The authors thank Christiane Pahrmann for her technical assistance. This study was funded by the Deutsche Stiftung fuer Herzforschung (F/28/14). D.W. was supported by the travel award from the International Society for Heart and Lung Transplantation. T.D. received the Else Kro&#776;ner Excellence Stipend from the Else-Kro&#776;ner-Fresenius-Stiftung (2012_EKES.04). S.S. received research grants from the Deutsche Forschungsgemeinschaft (DFG; DE2133/2-1, T.D.&#160; and SCHR992/3- 1, SCHR992/4-1, S.S.).

Animals received humane care in compliance with the Guide for the Principles of Laboratory Animals, prepared by the Institute of Laboratory Animal Resources and published by the National Institutes of Health. All animal protocols were approved by the responsible local authority (&#34;Amt f&#252;r Gesundheit und Verbraucherschutz, Hansestadt (Office for Health and Consumer Protection) Hamburg&#34;).
1. Animal Care
<ol>
	<li>Obtain Lewis rats (LEW/Crl) rats and ROSA/luciferase-LEW transgenic r...

<ol>
	<li>Sabik, J. F., 3rd, <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Understanding+saphenous+vein+graft+patency.">Understanding saphenous vein graft patency.</a> Circulation. 124, 273-275 (2011).</li><li>Goldman, 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=Long-term+patency+of+saphenous+vein+and+left+internal+mammary+artery+grafts+after+coronary+artery+bypass+surgery:+results+from+a+Department+of+Veterans+Affairs+Cooperative+Study.">Long-term patency of saphenous vein and left internal mammary artery grafts after coronary artery bypass surgery: results from a Department of Veterans Affairs Cooperative Study.</a> J Am Coll Cardiol. 44, 2149-2156 (2004).</li><li>Fitzgibbon, 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=Coronary+bypass+graft+fate+and+patient+outcome:+angiographic+follow-up+of+5,065+grafts+related+to+survival+and+reoperation+in+1,388+patients+during+25+years.">Coronary bypass graft fate and patient outcome: angiographic follow-up of 5,065 grafts related to survival and reoperation in 1,388 patients during 25 years.</a> J Am Coll Cardiol. 28, 616-626 (1996).</li><li>O'Brien, J. E., 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=Wound+healing+around+and+within+saphenous+vein+bypass+grafts.">Wound healing around and within saphenous vein bypass grafts.</a> J Thorac Cardiovasc Surg. 114, 38-45 (1997).</li><li>El-Kurdi, M. 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=Ovine+femoral+artery+bypass+grafting+using+saphenous+vein:+a+new+model.">Ovine femoral artery bypass grafting using saphenous vein: a new model.</a> J Surg Res. 193, 458-469 (2015).</li><li>Yuda, 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=Angiotensin+II+receptor+antagonist,+L-158,809,+prevents+intimal+hyperplasia+in+dog+grafted+veins.">Angiotensin II receptor antagonist, L-158,809, prevents intimal hyperplasia in dog grafted veins.</a> Life Sci. 68, 41-48 (2000).</li><li>McCann, R. L., Hagen, P. O., Fuchs, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aspirin+and+dipyridamole+decrease+intimal+hyperplasia+in+experimental+vein+grafts.">Aspirin and dipyridamole decrease intimal hyperplasia in experimental vein grafts.</a> Ann Surg. 191, 238-243 (1980).</li><li>Thomas, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+for+studying+vein+graft+failure+and+therapeutic+interventions.">Animal models for studying vein graft failure and therapeutic interventions.</a> Curr Opin Pharmacol. 12, 121-126 (2012).</li><li>Hu, Y., Xu, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+mouse+model+of+vein+bypass+graft+atherosclerosis.">New mouse model of vein bypass graft atherosclerosis.