Name: non-enzymatic, serum-free tissue culture of pre-invasive breast lesions for spontaneous generation of mammospheres
Uploaded: 2014-11-08T14:00:00
Description: Watch this Scientific Journal Video about Non-enzymatic, Serum-free Tissue Culture of Pre-invasive Breast Lesions for Spontaneous Generation of Mammospheres at JoVE.com

Dette arbeidet ble delvis støttet av (1) Department of Defense Breast Cancer Research Program (US Army Medical Research Acquisition Activity) award # W81XVVH-10-1-0781 til LAL og VE, og (2) Susan G. Komen Foundation stipend IR122224446 i Lal og VE. Patologi støtte og vev innbringende ble vennlig levert av Inova Fairfax Pathology avdeling, Dr. Hassan Nayer, Dr. Geetha A. Menezes, og Dr. Charles Bechert. Pasientsamtykke og prøve anskaffelser ble fagmessig guidet av Inova Fairfax Hospital klinisk forskningskoordinatorer Holly Gallimore, Heather Huryk, og Emil Kamar.

Menneskelig brystvev ble samlet inn fra pasienter som deltok i en forskningsstudie, med skriftlig informert samtykke, etter Department of Defense, George Mason University, og Inova Health System Institutional Review Board godkjente protokoller. 1. Forbered Nutrient Rich Medium med vekstfaktorer og antibiotika <ol><li> Forbered stamløsninger av insulin, epidermal vekstfaktor (EGF), streptomycin og gentamicin sulfat sulfat. <ol><li> Rekonstituer insulin i sterilt filtrert vann til en sluttk...

