Name: primary orthotopic glioma xenografts recapitulate infiltrative growth and isocitrate dehydrogenase i mutation
Uploaded: 2014-01-14T13:45:00
Description: Watch this Scientific Journal Video about Primary Orthotopic Glioma Xenografts Recapitulate Infiltrative Growth and Isocitrate Dehydrogenase I Mutation at JoVE.com

Vi står især i gæld til patienter på Vanderbilt University Medical Center, der leverede uvurderligt forskningsmateriale til Molecular Neurosurgical Tissue Bank. Vi takker dem, der etablerede og vedligeholder Tissue Bank, Reid C. Thompson MD (principal investigator), Cherryl Kinnard RN (forskningssygeplejerske) og Larry A. Pierce MS (manager). Histologiske tjenester blev delvist udført af Vanderbilt University Medical Center (VUMC) Translational Pathology Shared Resource (støttet af tildeling 5P30 CA068485 til Vanderbilt-Ingram Cancer Center). Dette arbejde blev støttet af tilskud til MKC fra NINDS (1R21NS070139), Burroughs Wellcome Fund og VMC udviklingsmidler. MKC er støttet af Department of Veterans Anliggender, Veterans Health Administration, Office of Research and Development, Biomedical Laboratory Research and Development gennem tilskud 1 I01 BX000744-01. Indholdet repræsenterer ikke synspunkter Department of Veterans Anliggender eller den amerikanske regering.

1. Forberedelse af tumorcelle suspension
Bemærk: Der kræves passende institutionelle godkendelser til brug af patientmateriale og -dyr for at etablere og vedligeholde primære ortopiske gliom xenografter. På Vanderbilt University Medical Center indsamles resected tumormateriale, der overstiger det, der kræves til diagnostiske formål, med patientens samtykke til et forskningsvævslager. Prøverne er mærket med et randomiseret 5-cifret REDcap-databasenummer, og alle patient...

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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=Cyclopamine-mediated+hedgehog+pathway+inhibition+depletes+stem-like+cancer+cells+in+glioblastoma.">Cyclopamine-mediated hedgehog pathway inhibition depletes stem-like cancer cells in glioblastoma.</a> Stem Cells. 25, 2524-2533 (2007).</li><li>Ehtesham, 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=Ligand-dependent+activation+of+the+hedgehog+pathway+in+glioma+progenitor+cells.">Ligand-dependent activation of the hedgehog pathway in glioma progenitor cells.</a> Oncogene. 26, 5752-5761 (2007).</li><li>Kelly, J. 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Dyrkede cellelinjer, xenografts og genetisk manipulerede mus er de mest almindelige metoder til modellering af gliomer, og der er forskellige fordele og begrænsninger for hvert modelsystem3,13,14. Relevante fordele ved primære ortopiske glioma xenografts omfatter infiltrerende vækst, der kendetegner diffuse gliomer og fastholdelse af genetiske ændringer og vigtige signalmekanismer, der kan være overordentlig vanskeligt at opretholde i dyrkede gliomceller. For eksempel kan isocitrat dehydrogenase ...

