The evolution of the device started with somewhat different cutting prototype demonstrated at the Imperial Cancer Research Fund (ICRF) mouse club by Bass Hassan, following this there were several discussions with Rob Rudling and Nikki Mandir in the containment facility at ICRF Clare Hall. Jo Kitau of the mechanical workshop at ICRF constructed the initial versions and Andy Lucas designed and perfected the final model in the CRUK workshop in Oxford.

Die verschiedenen experimentellen Verfahren und die humane Sterbehilfe werden von der gastgebenden Einrichtungen Ethikkommission und von den zuständigen Aufsichtsbehörden genehmigt. Sterbehilfe ist in der Regel unter Verwendung von CO 2 Ersticken durch Genickbruch folgte erreicht. 1. Vorrichtung <ol><li> Maschine der Hauptteil des Bodens der Vorrichtung von aus einem festen Block aus hochdichtem Acetyl-Harz, wie in diesem Manuskript beschrieben. An jedem Ende zu erstellen 3 mm hoch erhöhten Streifen, und ein Teil Maschine aus diesen 2,5 mm Haine, um die Stangen zu nehmen (siehe Abbildung 2 und Abbildung 3). </li><li> Machen Stangen aus einem 2,4 mm Durchmesser aus Edelstahl mit abgerundeten Enden. HINWEIS: Der Hauptrahmen des Deckels wurde aus Duralumin Platte gefräst. Eine begrenzte Anzahl von fertigen Vorrichtungen sind von Dr. Goodlad erhältlich. </li></ol> 2. Verwendung des Geräts <ol><li> Sezieren den Dünndarm und Dickdarm aus der Bauchhöhle mit einer Schereund behandschuhten Fingern. Entfernen Sie das Mesenterium, indem Sie sie mit einer gebogenen Pinzette und leichtes Ziehen sie weg aus dem Darm. </li><li> Spülen Sie den Darm mit kalter phosphatgepufferter Kochsalzlösung (PBS, pH 7,4) mit einer Pipettenspitze Gilson-Typ, der sich an seinem breiteren Ende geschnitten worden war, so dass er auf ein 10 oder 20 ml Luer-Fitting-Kunststoffspritze passen. </li><li> Legen Sie den Dünndarm (SB) auf einem Blatt Papier Handtuch und teilen sich in drei gleiche Abschnitte in der Länge (proximalen, mittleren und distalen - SB1, SB2 und SB3) mit einer Schere. Entfernen Sie den Blinddarm und legen Sie den Doppelpunkt. </li><li> Machen Sie mehrere sehr kleine Schnitte in das Darmsegmente mit einer Schere, damit die Flüssigkeit entweichen und legen Sie ein Blatt Papier Handtuch über den Darm und sanft laufen mit dem Finger über die Vorbereitungen zu verdrängen die Flüssigkeit und trocken tupfen. </li><li> Heben Sie die Abschnitte und legen Sie die Edelstahlstäbe, die zuvor durch Eintauchen in ein Becherglas mit phosphatgepufferter Kochsalzlösung benetzt wurden. HINWEIS: Der Darm kann sein,umgewandelt, wenn dies gewünscht (innen nach außen gedreht). </li><li> Beschriften Sie ein Stück Karton oder Filterpapier auf Maß geschnitten, um in die Basis des Geräts passen. Bleistift ist am besten, da sie unbeeinflusst von nahezu allen Lösungen ist. Autopsy Datum, experimentellen Code und Tieridentifikationsnummer vorgeschlagen. </li><li> Legen Sie die Karte oder Filterpapier in