</a> Heart Lung Circ. 11, 182-188 (2002).</li><li>Salzberg, S. P., 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+neointimal+formation+after+surgical+vein+grafting+in+a+murine+model+of+type+2+diabetes.">Increased neointimal formation after surgical vein grafting in a murine model of type 2 diabetes.</a> Circulation. 114, I302-I307 (2006).</li><li>Abbott, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laboratory+animals:+the+Renaissance+rat.">Laboratory animals: the Renaissance rat.</a> Nature. 428, 464-466 (2004).</li><li>Lindblad-Toh, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genome+sequencing:+three's+company.">Genome sequencing: three's company.</a> Nature. 428, 475-476 (2004).</li><li>Yu, P., Nguyen, B. T., Tao, M., Campagna, C., Ozaki, C. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rationale+and+practical+techniques+for+mouse+models+of+early+vein+graft+adaptations.">Rationale and practical techniques for mouse models of early vein graft adaptations.</a> Journal of vascular surgery. 52, 444-452 (2010).</li><li>Olver, D. T., Lacefield, J. C., Shoemaker, K. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+of+bidirectional+flow+in+the+sciatic+vasa+nervorum.">Evidence of bidirectional flow in the sciatic vasa nervorum.</a> Microvascular research. 94, 103-105 (2014).</li><li>Stubbendorff, 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=Inducing+myointimal+hyperplasia+versus+atherosclerosis+in+mice:+an+introduction+of+two+valid+models.">Inducing myointimal hyperplasia versus atherosclerosis in mice: an introduction of two valid models.</a> Journal of visualized experiments : JoVE. , (2014).</li><li>Conradi, 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=Immunobiology+of+fibrin-based+engineered+heart+tissue.">Immunobiology of fibrin-based engineered heart tissue.</a> Stem cells translational medicine. 4, 625-631 (2015).</li><li>Dashwood, M. R., Tsui, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term='No-touch'+saphenous+vein+harvesting+improves+graft+performance+in+patients+undergoing+coronary+artery+bypass+surgery:+A+journey+from+bedside+to+bench.">'No-touch' saphenous vein harvesting improves graft performance in patients undergoing coronary artery bypass surgery: A journey from bedside to bench.</a> Vascular Pharmacology. 58, 240-250 (2013).</li><li>Bekler, H. I., Rosenwasser, M. P., Akilina, Y., Bulut, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+use+of+an+absorbable+collagen+cover+(NeuraWrap)+improves+patency+of+interpositional+vein+grafts.">The use of an absorbable collagen cover (NeuraWrap) improves patency of interpositional vein grafts.</a> Acta orthopaedica et traumatologica turcica. 44, 157-161 (2010).</li><li>Geurts, A. 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=Knockout+rats+via+embryo+microinjection+of+zinc-finger+nucleases.">Knockout rats via embryo microinjection of zinc-finger nucleases.</a> Science. 325, 433 (2009).</li><li>Moreno, 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=Creation+and+characterization+of+a+renin+knockout+rat.">Creation and characterization of a renin knockout rat.</a> Hypertension. 57, 614-619 (2011).</li></ol>

This video demonstrates a rat vein interposition model to investigate vein graft loss and to allow for the exploration of the underlying pathological processes and the testing of new drugs or therapeutic options.
Anesthesia is a crucial aspect of surgical procedures. A continuous inhalative anesthesia system is recommended, as this is a safe and easy method, especially during prolonged operations. This can be of great importance during the training phase, when the operation takes more than 1 h...