Maligne gliomer er primære glia tumorer i centralnervesystemet, der forekommer i hjernen og lejlighedsvis rygmarven. Gliomer er klassificeret af Verdenssundhedsorganisationen (WHO) i henhold til histologiske lighed med astrocytter, oligodendrocytter eller ependymale celler og derefter numerisk klassificeret (I til IV) for patologiske træk ved malignitet. De mest almindelige histologiske undertyper er astrocytomas, oligodendrogliomas og blandede oligoastrocytomas. Maligne gliomer, der omfatter WHO-kvaliteter II til IV, er kendetegnet ved invasiv vækst og genstridige til nuværende behandlingsformer. Hvert år i USA, ca 15.750 personer er diagnosticeret med en ondartet gliom og en anslået 12.740 patienter bukke under for denne sygdom. Disse statistikker fremhæver den særligt dødelige karakter af ondartede gliomer og det vigtige behov for øget terapeutisk effekt.
Kræft modeller er afgørende for at undersøge tumor biologi og behandlinger. Humane kræftcellelinjer udgør et vigtigt første skridt for in vitro-manipulationer og in vivo xenograftingstudier (sekundære xenografts)1. Men, standard kræft cellekulturer gennemgå fænotypiske og genotypiske transformation2-4, der ikke kan genoprettes i sekundære xenografts5. Desuden kan genetiske ændringer såsom isocitrat dehydrogenase (IDH) mutationer6, forskellige stamcellepopulationer7 og afhængighed af centrale signalveje8 gå tabt i kræftcellekulturer. Genomiske profiler kan opretholdes i bedre grad i kræft sfære kultur, men stadig undlader at spejle fuldt ud genotype af primære tumorer2,3. Direkte ortopisk transplantation negerer indflydelsen fra in vitro-kulturen, giver et ordentligt mikromiljø og bevarer integriteten af tumorinitierende celler9,10. Derfor udgør primære xenografter en kraftfuld og relevant præklinisk model til streng test af målrettede midler for at hjælpe med at designe fremtidige kliniske forsøg5,11,12.

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			Phosphate buffered saline
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			<td style="width:142px;">
			Life Technologies
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			<td style="width:140px;">
			14040-133
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			<td style="width:211px;">
			Papain dissociation system
			</td>
			<td style="width:142px;">
			Worthington Biochemical Corp.
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			<td style="width:140px;">
			LK003150
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			<td style="width:97px;">
			
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			<td style="width:211px;">
			Trypan blue solution 0.4%
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			Life Technologies
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			15250061
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			Ketamine HCl
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			Obtained from institutional pharmacy or local veterinary supply company
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			Xylazine HCl
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			Ketoprofen
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			Ophthalmic ointment
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			Povidone-iodine
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			<td style="width:142px;">
			Fisher Scientific
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			190061617
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			<td style="width:97px;">
			
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			Cryopreservation medium and proliferation supplement
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			StemCell Technologies
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			05751
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			0.2% Heparin sodium salt in PBS
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			<td style="width:142px;">
			StemCell Technologies
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			<td style="width:140px;">
			07980
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			Penicillin-streptomycin
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			<td style="width:142px;">
			Life Technologies
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			15140-122
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			Dimethyl sulfoxide
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			Sigma-Aldrich
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			D6250-5X10ML
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			NOD.Cg-Prkdcscid I/2rgtm1Wjl/SzJ mice
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			The Jackson Laboratory
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			005557
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			NSG mice
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			Anti-human vimentin antibody
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			Dako
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			M7020
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			Use 1:200 to 1:800
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			Anti-human IDH1 R132H antibody
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			<td style="width:142px;">
			Dianova
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			<td style="width:140px;">
			DIA-H09
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			<td style="width:97px;">
			Use 1:100 to 1:400
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	</tbody>
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			Comments
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			Centrifuge with swinging bucket rotor
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			Pipetter with dispensing speed control
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			Disposable hemocytometer
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			22-600-100
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			<td style="width:210px;">
			Sterile surgical gloves
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			11-388128
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			<td style="width:97px;">
			
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			Disposable gown
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			18-567
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			Surgical mask
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			19-120-1256
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			Tuberculin syringe
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			<td style="width:140px;">
			BD
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			305620
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			Alcohol pads
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			22-246-073
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			Portable electronic scale
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			<td style="width:140px;">
			Fisher Scientific
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			<td style="width:143px;">
			01-919-33
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			Zoom stereomicroscope
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			Surgical clipper
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			<td style="width:140px;">
			Stoelting
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			<td style="width:143px;">
			51465
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			<td style="width:97px;">
			