die Basis des Geräts. Setzen Sie die Stangen und Darm in die Schlitze in der Basis des Gerätes. Sicherzustellen, dass das proximale Ende der Segmente in einer standardisierten Weise positioniert. Die proximalen Enden sollten in der Nähe der Karte Etiketten sein. </li><li> Legen Sie das Oberteil des Gerätes über die Basis. Mit den abgewinkelten Bars, ein Skalpell zu führen, um die Abschnitte längs geschnitten. HINWEIS: Es ist hilfreich, wenn das Gewebe vorsichtig und mit einem Finger gehalten werden, um sicherzustellen, dass sie nicht mit dem Messer zu bewegen. </li><li> Entfernen Sie das Oberteil des Gerätes. Das Filterpapier und Gewebe vorsichtig entfernen (immer noch auf ihre Stäbe). HINWEIS: Ein Stück steifen Karte unter dem fil platziertter Papier Hebehilfe. </li><li> Leicht feucht (mit einer behandschuhten Fingerspitze in PBS getaucht) der Segmente. Rollen Sie die Stange, von Seite zu Seite zu öffnen, die gut und breitete sie flach. Das Gewebe wird dann auf das Filterpapier haftet. </li><li> Visuell untersuchen die Vorbereitungen für alle makroskopischen Veränderungen. </li><li> Übertragen des Gewebes auf das Filterpapier haftet in eine flache Wanne (oder Sandwichbox) enthaltenden Fixiermittels. HINWEIS: Nachdem das Gewebe hält sich an das Filterpapier oder auf die Karte wird es bleiben, fest angebracht. Eine Vielzahl von Fixierungsmitteln kann verwendet werden, um die Herstellung zu bewahren. Es wird empfohlen, Carnoy-Flüssigkeit sollte für die Untersuchung von Darmzellwucherungen 6, die die ideale Fixierung für den besten Zellproliferation Verfahren verwendet wird; jedoch Formalin oder viele andere Fixierungsmittel verwendet werden, wenn andere Endpunkte erforderlich sind. </li><li> Nach der Fixierung (in der Regel 3 h), übertragen Sie die Präparate in 70% Ethanol. Die Proben können dann gelagert, bis sie benötigt werden. Die Vorbereitungen cen face eine untersucht und / oder für die Herstellung von histologischen Proben für H &amp; E oder andere histologische Färbetechniken Immunhistochemie und / oder in situ-Hybridisierung verwendet werden. HINWEIS: Längerer Fixierung kann immunhistochemische Reaktivität und / oder in situ Hybridisierung Reaktivität zu reduzieren. Polypropylen-Sandwich-Boxen sind ideal für die Speicherung einer großen Anzahl der Zubereitungen. </li><li> Bei Bedarf die fixierten Gewebe aus den verschiedenen Darmabschnitten kann in &quot;Rouladen&quot; gemacht und mit dem gesamten Bereich der histologischen Methoden 6 einschließlich der oben beschriebenen gefärbt werden. Um einen &quot;Swiss roll&quot; machen die Darmsegmente werden aus dem Filterpapier gerollt um einen Cocktail-Stick oder einem Zahnstocher, mit einer entomologischen Stift gesichert und für Wachs Einbetten 6 verarbeitet entfernt. </li><li> Falls erforderlich, beflecken den Großgewebe mit Methylenblau zu abweichenden Kryptenherden 7 visualisieren. </li><li> Note die festen Zubereitungen, die aufFreizeit. </li></ol>

