Name: murine endoscopy for in vivo multimodal imaging of carcinogenesis and assessment of intestinal wound healing and inflammation
Uploaded: 2014-08-26T14:00:00
Description: Watch this Scientific Journal Video about Murine Endoscopy for In Vivo Multimodal Imaging of Carcinogenesis and Assessment of Intestinal Wound Healing and Inflammation at JoVE.com

Vi takker Sonja Dufentester og Elke Weber for ekspert teknisk assistanse. Vi takker Faekah Gohar for korrekturlesing manuskriptet og Stefan Brückner for medisinsk informatikk støtte. Dette arbeidet ble støttet av en tverrfaglig stipend fra Else-Kröner-Fresenius-Stiftung (2012_A94). D. Bettenworth ble støttet av en stipendiatstilling fra Det medisinske fakultet, Westfälische Wilhelms-Universität Münster. M. Brückner ble støttet av en &quot;Gerok&quot; rotasjonsposisjonen til Deutsche Forschungsgemeinschaft (DFG SFB1009B8). Vi takker Heike Blum for illustrasjon av musen tegnefilm.

Alle dyreforsøk ble godkjent av Landesamt für Natur, Umwelt und Verbraucherschutz (LANUV) i henhold til tysk Dyrevernloven. 1. Materialer og Forsøksoppsett <ol><li> Dyr omsorg <ol><li> Bruk kvinnelig eller mannlig mus av noen belastning som veier 20 til 25 g og huse dem i henhold til lokale dyr omsorg lovgivning. </li><li> Mate mus med spesielle chow for gnagere og bruke alfalfa-free chow minst tre dager før fluorescens undersøkelser for å minimere Endoluminal auto-fluorescens. </li>...

<ol>
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Epithelial sårheling er en pågående prosess. Kontinuerlig fysiologisk eksfoliering av overflate celler i mage-tarmslimhinnen oppstår som krever hyppig regenerering av epitelceller 16. Følgelig har svekket sårheling en enorm innvirkning på flere sykdommer, inkludert gastrointestinale sår og 17 anastomotic lekkasje 18. Evaluering av molekyl bakgrunn, så vel som potensielle medikamentkandidater for å stimulere epitel helbredelse kan bare ufullstendig utført i cellekultursystemer ...

