Name: angiogenesis in the ischemic rat lung
Uploaded: 2013-02-08T13:00:00
Description: Watch this Scientific Journal Video about Angiogenesis in the Ischemic Rat Lung at JoVE.com

Vi anerkender det arbejde, som Dr. Adlah Sukkar, MD i at hjælpe med støbning af lungen. Dette arbejde er blevet finansieret af NHLBI, HL088005.

Alle protokoller udført på rotter er blevet godkendt af Johns Hopkins University Animal Care og brug Udvalg og i overensstemmelse med NIH retningslinjer. Når det er muligt, bør dyret kirurgisk prepped i et område adskilt fra det kirurgiske område for at minimere forurening af det kirurgiske sted. 1. Anæstesi / analgesi <ol><li> Sted rotte (Sprague Dawley-hanrotter, 125-150 g, Harlan, Indianapolis, IN) i et induktionskammer infunderes med 3% isofluran. </li><li> Placer bedøvet rotte...

<ol>
	<li>Weibel, E. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+stages+in+the+development+of+collateral+circulation+to+the+lung+in+the+rat.">Early stages in the development of collateral circulation to the lung in the rat.</a> Circulation Research. 8, 353-376 (1960).</li><li>Li, X., Wilson, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+vascularity+of+the+bronchial+mucosa+in+mild+asthma.">Increased vascularity of the bronchial mucosa in mild asthma.</a> Am. J. Respir. Crit. Care Med. 156, 229-233 (1997).</li><li>Turner-Warwick, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precapillary+systemic-pulmonary+anastomoses.">Precapillary systemic-pulmonary anastomoses.</a> Thorax. 18, 225-237 (1963).</li><li>Muller, K. M., Meyer-Schwickerath, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bronchial+arteries+in+various+stages+of+bronchogenic+carcinoma.">Bronchial arteries in various stages of bronchogenic carcinoma.</a> Pathol. Res. Pract. 163, 34-46 (1978).</li><li>Remy-Jardin, M., Duhamel, 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=Systemic+Collateral+Supply+in+Patients+with+Chronic+Thromboembolic+and+Primary+Pulmonary+Hypertension:+Assessment+with+Multi-Detector+Row+Helical+CT+Angiography.">Systemic Collateral Supply in Patients with Chronic Thromboembolic and Primary Pulmonary Hypertension: Assessment with Multi-Detector Row Helical CT Angiography.</a> Radiology. , 274-281 (2005).</li><li>Karsner, H., Ghoreyeb, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+in+infarction:+The+circulation+in+experimental+pulmonary+embolism.">Studies in infarction: The circulation in experimental pulmonary embolism.</a> J. Exp. Med. 18, 507-522 (1913).</li><li>Endrys, J., Hayat, N., 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=Comparison+of+bronchopulmonary+collaterals+and+collateral+blood+flow+in+patients+with+chronic+thromboembolic+and+primary+pulmonary+hypertension.">Comparison of bronchopulmonary collaterals and collateral blood flow in patients with chronic thromboembolic and primary pulmonary hypertension.</a> Heart. 78, 171-176 (1997).</li><li>Virchow, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Uber+die+Standpunkte+in+den+Wissenschaftlichen+Medizin.">Uber die Standpunkte in den Wissenschaftlichen Medizin.</a> Virchow Archiv. 1, 1-19 .</li><li>Fadel, E., Mazmanian, G. 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=Lung+reperfusion+injury+after+chronic+or+acute+unilateral+pulmonary+artery+occlusion.">Lung reperfusion injury after chronic or acute unilateral pulmonary artery occlusion.</a> Am. J. Respir. Crit. Care Med. 157, 1294-1230 (1998).</li><li>Charan, N. B., Carvalho, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Angiogenesis+in+bronchial+circulatory+system+after+unilateral+pulmonary+artery+obstruction.">Angiogenesis in bronchial circulatory system after unilateral pulmonary artery obstruction.</a> J. Appl. Physiol. 82, 284-291 (1997).</li><li>Shi, W., Hu, F., 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=Altered+reactivity+of+pulmonary+vessels+in+postobstructive+pulmonary+vasculopathy.">Altered reactivity of pulmonary vessels in postobstructive pulmonary vasculopathy.</a> J. Appl. Physiol. 88, 17-25 (2000).</li><li>Shi, W., Giaid, 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=Increased+reactivity+to+endothelin+of+pulmonary+arteries+in+long-term+post-obstructive+pulmonary+vasculopathy+in+rats.">Increased reactivity to endothelin of pulmonary arteries in long-term post-obstructive pulmonary vasculopathy in rats.</a> Pulm. Pharmacol. Ther. 11, 189-196 (1998).</li><li>Sukkar, A., Jenkins, 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=Inhibition+of+CXCR2+Attenuates+Bronchial+Angiogenesis+in+the+Ischemic+Rat+Lung.">Inhibition of CXCR2 Attenuates Bronchial Angiogenesis in the Ischemic Rat Lung.</a> J. Appl. Physiol. 104, 1470-1475 (2008).</li><li>Mitzner, W., Lee, W., 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=Angiogenesis+in+the+mouse+lung.">Angiogenesis in the mouse lung.</a> Am. J. Pathol. 157, 93-101 (2000).</li><li>Wagner, E. M., Jenkins, 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=Lung+and+vascular+function+during+chronic+severe+pulmonary+ischemia.">Lung and vascular function during chronic severe pulmonary ischemia.</a> J. Appl. Physiol. 110, 538-544 (2011).</li><li>Remy-Jardin, M., Bouaziz, N., 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=Bronchial+and+nonbronchial+systemic+arteries+at+multi-detector+row+CT+angiography:+comparison+with+conventional+angiography.">Bronchial and nonbronchial systemic arteries at multi-detector row CT angiography: comparison with conventional angiography.</a> Radiology. 233, 741-749 (2004).</li><li>Baluk, P., Tammela, T., 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=Pathogenesis+of+persistent+lymphatic+vessel+hyperplasia+in+chronic+airway+inflammation.">Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation.</a> J. Clin. Invest. 115, 247-257 (2005).</li><li>Bailey, S. R., Boustany, 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=Airway+vascular+reactivity+and+vascularisation+in+human+chronic+airway+disease.">Airway vascular reactivity and vascularisation in human chronic airway disease.</a> Pulm. Pharmacol. Ther. 22, 417-425 (2009).</li></ol>

