliquots of the solution are made and stored at 4&#176;C</td>
		</tr>
		<tr>
			<td>25G Needle</td>
			<td>BD, Medstore, University of Toronto</td>
			<td>2548-CABD305127</td>
			<td></td>
		</tr>
		<tr>
			<td>6-Aminocaproic Acid, &#8805;99% (titration), Powder</td>
			<td>Sigma - Aldrich</td>
			<td>A2504-100G</td>
			<td>A 50 mg / mL stock solution is generated by dissolving 5 mg of 6-aminocaproic acid powder in 100 mL of autoclaved, distilled water. The solution is vaccum filtered and 10 mL aliquots are stored at 4&#176;C</td>
		</tr>
		<tr>
			<td>6.35 mm ID Tubing</td>
			<td>VWR</td>
			<td>60985-528</td>
			<td></td>
		</tr>
		<tr>
			<td>AB1167 Myoblast Cell Line</td>
			<td>Institut de Myologie (Paris, France)</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Arbitrary Waveform Generator</td>
			<td>Rigol</td>
			<td>DG1022Z</td>
			<td></td>
		</tr>
		<tr>
			<td>Basement Membrane Extract (Geltrex)</td>
			<td>Thermo Fisher Scientific</td>
			<td>A14132-02</td>
			<td>Stored as aliquots of 50 &#181;L or 100 &#181;L at -80&#176;C</td>
		</tr>
		<tr>
			<td>Benchtop Vacuum Chamber</td>
			<td>Sigma - Aldrich</td>
			<td>D2672</td>
			<td></td>
		</tr>
		<tr>
			<td>BNC to Aligator Clip Cable</td>
			<td></td>
			<td></td>
			<td>Ordered from Amazon</td>
		</tr>
		<tr>
			<td>Culture Plastics</td>
			<td>Sarstedt</td>
			<td></td>
			<td>Includes culture plates, serological pipettes, etc</td>
		</tr>
		<tr>
			<td>Dimethyl Sulfoxide</td>
			<td>Sigma - Aldrich</td>
			<td>D8418-250ML</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS, Powder, No Calcium, No Magnesium</td>
			<td>Thermo Fisher Scientific</td>
			<td>21600069</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s Modified Eagle Medium (DMEM) (1X)</td>
			<td>Gibco</td>
			<td>11995-065</td>
			<td>This is a high glucose DMEM with L-glutamine and sodium pyruvate</td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum</td>
			<td>Fisher Scientific</td>
			<td>10437028</td>
			<td></td>
		</tr>
		<tr>
			<td>Fibrinogen from Bovine Plasma</td>
			<td>Sigma - Aldrich</td>
			<td>F8630-5G</td>
			<td>Aliquots ranging from 7 - 10 mg of fibrinogen powder are made and stored at -20&#176;C</td>
		</tr>
		<tr>
			<td>Filtropur Syringe Filter, 0.22um Pore Size</td>
			<td>Sarstedt</td>
			<td>83.1826.001</td>
			<td></td>
		</tr>
		<tr>
			<td>Horse Serum</td>
			<td>Gibco</td>
			<td>16050-122</td>
			<td></td>
		</tr>
		<tr>
			<td>Human Recombinant Insulin</td>
			<td>Sigma - Aldrich</td>
			<td>91077C</td>
			<td>Stock solution is 100X and made by dissolving 1 mg of human recombinant insulin in 1 mL of DMEM and 1 &#181;L of NaOH 10N. Solution is filtered and stored as 1 mL aliquots at 4&#176;C</td>
		</tr>
		<tr>
			<td>Image Acquisition Software</td>
			<td>Olympus</td>
			<td>cellSens Dimension</td>
			<td></td>
		</tr>
		<tr>
			<td>Image Processing Software</td>
			<td>National Institutes of Health</td>
			<td>ImageJ</td>
			<td></td>
		</tr>
		<tr>
			<td>Isotemp Oven</td>
			<td>Thermo Fisher Scientific</td>
			<td>201</td>
			<td></td>
		</tr>
		<tr>
			<td>Microscope</td>
			<td>Olympus</td>