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			Scalpel handle
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			<td style="width:140px;">
			Fine Science Tools
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			<td style="width:143px;">
			10003-12
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			Scalpel blades, #10
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			<td style="width:210px;">
			Stereotaxic instrument
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			<td style="width:140px;">
			Stoelting
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			<td style="width:143px;">
			51730
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			<td style="width:97px;">
			
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			<td style="width:210px;">
			High-speed drill
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			<td style="width:140px;">
			Stoelting
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			<td style="width:143px;">
			51449
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">Drill bit, 0.6 mm</td>
			<td style="width:140px;">Stoelting</td>
			<td style="width:143px;">514552</td>
			<td style="width:97px;"></td>
		</tr>
		<tr>
			<td style="width:210px;">
			Hamilton syringe
			</td>
			<td style="width:140px;">
			Hamilton
			</td>
			<td style="width:143px;">
			80336
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Autoclip, 9 mm
			</td>
			<td style="width:140px;">
			BD
			</td>
			<td style="width:143px;">
			427630
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Circulating water warming pad
			</td>
			<td style="width:140px;">
			Kent Scientific
			</td>
			<td style="width:143px;">
			TP-700
			TP-1215EA
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Hot bead dry sterilizer
			</td>
			<td style="width:140px;">
			Kent Scientific
			</td>
			<td style="width:143px;">
			INS300850
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Surgical scissors
			</td>
			<td style="width:140px;">
			Fine Science Tools
			</td>
			<td style="width:143px;">
			14101-14
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Fine scissors
			</td>
			<td style="width:140px;">
			Fine Science Tools
			</td>
			<td style="width:143px;">
			14094-11
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Spring scissors
			</td>
			<td style="width:140px;">
			Fine Science Tools
			</td>
			<td style="width:143px;">
			15018-10
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Dumont forceps
			</td>
			<td style="width:140px;">
			Fine Science Tools
			</td>
			<td style="width:143px;">
			11251-30
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Semimicro spatulas
			</td>
			<td style="width:140px;">
			Fisher Scientific
			</td>
			<td style="width:143px;">
			14374
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Mouse brain slicer matrix
			</td>
			<td style="width:140px;">
			Zivic Instruments
			</td>
			<td style="width:143px;">
			BSMAS002-1
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
		<tr>
			<td style="width:210px;">
			Cryogenic storage vials
			</td>
			<td style="width:140px;">
			Fisher Scientific
			</td>
			<td style="width:143px;">
			12-567-501
			</td>
			<td style="width:97px;">
			
			</td>
		</tr>
	</tbody>
</table>

primary orthotopic glioma xenografts recapitulate infiltrative growth and isocitrate dehydrogenase i mutation

Maligne gliomer udgør en heterogen gruppe af stærkt infiltrerende glial neoplasmer med forskellige kliniske og molekylære træk. Primære ortopiske xenografter opsummerer de histopatologiske og molekylære træk ved ondartede gliomundertyper i prækliniske dyremodeller. Til model WHO kvaliteter III og IV ondartede gliomer i transplantation assays, menneskelige tumorceller er xenografted i en orthotopic site, hjernen, af immunkompromitterede mus. I modsætning til sekundære xenografts, der bruger dyrkede tumorceller, er humane glioma celler dissociated fra resected prøver og transplanteret uden forudgående passage i vævskultur til at generere primære xenografts. Proceduren i denne rapport detaljer tumor prøve forberedelse, intrakraniel transplantation i immunkompromitterede mus, overvågning for tumor engraftment og tumor høst for efterfølgende passage i recipient dyr eller analyse. Tumor celle forberedelse kræver 2 timer og kirurgisk procedure kræver 20 min / dyr.