<ol>
	<li>Goodlad, R. A., Alison, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=+Cancer+Handbook."> Cancer Handbook.</a> , 243-262 (2007).</li><li>Ward, J. M., Treuting, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+intestinal+epithelial+carcinogenesis:+pathology+and+preclinical+models.">Rodent intestinal epithelial carcinogenesis: pathology and preclinical models.</a> Toxicol Pathol. 42, 148-161 (2014).</li><li>Sundberg, J. P., Hogenesch, H., Nikitin, A. Y., Treuting, P. M., Ward, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Training+mouse+pathologists:+ten+years+of+workshops+on+the+Pathology+of+Mouse+Models+of+Human+Disease.">Training mouse pathologists: ten years of workshops on the Pathology of Mouse Models of Human Disease.</a> Toxicol Pathol. 40, 823-825 (2012).</li><li>Wasan, H. S., Novelli, M., Bee, J., Bodmer, W. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dietary+fat+influences+on+polyp+phenotype+in+multiple+intestinal+neoplasia+mice.">Dietary fat influences on polyp phenotype in multiple intestinal neoplasia mice.</a> Proc Natl Acade Sci USA. 94, 3308-3313 (1997).</li><li>Rudling, R., Hassan, A. B., Kitau, J., Mandir, N., Goodlad, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+device+to+rapidly+prepare+whole+mounts+of+murine+intestine.">A simple device to rapidly prepare whole mounts of murine intestine.</a> Cell Prolif. 39, 415-420 (2006).</li><li>Alferez, D., Goodlad, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=To+best+measure+cell+proliferation+in+samples+from+the+intestine.">To best measure cell proliferation in samples from the intestine.</a> Cell Prolif. 40, 231-240 (2007).</li><li>Park, H. S., Goodlad, R. A., Wright, N. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+incidence+of+aberrant+crypt+foci+and+colonic+carcinoma+in+dimethylhydrazine-treated+rats+varies+in+a+site-specific+manner+and+depends+on+tumor+histology.">The incidence of aberrant crypt foci and colonic carcinoma in dimethylhydrazine-treated rats varies in a site-specific manner and depends on tumor histology.</a> Cancer Res. 57, 4507-4510 (1997).</li><li>Park, H. S., 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=Effects+of+epidermal+growth+factor+and+dimethylhydrazine+on+crypt+size,+cell+proliferation,+and+crypt+fission+in+the+rat+colon.+Cell+proliferation+and+crypt+fission+are+controlled+independently.">Effects of epidermal growth factor and dimethylhydrazine on crypt size, cell proliferation, and crypt fission in the rat colon. Cell proliferation and crypt fission are controlled independently.</a> Am J Pathol. 151, 843-852 (1997).</li><li>Femia, A. P., Dolara, P., Caderni, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mucin-depleted+foci+(MDF)+in+the+colon+of+rats+treated+with+azoxymethane+(AOM)+are+useful+biomarkers+for+colon+carcinogenesis.">Mucin-depleted foci (MDF) in the colon of rats treated with azoxymethane (AOM) are useful biomarkers for colon carcinogenesis.</a> Carcinogenesis. 25, 277-281 (2004).</li><li>Ruehl-Fehlert, 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=Revised+guides+for+organ+sampling+and+trimming+in+rats+and+mice--part+1.">Revised guides for organ sampling and trimming in rats and mice--part 1.</a> Exp Toxicol Pathol. 55, 91-106 (2003).</li><li>Goodlad, R. 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=Inhibiting+vascular+endothelial+growth+factor+receptor-2+signaling+reduces+tumor+burden+in+the+ApcMin/++mouse+model+of+early+intestinal+cancer.">Inhibiting vascular endothelial growth factor receptor-2 signaling reduces tumor burden in the ApcMin/+ mouse model of early intestinal cancer.</a> Carcinogenesis. 27, 2133-2139 (2006).</li><li>Mandir, N., Englyst, H., Goodlad, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Resistant+carbohydrates+stimulate+cell+proliferation+and+crypt+fission+in+wild-type+mice+and+in+the+Apc(Min/+)+mouse+model+of+intestinal+cancer,+association+with+enhanced+polyp+development.">Resistant carbohydrates stimulate cell proliferation and crypt fission in wild-type mice and in the Apc(Min/+) mouse model of intestinal cancer, association with enhanced polyp development.</a> Br J Nutr. 100, 711-721 (2008).</li></ol>

