Name: germ cell transplantation and testis tissue xenografting in mice
Uploaded: 2012-02-06T12:00:00
Description: Watch this Scientific Journal Video about Germ Cell Transplantation and Testis Tissue Xenografting in Mice at JoVE.com

Werk van de auteurs laboratorium werd ondersteund door USDA / CSREES / NRICGP (2007-35203-18213); NIH / NCRR (2 R01 RR17359-06), NIH / NIEHS (1 R21 ES014856-01A2) en Alberta innoveert - Health Solutions.

DEEL A. Germ cel transplantatie bij muizen 1. Voorbereiding van de ontvanger muizen <ol><li> Ontvangers moeten immunologisch tolerant (hetzij genetisch aangepast aan donoren of immuun-deficiënte) aan de donor testis cellen. </li><li> De afnemers moeten worden hetzij van nature, zonder spermatogenese (bijv. W / W v muizen) of helemaal leeg van endogene kiemcellen. Germ cel depletie kan worden bereikt door bestralen of chemotherapeutische medicijnen zoals Busulfan. In dit ...

<ol>
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1. Germ celtransplantatie Germ celtransplantatie vormt het enige functionele assay voor ondubbelzinnige bevestiging van de aanwezigheid van spermatogoniale stamcellen (SSC&#39;s) in een cel populatie. Alleen SSC&#39;s kunnen naar huis aan en de SSC niche in de kelder membraan en koloniseren initiëren donor-afgeleide spermatogenese. Germ celtransplantatie maakte het mogelijk om te studeren en SSC&#39;s te manipuleren in een ongekende manier. De techniek is gebruikt om transgene dieren te produ...

<table border="1"><tbody><tr><td> Naam van het serum </td><td> Vennootschap </td><td> Catalogusnummer </td></tr><tr><td> Collagenase (type IV) </td><td> Sigma </td><td> C5138 </td></tr><tr><td> Trypsine-EDTA </td><td> Invitrogen </td><td> 25200-056 </td></tr><tr><td> DNasel </td><td> Sigma </td><td> DN25 </td></tr><tr><td> DMEM </td><td> Invitrogen </td><td> 31053-028 </td></tr><tr><td> Trypaan blauwe vlek </td><td> Invitrogen </td><td> 15250-061 </td></tr><tr><td> Nylon mesh cel zeef </td><td> BD Biosciences </td><td> 352.340 (40 pm) 352350 (70μm) </td></tr><tr><td> Busulfan </td><td> Sigma </td><td> B2635 </td></tr><tr><td> Dunwandige glazen capillairen </td><td> Wereld precisie-instrument </td><td> TW 100-3 </td></tr><tr><td> BD intramedic plyethylene buis (PE100) </td><td> BD </td><td> CA-63018-725 </td></tr><tr><td> Ethicon 6-0 Silk Suture </td><td> Ethicon </td><td> 706G </td></tr><tr><td> Wond clips </td><td> BD </td><td> 427631 </td></tr><tr><td> Sigmacote </td><td> Sigma </td><td> SL2 </td></tr><tr><td> X-gal </td><td> Sigma </td><td> B4252 </td></tr><tr><td> Kaliumferrocyanide </td><td> Sigma </td><td> P9387 </td></tr><tr><td> Kaliumferricyanide </td><td> Sigma </td><td> P3667 </td></tr><tr><td> magnesium chloride </td><td> Sigma </td><td> 208337 </td></tr><tr><td> natrium deoxycholaat </td><td> Sigma </td><td> D6750 </td></tr><tr><td> N, N-Dimethylformamide </td><td> Sigma </td><td> D4551 </td></tr><tr><td> Igepal CA-630 </td><td> Sigma </td><td> 18896 </td></tr></tbody></table>

germ cell transplantation and testis tissue xenografting in mice

Germ celtransplantatie is ontwikkeld door Dr Ralph Brinster en collega&#39;s van de Universiteit van Pennsylvania in 1994 1,2. Deze baanbrekende studies toonden aan dat micro-injectie van kiemcellen van vruchtbare donor muizen in de testes van de onvruchtbare ontvangende muizen resulteert in een donor-afgeleide spermatogenese en sperma productie door de ontvanger dier 2. Het gebruik van donor mannen dat het bacteriële β-galactosidase gen toegestaan ​​identificatie van donor-afgeleide spermatogenese en de overdracht van de donor haplotype aan het nageslacht door ontvangende dieren 1. Verrassend, na transplantatie in het lumen van de tubuli seminiferis, getransplanteerde kiemcellen konden van de luminale compartiment verplaatsen naar de kelder membraan waar spermatogonia zich bevinden 3. Het is algemeen aanvaard dat alleen SSC&#39;s kunnen de nis en herstellen spermatogenese bij de ontvanger testis koloniseren. Daarom kiemceltransplantatie zorgt voor een functionele benadering van de stamcel niche in de testis te bestuderen en vermeende spermatogoniale stamcellen te karakteriseren. Tot op heden is kiemcel transplantatie gebruikt om de basis stamcelbiologie toe te lichten, om transgene dieren produceren door middel van genetische manipulatie van geslachtscellen voorafgaand aan de transplantatie 4,5 tot Sertoli cel-kiemcel interactie 6,7, SSC homing en kolonisatie 3 te bestuderen, 8, evenals SSC zelfvernieuwing en differentiatie 9,10. Germ celtransplantatie is ook mogelijk in grote soorten 11. In deze, de belangrijkste toepassingen zijn het behoud van de vruchtbaarheid, de verspreiding van de elite van de genetica bij dieren, en het genereren van transgene dieren als de studie van de spermatogenese en SSC biologie met deze techniek is logistiek moeilijker en duurder dan in knaagdieren. Transplantatie van kiemcellen van grote soorten in de testes van muizen resulteert in colonization van donor cellen en spermatogonia expansie, maar niet in hun volledige differentiatie vermoedelijk wegens onverenigbaarheid van de ontvanger somatische cel compartiment met de kiem cellen van fylogenetisch verre species 12. Een alternatieve benadering is de transplantatie van kiemcellen van grote soorten samen met hun omringende somatische compartiment. We hebben eerst gemeld in 2002, dat kleine fragmenten van testis weefsel van onvolwassen mannetjes getransplanteerd onder de dorsale huid van immunodeficiënte muizen in staat zijn om te overleven en te ondergaan volle ontwikkeling met de productie van de bevruchting bevoegde sperma 13. Sindsdien testis weefsel xenografting is gebleken om succesvol te zijn in vele soorten en naar voren gekomen als een waardevol alternatief voor de testis ontwikkeling en spermatogenese van de grote dieren in muizen 14 te bestuderen.

Watch this Scientific Journal Video about Germ Cell Transplantation and Testis Tissue Xenografting in Mice at JoVE.com

Germ Cell Transplantation and Testis Tissue Xenografting in Mice

Protocollen voor kiemcellen transplantatie en testis weefsel xenografting worden beschreven. Germ celtransplantatie resulteert in een donor-afgeleide spermatogenese in de ontvangende testes en vertegenwoordigt een functionele reconstitutie test voor de identificatie van spermatogoniale stamcellen (SSC&#39;s). Testis weefsel xenografting reproduceert testis ontwikkeling en spermatogenese van verschillende soorten donor in de ontvangende muizen.

Germ Cell Transplantation en de testis weefsel Xenografting in Muizen

Germ cell transplantation was developed by Dr. Ralph Brinster and colleagues at the University of Pennsylvania in 19941,2. These ground-breaking studies ...

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