Coronary artery diseases and their complications are among the leading causes of death worldwide. Current therapeutic strategies focus on re-establishing the blood flow, either by dilating the narrowed vessel or by creating a bypass. Coronary artery bypass grafting (CABG) using vein autografts was first described in 1968 and has been refined over the years. Apart from the revascularization of the left anterior descending coronary artery, saphenous vein conduits are most commonly used1. However, graft patency remains the Achilles heel of saphenous vein grafts (SVG). One year after surgery, graft patency is 85%, dropping to 61% after ten years2,3. Unveiling the pathophysiological mechanisms and causes of SVG patency loss is therefore an important task.
This video demonstrates a rat vein interposition model to investigate vein graft loss. The overall goals of this method are to explore the underlying pathobiological and -physiological processes during disease progression and to develop a suitable model for drug or therapeutic option testing. By transplanting the superficial epigastric vein into the arterial system, this model closely mimics the clinical setting of coronary artery bypass grafting. Surgical trauma, ischemia, and wall stress are important triggers of pathological vascular changes and are imitated in the model described.
Different models and species are available to investigate vein graft patency loss. Large animal models, such as pigs4, sheep5, dogs6, and monkeys7, resemble human vessel and anatomical structures and thus enable complex therapeutic strategies, such as bypass stenting or new surgical techniques, to be tested8. However, special housing, equipment, and staff are required. In addition, high costs and the need for an additional anesthetist during surgery impede their broader application. Small animals, including rats, are easy to handle, do not require special housing, and have manageable costs. Compared to mouse models9,10, rat models have the advantage of better operability and therefore less variability in the outcome. Rats are physiologically and genetically more similar to humans than mice11,12. In addition, most wild-type mice only develop limited myointima13, which make mouse models prone to type II errors. The histology of the main mouse veins, such as the inferior vena cava, only consists of a few cell layers and renders early evaluation difficult13. A further disadvantage is the small amount of tissue available for subsequent analysis after graft recovery.
The model described in this video is reproducible, inexpensive, and easy to perform, and it can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, such as viral vectors for gene therapy, in an economical fashion.