The authors have nothing to disclose.

Das Gerät erheblich beschleunigt den Prozess, so daß zwei Bediener kann vollständig eine Maus Autopsie innerhalb von 6 min. Dazu gehört auch eine Sichtkontrolle auf offensichtliche makroskopischen Veränderungen und einem Gewicht von und zur Festsetzung der wichtigsten Organe. Es ermöglicht auch die Betreiber zu spülen, trocknen und wiegen den Magen, Blinddarm, Dünndarm und Dickdarm und bereiten die gesamte Reittiere. Der abgeflachte Gewebe sehr schnell behoben werden und somit zu vermeiden Abbau Artefakte. <...

Die Maus und die genetisch veränderte Maus 1-3 sind eine unschätzbare Modell für die Untersuchung von verschiedenen Pathologien, insbesondere Krebs und seine Beziehung zu Entzündungen im Magen-Darm-Trakt. Dies erfordert oft die Herstellung von Gesamtpräparate des Dünndarms und / oder Dickdarm. Die Herstellung von Gesamtpräparate mit einer Genauigkeit Paar versetzter Schere gemacht werden, aber es kann etwa 20 min pro Maus 4 nehmen. Dies ist nicht sehr praktisch für Vergleichsstudien, die eine erhebliche Menge an Mäusen benötigen (etwa 10 pro Gruppe für die statistische Analyse). Darüber hinaus ist die Länge der Zeit erhöht das Risiko von Gewebeabbau. Ein Versuch, dieses Problem zu lindern führte zu der Studie von verschiedenen Schneidvorbereitung Hilfen. Die erste auf einer Reihe von Metallplatten basiert, mit Schneidhainen, aber die Platten verdeckt das Gewebe und die Schneid war daher schwierig zu kontrollieren. Ein Konventionel sechseckigen Bleistift schließlich Inspiration zur Verfügung gestellt für die Schneideinrichtung, wie wir könnten dann sehen, wie eine halbe Stiftform wäre eine gute Führung für ein Skalpell (Abbildung 1) zu machen. Dies führte zu einem Design für einen Rahmen mit vier Schneidführungen 5. Die Wirksamkeit dieser Vorrichtung war dramatisch wie die Präparate in Figur 4 veranschaulichen den Unterschied zwischen der Scheren Zubereitung und der Vorrichtung Zubereitung. Die Oberseiten der Originalgeräte wurde aus mehreren Metallstücke, aber späteren aufgebaut, und das aktuelle Modell wurde aus einem massiven Stück Duraluminium gefertigt. Die Basis wurde von höchster Qualität mit hoher Dichte Harz bearbeitet. Das Gerät hat sich als von großem Nutzen für die Herstellung von ganzen Bergen von Mausdarm, da es erheblich beschleunigt und verbessert die Qualität und Konsistenz der Präparate, die eine breite Palette von Anwendungen.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Delrin</td>
			<td>Ryan Plastics, Earls Barton, UK</td>
			<td>N/A</td>
			<td>High-density acetal resin similar material would suffice</td>
		</tr>
		<tr>
			<td>Duralumin plate</td>
			<td>Metal Supplies Ltd, Park Road,Dukinfield. SK16 5LP UK</td>
			<td>N/A</td>
			<td>Aluminium allow dating back to 1909 so alterative suppliers are available&#160;</td>
		</tr>
		<tr>
			<td>Finished device(s) ready for use</td>
			<td>Contact Dr Goodlad&#160; r.goodlad@imperial.ac.uk</td>
			<td>N/A</td>
			<td>Contact&#160; r.goodlad@imperial.ac.uk&#160; for supply details</td>
		</tr>
	</tbody>
</table>

a simple device to rapidly prepare whole mounts of the mouse intestine

Herstellung Gesamtpräparate der Maus-Dünndarm und Dickdarm zur nachfolgenden Analyse oder Quantifizierung kann zeitaufwendig und schwierig sein. Wir beschreiben die Verwendung einer einfachen Vorrichtung zu schneiden und zu &#39;roll&#39; Maus Darm ganze Berg Zubereitungen von Superior und gleichbleibende Qualität zu dem, was von Hand erreicht werden kann schnell vorzubereiten. Die Vorrichtung umfasst eine Basis, die 4 Edelstahlstangen und eine Oberseite, die eine Schneidführung wirkt hält. Die Stäbe werden in das Lumen des Dünndarms [gedrittelt] und den Dickdarm eingelegt. Die Stäbe und die Proben werden dann in einem Stück Filterpapier oder einer Karte in die Halteschlitze in der Basis der Vorrichtung platziert. Die Oberseite des Gerätes wird dann positioniert und dient als Schnittführung. Die beiden Abwinkelungen in der Mitte des Oberteils werden verwendet, um ein Messer oder Skalpell führen und schneiden den Darm in Längsrichtung auf der Oberseite der Stangen. Sobald die Darmabschnitte geschnitten worden sind, wird der Kopf abgetrennt und die Karte, tissue und Stangen leicht aus der Vorrichtung entnommen und auf die Bank gelegt. Die Stäbe werden dann sanft zur Seite gerollt, um zu glätten und kleben die Darmsegmente auf die darunterliegende Stück Filterpapier oder Karte. Die endgültige Präparation kann dann untersucht oder befestigt werden und für eine spätere Analyse gelagert. Die Zubereitungen sind von unschätzbarem Wert für die Untersuchung von Veränderungen der intestinalen normalen oder genetisch modifizierte Mausmodellen. Die Vorbereitungen für die Untersuchung und Quantifizierung der Auswirkungen der Entzündung (Colitis), Schäden, Krebsvorstufen verwendet (anomale Kryptenherden (ACFs) und Mucin erschöpft Brennpunkten (MDF)) und Polypen oder Tumoren.

Polypen sind einfach, unter einem Stereomikroskop mit einer Kaltlichtquelle zu identifizieren, um die Seite zu beleuchten das Gewebe (Abbildung 4). Es ist hilfreich, um die Karte mit Bleistiftmarkierungen kennzeichnen, wenn ein Polyp, damit Scoring kann später diskutiert und verschiedene Betreiber kann Einigung beobachtet. Das Gewebe kann auch Groß sein befleckt [mit Methylenblau] zu identifizieren anomale Kryptenherden 8 oder Mucin erschöpft Brennpunkten (MDF) 9 durch Beleuchtu...