<ol>
	<li>Shaughnessy, J. A., 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=Treatment+and+prevention+of+intraepithelial+neoplasia:+an+important+target+for+accelerated+new+agent+development.">Treatment and prevention of intraepithelial neoplasia: an important target for accelerated new agent development.</a> Clin Cancer Res. 8 (2), 314-346 (2002).</li><li>Lee, J., 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=Tumor+stem+cells+derived+from+glioblastomas+cultured+in+bFGF+and+EGF+more+closely+mirror+the+phenotype+and+genotype+of+primary+tumors+than+do+serum-cultured+cell+lines.">Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines.</a> Cancer Cell. 9 (5), 391-403 (2006).</li><li>Behbod, F., 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=An+intraductal+human-in-mouse+transplantation+model+mimics+the+subtypes+of+ductal+carcinoma+in+situ.">An intraductal human-in-mouse transplantation model mimics the subtypes of ductal carcinoma in situ.</a> Breast Cancer Res. 11 (5), (2009).</li><li>Debnath, J., Muthuswamy, S. K., Brugge, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morphogenesis+and+oncogenesis+of+MCF-10A+mammary+epithelial+acini+grown+in+three-dimensional+basement+membrane+cultures.">Morphogenesis and oncogenesis of MCF-10A mammary epithelial acini grown in three-dimensional basement membrane cultures.</a> Methods. 30 (3), 256-268 (2003).</li><li>Espina, V., 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=Malignant+precursor+cells+pre-exist+in+human+breast+DCIS+and+require+autophagy+for+survival.">Malignant precursor cells pre-exist in human breast DCIS and require autophagy for survival.</a> PLoS One. 5 (4), (2010).</li><li>Dontu, G., 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=In+vitro+propagation+and+transcriptional+profiling+of+human+mammary+stem/progenitor+cells.">In vitro propagation and transcriptional profiling of human mammary stem/progenitor cells.</a> Genes Dev. 17 (10), 1253-1270 (2003).</li><li>Farnie, G., 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=Novel+cell+culture+technique+for+primary+ductal+carcinoma+in+situ:+role+of+Notch+and+epidermal+growth+factor+receptor+signaling+pathways.">Novel cell culture technique for primary ductal carcinoma in situ: role of Notch and epidermal growth factor receptor signaling pathways.</a> J Natl Cancer Inst. 99 (8), 616-627 (2007).</li><li>Wicha, M. S., Liotta, L. A., Garbisa, S., Kidwell, W. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Basement+membrane+collagen+requirements+for+attachment+and+growth+of+mammary+epithelium.">Basement membrane collagen requirements for attachment and growth of mammary epithelium.</a> Exp Cell Res. 124 (1), 181-190 (1979).</li><li>Espina, V., Liotta, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=What+is+the+malignant+nature+of+human+ductal+carcinoma+in+situ.">What is the malignant nature of human ductal carcinoma in situ.</a> Nat Rev Cancer. 11 (1), 68-75 (2011).</li><li>Gong, C., 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=Beclin+1+and+autophagy+are+required+for+the+tumorigenicity+of+breast+cancer+stem-like/progenitor+cells.">Beclin 1 and autophagy are required for the tumorigenicity of breast cancer stem-like/progenitor cells.</a> Oncogene. 32 (18), 2261-2272 (2012).</li><li>Simon, B., 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=Epithelial+glycoprotein+is+a+member+of+a+family+of+epithelial+cell+surface+antigens+homologous+to+nidogen,+a+matrix+adhesion+protein.">Epithelial glycoprotein is a member of a family of epithelial cell surface antigens homologous to nidogen, a matrix adhesion protein.</a> PNAS. 87 (7), 2755-2759 (1990).</li><li>Ma, X. J., 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=Gene+expression+profiles+of+human+breast+cancer+progression.">Gene expression profiles of human breast cancer progression.</a> Proc Natl Acad Sci U S A. 100 (10), 5974-5979 (2003).</li><li>Schnitt, S. J., Harris, J. R., Smith, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developing+a+prognostic+index+for+ductal+carcinoma+in+situ+of+the+breast.+Are+we+there+yet.">Developing a prognostic index for ductal carcinoma in situ of the breast. Are we there yet.</a> Cancer. 77 (11), 2189-2192 (1996).</li><li>Sgroi, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preinvasive+breast+cancer.">Preinvasive breast cancer.</a> Annu Rev Pathol. 5, 193-221 (2010).</li><li>Asch, H. L., Asch, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterogeneity+of+keratin+expression+in+mouse+mammary+hyperplastic+alveolar+nodules+and+adenocarcinomas.">Heterogeneity of keratin expression in mouse mammary hyperplastic alveolar nodules and adenocarcinomas.</a> Cancer Res. 45 (6), 2760-2768 (1985).</li><li>LaBarge, M. A., Petersen, O. W., Bissell, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Of+microenvironments+and+mammary+stem+cells.">Of microenvironments and mammary stem cells.</a> Stem Cell Rev. 3 (2), 137-146 (2007).</li><li>Smart, C. E., 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=In+vitro+analysis+of+breast+cancer+cell+line+tumourspheres+and+primary+human+breast+epithelia+mammospheres+demonstrates+inter-+and+intrasphere+heterogeneity.">In vitro analysis of breast cancer cell line tumourspheres and primary human breast epithelia mammospheres demonstrates inter- and intrasphere heterogeneity.</a> PLoS One. 8 (6), (2013).</li><li>Vaillant, F., Asselin-Labat, M. L., Shackleton, M., Lindeman, G. J., Visvader, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+emerging+picture+of+the+mouse+mammary+stem+cell.">The emerging picture of the mouse mammary stem cell.</a> Stem Cell Rev. 3 (2), 114-123 (2007).</li><li>Kim, D. 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=Actin+cap+associated+focal+adhesions+and+their+distinct+role+in+cellular+mechanosensing.">Actin cap associated focal adhesions and their distinct role in cellular mechanosensing.</a> Sci Rep. 2, 555 (2012).</li><li>Espina, V., Wysolmerski, J., Edmiston, K., Liotta, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Attacking+breast+cancer+at+the+preinvasion+stage+by+targeting+autophagy.">Attacking breast cancer at the preinvasion stage by targeting autophagy.</a> Women's Health. 9 (2), 1-14 (2013).</li><li>D'amonte, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mammary+carcinoma+behavior+is+programmed+in+the+precancer+stem+cell.">Mammary carcinoma behavior is programmed in the precancer stem cell.</a> Breast Cancer Res. 10 (3), (2008).</li></ol>

Kulturen systemet beskrevet her utgjør en ny modell for å generere levende pre-invasive neoplastiske bryst celler for grunnleggende og translasjonsforskning studier. I det siste har pre-ondartet brystkreft progresjon vanligvis blitt studert ved hjelp av tre forskjellige metoder. Den første metoden er histopatologisk og genetisk analyse av frosne microdissected eller faste humane prøvene 12-14. Den andre metoden benytter musemodeller som inneholder hyperplastiske alveolære knuter (HAN lesjoner) som er ten...

