Name: right ventricular systolic pressure measurements in combination with harvest of lung and immune tissue samples in mice
Uploaded: 2013-01-16T12:00:00
Description: Watch this Scientific Journal Video about Right Ventricular Systolic Pressure Measurements in Combination with Harvest of Lung and Immune Tissue Samples in Mice at JoVE.com

Dit werk werd gefinancierd door de National Institutes of Health 1R21HL092370-01 (GG), 1R01 HL095764-01 (GG); R01HL082694 (JW); American Heart Association, oprichters affiliate (0855943D, GG); Stony Wold - Herbert Fonds, New York (SHP).

1. Voorbereiding <ol><li> Bereid de volgende oplossingen en buizen (Tabel 1) als volgt: <ol><li> Hanks oplossing, geen calcium, magnesium of indicator met penicilline (100 U / ml) / streptomycine (100 mg / ml). </li><li> Fosfaat gebufferde zoutoplossing (PBS), 1x, geen calcium, magnesium no. </li><li> Ethanol, 70%, 500 ml maken. </li><li> Gebufferde formaldehyde, 7-10% met PBS, maken 500 ml. </li><li> Anesthesie-oplossingen: <ol><li> Avertin. Voeg voorzichtig 5 ml 2-Methyl-2-butanol met 5 g 2,2,2-trib...

<ol>
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J. 21, 770-773 (2003).</li><li>West, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mice+expressing+BMPR2R899X+transgene+in+smooth+muscle+develop+pulmonary+vascular+lesions.">Mice expressing BMPR2R899X transgene in smooth muscle develop pulmonary vascular lesions.</a> Am. J. Physiol. Lung Cell Mol. Physiol. 295, L744-L755 (2008).</li><li>Tu, L., 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=Autocrine+fibroblast+growth+factor-2+signaling+contributes+to+altered+endothelial+phenotype+in+pulmonary+hypertension.">Autocrine fibroblast growth factor-2 signaling contributes to altered endothelial phenotype in pulmonary hypertension.</a> Am. J. Respir. Cell Mol. 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J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Attenuation+of+leukocyte+recruitment+via+CXCR1/2+inhibition+stops+the+progression+of+PAH+in+mice+with+genetic+ablation+of+endothelial+BMPR-II.">Attenuation of leukocyte recruitment via CXCR1/2 inhibition stops the progression of PAH in mice with genetic ablation of endothelial BMPR-II.</a> Blood. 118, 4750-4758 (2011).</li><li>Fujita, 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=Pulmonary+hypertension+in+TNF-alpha-overexpressing+mice+is+associated+with+decreased+VEGF+gene+expression.">Pulmonary hypertension in TNF-alpha-overexpressing mice is associated with decreased VEGF gene expression.</a> J. Appl. Physiol. 93, 2162-2170 (2002).</li><li>Motley, H. L., Cournand, A., Werko, L., Himmelstein, A., Dresdale, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Influence+of+Short+Periods+of+Induced+Acute+Anoxia+Upon+Pulmonary+Artery+Pressures+in+Man.">The Influence of Short Periods of Induced Acute Anoxia Upon Pulmonary Artery Pressures in Man.</a> Am. J. Physiol. 150, 315-320 (1947).