Watch this Scientific Journal Video about A Simple Device to Rapidly Prepare Whole Mounts of the Mouse Intestine at JoVE.com

A Simple Device to Rapidly Prepare Whole Mounts of the Mouse Intestine

The use of a simple device to cut and &#8216;roll&#8217; mouse intestines to rapidly prepare whole mount preparations is described.

Eine einfache Vorrichtung, um schnell vorbereiten Whole Mounts des Mausdarm

Preparing whole mounts of the mouse small intestine and colon for subsequent analysis or quantification can be time consuming and difficult. We describe ...

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19.4K Aufrufe.  National Institutes of Health. The use of a simple device to cut and ‘roll’ mouse intestines to rapidly prepare whole mount preparations is described.

Serie: A Simple Device to Rapidly Prepare Whole Mounts of the Mouse Intestine

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&beta; using a Whole Blood Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues as well as altered cytokine homeostasis. The innate immune system plays a crucial role in recognizing pathogens and other endogenous danger stimuli. One of the major cytokines released by innate immune cells is Interleukin (IL)-1. Therefore, we utilize a whole blood stimulation assay in order to measure the secretion of inflammatory cytokines and specifically of the pro-inflammatory cytokine IL-1&beta; 1, 2, 3.
Patients with genetic dysfunctions of the innate immune system causing autoinflammatory syndromes show an exaggerated release of mature IL-1&beta; upon stimulation with LPS alone. In order to evaluate the innate immune component of patients who present with inflammatory-associated pathologies, we use a specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns (PAMPs), such as the gram-negative bacterial endotoxin, lipopolysaccharide (LPS). These PAMPs are recognized by pathogen recognition receptors (PRRs), which are found on the cells of the innate immune system 4, 5, 6, 7. A primary signal, LPS, in conjunction with a secondary signal, ATP, is necessary for the activation of the inflammasome, a multiprotein complex that processes pro-IL-1&beta; to its mature, bioactive form 4, 5, 6, 8, 9, 10.
The whole blood assay requires minimal sample manipulation to assess cytokine production when compared to other methods that require labor intensive isolation and culturing of specific cell populations. This method differs from other whole blood stimulation assays; rather than diluting samples with a ratio of RPMI media, we perform a white blood cell count directly from diluted whole blood and therefore, stimulate a known number of white blood cells in culture 2. The results of this particular whole blood assay demonstrate a novel technique useful in elucidating patient cohorts presenting with autoinflammatory pathophysiologies.

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&amp;#946; using a Whole Blood Stimulation Assay

We describe a simple immunoassay to measure the production of pro-inflammatory cytokines, such as IL-1 beta production, in patients presenting with autoinflammatory phenotypes. By activating cells in whole blood cultures with pathogen-associated molecular patterns, specifically with lipopolysaccharide, cytokine secretion can be conveniently evaluated in whole blood supernatants.

Genaue und einfache Messung der pro-inflammatorischen Zytokins IL-1β mit einem Vollblut Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues ...

Forschung

Medizin

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

Eine einfache Methode der Maus Lung Intubation

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Isolating and Using Sections of Bovine Mesenteric Artery and Vein as a Bioassay to Test for Vasoactivity in the Small Intestine

Mammalian gastrointestinal systems are constantly exposed to compounds (desirable and undesirable) that can have an effect on blood flow to and from that system. Changes in blood flow to the small intestine can result in effects on the absorptive functions of the organ. Particular interest in toxins liberated from feedstuffs through fermentative and digestive processes has developed in ruminants as an area where productive efficiencies could be improved. The video associated with this article describes an in vitro bioassay developed to screen compounds for vasoactivity in isolated cross-sections of bovine mesenteric artery and vein using a multimyograph. Once the blood vessels are mounted and equilibrated in the myograph, the bioassay itself can be used: as a screening tool to evaluate the contractile response or vasoactivity of compounds of interest; determine the presence of receptor types by pharmacologically targeting receptors with specific agonists; determine the role of a receptor with the presence of one or more antagonists; or determine potential interactions of compounds of interest with antagonists. Through all of this, data are collected real-time, tissue collected from a single animal can be exposed to a large number of different experimental treatments (an in vitro advantage), and represents vasculature on either side of the capillary bed to provide an accurate picture of what could be happening in the afferent and efferent blood supply supporting the small intestine.