Dyremodeller har sterkt beriket vår forståelse av mange tarm patologier. Laboratoriet mus (Mus musculus) har dukket opp som en førsteklasses dyremodell i biomedisinsk forskning på grunn av sitt rike genetisk og genomisk informasjon og er lett tilgjengelig i transgene og knockout stammer. I tillegg til å bedre forståelsen sykdom patogenesen, er dyremodeller også viktigere brukt for å teste narkotika kandidater samt prekliniske diagnostiske eller terapeutiske intervensjoner. Men til tross for ulike musemodeller som etterligner menneskelig sykdom, mange diagnostiske og intervensjons alternativer som rutinemessig brukes i pasientbehandling er ikke tilgjengelige for mus. Følgelig til overvåkingsstrategier overvåke løpet av murine sykdom eller effekten av terapeutiske intervensjoner er ofte begrenset til indirekte observasjoner eller post mortem analyser. Mens ikke-invasive prosedyrer finnes for overvåking mus vitalitet som sykdomsaktivitet indekser, quantification av vekttap eller gevinst, blod, urin og avføring analyser, dette er bare indirekte indikatorer, og er partisk av interindividuell variasjon. I tillegg, post mortem analyser hindre langsgående observasjoner ved gjentatte tidspunkter. Sofistikerte imaging teknikker for å overvåke sykdomsaktivitet hos mus har bare nylig blitt introdusert 1,2. Selv om disse bildeteknikker gir mulighet for gjentatte analyser, de bare gir et beskrivende og ofte upresis syn på tarmen, ikke muliggjøre direkte slimhinne visualisering eller tillate diagnostiske eller terapeutiske intervensjoner som biopsi erverv eller aktuell og intramucosal anvendelse av legemiddelkandidater. Nylig har høy oppløsning endoskop systemer for bruk i levende mus blitt utviklet 3,4. For første gang disse endoskopiske teknikker tillater direkte visualisering av endoluminale colonic sykdom patologi som sårtilheling eller tarmbetennelse gi objektiv, real-time status slik at longitudinelle studier i samme dyr ved gjentatte tidspunkter. Bortsett fra slik at gjentatte biopsier i en individuell mus, kan endoskop systemer også brukes til å påvirke en distinkt terapeutisk tumor eller lokalisert inflammasjon ved å tillate direkte anvendelse av et stoff til området av interesse. Videre, som terapeutiske og kontrollstoffer kan leveres direkte til området av interesse, kan dette utføres på samme mus, med unntak av inter-individuell variasjon. Disse systemene har nå vært ansatt for vurdering av tykktarmsbetennelse, sårheling, laparoskopisk leverbiopsi og orthotopic induksjon av leversvulster 8 og tumor utvikling ved hjelp av ulike scoring systemer som murine endoskopisk indeks over kolitt alvorlighetsgrad (MEICS) 5-7. MEICS består av fem parametere for å vurdere betennelse: fortykkelse av tarmveggen, endringer i den vaskulære mønster, nærvær av fibrin, kornete av de mucosal overflate, og avføring konsistens. I denne protokollen beskriver vi bruk av stive endoskopi i murine modeller av intestinal sårtilheling, betennelse og kreft i tykktarmen. Først demonstrerer vi endoskopisk evaluering av sårtilheling og i tykk betennelser samt langsgående vurdering av kolitt aktivitet og studiet av cancerogenesis i murine colon. Utover det beskrivende bruk av murine endoskopi, vi gir detaljerte instruksjoner om bruk av endoskopisk instrumentering for å få biopsier, og aktuell og intramucosal anvendelse av ulike komponenter av interesse (f.eks legemiddelkandidater eller kreftceller). Til slutt, vi demonstrere bruken av murine fluorescens endoskopi, som sysselsetter avanserte molekylære bildeteknikker, i innstillingen av kolorektal tumorer.

