Name: a functional motor unit in the culture dish: co-culture of spinal cord explants and muscle cells
Uploaded: 2012-04-12T12:00:00
Description: Watch this Scientific Journal Video about A Functional Motor Unit in the Culture Dish: Co-culture of Spinal Cord Explants and Muscle Cells at JoVE.com

Vores arbejde er støttet af den schweiziske National Science Foundation (SNF), det schweiziske initiativ i Systembiologi (SystemsX.ch), den Muskelsvindfonden Association USA (MDA), de Association Française contre les Myopati (AFM), De Forenede mitochondrial sygdom Foundation (umdf), den Gebert-Ruf Fonden Sjældne sygdomme Programme (GRF), den schweiziske Selskab for Forskning i de muskelsygdomme (SSEM / FSRMM), den schweiziske Life &quot;Jubiläumsstiftung für Volksgesundheit und Medizinische Forschung&quot;, Roche Grundforskningsfond og universitetet af Basel.

1. Fremstillinq af den primære human muskel Cell Culture <ol><li> Etabler de menneskelige muskel cellekulturer efter explantation-re-explantation teknik 4. Først fjerne ikke-muskelvæv fra biopsier. Derefter indkapsle 1 mM 3 muskel eksplantater i et koagel af plasma, som tillader fibroblaster kan udledes af de eksplantater. </li><li> De indlejrede eksplantater overføres til en plasma-gelatine overtrukket skål og lod dem vokse i normalt vækstmedium (MEM suppleret med 25% medium 199, 10% føta...

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En fysiologisk in vitro værktøj til at studere muskelcelle funktion i normal og patologisk forbindelse er af stor interesse for myologists, fordi musklerne cellekulturer normalt ikke gentage betydningen af multiple celler og celle-typen forbindelser. Tilsætning af renset motoriske neuroner til muskelceller, er ikke tilstrækkeligt til at opnå en funktionel motorenhed, eftersom tilstedeværelsen af Schwann-celler er nødvendig for innervation at være effektiv 5 og det er ikke topologisk (3D + sæ...

<table border="1"><tbody><tr><td> Navn reagens </td><td> Firma </td><td> Katalognummer </td></tr><tr><td> HBSS </td><td> Gibco / Invitrogen </td><td> 14170 </td></tr><tr><td> MEM </td><td> Gibco / Invitrogen </td><td> 31095 </td></tr><tr><td> Medium 199 </td><td> Gibco / Invitrogen </td><td> 31153 </td></tr><tr><td> Føtalt bovint serum </td><td> Føtal Klon Perbio </td><td> SH30066.03 </td></tr><tr><td> Insulin </td><td> Sigma </td><td> I9278 </td></tr><tr><td> Human EGF </td><td> Sigma </td><td> E9644 </td></tr><tr><td> Human FGF </td><td> SiGMA </td><td> F0291 </td></tr><tr><td> Penicillin / streptomycin-opløsning </td><td> Gibco </td><td> 15140 </td></tr></tbody></table>

a functional motor unit in the culture dish: co-culture of spinal cord explants and muscle cells

Humane primære muskelceller dyrket aneurally i monolag sjældent kontrakt spontant fordi, i fravær af en nerve komponent, celledifferentiering begrænset og motorisk neuron stimulering mangler en. Disse begrænsninger hæmmer in vitro-undersøgelse af mange neuromuskulære sygdomme i dyrkede muskelceller. Vigtigt, kan de eksperimentelle begrænsninger monolayered, dyrkede muskelceller overvindes ved funktionel innervation af myofibre med rygmarv eksplantater i co-kulturer. Her viser vi de forskellige trin, der kræves for at opnå en effektiv og korrekt innervation af humane primære muskelceller, hvilket fører til fuldstændig differentiering og fiber kontraktion ifølge fremgangsmåden udviklet af Askanas 2. At gøre dette, er muskelceller co-dyrket med rygmarvsneuroner eksplantater af rotteembryoer ved ED 13,5, med den dorsale rodganglier stadig fastgjort til rygmarven skiver. Efter et par dage, fi musklenmedlemmer begynder at kontrakt og bliver til sidst på tværs af tværstribede gennem innervation af funktionelle neurites fremspringende fra rygmarven eksplantater, der forbinder til muskelceller. Denne struktur kan opretholdes i mange måneder, simpelthen ved regelmæssig udskiftning af dyrkningsmediet. De anvendelser af denne uvurderligt værktøj er mange, da den udgør en funktionel model for tværfaglige analyser af menneskelig muskel udvikling og innervation. Faktisk forekommer en fuldstændig de novo neuromuskulære junction installation i en dyrkningsskål, der tillader en nem måling af mange parametre ved hvert trin i en grundlæggende og fysiologisk sammenhæng. Blot for at nævne et par eksempler, genomiske og / eller proteom undersøgelser kan udføres direkte på co-kulturer. Endvidere kan præ-og post-synaptiske virkninger specifikt og separat vurderet ved den neuromuskulære junction, fordi begge komponenter kommer fra forskellige arter,rotte og human hhv. Nerven-muskel co-kultur kan også udføres med humane muskelceller isoleret fra patienter, der lider af muskelvæv eller neuromuskulære sygdomme 3, og kan således anvendes som et screeningsværktøj til lægemiddelkandidater. Endelig er ikke særligt udstyr, men en regulær BSL2 facilitet bruges til at danne en funktionel motorenheden i en kultur fad. Denne metode er således værdifuldt for både muskler samt de neuromuskulære forskningsmiljøer for fysiologiske og mekanistiske studier af neuromuskulære funktion, i en normal og sygdom sammenhæng.

Watch this Scientific Journal Video about A Functional Motor Unit in the Culture Dish: Co-culture of Spinal Cord Explants and Muscle Cells at JoVE.com

A Functional Motor Unit in the Culture Dish: Co-culture of Spinal Cord Explants and Muscle Cells

Dyrkede muskelceller er en utilstrækkelig model til rekapitulere innerverede muskel<em&gt; In vivo</em&gt;. En funktionel motor enhed kan reproduceres<em&gt; In vitro</em&gt; Ved innervation af differentierede humane primære muskelceller ved hjælp af rotte foster rygmarv eksplantaterne. Denne artikel beskriver hvordan co-kulturer af rygmarv eksplantater og muskelceller er etableret.

En funktionel Motor enhed i Kultur Dish: Co-kultur for Rygmarvsskader eksplantater og muskelceller

Human primary muscle cells cultured  aneurally in monolayer rarely contract spontaneously because, in the absence of  a nerve component, cell ...

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