			<td>IX83</td>
			<td></td>
		</tr>
		<tr>
			<td>Microscope - Camera Mount</td>
			<td>Labcam</td>
			<td>Labcam for iPhone</td>
			<td>Ordered from Amazon</td>
		</tr>
		<tr>
			<td>Penicillin-Streptomycin (10,000 U/mL)</td>
			<td>Gibco</td>
			<td>15140-122</td>
			<td></td>
		</tr>
		<tr>
			<td>Plastic Disposable Syringes, 1cc</td>
			<td>BD</td>
			<td>2606-309659</td>
			<td></td>
		</tr>
		<tr>
			<td>Plastic Disposable Syringes, 50cc</td>
			<td>BD</td>
			<td>2612-309653</td>
			<td></td>
		</tr>
		<tr>
			<td>Pluronic F-127, Powder, BioReagent</td>
			<td>Sigma - Aldrich</td>
			<td>P2443-250G</td>
			<td>A 5% stock solution of pluronic acid is made by dissolving 5 g of pluronic acid powder in 100 mL of chilled, autoclaved, distilled water. The solution is vaccum filtered and 10 mL aliquots are stored at 4&#176;C</td>
		</tr>
		<tr>
			<td>Polydimethylsiloxane (Sylgard 184 Silicone Elastomer Kit)</td>
			<td>Dow</td>
			<td>4019862</td>
			<td>Kits are also available at Thermo Fisher Scientific, Sigma - Aldrich, etc.</td>
		</tr>
		<tr>
			<td>Polyurethane Negative Mold</td>
			<td>In House</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Release Agent</td>
			<td>Mann Release Technologies</td>
			<td>200</td>
			<td></td>
		</tr>
		<tr>
			<td>Rotary Vane Vacuum Pump</td>
			<td>Edwards</td>
			<td>A65401906</td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel</td>
			<td>Almedic, Medstore, University of Toronto</td>
			<td>2586-M36-0100</td>
			<td></td>
		</tr>
		<tr>
			<td>Single Edge Razor Blade</td>
			<td>VWR</td>
			<td>55411-050</td>
			<td></td>
		</tr>
		<tr>
			<td>Skeletal Muscle Cell Basal Medium</td>
			<td>Promocell</td>
			<td>C-23260</td>
			<td>30 mL aliquotes are generated and at stored at 4&#176;C.</td>
		</tr>
		<tr>
			<td>Skeletal Muscle Cell Growth Medium (Ready-to-use)</td>
			<td>Promocell</td>
			<td>C-23060</td>
			<td>42 mL aliquots are generated and stored at 4&#176;C.</td>
		</tr>
		<tr>
			<td>Smartphone (iPhone)</td>
			<td>Apple</td>
			<td>SE</td>
			<td></td>
		</tr>
		<tr>
			<td>Standard Duty Dry Vacuum Pump</td>
			<td>Welch</td>
			<td>2546B-01</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterilization Bag</td>
			<td>Alliance</td>
			<td>211-SCM2</td>
			<td></td>
		</tr>
		<tr>
			<td>Thimble</td>
			<td>Igege</td>
			<td></td>
			<td>Ordered from Amazon</td>
		</tr>
		<tr>
			<td>Thrombin from human plasma</td>
			<td>Sigma - Aldrich</td>
			<td>T6884-250UN</td>
			<td>100 units of thrombin is dissolved in 1 mL of a 0.1% BSA solution. 10 &#181;L aliquots are prepared and stored at - 20&#176;C.</td>
		</tr>
		<tr>
			<td>Tin coated copper wire</td>
			<td>Arco</td>
			<td>B8871K48</td>
			<td>Ordered from Amazon</td>
		</tr>
		<tr>
			<td>Trypan Blue Solution, 0.4%</td>
			<td>Thermo Scientific</td>
			<td>15250061</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin-EDTA, 0.25%</td>
			<td>Thermo FIsher Scientific</td>
			<td>25200072</td>
			<td></td>
		</tr>
		<tr>
			<td>Vacuum Chamber 2</td>
			<td>SP Bel-Art</td>
			<td>F42027-0000</td>
			<td></td>
		</tr>
	</tbody>
</table>