Spredning av epitelceller innenfor rammen av bryst kanaler og alveolene (ductal carcinoma in situ) er anerkjent som en obligat forløper til invasiv ductal og lobular brystkreft. Ikke desto mindre er de molekylære mekanismer og dynamikk cellepopulasjon som tillater progresjon til invasiv kreft dårlig forstått. Belyse overlevelsesmekanismer som brukes av pre-invasive brystkreft celler, eller en hvilken som helst pre-invasive svulster, kan avsløre terapeutiske strategier for å drepe, eller hindre, pre-invasive svulster 1. Imidlertid har enkle lavkost metoder for funksjonelt studerer menneskelige pre-invasive lesjoner vært mangelfull. Selv om in vitro-monolag-kultur av transformerte cellelinjer er en etablert fremgangsmåte laboratorie, genotype og fenotype av disse udødeliggjorte cellelinjer unnlater å rekapitulere molekyl status av primære humane tumorceller 2. Videre, til og med den ikke-tumorigen MCF-10A-cellelinjen, som Recapitulates 3-D melkekjertlene arkitektur, unnlater å representere funksjonell fenotype og molekylære egenskapene til den enkelte pasients pre-invasiv bryst lesjon 3,4 tilstrekkelig. For å bestemme om stammeliknende neoplastiske celler i stand til invasjons eksistert innen duktalt karsinom in situ (DCIS) cellepopulasjon, har vi utviklet en metode for innsamling og dyrking sterilt humant brystvev ved tidspunktet for kirurgi (figur 1) 5. Vår ex vivo bryst organoid kultursystemet ikke er avhengig av enzymatisk vev avbrudd, basalmembranekstrakt matrise eller fibroblast uttømming, for å isolere og som forplanter seg mammosphere-dannende celler fra friske humane bryst duktalt karsinom vev 6-8. Vår nye systemet er basert på prinsippet om celle streaming / migrasjon 5. De bare bryst kanaler, og omkringliggende stroma blir neddykket i et minimalt volum av serumfritt næringsmedium (e barenough å dekke kanal fragmenter) for å maksimere gassutveksling, med snittflaten av kanalen eksponert til kulturmediet, men på ingen bestemt retning i kolben (figur 1E-F). Denne kulturen systemet tillater celler til å migrere ut av kanalen og inn / ut mot autologe stroma og kulturflaske. Den næringsmedium, supplert bare med epidermal vekstfaktor (EGF), insulin, og antibiotika, støtter veksten av blandede cellepopulasjoner som kommer fra organoid. Den vevskulturkolbe har et avtagbart, lukkbar lokk som gjør det mulig for organoids og / eller celler som skal høstes uten å forstyrre hele kolbe eller nabo organoids, og samtidig opprettholde et sterilt miljø fuktet.