Venstre lungepulsåren ligering i alle testede arter fører til robust systemisk neovaskularisering af den iskæmiske lunge. Vi har præsenteret detaljerne i den kirurgiske metode i en rottemodel. Vores resultater produceret af vaskulær støbning, histopatologi og in vivo mærkning viser, at bronchiale arterier proliferere og perfundere det pulmonale parenkym. Således kan mekanismer bronkial angiogenese undersøges i en dyremodel, der svarer til den humane tilstand af kronisk pulmonal tromboembolisme. Desuden...

Systemisk angiogenese i lungen er velkendt. Ved sygdomstilstande, såsom astma 2, interstitiel lungefibrose 3, cancer 4, og kronisk pulmonær thromboembolisme 5, den systemiske vaskulatur i og omgiver lunge proliferates og invaderer pulmonal parenkym. Imidlertid dyremodeller for at studere denne differentielle aktivering af den systemiske og ikke den pulmonale cirkulation er få. Måske den mest reproducerbar model af systemisk neovaskularisering i lungen af ​​den voksne pattedyr er den, der forekommer efter induktion af kronisk pulmonal iskæmi. Reaktionen på venstre pulmonal obstruktion i mennesker 5-7, hunde 8, svin 9, får 10, marsvin 11, rotter 1, 12, 13, og mus 14 er den hurtige udbredelse af de bronchiale arterie samt interkostale arterier. De mekanismer, der er ansvarlige for systemisk neovascularining af lungen efter pulmonær iskæmi er stort set ukendt og er ikke blevet bredt undersøgt. Tidsforløbet for bronkial angiogenese hos rotter efter venstre pulmonal obstruktion er nøje beskrevet i histologisk arbejde Weibel 1. Udvide dette arbejde hos rotter, har vores laboratorium fokuseret på både vækstfaktorer vigtige i denne proces og den fysiologiske resultat af denne neovaskulatur i lungen. Resultaterne påviser CXC-kemokin Cinc-3 hæves tidligt efter iskæmi og behandling af rotter med en neutraliserende antistof mod CXCR2, receptoren for Cinc-3, dæmpes angiogenese 13. Den nyetablerede bronchial vaskulatur 14 dage efter indtræden af pulmonal iskæmi viste sig at være unormal markant forøgede protein permeabilitet 15. Venstre lunge funktion var ikke normal viste reduceret spredende kapacitet og et fald i lungevolumen 15. Selv om neovaskulatur kan have contributed til bevarelse af lungevæv i kronisk pulmonal iskæmi, synes ikke at være normal og kan bidrage til en vedvarende fald i lungefunktion. Måske en af ​​de mest mærkværdige aspekter af denne model vedrører den rumlige fordeling af prolifererende blodkar. Til trods for frigivelsen af ​​vækstfaktorer i det pulmonale parenkym grund af iskæmi, stammer den neovaskulatur i relativt store opstrøms bronchiale arterier. Den normale bronkial arterie opstår som en lille gren fra aorta og invaderer luftvejene træet ved carina. Således den mekanisme, hvormed vækstfaktorer inducerer den indledende fase af arteriogenese er ikke klar. Vi foreslår, at