assessing functional metrics of skeletal muscle health in human skeletal muscle microtissues

Tre-dimensionelle (3D) in vitro modeller af skeletmuskulatur er et værdifuldt fremskridt i biomedicinsk forskning, da de giver mulighed for at studere skeletmuskulatur reformation og funktion i et skalerbart format, der er modtagelige for eksperimentelle manipulationer. 3D-muskelkultursystemer er ønskelige, da de gør det muligt for forskere at studere skeletmuskulatur ex vivo i forbindelse med menneskelige celler. 3D in vitro modeller nøje efterligne aspekter af den indfødte vævsstruktur af voksne skeletmuskulatur. Men deres universelle anvendelse er begrænset af tilgængeligheden af platforme, der er enkle at fremstille, omkostninger og brugervenlige, og giver relativt store mængder af human skelet muskelvæv. Da skeletmuskulaturen spiller en vigtig funktionel rolle, der er svækket over tid i mange sygdomstilstande, er en eksperimentel platform for mikrotissuestudier mest praktisk, når minimalt invasive calciumtransient- og kontraktile kraftmålinger kan udføres direkte inden for selve platformen. I denne protokol beskrives fremstillingen af en 96-velkendt platform kendt som 'MyoTACTIC' og en masseproduktion af 3D-humane skeletmuskulaturmikrotiss (hMMT'er). Derudover rapporteres metoderne til en minimalt invasiv anvendelse af elektrisk stimulation, der muliggør gentagne målinger af skeletmuskulaturkraft og calciumhåndtering af hver mikrotissue over tid.

Beskrevet heri er metoder til at kaste en 96-godt PDMS-baserede MyoTACTIC kultur platform fra en PU skimmel, at fremstille arrays af hMMT replika væv, og til at analysere to aspekter af hMMT funktion inden for kultur enhed-kraft generation og calcium håndtering. Figur 1 giver et skematisk overblik over forberedelsen af myoTACTIC kultur brønde før hMMT såning. PDMS er en meget anvendt silikonebaseret polymer, der let kan formes til at skabe komplekse enheder32. En...

Watch this Scientific Journal Video about Assessing Functional Metrics of Skeletal Muscle Health in Human Skeletal Muscle Microtissues at JoVE.com

Assessing Functional Metrics of Skeletal Muscle Health in Human Skeletal Muscle Microtissues

Dette manuskript beskriver en detaljeret protokol til fremstilling af arrays af 3D menneskelige skeletmuskulaturmikrotiss og minimalt invasive downstream in situ-analyser af funktion, herunder kontraktilkraft og calciumhåndteringsanalyser.

Vurdering af funktionelle målinger af skeletmuskulatursundhed i humane skeletmuskulaturmikrotisser

Three-dimensional (3D) in vitro models of skeletal muscle are a valuable advancement in biomedical research as they afford the opportunity to study ...