<table border="0" cellpadding="0" cellspacing="0" width="1024">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">Name of Reagent/ Equipment</td>
			<td style="width:187px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:365px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">Ethanol</td>
			<td style="width:187px;">Fisher</td>
			<td style="width:119px;">A405-P</td>
			<td>prepare a 70% solution in Type 1 reagent grade water</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">18 M&#937;-cm water, sterile filtered</td>
			<td style="width:187px;"></td>
			<td style="width:119px;"></td>
			<td>sterile filtered, Type 1 reagent grade water</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10cc plastic disposable syringe, sterile</td>
			<td style="width:187px;">BD</td>
			<td style="width:119px;">305482</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">0.2&#181;m polyethersulfone (PES) syringe filter, sterile</td>
			<td style="width:187px;">Thermo Scientific</td>
			<td style="width:119px;">194-2520</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">15ml polypropylene conical tubes, sterile</td>
			<td style="width:187px;">Fisher</td>
			<td>14-959-49B</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">50ml polypropylene conical tubes, sterile</td>
			<td style="width:187px;">Fisher</td>
			<td>05-539-6</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">1.5ml low retention microcentrifuge tubes,sterile</td>
			<td style="width:187px;">Fisher</td>
			<td style="width:119px;">02-681-331</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">nutrient medium, DMEM-F12/HEPES</td>
			<td style="width:187px;">Invitrogen</td>
			<td style="width:119px;">11330-032</td>
			<td>with L-glutamine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Insulin, human recombinant</td>
			<td style="width:187px;">Roche</td>
			<td style="width:119px;">11376497001</td>
			<td>10mg/ml stock</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Epidermal Growth Factor (EGF), human recombinant</td>
			<td style="width:187px;">Millipore</td>
			<td style="width:119px;">GF144</td>
			<td>100&#181;g/ml stock</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Streptomycin sulfate</td>
			<td style="width:187px;">Sigma-Aldrich</td>
			<td style="width:119px;">S1567</td>
			<td>10mg/ml stock</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Gentamicin sulfate</td>
			<td style="width:187px;">Sigma-Aldrich</td>
			<td style="width:119px;">G19114</td>
			<td>10mg/ml stock</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Filtration flask and filter top, sterile</td>
			<td style="width:187px;">Millipore</td>
			<td>SCGPU02RE</td>
			<td>0.22&#181;m PES membrane</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">25ml sterile, disposable pipettes</td>
			<td style="width:187px;">Fisher</td>
			<td>4489</td>
			<td>paper-plastic wrapped</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10ml sterile, disposable pipettes</td>
			<td style="width:187px;">Fisher</td>
			<td style="width:119px;">4488</td>
			<td>paper-plastic wrapped</td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:353px;">Tissue marking dyes (black, blue, red, green, yellow and orange)</td>
			<td style="width:187px;">CDI</td>
			<td style="width:119px;">MD2000</td>
			<td style="width:365px;">after opening use only with single-use, sterile cotton tipped applicators, or use once and discard</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">Cotton tipped applicators, sterile</td>
			<td style="width:187px;">Fisher</td>
			<td>23-400-115</td>
			<td>single use only</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">Gauze pads, 10x10cm, sterile</td>
			<td style="width:187px;">Fisher</td>
			<td style="width:119px;">2187</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Plastic transfer pipettes, sterile, disposable</td>
			<td style="width:187px;">Samco</td>
			<td>202-20S</td>
			<td></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;">Vinegar, white distilled</td>
			<td style="width:187px;">household use</td>
			<td></td>
			<td style="width:365px;">5% acetic acid; after opening use only with sterile pipettes</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">#10 scalpels, sterile, disposable&#160;</td>
			<td style="width:187px;">Thermo Scientific</td>
			<td>31-200-32&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">petri dish, sterile</td>
			<td style="width:187px;">Fisher</td>
			<td>FB0875713A</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">TPP 115cm2 flask, with removable lid</td>
			<td style="width:187px;">MidSci</td>
			<td style="width:119px;">90652</td>
			<td>screw cap with filter</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;width:353px;">CO2 incubator</td>
			<td style="width:187px;">Fisher</td>
			<td>13-998-074</td>
			<td>5% CO2, 37 oC, humidified chamber</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">inverted light microscope</td>
			<td style="width:187px;">Olypmus</td>
			<td>IX51</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">8M urea</td>
			<td style="width:187px;">Fisher</td>
			<td style="width:119px;">BP169-500</td>
			<td>optional, for mass spectrophotometric analysis of cultured cells</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">2X SDS tris-glycine buffer</td>
			<td style="width:187px;">Life Technologies</td>
			<td>LC2676</td>
			<td>optional, for proteomic analysis of cultured cells</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:353px;">Cytocentrifuge</td>
			<td style="width:187px;">Thermo Scientific</td>
			<td>A78300003</td>
			<td>optional, for preparing cell smears</td>
		</tr>
	</tbody>
</table>

non-enzymatic, serum-free tissue culture of pre-invasive breast lesions for spontaneous generation of mammospheres

Bryst duktalt carcinoma in situ (DCIS), per definisjon, er spredning av neoplastiske epitelceller innenfor rammen av bryst duct, uten å bryte collagenous basalmembran. Mens DCIS er en non-obligat forløper til invasiv brystkreft, er de molekylære mekanismer og cellepopulasjoner som tillater progresjon til invasiv kreft ikke fullt ut kjent. For å bestemme om progenitor-celler i stand til invasjons eksisterte i DCIS cellepopulasjon, har vi utviklet en metode for innsamling og dyrking sterilt humant brystvev ved tidspunktet for kirurgi, enzymatisk uten avbrudd av vev. Sterile brystvev som inneholder duktale segmenter er høstet fra kirurgisk excised brystvev følgende rutine patologisk undersøkelse. Vev som inneholder DCIS er plassert i næringsrikt, antibiotika-inneholdende, serumfritt medium, og transportert til laboratoriet vevskultur. Brystvevet er videre dissekertd for å isolere de forkalkede områder. Multiple brystvev stykker (organoids) er plassert i et minimalt volum av serumfritt medium i en kolbe med et avtagbart lokk, og dyrket i en fuktet CO2-inkubator. Epiteliale og fibroblast cellepopulasjoner dukke opp fra organoid etter 10 - 14 dager. Mammospheres spontant danne på og rundt epitelcelle monolayer. Spesifikke cellepopulasjoner kan høstes direkte fra flasken uten å forstyrre nabocellene. Våre ikke-enzymatisk vev kultur system avslører pålitelig cytogenetisk unormale, invasive stamceller fra friske menneskelige DCIS lesjoner.