The small intestine is frequently exposed to toxins that can influence blood flow and negatively impact nutrient absorption. Using a multimyograph and mesenteric artery and vein isolates, compounds or toxins of interest can be screened for vasoactivity.

Isolierung und Verwendung von Rinder Abschnitte A. und V. mesenterica als Bioassay für Vasoaktivität im Dünndarm Testen

Mammalian gastrointestinal systems are constantly exposed to compounds (desirable and undesirable) that can have an effect on blood flow to and from that ...

A Simple Mechanical Procedure to Create Limbal Stem Cell Deficiency in Mouse

Limbal stem cell deficiency (LSCD) is a state of malfunction or loss of limbal epithelial stem cells, after which the corneal epithelium is replaced with conjunctiva. Patients suffer from recurrent corneal defects, pain, inflammation, and loss of vision.
Previously, a murine model of LSCD was described and compared to two other models. The goal was to produce a consistent mouse model of LSCD that both mimics the phenotype in humans and lasts long enough to make it possible to study the disease pathophysiology and to evaluate new treatments. Here, the technique is described in more detail.
A motorized tool with a rotating burr has been designed to remove the rust rings from the corneal surface or to smooth the pterygium bed in patients. It is a suitable device to create the desired LSCD model. It is a readily available, easy-to-use tool with a fine tip that makes it appropriate for working on small eyes, as in mice. Its application prevents unnecessary trauma to the eye and it does not result in unwanted injuries, as often is the case with chemical injury models. As opposed to a blunt scraper, it removes the epithelium with the basement membrane. In this protocol, the limbal area was abraded two times, and then the whole corneal epithelium was shaved from limbus to limbus. To avoid stroma injury, care was taken not to brush the corneal surface once the epithelium was already removed.

The following article provides an easy, reproducible technique to effectively create a sustainable mouse model of limbal stem cell deficiency (LSCD). This animal model is useful in testing and comparing the efficacy of treatments for limbal stem cell diseases.

Eine einfache mechanische Verfahren Limbusstammzellinsuffizienzen in der Maus zum Erstellen

Limbal stem cell deficiency (LSCD) is a state of malfunction or loss of limbal epithelial stem cells, after which the corneal epithelium is replaced with ...

Ultrasound-guided Intracardiac Injection of Human Mesenchymal Stem Cells to Increase Homing to the Intestine for Use in Murine Models of Experimental Inflammatory Bowel Diseases

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have immunosuppressive potential and have been shown to suppress inflammation in mouse models of intestinal inflammation, even though the route of administration can limit their homing and effectiveness 1,3,4,5. Local application of MSCs to colonic injury models has shown greater efficacy at ameliorating inflammation in the colon. However, there is paucity of data on techniques to enhance the localization of human bone marrow-derived MSCs (hMSCs) to the small intestine, the site of inflammation in the SAMP-1/YitFc (SAMP) model of experimental Crohn&#39;s disease. This work describes a novel technique for the ultrasound-guided intracardiac injection of hMSCs in SAMP mice, a well-characterized spontaneous model of chronic intestinal inflammation. Sex- and age-matched, inflammation-free AKR/J (AKR) mice were used as controls. To analyze the biodistribution and the localization, hMSCs were transduced with a lentivirus containing a triple reporter. The triple reporter consisted of firefly luciferase (fl), for bioluminescent imaging; monomeric red fluorescent protein (mrfp), for cell sorting; and truncated herpes simplex virus thymidine kinase (ttk), for positron emission tomography (PET) imaging. The results of this study show that 24 h after the intracardiac administration, hMSCs localize in the small intestine of SAMP mice as opposed to inflammation-free AKR mice. This novel, ultrasound-guided injection of hMSCs in the left ventricle of SAMP mice ensures a high success rate of cell delivery, allowing for the rapid recovery of mice with minimal morbidity and mortality. This technique could be a useful method for the enhanced localization of MSCs in other models of small-intestinal inflammation, such as TNF&#916;RE6. Future studies will determine if the increased localization of hMSCs by intra-arterial delivery can lead to increased therapeutic efficacy.

Murine studies in models of colonic inflammation have demonstrated that a small percentage (1 - 5%) of mesenchymal stem cells (MSC) injected intravenously or intraperitoneally home to the inflamed colon1,2. This study shows that ultrasound-guided intracardiac injections of MSCs result in increased localization to the intestine.

Ultraschall-geführte intrakardiale Injektion humaner mesenchymaler Stammzellen Homing in den Darm zur Verwendung in murinen Modellen von experimentellen entzündlichen Darmerkrankungen erhöhen

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have ...