Vores protokol beskriver detaljerede metoder til at fremstille og analysere en 3D human skeletmuskulatur mikrotissue kultur. Muskelmikrotiss kan anvendes til undersøgelser af grundlæggende muskelbiologi, sygdom modellering, eller for kandidat molekyle test. Den største fordel ved denne teknik er, at den tillader in situ-vurdering af kontraktilkraft og calciumtransienter.På grund af dette kan du foretage langsgående undersøgelser af muskelvævsfunktion. Demonstrere proceduren vil være Brennen Musgrave og Heta Lad, kandidatstuderende fra mit laboratorium. To til tre timer før celle såning, placere en seks godt mild taktik plade del i en 10 centimeter celle kultur skål.Forbered hver enkelt myotactic kultur godt ved at tilføje 100 mikroliter af en 5% pluronic F-127 løsning. Brug låget til at dække brøndene, og påfør en paraffinfilm til at forsegle skålen. Centrifuge fadet på 1, 550 gange G i et minut i en centrifuge udstyret med en plade spinner adapter.Opbevar kulturskålen ved fire grader Celsius, indtil cellerne er klar til såning. Derefter langsomt tø 150 mikroliter aliquot af kælderen membran ekstrakt, og 110 mikroliter aliquot af thrombin på is i kultur hætten. Arbejder i cellekultur hætte, tilføje 700 mikroliter af 0,9% saltvandsopløsning til en 1,5 milliliter mikro centrifuge rør, der indeholder syv milligram pulveriseret fibrinogen.Placer røret i en 37 grader Celsius celle kultur inkubator i tre til fem minutter uden hvirvler. Fjern og svirp røret forsigtigt, og pulsen spinder derefter den opløste opløsning i en mikrocentrifuge, før den returneres til kulturhætten. Fibrinogenopløsningen filtreres ved hjælp af en sprøjte på 1 milliliter udstyret med et 0,22 mikrometer sprøjtefilter.Overfør derefter den opløste fibrinogenopløsning til is ved siden af kældermembranekstraktet og thrombin aliquots. Forbered og forvarm vævsvækstmedierne ved 37 grader Celsius, som vil blive introduceret til kulturtjerne efter såning af vævene. Fjern cellekulturpladerne fra inkubatoren og aspirer kulturmediet.Vask derefter cellerne en gang ved at tilføje fem milliliter DPBS i hver kulturplade. Aspirere DPBS og løsne cellerne ved at tilføje en milliliter af 0,25% trypsin EDTA i hver kultur skål. Placer pladen i cellekulturinkubatoren i tre minutter, hold trypsinen ved at tilføje tre milliliter vaskemedium til kulturskålen og overfør derefter cellerne til et passende størrelse konisk rør.Pellet cellerne ved centrifugering ved 400 gange G i 10 minutter. Aspirere medierne omhyggeligt uden at beskadige celle pellet, derefter suspendere cellerne i en milliliter af vaskemediet. Tæl cellerne ved hjælp af et hæmocytometer og trypan blå farvestof under lyse feltmikroskopi.At frø seks væv, forberede nok celler og ekstracellulære matrix for otte væv, og dermed tegner sig for tab og boble dannelse. Overfør 1,2 millioner celler til et nyt konisk rør, og forøg derefter lydstyrken til 10 millimeter med vaskemedium. Pellet cellerne ved centrifugering ved 400 gange G i 10 minutter.Forbered 150 mikroliter ECM blanding i en 1,5 milliliter mikro centrifuge rør, og gemme blandingen på is, indtil brug. Aspirere medierne fra det koniske rør, der indeholder cellerne, og pas på at undgå cellepillen. Svirp kraftigt enden af røret med en behandsket finger, indtil pellet fremstår som en celle gylle.Overfør 120 mikroliter af ECM-opløsningen til røret, der indeholder celleslammen, og pipettes forsigtigt op og ned for helt at suspendere cellerne i ECM for at generere en enkelt celleaffjedring. Undgå at skabe bobler, og placer derefter cellens ECM-affjedring på is, indtil den bruges. Placer den nedkølede 10 centimeter skål, der indeholder myotactic brønde på toppen af en ispose inde i cellekultur hætte, og aspirere pluronic F-127 løsning fra hver brønd.Lad resterende pluronic F-127 løsning til at frigive fra porøse PDMS, og bosætte sig til bunden af brønden ved at lade brønde sidde i fem minutter på is, derefter aspirere igen. Pipette cellen ECM suspension omhyggeligt at re-suspendere cellerne, derefter overføre 105 mikroliter af suspensionen i en frisk, pre-kølet 1,5 milliliter rør ved at gribe det tæt på toppen. Tilsæt 0,84 mikroliter af 100 enheder pr. milliliter thrombin lageropløsning til de 105 mikroliter af celle ECM suspension.Pipette hurtigt og grundigt at blande, undgå indførelsen af bobler. Tilsæt 15 mikroliter af cellens ECM-blanding til hver brønd uden at trykke pipetten ind i bunden af brønden. Med to lette bevægelser spredes celleaffjedringen bag hver stolpe i brønden.Placer låget på 10 centimeter kulturpladen og overfør den til en 37 grader Celsius vævskultur inkubator i cirka fem minutter. Tilsæt 200 mikroliter af forvarmede vævsvækstmedier til hver myotactic godt efter cellen ECM blanding har polymeriseret. Udskift låget på 10 centimeter skålen og opbevar det i inkubatoren indtil differentiering.I løbet af en 14 dages kulturperiode viste myoblasts aspekterne af indfødte væv ved selvorganisering i den milde taktik godt at danne en 3D-mikrotissue med multi-nukleated striated myotubes. Myotubes har sarkom striationer, som blev visualiseret ved immunstaining for sarkomisk alpha actinin. For at opnå veljusterede myotubes bør tekniske fejl som bobledannelse under pipettering eller beskadigende pluronic belægning under aspiration undgås.En repræsentativ forskydning af den myotactic brøndpladepost som reaktion på lav og højfrekvent elektrisk stimulation, vises her, hvilket indikerer, at myotubes reagerer på varierende elektriske stimuli og trækker sig sammen i overensstemmelse hermed. Kvantificering af postforskydningen giver mulighed for konvertering til absolut kontraktile kraft af HMMTs. Calcium transience på HMMTs lavet af GCaMP6-transduced myoblasts blev også analyseret som reaktion på lav og høj frekvens stimulation.Kvantificering af fluorescerende intensitet kan bruges som måling for calcium håndtering egenskaber HMMTs. De fleste mennesker, der udfører denne væv såning for første gang, kæmper med at tilføje cellen ekstracellulære matrix til brønde, og med boble dannelse. Vi anbefaler at øve teknikken på nogle ekstra mikrotissue kultur brønde med en meget enkel tyktflydende løsning før det første forsøg med celler.