Arbeidsflyt for å fremskaffe og dyrking steril bryst duktalt carcinoma in situ vev Brystvev sterilitet opprettholdes fra operasjonsstuen til cellekultur lab via mindre endringer i typiske sykehus patologi arbeidsflyt (figur 1). Vev blir transportert i en steril beholder med en plastfilm deksel som gjør det mulig radiologisk vurdering og samtidig opprettholde sterilitet vev. Gross vev behandling av et bryst lumpectomy eller mastektomi prøven er utført med sterile ha...

Watch this Scientific Journal Video about Non-enzymatic, Serum-free Tissue Culture of Pre-invasive Breast Lesions for Spontaneous Generation of Mammospheres at JoVE.com

Non-enzymatic, Serum-free Tissue Culture of Pre-invasive Breast Lesions for Spontaneous Generation of Mammospheres

Primære cellekultur ved hjelp av intakte vev organoids gir et modellsystem som etterligner multi-mobilnettet in vivo mikromiljøet. Vi utviklet et serum-free primære brystepitel vevskultur modell som foreviger blandede cellekultur linjene og utstillinger differensierte morfologi, uten enzymatisk vev avbrudd. Bryst organoids være levedyktig i&gt; 6 måneder.

Ikke-enzymatisk, Serum-free Tissue Culture of Pre-invasive brystlesjonene for Spontan generasjon Mammospheres

Breast ductal carcinoma in situ (DCIS), by definition, is proliferation of neoplastic epithelial cells within the confines of the breast duct, without ...

Research

JoVE Journal

Biology

Actin Co-Sedimentation Assay; for the Analysis of Protein Binding to F-Actin

The actin cytoskeleton within the cell is a network of actin filaments that allows the movement of cells and cellular processes, and that generates ...

Mouse Mammary Epithelial Cells form Mammospheres During Lactogenic Differentiation

A phenotypic measure commonly used to determine the degree of lactogenic differentiation in mouse mammary epithelial cell cultures is the formation of ...

Generation of Single-Cell Suspensions from Mouse Neural Tissue

Within the nervous system, hundreds of neuronal and glial cell types have been described. Each specific cell type in the brain or spinal cord has a ...

Preparation of Pooled Human Platelet Lysate (pHPL) as an Efficient Supplement for Animal Serum-Free Human Stem Cell Cultures

Preparation of Pooled Human Platelet Lysate pHPL as an Efficient Supplement for Animal Serum-Free Human Stem Cell Cultures

Platelet derived growth factors have been shown to stimulate cell proliferation efficiently in vivo1,2 and in vitro. This effect has been reported for ...

Electrospinning Fibrous Polymer Scaffolds for Tissue Engineering and Cell Culture

As the field of tissue engineering evolves, there is a tremendous demand to produce more suitable materials and processing techniques in order to address ...

Long-term Culture of Human Breast Cancer Specimens and Their Analysis Using Optical Projection Tomography

Breast cancer is a leading cause of mortality in the Western world. It is well established that the spread of breast cancer, first locally and later ...

Mouse Embryonic Development in a Serum-free Whole Embryo Culture System

Mus embryoutvikling i en Serum-fri Whole Embryo Kultur System

Mid-gestation stage mouse embryos were cultured utilizing a serum-free culture medium prepared from commercially available stem cell media supplements in ...

Generation and Culture of Blood Outgrowth Endothelial Cells from Human Peripheral Blood

Generasjon og Culture of Blood utvekst endotelceller fra humant perifert blod

Historically, the limited availability of primary endothelial cells from patients with vascular disorders has hindered the study of the molecular ...

A Tissue Culture Model of Estrogen-producing Primary Bovine Granulosa Cells

En vev kultur modell av Estrogen-produserende primære bovin Granulosa celler

Ovarian granulosa cells (GC) are the major source of estradiol synthesis. Induced by the preovulatory luteinizing hormone (LH) surge, cells of the theca ...

Tissue Collection of Bats for -Omics Analyses and Primary Cell Culture

Tissue innsamling av ball for-omics analyser og primær Cell kultur

As high-throughput sequencing technologies advance, standardized methods for high quality tissue acquisition and preservation allow for the extension of ...