Ileectomy-induced Bile Overaccumulation in Mouse Intestine

Intestinal resection is a common therapeutic approach for human diseases such as obesity, inflammatory bowel disease, Crohn's disease, and colon cancer that often results in severe short bowel syndrome-like adverse effects including bile acid diarrhea, dehydration, electrolyte disturbances, and nutrient malabsorption. Here we introduce a murine ileal resection model, termed ileectomy, to evaluate tissue communication and the maintenance of systemic homeostasis. After ileal resection, circulating blood is permanently devoid of the ileum-specific endocrine hormone fibroblast growth factor 15 (FGF15), which releases its endocrinal inhibition of bile acid synthesis in the liver. In combination with the increased production and abolished reabsorption of bile acids after removing the ileum, mice that underwent surgery suffer from bile salt overaccumulation in the intestine and associated diarrhea, morbidity, and mortality. Novel usage of the surgery model introduced in this study may provide mechanistic and functional insights into ileal control of systemic metabolic regulation in physiology and disease.

Small intestine-dependent bile acid reabsorption and feedback inhibition of hepatic bile acid synthesis is important for systemic homeostasis and health. In this study, we describe a mouse model for ileal resection to evaluate ileectomy-induced bile malabsorption, overaccumulation, and toxicity in mouse intestine.

Overaccumulation Ileectomy-induzierten Galle im Darm der Maus

Intestinal resection is a common therapeutic approach for human diseases such as obesity, inflammatory bowel disease, Crohn's disease, and colon cancer ...

A Clinical Trial Assessing the Safety, Efficacy, and Delivery of Olive-Oil-Based Three-Chamber Bags for Parenteral Nutrition

Limited evidence exists to precisely estimate efficacy and safety differences between parenteral nutrition (PN) prepared using olive-oil-based three-chamber bags (3CBs) and soybean-oil-based compounded bags (CoBs) in hospitalized adult patients. We designed a multicenter, randomized, prospective, open-label, noninferiority protocol to compare the efficacy, safety, and distribution of olive-oil-based 3CBs and soybean-oil-based CoB formulations in adult Chinese patients requiring PN during surgical intervention. Subjects were randomized to receive either one of the study treatments using an interactive voice or web-based recognition system in accordance with the randomization code. Randomization was further stratified based on the study site and surgical category. Both treatment groups received similar amounts of calories and protein. In addition, the two study treatments contained a similar composition of the amino-acid component. The only difference between the two PN formulations was the lipid constitution. The duration of administration of study treatments was a minimum of 5 days up to a maximum of 14 days after the surgical procedure. The primary efficacy endpoint was serum prealbumin levels on day 5 of the study. Noninferiority was proved if the anti-log of the lower bound of the 95% confidence interval (CI) of the treatment difference was at least 0.80. Other efficacy measures included treatment preparation time; duration to achieve tolerability of oral nutrition; associated infectious complications; length of hospitalization; and laboratory assessment of markers of nutrition, inflammation, metabolism, and oxidative stress. A total of 458 patients were enrolled in the study. The results showed that olive-oil-based 3CBs were non-inferior to soybean-based CoBs, besides being well tolerated. The infection rate was found to be significantly lower in the olive-oil-based 3CB group. Thus, this study may be used as a reference for future research on lipid emulsion and 3CBs.

Here, we present a protocol for a study comparing the efficacy, safety, and delivery of olive-oil-based 3CB and soybean-oil-based CoB formulations in adults requiring parenteral nutrition. The results revealed that olive-oil-based 3CBs is non-inferior and well tolerated compared to soybean formulations.

Eine klinische Studie zur Bewertung der Sicherheit, Wirksamkeit und Lieferung von Dreikammerbeuteln auf Olivenölbasis für die parenterale Ernährung

Limited evidence exists to precisely estimate efficacy and safety differences between parenteral nutrition (PN) prepared using olive-oil-based ...

