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 ekspert teknisk bistand. Vi takker Faekah Gohar for korrekturlæsning af manuskriptet og Stefan Brückner for medicinsk informatik støtte. Dette arbejde blev støttet af en tværfaglig bevilling fra Else-krone-Fresenius-Stiftung (2012_A94). D. Bettenworth blev understøttet af en forskningsstipendiat fra Det Sundhedsvidenskabelige Fakultet, Westfälische Wilhelms-Universität Münster. M. Brückner blev understøttet af en &quot;Gerok&quot; Drejeplan af Deutsche Forschungsgemeinschaft (DFG SFB1009B8). Vi takker Heike Blum til illustration af musen tegneserie.

Alle dyreforsøg blev godkendt af Landesamt für Natur, Umwelt und Verbraucherschutz (LANUV) i henhold til den tyske Animal Protection Law. 1. Materialer og forsøgsopstilling <ol><li> Animal pleje <ol><li> Brug kvindelig eller mandlig mus af en stamme, der vejer 20 til 25 g og huse dem i henhold til lokal pasning af dyr lovgivning. </li><li> Feed mus med særlige chow til gnavere og anvende lucerne-fri chow mindst tre dage før fluorescens prøver at minimere endoluminal auto-fluorescens....

<ol>
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Epitelial sårheling er en løbende proces. Kontinuerlig fysiologiske eksfoliering af overfladeceller inden mavetarmslimhinden opstår kræver hyppig regenerering af epitelceller 16. Derfor forringet sårheling har en enorm betydning på flere sygdomme, herunder gastrointestinale sår og 17 anastomotisk lækage 18. Evaluering af molekylære baggrund samt potentielle lægemiddelkandidater til at stimulere epitelial heling må kun ufuldstændigt udført i cellekultur systemer in vitro<...

Dyremodeller har i høj grad beriget vores forståelse af talrige intestinale patologier. Laboratoriet mus (Mus musculus), har vist sig som en førsteklasses dyremodel i biomedicinsk forskning på grund af sin rigelige genetiske og genomisk information og er let tilgængelige i transgene og knockout-stammer. Ud over at øge forståelsen sygdom patogenese er dyremodeller også vigtigt bruges til at teste lægemiddelkandidater samt prækliniske diagnostiske eller terapeutiske indgreb. Men på trods af de mange forskellige musemodeller efterligner menneskelig sygdom, mange diagnostiske og interventionelle muligheder, der rutinemæssigt anvendes i patientbehandlingen er ikke tilgængelige for mus. Derfor tilsynsforanstaltninger strategier overvåge forløbet af murine sygdom eller effekten af terapeutiske indgreb er ofte begrænset til indirekte observationer eller post mortem analyser. Mens ikke-invasive procedurer for overvågning mus vitalitet lignende sygdom aktivitet indeks, quantification af vægttab eller gevinst, blod, urin og afføring analyser, er disse kun indirekte indikatorer og er forspændt af inter-individuel variation. Derudover post mortem analyser forhindre langsgående observationer ved gentagne tidspunkter. Avancerede billeddiagnostiske teknikker til at overvåge sygdomsaktivitet hos mus først for nylig blevet indført 1,2. Selv om disse billeddiagnostiske teknikker gør det muligt for gentagne analyser, de kun giver en beskrivende og ofte upræcise syn på tarmen, skal du ikke aktivere direkte slimhinde-visualisering eller tillade diagnostiske eller terapeutiske indgreb såsom biopsi erhvervelse eller aktuel og intramucosal anvendelse af lægemiddelkandidater. For nylig har høj opløsning endoskopiske systemer til anvendelse i levende mus blevet udviklet 3,4. For første gang disse endoskopiske teknikker gør det muligt direkte visualisering af endoluminale colon sygdom patologier såsom sårheling eller tarmbetændelse levere objektiv, real-time status tillader langsgående studier i det samme dyr ved gentagne tidspunkter. Bortset fra at tillade gentagne biopsier i en enkelt mus, kan endoskopiske systemer også anvendes til terapeutisk at påvirke en særskilt tumor eller lokaliseret betændelse ved at tillade en direkte anvendelse af et stof til området af interesse. Desuden som terapeutiske og bekæmpelsesmidler kan leveres direkte til området af interesse, kan dette udføres i samme mus, bortset fra inter-individuel variation. Disse systemer er nu blevet anvendt til vurdering af colon inflammation, sårheling, laparoskopiske leverbiopsier og ortotopisk induktion af levertumorer 8 og tumor udvikling ved hjælp af forskellige pointsystemer, såsom det murine endoskopisk indeks colitis sværhedsgrad (MEICS) 5-7. MEICS består af fem parametre til vurdering betændelse: fortykkelse af tyktarmen væggen, ændringer i det vaskulære mønster, tilstedeværelse af fibrin, granulering af MUCOSal overflade, og afføring konsistens. I denne protokol vi beskrive brugen af ​​stive endoskopi i murine modeller af tarm sårheling, inflammation og tyktarmskræft. Først, vi demonstrere endoskopisk evaluering af sårheling og colon inflammation samt langsgående vurdering af colitis aktivitet og studiet af cancerogenesis i den murine kolon. Ud over den beskrivende brug af murine endoskopi, vi give detaljerede anvisninger om brugen af endoskopisk instrumentering at opnå biopsier, og den aktuelle og intramucosal anvendelse af forskellige komponenter af interesse (f.eks lægemiddelkandidater eller tumorceller). Endelig vil vi demonstrere brugen af ​​murine fluorescens endoskopi, som beskæftiger sofistikerede molekylære billeddiagnostiske teknikker i fastsættelsen af ​​kolorektale tumorer.