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			<td height="21" style="height:21px;width:469px;">Name</td>
			<td style="width:423px;">Company</td>
			<td style="width:320px;">Catalogue Number</td>
			<td style="width:255px;">Comment</td>
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		<tr height="20">
			<td height="20" style="height:20px;width:469px;"></td>
			<td style="width:423px;"></td>
			<td style="width:320px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Reagents</td>
			<td style="width:423px;"></td>
			<td style="width:320px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Alfalfa-free diet</td>
			<td>Harlan Laboritories, Madison, USA</td>
			<td style="width:320px;">2014</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Azoxymethane (AOM)</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">A5486</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Bepanthen eye ointment</td>
			<td style="width:423px;">Bayer, Leverkusen, Germany</td>
			<td style="width:320px;">80469764</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dextran sulphate sodium (DSS)</td>
			<td>TdB Consulatancy, Uppsala, Sweden</td>
			<td>DB001</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:469px;">Eosin</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">E 4382</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:469px;">Ethylenediaminetetraacetic acid (EDTA)&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">E 9884</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Falcon Tube 50ml</td>
			<td>BD Biosciences, Erembodegem, Belgium</td>
			<td style="width:320px;">352070</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Florene 100V/V</td>
			<td>Abbott, Wiesbaden, Germany</td>
			<td style="width:320px;">B506</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fluorescein-Isothiocyanat (FITC)&#8211;dextrane&#160;</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">FD4-250MG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Haematoxylin&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">HHS32-1L</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Isopentane (2- Methylbutane)</td>
			<td style="width:423px;">Sigma - Aldrich, Deisenhofen, Germany</td>
			<td style="width:320px;">M32631-1L</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:469px;">Methylene blue</td>
			<td style="width:423px;">Merck, Darmstadt, Germany</td>
			<td style="width:320px;">1159430025</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">O.C.T. Tissue Tek compound&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:423px;">Sakura, Zoeterwonde, Netherlands</td>
			<td style="width:320px;">4583</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Omnican F - canula</td>
			<td style="width:423px;">Braun, Melsungen, Germany</td>
			<td style="width:320px;">9161502</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Phosphate buffered saline, PBS</td>
			<td style="width:423px;">Lonza, Verviers, Belgium</td>
			<td style="width:320px;">4629</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sodium Chloride 0,9%</td>
			<td>Braun, Melsungen, Germany</td>
			<td style="width:320px;">5/12211095/0411</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Standard diet</td>
			<td>Altromin, Lage, Germany</td>
			<td style="width:320px;">1320</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tissue-Tek Cryomold</td>
			<td>Sakura, Leiden, Netherlands</td>
			<td style="width:320px;">4566</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:469px;">Vitro &#8211; Clud&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:423px;">R. Langenbrinck, Teningen, Germany</td>
			<td style="width:320px;">04-0002&#160;</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td style="width:320px;"></td>
			<td></td>
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		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td style="width:320px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Equipment</td>
			<td></td>
			<td style="width:320px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">AIDA Control</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20 096020</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Bandpass filter</td>
			<td>Semrock, Rochester, USA</td>
			<td style="width:320px;">HC 716/40</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Bandpass filter</td>
			<td>Semrock, Rochester, USA</td>
			<td style="width:320px;">HC 809/81</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Biopsy Forceps, 3 Fr., 28cm</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">61071ZJ</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dell Monitor</td>
			<td>Dell, Frankfurt am Main, Germany</td>
			<td style="width:320px;">U2412Mb</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Examination Sheath, 9 Fr.</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">61029D</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Examination Sheath, 9 Fr.</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">61029C</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fiber Optic Light Cable, 3.5mm</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">69495NL</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fluorescein Blue Filter System</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20100032</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fluorescein Barrier Filter</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20100033</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Foot switch</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20010430</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">HOPKINS Telescope, 1.9mm, Length 10cm</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">1830231</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">SCB D-light P&#160;</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20 133720</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">SCB tricam SL II</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20 2230 20</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tubing set instruments VETPUMP II</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">69811</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tricam PDD PAL</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">20221037</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">UniVet Porta</td>
			<td>Groppler Medizintechnik, Deggendorf, Germany</td>
			<td style="width:320px;">BKGM 0451</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Vetpump 2</td>
			<td>Karl Storz - Endoskope, Tuttlingen, Germany</td>
			<td style="width:320px;">69321620</td>
			<td></td>
		</tr>
	</tbody>
</table>

murine endoscopy for in vivo multimodal imaging of carcinogenesis and assessment of intestinal wound healing and inflammation

Musemodeller er mye brukt til å studere patogenesen av menneskelige sykdommer og å vurdere diagnostiske prosedyrer samt terapeutiske intervensjoner preklinisk. Imidlertid gjelder vurdering av patologiske forandringer krever ofte histologisk analyse, og da utføres ex vivo, nødvendig død av dyret. Derfor i konvensjonelle eksperimentelle innstillinger, intra-individuelle oppfølgingsundersøkelse er sjelden mulig. Dermed utvikling av murine endoskopi i levende mus gjør at etterforskerne for første gang til både direkte visualisere gastrointestinal slimhinne og også gjenta prosedyren for å overvåke endringer. Mange søknader om in vivo murine endoskopi eksisterer, inkludert studere tarmbetennelse eller sårtilheling, skaffe slimhinne biopsier gjentatte ganger, og til lokalt administrere diagnostiske eller terapeutiske midler som bruker miniatyr injeksjonskateter. I det siste har molekylær avbildning utvidet billeddiagnostikk modalities tillater spesifikk påvisning av bestemte mål-molekyler med bestemte photoprobes. I konklusjonen, har murine endoskopi dukket opp som en roman cutting-edge teknologi for diagnostisk eksperimentelle in vivo avbildning og kan ha betydelig innvirkning på preklinisk forskning på ulike felt.

In vivo overvåking av intestinal sårtilheling Under rutinemessig endoskopi, ble det mukosale sår fremkalt mekanisk ved miniatyr biopsitang med en diameter på 3 Fransk (lik 1 mm; figur 1A). Deretter ble sårheling overvåkes av daglige endoskopiske undersøkelser og kvantifisert ved måling av rest såret området ved hjelp av bilderedigeringsprogramvare, for eksempel, ImageJ (Figur 1B). Den enkelte lukking av sår over tid er uttrykt ved kvo...