</li><li>Liljestrand, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+Pulmonary+Arterial+Blood+Pressure.">Regulation of Pulmonary Arterial Blood Pressure.</a> Arch. Intern. Med. 81, 162-172 (1948).</li><li>Euler, U. S. V., Liljestrand, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Observations+on+the+pulmonary+arterial+blood+pressure+in+the+cat.">Observations on the pulmonary arterial blood pressure in the cat.</a> Acta Physiol. Scand. 12, 301-320 (1946).</li><li>Van den Broeck, W., Derore, A., Simoens, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anatomy+and+nomenclature+of+murine+lymph+nodes:+Descriptive+study+and+nomenclatory+standardization+in+BALB/cAnNCrl+mice.">Anatomy and nomenclature of murine lymph nodes: Descriptive study and nomenclatory standardization in BALB/cAnNCrl mice.</a> Journal of immunological. 312, 12-19 (2006).</li><li>Rabinovitch, 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=Angiotensin+II+prevents+hypoxic+pulmonary+hypertension+and+vascular+changes+in+rat.">Angiotensin II prevents hypoxic pulmonary hypertension and vascular changes in rat.</a> Am. J. Physiol. 254, 500-508 (1988).</li><li>Rabinovitch, M., Gamble, W., Nadas, A. S., Miettinen, O. S., Reid, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+pulmonary+circulation+after+chronic+hypoxia:+hemodynamic+and+structural+features.">Rat pulmonary circulation after chronic hypoxia: hemodynamic and structural features.</a> Am. J. Physiol. 236, 818-827 (1979).</li><li>Rabinovitch, 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=Changes+in+pulmonary+blood+flow+affect+vascular+response+to+chronic+hypoxia+in+rats.">Changes in pulmonary blood flow affect vascular response to chronic hypoxia in rats.</a> Circ. Res. 52, 432-441 (1983).</li><li>Kugathasan, L., 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=The+angiopietin-1-Tie2+pathway+prevents+rather+than+promotes+pulmonary+arterial+hypertension+in+transgenic+mice.">The angiopietin-1-Tie2 pathway prevents rather than promotes pulmonary arterial hypertension in transgenic mice.</a> J. Exp. Med. 206, 2221-2234 (2009).</li><li>Bearer, C., Emerson, R. K., ORiordan, M. A., Roitman, E., Shackleton, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maternal+tobacco+smoke+exposure+and+persistent+pulmonary+hypertension+of+the+newborn.">Maternal tobacco smoke exposure and persistent pulmonary hypertension of the newborn.</a> Environ. Health Persp. , 105-202 (1997).</li><li>Graham, B. 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De experimentele stroom hier beschreven zorgt voor een snelle en gelijktijdige meting van systolische rechterventriculaire druk en oogsten van monsters voor analyse van de reacties in de longen, het hart en het immuunsysteem in muizen. De procedure combineert hart fysiologie metingen, micro-dissectie en de daaropvolgende weefsel oogst voor levende cellen onderzoeken is een histologische analyse, of omics-analyse van de weefsels. De hele procedure duurt minder dan 20 minuten per muis. Door het werk-gebied georganiseerde ...