Dissociated glioma celler transplanteres direkte ind i hjernen hos immunkompromitterede mus for at opnå primære ortopiske xenograft linjer. Hver tumorprøve tildeles et randomiseret tal før transplantation som en del af afidentifikationsprocessen for at fjerne beskyttede sundhedsoplysninger. Vi bruger et 5-cifret REDcap-databasenummer til dette formål. Figur 1 illustrerer processen og nomenklaturen for etablering af en xenograftlinje fra et glioblastoma (GBM 17182) med isocitratdehydrogenase 1 (IDH1)...

Watch this Scientific Journal Video about Primary Orthotopic Glioma Xenografts Recapitulate Infiltrative Growth and Isocitrate Dehydrogenase I Mutation at JoVE.com

Primary Orthotopic Glioma Xenografts Recapitulate Infiltrative Growth and Isocitrate Dehydrogenase I Mutation

Maligne gliomer udgør en heterogen gruppe af stærkt infiltrerende glial neoplasmer med forskellige kliniske og molekylære træk. Primære ortopiske xenografter opsummerer de histopatologiske og molekylære træk ved ondartede gliomundertyper i prækliniske dyremodeller.

Primær ortopisk Glioma Xenografts Rekapitulere infiltrerende vækst og isocitrate Dehydrogenase I Mutation

Malignant gliomas constitute a heterogeneous group of highly infiltrative glial neoplasms with distinct clinical and molecular features. Primary ...

Research

JoVE Journal

Medicine

An Orthotopic Model of Murine Bladder Cancer

In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and ...

Orthotopic Small Bowel Transplantation in Rats

Orthotopisk tyndtarmstransplantation i rotter

Small  bowel transplantation has become an accepted clinical option for patients with  short gut syndrome and failure of parenteral nutrition ...

Orthotopic Liver Transplantation in Rats

Ortotopisk levertransplantation i rotter

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat ...

Imaging Glioma Initiation In Vivo Through a Polished and Reinforced Thin-skull Cranial Window

Imaging Glioma Initiation In Vivo Through a Polished and Reinforced Thin-skull Cranial Window

Imaging gliom Indledning<em&gt; In Vivo</em&gt; Gennem en poleret og Reinforced Thin-kranium Cranial Window

Glioma is the one of the most lethal forms of human cancer. The most effective glioma therapy to date-surgery followed by radiation treatment-offers ...

Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

Føtal Ekkokardiografi og Pulserende-bølge Doppler Ultrasound i en kanin Model of Intrauterin vækst Begrænsning

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ...

Bioluminescent Orthotopic Model of Pancreatic Cancer Progression

Bioluminescent ortotopisk Model af kræft i bugspytkirtlen Progression

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%.  New therapeutic options are critically needed and depend on improved ...

Generation of Subcutaneous and Intrahepatic Human Hepatocellular Carcinoma Xenografts in Immunodeficient Mice

Generering af Subkutan og Intrahepatisk Menneskelige hepatocellulært carcinom xenotransplantater i immunsvækkede mus

In  vivo experimental models of hepatocellular carcinoma (HCC) that recapitulate  the human disease provide a valuable platform for research into disease  ...

Minimally Invasive Establishment of Murine Orthotopic Bladder Xenografts

Minimalt invasiv Etablering af murine ortotopisk Blære xenotransplantater

Orthotopic bladder cancer xenografts are the gold standard to study molecular cellular manipulations and new therapeutic agents&#160;in vivo. Suitable ...

Orthotopic Injection of Breast Cancer Cells into the Mammary Fat Pad of Mice to Study Tumor Growth.

Ortotopisk Injektion af brystkræft celler i brystfedtpuden af ​​mus til at studere tumorvækst.

Breast cancer growth can be studied in mice using a plethora of models. Genetic manipulation of breast cancer cells may provide insights into the ...

Creating Anatomically Accurate and Reproducible Intracranial Xenografts of Human Brain Tumors

Oprettelse Anatomisk nøjagtig og reproducerbar Intrakraniel xenotransplantater of Human hjernetumorer

Orthotopic tumor models are currently the best way to study the characteristics of a tumor type, with and without intervention, in the context of a live ...