<table border="0" cellpadding="0" cellspacing="0" width="1467">
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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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			<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>
		</tr>
		<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 almindeligt anvendt til at studere patogenesen af ​​sygdomme, samt at evaluere diagnostiske procedurer samt terapeutiske interventioner præklinisk. Men gyldig vurdering af patologiske forandringer ofte kræver histologisk analyse, og når de udføres ex vivo, nødvendiggør dyrets død. Derfor vil der i konventionelle eksperimentelle indstillinger, er sjældent mulig intraindividuelle opfølgende undersøgelser. Således udvikling af muse endoskopi i levende mus giver efterforskere for første gang til både direkte visualisere den gastrointestinale mucosa og også gentage proceduren for at kontrollere for ændringer. Talrige anvendelser til in vivo muse endoskopi findes, herunder at studere tarmbetændelse eller sårheling, opnåelse af slimhinder biopsier gentagne gange, og til lokalt at administrere diagnostiske eller terapeutiske midler ved hjælp af miniature injektion katetre. Senest har molekylær billeddannelse udvidet diagnostisk billeddannelse modalities muliggør specifik påvisning af forskellige målmolekyler med særlige fotoprober. Sammenfattende har muse endoskopi opstået som en ny banebrydende teknologi til diagnostisk eksperimentel in vivo billeddannelse og kan få alvorlige konsekvenser på præklinisk forskning på forskellige områder.

In vivo overvågning af intestinal sårheling Under rutinemæssig endoskopi blev slimhindeirriterende sår induceret mekanisk ved miniature biopsitænger med en diameter på 3 fransk (lig med 1 mm; figur 1a). Efterfølgende blev sårheling overvåget af daglige endoskopiske undersøgelser og kvantificeres ved måling af den resterende sår området ved hjælp af et billedredigeringsprogram, f.eks ImageJ (figur 1B). Den enkelte sår lukning over ...

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

Murin Endoskopi til<em&gt; In Vivo</em&gt; Multimodal Imaging af Carcinogenesis og vurdering af Intestinal sårheling og betændelse

Små dyr billeddiagnostiske teknikker tillade serielle diagnostiske undersøgelser og terapeutiske interventioner in vivo. For nylig, er omfanget af ansøgninger betydeligt udvidet og omfatter i dag vurdering af colon tumor udvikling, sårheling og overvågning af betændelse. Denne protokol illustrerer disse forskellige mulige anvendelser af 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

Functional Assessment of tarmmotilitet og tarmvæggen Inflammation i Gnavere: Analyser i en standardiseret model af 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

Musemodel af Sårheling

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 murin model af Irreversible og reversibel Ensidig ureteric Obstruktion

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 Microscopy Præcist Vurderer tarmbetændelse og Epithelial Sårheling

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

Udvikling af en Direct Pulp-capping Model for Vurdering af pulpa Sårheling og Reparative Dentin Formation 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-medieret tomografi til påvisning og kvantificering af makrofag-relaterede Murine tarmbetændelse

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 baseret metoder til vurdering af epitelcelle Migration under 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

Hornhindens epitel slid med okulær Burr som en Model for hornhinden sårheling

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 ...