rotten, med en vaskulær anatomi ligner mennesker, giver en enestående mulighed for at studere de mekanismer, der er ansvarlige for systemisk angiogenese under pulmonal iskæmi. Selvom fuldstændig obstruktion af venstre lungepulsåren er en sjælden forekomst i mennesker,forøget bronchial vaskularisering synes at være tilsvarende induceres hos patienter uanset stedet og størrelsen af lungepulsåren obstruktion 16. Således tilvejebringer vi en detaljeret beskrivelse af den kirurgiske metode til at ligere den venstre lungepulsåren hos rotter og et middel til at bestemme omfanget af angiogenese.

<table border="1"><tbody><tr><td> Reagenser: </td><td> Firma </td><td></td><td></td></tr><tr><td> buprenorphin-hydrochlorid, Puralube </td><td> Butler Schein </td><td></td><td></td></tr><tr><td> bupivicain </td><td> APP Pharmaceuticals </td><td></td><td></td></tr><tr><td> Povidon-iod vatpind </td><td> Dynarex Corporation </td><td></td><td></td></tr><tr><td> polypropylen sutur size 6-0, 3/8 cirkel revers skære nål </td><td> Myco Medical </td><td></td><td></td></tr><tr><td> PE20 rør </td><td> Becton Dickinson </td><td></td><td></td></tr><tr><td> 15 um røde fluorescerende polystyrenpartikler </td><td> Invitrogen </td><td></td><td></td></tr><tr><td> 1 ml Hamilton glas injektionssprøjte </td><td> Hamilton Company </td><td> &lt;/ Td&gt; <td></td></tr><tr><td> Udstyr: </td><td></td><td></td><td></td></tr><tr><td> Genie Plus sprøjtepumpe </td><td> Kent Scientific </td><td></td><td></td></tr><tr><td> Fluorescensspektrofotometer </td><td> Digilab </td><td></td><td></td></tr><tr><td> Rodent Ventilator Model 683 </td><td> Harvard Apparatus </td><td></td><td></td></tr><tr><td></td><td></td><td></td><td> Tabel 1. Table af specifikke reagenser og udstyr. </td></tr></tbody></table>

angiogenesis in the ischemic rat lung

Den voksne lunge perfuseres af både systemiske bronchial arterie og hele venøst ​​tilbageløb strømmer gennem de pulmonale arterier. I de fleste lunge patologier, er det den mindre systemisk vaskulatur der opfylder et behov for øget lunge perfusion og viser robust neovaskularisering. Pulmonal vaskulær iskæmi induceret af pulmonal obstruktion har vist sig at føre til hurtig systemisk arteriel angiogenese hos mennesker såvel som i flere dyremodeller. Selv om den histologiske vurdering af tidsforløbet af bronkial arterie proliferation hos rotter omhyggeligt blev beskrevet af Weibel 1, mekanismer ansvarlige for denne organiserede vækst af nye fartøjer er ikke klare. Vi giver kirurgiske detaljer inducere venstre pulmonal iskæmi i rotten, der fører til bronkial neovaskularisering. Kvantificering af omfanget af angiogenese udgør en yderligere udfordring på grund af tilstedeværelsen af ​​de to kar i lungen. Metoderat bestemme funktionelle angiogenese baseret på mærkede mikrokugle injektioner er tilvejebragt.