<table border="1"><tbody><tr><td> Naam </td><td> Vennootschap </td><td> Catalogusnummer </td><td> Opmerkingen (optioneel) </td></tr><tr><td></td><td></td><td></td><td> Reagentia </td></tr><tr><td> 2-Methyl-2-butanol </td><td> Sigma-Aldrich </td><td> 152463 </td><td></td></tr><tr><td> 2,2,2-tribroomethanol </td><td> Sigma-Aldrich </td><td> T48402 </td><td></td></tr><tr><td> desinfecterende zeep (Dekking Spray TB plus Steris) </td><td> Fisher Scientific </td><td> 1629-08 </td><td></td></tr><tr><td> Ethyl Alcohol, 200 Proof, Absolute, Watervrij ACS / USP Grade </td><td> PHARMCO-AAPER </td><td> 111000200 </td><td> Verdunnen tot 70% met gedestilleerd water </td></tr><tr><td> Formaldehydeoplossing </td><td> Sigma-Aldrich </td><td> F1635-500ML </td><td> Verdunnen tot een 7-10% formaldehyde concentratie op een PBS concentratie van 1x gebruik van PBS voorraad-oplossing en water </td></tr><tr><td> Hanks oplossing, geen calcium, magnesium </td><td> Fisher Scientific </td><td> 21-022-CV </td><td></td></tr><tr><td> Oktober </td><td> Tissue-Tek </td><td> 4583 </td><td></td></tr><tr><td> Penicilline (10.000 U / ml) / streptomycine (10.000 mg / ml) oplossing </td><td> Thermo Scientific </td><td> SV30010 </td><td></td></tr><tr><td> Fosfaat gebufferde zoutoplossing (PBS), geen calcium, magnesium no, 1x en 10x oplossingen </td><td> Fisher Scientific </td><td></td><td></td></tr><tr><td> Natriumpentobarbital 26% </td><td> Fort Dodge Animal Health </td><td> NDC 0856-0471-01 </td><td></td></tr><tr><td></td><td></td><td></td><td> Labware </td></tr><tr><td> Platen 12, 24, 96 well </td><td> Valk </td><td></td><td></td></tr&gt; <tr><td> Transfer Pipet </td><td> Fisher Scientific </td><td> 13-711-9BM </td><td></td></tr><tr><td> Tube, EDTA gecoat </td><td> Sarstedt </td><td> 2,013-08 </td><td></td></tr><tr><td> Buizen 0,65 ml en 1,7 ml micro-centrifuge </td><td> VWR </td><td></td><td></td></tr><tr><td> Buizen 12 x 75 mm polypropyleen </td><td> Fisher Scientific </td><td> 14-956-1D </td><td></td></tr><tr><td> Buizen, diverse maten, polypropyleen </td><td> Fisher Scientific </td><td></td><td></td></tr><tr><td></td><td></td><td></td><td> Instrumenten </td></tr><tr><td> Tang, Dumon # 5 Fine </td><td> Fine Science Tools </td><td> 11.254-20 </td><td></td></tr><tr><td> Tang, extra fijne Graefe-0.5mm tips gebogen </td><td> Fine Science Tools </td><td> 11.152-10 </td><td></td></tr><tr><td> Tang, extra fine Graefe-0.5mm tips rechtstreeks </td><td> Fine Science Tools </td><td> 11.150-10 </td><td></td></tr><tr><td> Canule 18ga, 19 ga </td><td> BD </td><td> Precisie Glide Naalden </td><td> Gesneden optimale lengte, afgestompt en buiten geraspte op een grove buiten oppervlak te creëren </td></tr><tr><td> Schaar, Dissector schaar-slim bladen 9cm </td><td> Fine Science Tools </td><td> 14.081-09 </td><td></td></tr><tr><td> Hechtdraad voor BAL, gevlochten zijden hechtdraad, 4-0 </td><td> Fine Science Tools </td><td> SP116 </td><td></td></tr><tr><td> Hechtdraad voor rechts hartkatheterisatie, gevlochten zijden hechtdraad, 6-0 </td><td> Teleflex Medical </td><td> 18.020-60 </td><td></td></tr><tr><td> Spuit, 1 ml </td><td> BD </td><td> 309659 </td><td></td></tr><tr><td></td><td></td><td></td><td> Uitrusting </td></tr><tr><td> Versterker, PowerLab 4/30 </td&gt; <td> ADInstrument </td><td> Model ML866 </td><td></td></tr><tr><td> Catheter, druk F1.4 </td><td> Millar Instruments, Inc </td><td> 840-6719 </td><td></td></tr><tr><td> Dissectiemicroscoop </td><td> Variscope </td><td></td><td></td></tr><tr><td> Pincet, Vannas lente schaar-2mm bladen </td><td> Fine Science Tools </td><td> 15.000-00 </td><td></td></tr><tr><td> Halogeen Verlichte Desk Vergrootglas </td><td> Fisher Scientific </td><td> 11-990-56 </td><td></td></tr><tr><td> Laptop computer </td><td> Asus </td><td> Modelnummer A52F i5-processor, 15 inch </td><td></td></tr><tr><td> Lichtbron </td><td> Amscope </td><td> HL-250-A </td><td></td></tr><tr><td> Pressure Control Unit </td><td> Millar Instruments, Inc </td><td> PCU-2000 </td><td></td></tr><tr><td> Software, Labchart-Pro V.7 </td><td> AD Instrumenten </td><td></td><td></td></tr></tbody></table>