Watch this Scientific Journal Video about Murine Endoscopy for In Vivo Multimodal Imaging of Carcinogenesis and Assessment of Intestinal Wound Healing and Inflammation at JoVE.com

Murine Endoscopy for In Vivo Multimodal Imaging of Carcinogenesis and Assessment of Intestinal Wound Healing and Inflammation

Murine Endoskopi for<em&gt; In Vivo</em&gt; Multimodal Imaging av Carcinogenesis og vurdering av Intestinal sårheling og Betennelse

Små dyr imaging teknikker tillate serie diagnostiske undersøkelser og terapeutiske intervensjoner i vivo. Nylig, har omfanget av søknader betydelig utvidet og i dag omfatter vurdering av colonic svulst utvikling, sårheling og overvåking av betennelse. Denne protokollen illustrerer disse ulike potensielle anvendelser av murine endoskopi.

Mouse models are widely used to study pathogenesis of human diseases and to evaluate diagnostic procedures as well as therapeutic interventions ...

Research

JoVE Journal

Medicine

Murine Endoscopy for In Vivo Multimodal Imaging of Carcinogenesis and Assessment of Intestinal Wound Healing and Inflammation

In vivo Near Infrared Fluorescence (NIRF) Intravascular Molecular Imaging of Inflammatory Plaque, a Multimodal Approach to Imaging of Atherosclerosis

In vivo Near Infrared Fluorescence NIRF Intravascular Molecular Imaging of Inflammatory Plaque, a Multimodal Approach to Imaging of Atherosclerosis

The vascular response to injury is a well-orchestrated inflammatory response triggered by the accumulation of macrophages within the vessel wall leading ...

Functional Assessment of Intestinal Motility and Gut Wall Inflammation in Rodents: Analyses in a Standardized Model of Intestinal Manipulation

Funksjonell Vurdering av intestinal motilitet og tarmveggen Betennelse i Gnagere: Analyser i en standardisert modell for Intestinal Manipulation

Inflammation of the gastrointestinal tract is a common reason for a variety of human diseases. Animal research models are critical in investigating the ...

Murine Model of Wound Healing

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

A Murine Model of Irreversible and Reversible Unilateral Ureteric Obstruction

En Muse Modell av Irreversible og Reversibel Ensidig ureter Obstruksjon

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. Presented here is a model of obstructive ...

Multimodal Quantitative Phase Imaging with Digital Holographic Microscopy Accurately Assesses Intestinal Inflammation and Epithelial Wound Healing

Multimodal Kvantitativ Phase Imaging med Digital Holografisk Mikros Nøyaktig Vurderer tarmbetennelse og epithelial Wound Healing

The incidence of inflammatory bowel disease, i.e., Crohn&#39;s disease and Ulcerative colitis, has significantly increased over the last decade. The ...

Development of a Direct Pulp-capping Model for the Evaluation of Pulpal Wound Healing and Reparative Dentin Formation in Mice

Utvikling av en direkte Pulp-capping modell for evaluering av pulpal Wound Healing og reparerende Dentin Dannelse i Mus

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is ...

Fluorescence-mediated Tomography for the Detection and Quantification of Macrophage-related Murine Intestinal Inflammation

Fluorescens-mediert tomografi av kvantifisering av Macrophage-relaterte Murine intestinale betennelsen

Murine models of disease are indispensable to scientific research. However, many diagnostic tools such as endoscopy or tomographic imaging are not ...

Microscopy Based Methods for the Assessment of Epithelial Cell Migration During In Vitro Wound Healing

Microscopy Based Methods for the Assessment of Epithelial Cell Migration During  In Vitro Wound Healing

Mikroskopi basert metoder for vurdering av epitelial celle migrasjon ved In Vitro sårheling

Cell migration is a mandatory aspect for wound healing. Creating artificial wounds on research animal models often results in costly and complicated ...

Corneal Epithelial Abrasion with Ocular Burr As a Model for Cornea Wound Healing

Hornhinnen Epithelial slitasje med okulær Burr som modell for hornhinnen sårtilheling

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to ...