Vaskulær stemmer: Resultater af virkningerne af venstre pulmonal iskæmi i rotten er vist i figur 1.. Vist er en methacrylat støbt af de bronchiale vaskulatur og den omfattende vaskularisering af venstre luftvejs træet 28 dage efter LPAL. At opnå dette stemmer, blev den systemiske vaskulatur injiceret med en methacrylat blanding (rød), retrograd i nedadgående aorta og luftrøret blev kanyleret og injiceret med en hvid silikonebaseret materiale. Denne vaskulære støbt tilvejebringer en bemærkelse...

Watch this Scientific Journal Video about Angiogenesis in the Ischemic Rat Lung at JoVE.com

Angiogenesis in the Ischemic Rat Lung

Lungen er perfuseres af både systemiske bronkial arterie og pulmonale arterier. I de fleste lunge patologier, er det mindre systemisk vaskulatur, der viser robust neovaskularisering. Ophør af pulmonal blodgennemstrømning fremmer frisk bronkial angiogenese. Vi giver kirurgiske detaljer inducere venstre pulmonal iskæmi, der fremmer bronkial neovaskularisering.

Angiogenese i den iskæmiske rottelunge

The  adult lung is perfused by both the systemic bronchial artery and the entire  venous return flowing through the pulmonary arteries. In most lung ...

Research

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Medicine

Multi-modal Imaging of Angiogenesis in a Nude Rat Model of Breast Cancer Bone Metastasis Using Magnetic Resonance Imaging, Volumetric Computed Tomography and Ultrasound

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell proliferation in the bone marrow cavity as well as for interaction of tumor and bone cells resulting in local bone destruction. Our aim was to develop a model of experimental bone metastasis that allows in vivo assessment of angiogenesis in skeletal lesions using non-invasive imaging techniques.
For this purpose, we injected 105 MDA-MB-231 human breast cancer cells into the superficial epigastric artery, which precludes the growth of metastases in body areas other than the respective hind leg1. Following 25-30 days after tumor cell inoculation, site-specific bone metastases develop, restricted to the distal femur, proximal tibia and proximal fibula1. Morphological and functional aspects of angiogenesis can be investigated longitudinally in bone metastases using magnetic resonance imaging (MRI), volumetric computed tomography (VCT) and ultrasound (US).
MRI displays morphologic information on the soft tissue part of bone metastases that is initially confined to the bone marrow cavity and subsequently exceeds cortical bone while progressing. Using dynamic contrast-enhanced MRI (DCE-MRI) functional data including regional blood volume, perfusion and vessel permeability can be obtained and quantified2-4. Bone destruction is captured in high resolution using morphological VCT imaging. Complementary to MRI findings, osteolytic lesions can be located adjacent to sites of intramedullary tumor growth. After contrast agent application, VCT angiography reveals the macrovessel architecture in bone metastases in high resolution, and DCE-VCT enables insight in the microcirculation of these lesions5,6. US is applicable to assess morphological and functional features from skeletal lesions due to local osteolysis of cortical bone. Using B-mode and Doppler techniques, structure and perfusion of the soft tissue metastases can be evaluated, respectively. DCE-US allows for real-time imaging of vascularization in bone metastases after injection of microbubbles7.
In conclusion, in a model of site-specific breast cancer bone metastases multi-modal imaging techniques including MRI, VCT and US offer complementary information on morphology and functional parameters of angiogenesis in these skeletal lesions.

In the pathogenesis of bone metastasis, angiogenesis is a crucial process and therefore represents a target for imaging and therapy. Here, we present a rat model of site-specific breast cancer bone metastasis and describe strategies to non-invasively image angiogenesis in vivo using magnetic resonance imaging, volumetric computed tomography and ultrasound.

Multi-modal Imaging af angiogenese i en nøgen rotte model af Breast Cancer knoglemetastaser Brug Magnetic Resonance Imaging, Volumetrisk Computed Tomography og Ultralyd

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell ...