Mechanisms Underlying Gut Hormone Secretion Using the Isolated Perfused Rat Small Intestine

The gut is the largest endocrine organ of the body, producing more than 15 different peptide hormones that regulate appetite and food intake, digestion, nutrient absorption and distribution, and post-prandial glucose excursions. Understanding the molecular mechanisms that regulate gut hormone secretion is fundamental for understanding and translating gut hormone physiology. Traditionally, the mechanisms underlying gut hormone secretion are either studied in vivo (in experimental animals or humans) or using gut hormone-secreting primary mucosal cell cultures or cell lines. Here, we introduce an isolated perfused rat small intestine as an alternative method for studying gut hormone secretion. The virtues of this model are that it relies on the intact gut, meaning that it recapitulates most of the physiologically important parameters responsible for the secretion in in vivo studies, including mucosal polarization, paracrine relationships and routes of perfusion/stimulus exposure. In addition, and unlike in vivo studies, the isolated perfused rat small intestine allows for almost complete experimental control and direct assessment of secretion. In contrast to in vitro studies, it is possible to study both the magnitude and the dynamics of secretion and to address important questions, such as what stimuli cause secretion of different gut hormones, from which side of the gut (luminal or vascular) is secretion stimulated, and to analyze in detail molecular sensors underlying the secretory response. In addition, the preparation is a powerful model for the study of intestinal absorption and details regarding the dynamics of intestinal absorption including the responsible transporters.

Here, we present a powerful and physiological model to study the molecular mechanisms underlying gut hormone secretion and intestinal absorption &#8212; the isolated perfused rat small intestine.

Mechanismen, die Hormonausschüttung der Darm mit der isolierten durchblutet Ratte Dünndarm

The gut is the largest endocrine organ of the body, producing more than 15 different peptide hormones that regulate appetite and food intake, digestion, ...

A Modified Simple Method for Induction of Myocardial Infarction in Mice

Myocardial infarction (MI) represents one of the leading causes of death. MI models are widely used for investigating the pathomechanisms of post-MI remodeling and evaluation of novel therapeutics. Different methods (e.g., isoproterenol treatment, cryoinjury, coronary artery ligation, etc.) have been used to induce MI. Compared with isoproterenol treatment and cryoinjury, coronary artery ligation may better reflect the ischemic response and chronic remodeling after MI. However, traditional methods for coronary ligation in mice are technically challenging. The current study describes a simple and efficient process for induction of MI in mice with readily available materials. The mouse chest skin was cut open under stable anesthesia. The heart was immediately externalized through the intercostal space after blunt separation of the pectoralis major and pectoralis minor. The left anterior descending branch (LAD) was ligated with a 6-0 suture 3 mm from its origin. Following LAD ligation, staining with 2,3,5-Triphenyltetrazolium chloride (TTC) indicated successful induction of MI and temporal changes of post-MI scar size. Meanwhile, survival analysis results showed overt mortality within 7 days after MI, mainly due to cardiac rupture. Moreover, post-MI echocardiographic assessment demonstrated successful induction of contractile dysfunction and ventricular remodeling. Once mastered, an MI model can be established in mice within 2-3 min with readily available materials.

Under adequate anesthesia, the mouse heart was externalized through the intercostal space, and myocardial infarction was successfully induced by ligating the left anterior descending artery (LAD) using materials readily available in most laboratories.

Eine modifizierte einfache Methode zur Induktion von Myokardinfarkt bei Mäusen

Myocardial infarction (MI) represents one of the leading causes of death. MI models are widely used for investigating the pathomechanisms of post-MI ...

Application of Optical Coherence Tomography to a Mouse Model of Retinopathy

Optical coherence tomography (OCT) offers a noninvasive method for the diagnosis of retinopathy. The OCT machine can capture retinal crosssectional images from which the retinal thickness can be calculated. Although OCT is widely used in clinical practice, its application in basic research is not as prevalent, especially in small animals such as mice. Because of the small size of their eyeballs, it is challenging to conduct fundus imaging examinations in mice. Therefore, a specialized retinal imaging system is required to accommodate OCT imaging on small animals. This article demonstrates a small-animal-specific system for OCT examination procedures and a detailed method for image analysis. The results of retinal OCT examination of very-low-density lipoprotein receptor (Vldlr) knockout mice and C57BL/6J mice are presented. The OCT images of C57BL/6J mice showed retinal layers, while those of Vldlr knockout mice showed subretinal neovascularization and retinal thinning. In summary, OCT examination could facilitate the noninvasive detection and measurement of retinopathy in mouse models.

Here, we describe an in vivo imaging technique using optical coherence tomography to facilitate the diagnosis and quantitative measurement of retinopathy in mice.

Anwendung der optischen Kohärenztomographie auf ein Mausmodell der Retinopathie

Optical coherence tomography (OCT) offers a noninvasive method for the diagnosis of retinopathy. The OCT machine can capture retinal crosssectional images ...