Name: stem cell transplantation in an in vitro simulated ischemia/reperfusion model
Uploaded: 2011-11-05T14:00:00
Description: Watch this Scientific Journal Video about Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model at JoVE.com

Dette arbeidet ble støttet av OTKA (Hungarian Scientific Research Fund) D45933, T049621,, Bolyai Tet (Hungarian Science and Technology Foundation) A4/04, Arg-17/2006 og SIN, Öveges Fellowships og TÁMOP 4.2.2-08 / 1 / KMR-2008-0004 og 4.2.1 / B 09/1/KMR-2010-0001. OTKA 83803. Vi ønsker å takke William Gesztes for å gi voice-over.

1. Forbereder H9c2 cardiomyoblast celler H9c2 rotte cardiomyoblasts ble hentet fra ATCC (Wesel, Tyskland) og utvidet i høy glukose (4,5 g / L) DMEM inneholder 10% føtale bovint serum, 4 mm L-glutamin, 100 U / ml penicillin og 100 mikrogram / ml streptomycin. Den H9c2 myoblast cellelinje er avledet fra embryonale rotte hjerte, er det brukt som en in vitro modell for både skjelett-og hjertemuskulatur 8,9. Forbered 12 godt plate av H9c2 celler: <ol><...

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Den demonstrerte protokollen er en in vitro tilnærming til mye mer komplekse utgaven av stamcelleterapi i hjerteinfarkt, med alle fordeler og ulemper ved en slik modell. Selvfølgelig kan det ikke reflektere komplekse (f.eks immunologiske) hendelser som finner sted under og etter hjerteinfarkt, men kan fokusere på de direkte effektene av den ekstra cellene på postischemic celler. Effekten av simulert iskemi på H9c2 cardiomyoblasts svært avhengig av tid på OGD på passasjen antall brukt celler og på O 2-konse...

<table border="1"><tbody><tr><td> Navn på reagens </td><td> Firma </td><td> Katalognummer </td><td> Kommentarer </td></tr><tr><td> calcein-AM </td><td> Molekylære prober </td><td> L3224, C3099 </td><td> <a href="http://www.invitrogen.com">http://www.invitrogen.com</a> </td></tr><tr><td> ethidium homodimer-2 </td><td> Molekylære prober </td><td> L3224, E3599 </td><td> <a href="http://www.invitrogen.com">http://www.invitrogen.com</a> </td></tr><tr><td> Vybrant Visste </td><td> Molekylære prober </td><td> V22887 </td><td> <a href="http://www.invitrogen.com">http://www.invitrogen.com</a> </td></tr></tbody></table> Tabell 1. Reagenser. <table border="1"><tbody><tr><td> Navn på utstyr </td><td> Firma </td><td> Kommentarer (valgfritt) </td></tr><tr><td> PeCon cell inkubering system for Zeiss mikroskoper </td><td> PeCon GmbH </td><td> <a href="http://www.pecon.biz/">www.pecon.biz/</a> </td></tr></tbody></table> Tabell 2. Equipment.

stem cell transplantation in an in vitro simulated ischemia/reperfusion model

Stamcelletransplantasjon protokoller finner veien inn i klinisk praksis 1,2,3. Få bedre resultater, noe som gjør protokollene mer robust, og finne nye kilder for implanterte cellene er i fokus for nyere forskning 4,5. Gransker effektiviteten av cellen terapi er ikke en lett oppgave og nye verktøy er nødvendig å undersøke mekanismene som er involvert i behandlingsprosessen 6. Vi har utformet en forsøksprotokoll av iskemi / reperfusjon slik at observasjon av cellulære forbindelser og selv subcellular mekanismene under iskemi / reperfusjon skade og etter stamcelletransplantasjon og å evaluere effekten av celleterapi. H9c2 cardiomyoblast celler ble plassert på celle kultur plater 7,8. Iskemi ble simulert med 150 minutter i en glukose frie medium med oksygen nivå under 0,5%. Deretter var normale media og oksygennivået i blodet gjeninnføres å simulere reperfusjon. Etter oksygen glukose deprivasjon,de skadede cellene ble behandlet med transplantasjon av merket human benmarg avledet stamceller ved å legge dem til kultur. Stamceller er foretrukket i kliniske studier, fordi de er lett tilgjengelig med minimal invasiv kirurgi, lett utvidbare og autolog. Etter 24 timer med co-dyrking, ble cellene farget med calcein og ethidium-homodimer å skille mellom levende og døde celler. Dette oppsettet tillot oss å undersøke intercellular tilkoblinger ved hjelp confocal fluoriserende mikroskopi og å kvantifisere overlevelsesraten postischemic celler ved flowcytometri. Konfokalmikroskopi viste interaksjonene mellom de to celle populasjoner som celle fusjon og dannelse av intercellulær nanorør. Flowcytometri analyse avslørte tre klynger av skadede celler som kan plottes på en graf og analysert statistisk. Disse populasjoner kan bli etterforsket separat og konklusjoner kan trekkes på disse data om effektiviteten av den simulerteterapeutisk tilnærming.

Watch this Scientific Journal Video about Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model at JoVE.com

Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

Vi viser hvordan du setter opp en<em&gt; In vitro</em&gt; Iskemi / reperfusjon modell og hvordan man skal evaluere effekten av stamcelleterapi på postischemic kardiale celler.

Stamcelletransplantasjon i en In vitro Simulert ischemia / reperfusjon Model

Stem cell transplantation protocols are finding their way into clinical practice1,2,3. Getting better results, making the protocols more robust, and ...

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Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

A Mouse Model of in Utero Transplantation

A Mouse Model of in Utero Transplantation

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus.  For example, in ...

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Ischemia-reperfusion induced acute kidney injury (IR-AKI) is widely used as a model of AKI in mice, but results are often quite variable with high, often ...

Neural Stem Cell Transplantation in Experimental Contusive Model of Spinal Cord Injury

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Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can ...

Oxygen-Glucose Deprivation and Reoxygenation as an In Vitro Ischemia-Reperfusion Injury Model for Studying Blood-Brain Barrier Dysfunction

Oxygen-Glucose Deprivation and Reoxygenation as an In Vitro Ischemia-Reperfusion Injury Model for Studying Blood-Brain Barrier Dysfunction

Oxygen-Glukose deprivasjon og reoksygenering som en<em&gt; In Vitro</em&gt; Iskemi-reperfusjonskade modell for å studere blod-hjernebarrieren Dysfunction

Ischemia-Reperfusion (IR) injury is known to contribute significantly to the morbidity and mortality associated with ischemic strokes. Ischemic ...

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies

Studerer Bukspyttkjertelkreft Stem Cell Kjennetegn for å utvikle nye behandlingsstrategier

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor ...

Murine Model of Intestinal Ischemia-reperfusion Injury

Murine modell av intestinal iskemi-reperfusjon Injury

Intestinal ischemia is a life-threatening condition associated with a broad range of clinical conditions including atherosclerosis, thrombosis, ...

Creation and Transplantation of an Adipose-derived Stem Cell (ASC) Sheet in a Diabetic Wound-healing Model

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Opprettelse og transplantasjon av en liggende under Adipose-avledet stilk cellen (ASC) ark i en diabetiker sårhelende modell

Artificial skin has achieved considerable therapeutic results in clinical practice. However, artificial skin treatments for wounds in diabetic patients ...

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In this study, a middle cerebral artery (MCA) occlusion mouse model is employed to study cerebral ischemia-reperfusion. A reproducible and reliable mouse ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

En pre-klinisk rotte modell for studier av ischemia-reperfusion skade i rekonstruktiv mikrokirurgi

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...