Retinal Detachment Model in Rodents by Subretinal Injection of Sodium Hyaluronate

Subretinal injection of sodium hyaluronate is a widely accepted method of inducing retinal detachment (RD). However, the height and duration of RD or the occurrence of subretinal hemorrhage can affect photoreceptor cell death in the detached retina. Hence, it is advantageous to create reproducible RDs without subretinal hemorrhage for evaluating photoreceptor cell death. We modified a previously reported method to create bullous and persistent RDs in a reproducible location with rare occurrence of subretinal hemorrhage. The critical step of this modified method is the creation of a self-sealing scleral incision, which can prevent leakage of sodium hyaluronate after injection into the subretinal space. To make the self-sealing scleral incision, a scleral tunnel is created, followed by scleral penetration into the choroid with a 30 G needle. Although choroidal hemorrhage may occur during this step, astriction with a surgical spear reduces the rate of choroidal hemorrhage. This method allows a more reproducible and reliable model of photoreceptor death in diseases that involve RD such as rhegmatogenous RD, retinopathy of prematurity, diabetic retinopathy, central serous chorioretinopathy, and age-related macular degeneration (AMD).

Creating experimental retinal detachments with a reproducible and sustained height of detachment, and without subretinal hemorrhage, is important for studying the pathophysiology of photoreceptor cell loss in retinal disease and evaluating potential therapeutic interventions. Here, we report such a method in detail.

Nethindeløsning Model i Gnavere ved subretinal Injektion af Natriumhyaluronats

Subretinal injection of sodium hyaluronate is a widely accepted method of inducing retinal detachment (RD). However, the height and duration of RD or the ...

Implantation of Fibrin Gel on Mouse Lung to Study Lung-specific Angiogenesis

Recent significant advances in stem cell research and bioengineering techniques have made great progress in utilizing biomaterials to regenerate and repair damage in simple tissues in the orthopedic and periodontal fields. However, attempts to regenerate the structures and functions of more complex three-dimensional (3D) organs such as lungs have not been very successful because the biological processes of organ regeneration have not been well explored. It is becoming clear that angiogenesis, the formation of new blood vessels, plays key roles in organ regeneration. Newly formed vasculatures not only deliver oxygen, nutrients and various cell components that are required for organ regeneration but also provide instructive signals to the regenerating local tissues. Therefore, to successfully regenerate lungs in an adult, it is necessary to recapitulate the lung-specific microenvironments in which angiogenesis drives regeneration of local lung tissues. Although conventional in vivo angiogenesis assays, such as subcutaneous implantation of extracellular matrix (ECM)-rich hydrogels (e.g., fibrin or collagen gels or Matrigel - ECM protein mixture secreted by Engelbreth-Holm-Swarm mouse sarcoma cells), are extensively utilized to explore the general mechanisms of angiogenesis, lung-specific angiogenesis has not been well characterized because methods for orthotopic implantation of biomaterials in the lung have not been well established. The goal of this protocol is to introduce a unique method to implant fibrin gel on the lung surface of living adult mouse, allowing for the successful recapitulation of host lung-derived angiogenesis inside the gel. This approach enables researchers to explore the mechanisms by which the lung-specific microenvironment controls angiogenesis and alveolar regeneration in both normal and pathological conditions. Since implanted biomaterials release and supply physical and chemical signals to adjacent lung tissues, implantation of these biomaterials on diseased lung can potentially normalize the adjacent diseased tissues, enabling researchers to develop new therapeutic approaches for various types of lung diseases.

Recapitulation of the organ-specific microenvironment, which stimulates local angiogenesis, is indispensable for successful regeneration of damaged tissues. This report demonstrates a novel method to implant fibrin gels on the lung surface of living mouse in order to explore how the lung-specific microenvironment modulates angiogenesis and alveolar regeneration in adult mouse.

Implantation af fibringel om Mouse Lung til Uddannelse Lunge-specifik Angiogenesis

Recent significant advances in stem cell research and bioengineering techniques have made great progress in utilizing biomaterials to regenerate and ...