Research

JoVE Journal

Bioengineering

Engineering Skeletal Muscle Tissues from Murine Myoblast Progenitor Cells and Application of Electrical Stimulation

Engineering skeletmuskel Væv fra murine myoblast progenitorceller og anvendelse af elektrisk stimulering

Engineered muscle tissues can be used for several different purposes, which include the production of tissues for use as a disease model in vitro, e.g. to ...

Transplantation of Induced Pluripotent Stem Cell-derived Mesoangioblast-like Myogenic Progenitors in Mouse Models of Muscle Regeneration

Transplantation af induceret pluripotente stamcelle-afledt Mesoangioblast-lignende Myogenic Stamfadere i Mouse Modeller af muskel regenerering

Patient-derived iPSCs could be an invaluable source of cells for future autologous cell therapy protocols. iPSC-derived myogenic stem/progenitor cells ...

Measurement of Maximum Isometric Force Generated by Permeabilized Skeletal Muscle Fibers

Måling af Maximum Isometrisk Kraft Dannet med permeabiliseres Skeletal muskelfibre

Analysis of the contractile properties of chemically skinned, or permeabilized, skeletal muscle fibers offers a powerful means by which to assess muscle ...

Engineered Vascularized Muscle Flap

Udviklet vaskulariserede Muskel Flap

One of the main factors limiting the thickness of a tissue construct and its consequential viability and applicability in vivo, is the control of oxygen ...

Computed Tomography and Optical Imaging of Osteogenesis-angiogenesis Coupling to Assess Integration of Cranial Bone Autografts and Allografts

Computed Tomography og optisk billeddannelse af Osteogenesis-angiogenese Kobling at Vurdere Integration af kranieknogler autografter og Allografter

A major parameter determining the success of a bone-grafting procedure is vascularization of the area surrounding the graft. We hypothesized that ...

Applications of In Vivo Functional Testing of the Rat Tibialis Anterior for Evaluating Tissue Engineered Skeletal Muscle Repair

Applications of In Vivo Functional Testing of the Rat Tibialis Anterior for Evaluating Tissue Engineered Skeletal Muscle Repair

anvendelser af<em&gt; In vivo</em&gt; Funktionel Test af Rat tibialis anterior for Evaluering manipuleret væv Skeletal Muscle Repair

Despite the regenerative capacity of skeletal muscle, permanent functional and/or cosmetic deficits (e.g., volumetric muscle loss (VML) resulting from ...

Cardiac Muscle-cell Based Actuator and Self-stabilizing Biorobot - PART 1

Kardial muskelcellebaseret aktuator og selvstabiliserende biorobot - DEL 1

Biological machines often referred to as biorobots, are living cell- or tissue-based devices that are powered solely by the contractile activity of living ...

Cardiac Muscle Cell-based Actuator and Self-stabilizing Biorobot - Part 2

Kardial muskelcellebaseret aktuator og selvstabiliserende biorobot - del 2

In recent years, hybrid devices that consist of a living cell or tissue component integrated with a synthetic mechanical backbone have been developed. ...

Semiautomated Longitudinal Microcomputed Tomography-based Quantitative Structural Analysis of a Nude Rat Osteoporosis-related Vertebral Fracture Model

Semiautomated langsgående Microcomputed CT-baseret kvantitative strukturel analyse af en nøgen rotte osteoporose Vertebral fraktur Model

Osteoporosis-related vertebral compression fractures (OVCFs) are a common and clinically unmet need with increasing prevalence as the world population ...

Melt Electrospinning Writing of Three-dimensional Poly(&#949;-caprolactone) Scaffolds with Controllable Morphologies for Tissue Engineering Applications

Melt Electrospinning Writing of Three-dimensional PolyÃƒÆ’Ã…Â½Ãƒâ€šÃ‚Âµ-caprolactone Scaffolds with Controllable Morphologies for Tissue Engineering Applications

Smelte Electrospinning skrivning af tre-dimensionelle Poly(ε-caprolactone) stilladser med kontrollerbare morfologier for Tissue Engineering applikationer

This tutorial reflects on the fundamental principles and guidelines for electrospinning writing with polymer melts, an additive manufacturing technology ...