right ventricular systolic pressure measurements in combination with harvest of lung and immune tissue samples in mice

De functie van de rechter hart is om bloed rond te pompen door de longen, waardoor het koppelen van rechter hart fysiologie en pulmonaire vasculaire fysiologie. Ontsteking is een gemeenschappelijke modifier van hart en longfunctie, door de uitwerking van cellulaire infiltratie, de productie van cytokines en groeifactoren, en door te remodeling processen 1. Vergeleken met de linker ventrikel, rechter ventrikel het een lage druk pomp die werkt in een relatief smalle zone van drukveranderingen. Verhoogde pulmonale arteriële bloeddruk gepaard gaan met hogere druk in de long vaatbed en pulmonale hypertensie 2. Pulmonale hypertensie wordt vaak geassocieerd met inflammatoire longziekten, zoals chronische obstructieve longziekte, of auto-immuunziekten 3. Omdat pulmonale hypertensie geeft een slechte prognose voor de kwaliteit van leven en de levensverwachting, is veel onderzoek gericht op het begrijpen van de mechanismen die might doelwit zijn van farmaceutische interventie 4. De voornaamste uitdaging voor de ontwikkeling van effectieve beheersinstrumenten voor pulmonale hypertensie blijft de complexiteit van de gelijktijdige begrip van moleculaire en cellulaire veranderingen in het rechter hart, de longen en het immuunsysteem. Hier geven we een procedurele workflow voor snelle en nauwkeurige meting van drukveranderingen in het rechter hart van muizen en de gelijktijdige oogst van monsters van hart, longen en immune weefsels. De methode is gebaseerd op de directe catheterisatie van de rechter ventrikel via de halsader in close-chested muizen eerst ontwikkeld in de late jaren 1990 als surrogaat maat voor de druk in de longslagader 5-13. De georganiseerde team-aanpak vergemakkelijkt een zeer snelle recht hartkatheterisatie techniek. Dit maakt het mogelijk de metingen uit te voeren in muizen die spontaan ademen kamerlucht. De organisatie van de work-flow in verschillende werk-gebiedenvermindert de vertraging en opent de mogelijkheid om gelijktijdig uit te voeren fysiologie experimenten en oogst immuunsysteem, hart en longen. De workflow hier geschetste procedure kan worden aangepast voor een groot aantal laboratoriumsituatie en studie ontwerpen van kleine, gerichte experimenten om grote drugtesten. De gelijktijdige aankoop van hartfysiologie gegevens die kunnen worden uitgebreid tot echocardiografie 5,14-17 en oogst van hart-, long-en immuunsysteem weefsels bevatten vermindert het aantal dieren dat nodig is om de gegevens die de wetenschappelijke kennisbasis vooruit te krijgen. De procedurele workflow hier gepresenteerde biedt ook een ideale basis voor het verwerven van kennis van de netwerken die immuun-, long-en hartfunctie te koppelen. Dezelfde principes hier beschreven kan worden aangepast aan andere of aanvullende organen bestuderen nodig.

Het primaire resultaat voor het verkrijgen van goede hart drukcurven wordt bereikt door de juiste positie van het rechter hart katheter. De vorm van de druk tijdcurve is kritiek omdat de juiste plaatsing van de katheter in de rechter ventrikel leidt tot druk plateaus (figuur 4). Spiky curves daarentegen blijkt een katheter die wordt bewogen door de ademhaling of hartslag beweging tegen de wand van de rechter hartkamer. Teneinde mogelijke problemen met de fase van overleving van de dieren, de standaardaf...

Watch this Scientific Journal Video about Right Ventricular Systolic Pressure Measurements in Combination with Harvest of Lung and Immune Tissue Samples in Mice at JoVE.com

Right Ventricular Systolic Pressure Measurements in Combination with Harvest of Lung and Immune Tissue Samples in Mice

Een specifieke en snelle protocol tegelijkertijd onderzoeken juiste hartfunctie, longontsteking en de immuunrespons wordt beschreven als leermiddel. Video en cijfers beschrijven fysiologie en microdissectie technieken in een georganiseerd team-aanpak die aanpasbaar is om te worden gebruikt voor kleine tot grote studies.

Rechter ventrikel systolische druk Metingen in combinatie met Harvest van Lung en Immune weefselmonsters in Muizen

The function of the right heart is to pump blood through the lungs, thus linking right heart physiology and pulmonary vascular physiology. Inflammation is ...

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Recurrent Escherichia coli Urinary Tract Infection Triggered by Gardnerella vaginalis Bladder Exposure in Mice

Recurrent Escherichia coli Urinary Tract Infection Triggered by Gardnerella vaginalis Bladder Exposure in Mice

Terugkerende Escherichia coli urineweginfectie veroorzaakt door Gardnerella vaginalis blaasblootstelling bij muizen

Recurrent urinary tract infections (rUTI) caused by uropathogenic Escherichia coli (UPEC) are common and costly. Previous articles describing models of ...

Monitoring Neutrophil Elastase and Cathepsin G Activity in Human Sputum Samples

Monitoring van neutrofiele Elastase en Cathepsine G-activiteit in menselijke Sputummonsters

Proteases are regulators of countless physiological processes and the precise investigation of their activities remains an intriguing biomedical ...

Gene Knock-in by CRISPR/Cas9 and Cell Sorting in Macrophage and T Cell Lines

Gene Knock-in door CRISPR/Cas9 en celsortering in macrofaag- en T-cellijnen

Functional genomics studies of the immune system require genetic manipulations that involve both deletion of target genes and addition of elements to ...