Method of Isolated Ex Vivo Lung Perfusion in a Rat Model: Lessons Learned from Developing a Rat EVLP Program

The number of acceptable donor lungs available for lung transplantation is severely limited due to poor quality. Ex-Vivo Lung Perfusion (EVLP) has allowed lung transplantation in humans to become more readily available by enabling the ability to assess organs and expand the donor pool. As this technology expands and improves, the ability to potentially evaluate and improve the quality of substandard lungs prior to transplant is a critical need. In order to more rigorously evaluate these approaches, a reproducible animal model needs to be established that would allow for testing of improved techniques and management of the donated lungs as well as to the lung-transplant recipient. In addition, an EVLP animal model of associated pathologies, e.g., ventilation induced lung injury (VILI), would provide a novel method to evaluate treatments for these pathologies. Here, we describe the development of a rat EVLP lung program and refinements to this method that allow for a reproducible model for future expansion. We also describe the application of this EVLP system to model VILI in rat lungs. The goal is to provide the research community with key information and &#8220;pearls of wisdom&#8221;/techniques that arose from trial and error and are critical to establishing an EVLP system that is robust and reproducible.

Method of Isolated Ex Vivo Lung Perfusion in a Rat Model: Lessons Learned from Developing a Rat EVLP Program

Ex-Vivo Lung Perfusion (EVLP) has allowed lung transplantation in humans to become more readily available by enabling the ability to assess organs and expand the donor pool. Here, we describe the development of a rat EVLP program and refinements that allow for a reproducible model for future expansion.

Metode til isoleret<em&gt; Ex vivo</em&gt; Lung Perfusion i en rotte Model: Erfaringer fra at udvikle en rotte EVLP Program

The number of acceptable donor lungs available for lung transplantation is severely limited due to poor quality. Ex-Vivo Lung Perfusion (EVLP) has allowed ...

Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history have&#160;remained poorly understood. Recently, we developed a reliable, reproducible animal model of PION and tested the treatment effect of some neurotrophic factors in this model1. The purpose of this video is to demonstrate our photochemically induced model of posterior ischemic optic neuropathy, and to evaluate its effects with retrograde labeling of retinal ganglion cells. Following surgical exposure of the posterior optic nerve, a photosensitizing dye, erythrosin B, is intravenously injected and a laser beam is focused onto the optic nerve surface. Photochemical interaction of erythrosin B and the laser during irradiation damages the vascular endothelium, prompting microvascular occlusion mediated by platelet thrombosis and edematous compression. The resulting ischemic injury yields a gradual but pronounced retinal ganglion cell dieback, owing to a loss of axonal input - a remote, injury-induced and clinically relevant outcome. Thus, this model provides a novel platform to study the pathophysiologic course of PION,&#160;and can be further optimized for testing therapeutic approaches for optic neuropathies as well as other CNS ischemic diseases.

The goal of this protocol is to photochemically induce ischemic injury to the posterior optic nerve in rat. This model is critical to studies of the pathophysiology of posterior ischemic optic neuropathy, and therapeutic approaches for this and other optic neuropathies, as well as of other CNS ischemic diseases.

Rotte model af fotokemisk-induceret posterior iskæmisk opticusneuropati

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft involves anastomosis of donor inferior vena cava (IVC) to recipient IVC, and donor ascending aorta (Ao) to recipient abdominal Ao. The right upper and middle lung lobes are preserved and function as conduits for blood flow from right heart to left heart.
There are several advantages to using this technique, and it lends itself to a broad range of applications. Because the graft is transplanted in a configuration that allows for dyamic volume-loading, cardiac function may be directly assessed in vivo. The use of pressure-volume conductance catheters permits characterization of load-dependent and load-independent hemodynamic parameters. The graft may be converted to a loaded configuration by applying a clamp to the recipient&#8217;s infra-hepatic IVC. We describe modified surgical techniques for both donor and recipient operations, and an ideal myocardial protection strategy. Depending on the experimental aim, this model may be adapted for use in both acute and chronic studies of graft function, immunologic status, and variable ventricular loading conditions. The conducting airways to the transplanted lung are preserved, and allow for acute lung re-ventilation. This facilitates analysis of the effects of the mixed venous and arterial blood providing coronary perfusion to the graft.
A limitation of this model is its technical complexity. There is a significant learning curve for new operators, who should ideally be mentored in the technique. A surgical training background is advantageous for those wishing to apply this model. Despite its complexity, we aim to present the model in a clear and easily applicable format. Because of the physiologic similarity of this model to orthotopic transplantation, and its broad range of study applications, the effort invested in learning the technique is likely to be worthwhile.

We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

Rotte Heterotop Abdominal Heart / Single-lunge transplantation i en Volume-loaded konfiguration

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft ...

Demonstration of the Rat Ischemic Skin Wound Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.

The rat, due to its size, availability, and rather docile behavior, has been utilized as a research model for many years. The goal of this protocol is to utilize the rat as an ischemic skin wound healing model to provide valuable insight into the pathophysiology of chronic wounds.

Demonstration af Rat iskæmisk hudsår Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and ...

A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods for choosing appropriate cuff sizes for the pulmonary vein (PV), pulmonary artery (PA), or bronchus (Br) are varied, thus making the determination of proper cuff size during rat lung transplantation an exercise of trial and error. By standardizing the cuffing technique to use the smallest effective cuff appropriate for the size of the vessel or bronchus, one can make the transplantation procedure safer, faster, and more successful. Since diameters of the PV, PA, and Br are related to the body weight of the rat, we present a strategy to choosing an appropriate size using a weight-based guide. Since lung volume is also related to body weight, we recommend that this relationship should also be considered when choosing the proper volume of air for donor lung inflation during warm ischemia as well as for the proper volume of PBS to be instilled during bronchoalveolar lavage (BAL) fluid collection. We also describe methods for 4th intercostal space dissection, wound closure, and sample collection from both the native and transplanted lobes.

Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection.

En rottelungetransplantationsmodel af varm iskæmi / reperfusionsskade: optimeringer for at forbedre resultaterne

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods ...

Securing the Rat in the Rat Jacket for Electroacupuncture

Wear a pair of gloves before proceeding to fit and secure the rat in the custom made rat jacket. Then using one hand hold the rat jacket's middle and back portion on the table. Using the other hand, gently hold the rat by its tail to pick it up from the feeding cage and place it into the jacket as the rat moves forward into the jacket and its head reaches the headgear point.Using both hands, carefully pull out its left and right upper limbs from the holes in the jacket. Securely wrap the tying ropes from the opening end of the jacket around the rat's trunk, and tie them tightly at the opposite end to adequately restrain its upper limbs. Tie the rope loose enough to prevent hypoxia or local limb ischemia, and tight enough to prevent the rat's escape.In a similar manner, secure its lower limbs.

Fastgørelse af rotten i rottejakken til elektroakupunktur

<meta charset="utf8"></head><body>Realtidsovervågning af cellulær dynamik ved muselungetransplantation

Intravital Two-Photon Microscopy of the Transplanted Mouse Lung

Complications after lung transplantation are largely related to the host immune system responding to the graft. Such immune responses are regulated by crosstalk between donor and recipient cells. A better understanding of these processes relies on the use of preclinical animal models and is aided by an ability to study intra-graft immune cell trafficking in real-time. Intravital two-photon microscopy can be used to image tissues and organs for depths up to several hundred microns with minimal photodamage, which affords a great advantage over single-photon confocal microscopy. Selective use of transgenic mice with promoter-specific fluorescent protein expression and/or adoptive transfer of fluorescent dye-labeled cells during intravital two-photon microscopy allows for the dynamic study of single cells within their physiologic environment. Our group has developed a technique to stabilize mouse lungs, which has enabled us to image cellular dynamics in na&#239;ve lungs and orthotopically transplanted pulmonary grafts. This technique allows for detailed assessment of cellular behavior within the vasculature and in the interstitium, as well as for examination of interactions between various cell populations. This procedure can be readily learned and adapted to study immune mechanisms that regulate inflammatory and tolerogenic responses after lung transplantation. It can also be expanded to the study of other pathogenic pulmonary conditions.

<meta charset="utf8"></head><body>Forfatter Spotlight: Fremme af lungetransplantationsimmunologi gennem intravital billeddannelse

Here, we present a protocol to intravitally image the transplanted mouse left lung using two-photon microscopy. This represents a valuable tool for studying cellular dynamics and interactions in real-time following murine lung transplantation.

Intravital to-fotonmikroskopi af den transplanterede muselunge

Complications after lung transplantation are largely related to the host immune system responding to the graft. Such immune responses are regulated by ...