<table border="0" cellpadding="0" cellspacing="0" width="720">
	<tbody>
		<tr height="17">
			<td height="17" style="height:17px;width:236px;">Rat LEW/Crl</td>
			<td style="width:183px;">Charles River</td>
			<td style="width:145px;">Stock number 004</td>
			<td style="width:156px;"></td>
		</tr>
		<tr height="51">
			<td height="51" style="height:51px;width:236px;">Rat LEW-Tg(Gt(ROSA)26Sor- 1 
			luc)11Jmsk</td>
			<td style="width:183px;">Institute of laboratory animals, Kyoto University, Japan</td>
			<td style="width:145px;">NBPR rat number 0299</td>
			<td style="width:156px;">http://www.anim.med.kyoto-u.ac.jp/NBR/</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">PFA 4%</td>
			<td>Electron Microscopy Sciences</td>
			<td>#157135S</td>
			<td align="right">20%</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">hair clipper</td>
			<td>WAHL</td>
			<td>8786-451A ARCO SE</td>
			<td></td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;">Forene</td>
			<td>AbbVie</td>
			<td style="width:145px;">PZN 10182054 Art.Nr.: B506</td>
			<td>Isoflurane</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">microsurgical clamp</td>
			<td>Fine Science Tools</td>
			<td>18055-04</td>
			<td>Micro-Serrefine - 4mm</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">clamp applicator</td>
			<td>Fine Science Tools</td>
			<td>18056-14</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">hair removal creme</td>
			<td>Rufin cosmetic</td>
			<td align="right">27618</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Povidone-Iodine</td>
			<td>Betadine Purdue Pharma</td>
			<td>NDC:67618-152</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">10-0 Ethilon suture</td>
			<td>Ethicon</td>
			<td>2814G</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">5-0 prolene suture</td>
			<td>Ethicon</td>
			<td>EH7229H</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Rimadyl</td>
			<td>Pfizer</td>
			<td style="width:145px;">400684.00.00</td>
			<td>Carprofen</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Novaminsulfon</td>
			<td>Ratiopharm</td>
			<td>PZN 03530402</td>
			<td>Metamizole</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Heparin</td>
			<td>Rotexmedica</td>
			<td>PZN: 3862340</td>
			<td>25.000 I.E./mL</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Xylocain 1%</td>
			<td>AstraZeneca</td>
			<td>PZN: 1137907</td>
			<td>Lidocain</td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;">EVICEL</td>
			<td>J&#38;J Med.Ethicon Biosur</td>
			<td style="width:145px;">PZN 7349697 Art. Nr.:EVK01DE</td>
			<td>fibrin glue</td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;">NaCl 0,9%</td>
			<td>B.Braun</td>
			<td style="width:145px;">PZN 06063042 Art. Nr.: 3570160</td>
			<td></td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:236px;">Vevo 770 high-resolution in vivo micro-imaging system</td>
			<td>VisualSonics</td>
			<td></td>
			<td>duplex sonography</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Ecogel 100 ultrasound gel</td>
			<td>Eco-med</td>
			<td>30GB</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">D-Luciferin Firefly, potassium salt</td>
			<td>Biosynth</td>
			<td>L-8220</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">PBS pH 7,4</td>
			<td>Gibco</td>
			<td>10010023</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Xenogen Ivis 200</td>
			<td>Perkin Elmer</td>
			<td></td>
			<td>bioluminescence imaging</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Weigerts iron hematoxylin Kit</td>
			<td>Merck</td>
			<td>1.15973.0002</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Resorcine-Fuchsine Weigert</td>
			<td>Waldeck</td>
			<td>2.00E-30</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Acid Fuchsin</td>
			<td>Sigma-Aldrich</td>
			<td>F8129-25G</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Ponceau S solution</td>
			<td>Serva Electrophoresis</td>
			<td>33427</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Azophloxin</td>
			<td>Waldeck</td>
			<td>1B-103</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:236px;">Molybdatophosphoric acid hydrate</td>
			<td>Merck</td>
			<td>1.00532.0100</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Orange G</td>
			<td>Waldeck</td>
			<td>1B-221</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Light Green SF</td>
			<td>Waldeck</td>
			<td>1B-211</td>
			<td>Trichrome staining</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Vitro-Clud</td>
			<td>Langenbrinck</td>
			<td>04-0001</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Glacial Acetic Acid</td>
			<td>Sigma-Aldrich</td>
			<td align="right">537020</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">37% HCl</td>
			<td>Sigma-Aldrich</td>
			<td>H1758</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Xylene</td>
			<td>Th. Geyer</td>
			<td align="right">3410</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Paraffin</td>
			<td>Leica biosystems</td>
			<td>REF 39602004</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Ethanol absolute</td>
			<td>Th. Geyer</td>
			<td align="right">2246</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Ethanol 96%</td>
			<td>Th. Geyer</td>
			<td align="right">2295</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Ethanol 70%</td>
			<td>Th. Geyer</td>
			<td align="right">2270</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Slide Rack</td>
			<td>Ted Pella</td>
			<td align="right">21057</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Staining dish</td>
			<td>Ted Pella</td>
			<td align="right">21075</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Bepanthen Eye and Nose ointment</td>
			<td>Bayer</td>
			<td align="right">1578675</td>
			<td>Eye ointment</td>
		</tr>
	</tbody>
</table>

vein interposition model: a suitable model to study bypass graft patency

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. Research models for bypass graft failure are essential for a better understanding of pathobiological and pathophysiological processes during graft patency loss. Large animal models, such as pigs or sheep, resemble human anatomical structures but require special facilities and equipment. This video describes a rat vein interposition model to investigate vein graft patency loss. Rats are inexpensive and easy to handle. Compared to mouse models, the convenient size of rats permits better operability and enables a sufficient amount of material to be obtained for further diverse analysis. In brief, the inferior epigastric vein of a donor rat is harvested and used to replace a segment of the femoral artery. Anastomosis is conducted via single stitches and sealed with fibrin glue. Graft patency can be monitored non-invasively using duplex sonography. Myointimal hyperplasia, which is the main cause for graft patency loss, develops progressively over time and can be calculated from histological cross sections.

The rat vein interposition model is suitable to study the development of myointima hyperplasia and vein graft failure. Animals recover well from the surgery and show excellent physical condition post-operation. Figure 1 shows the key surgical steps. After the skin incision along the linea inguinalis, the epigastric superficial vein and femoral artery are identified (Figure 1A). Harvesting of the graft should be performed carefully, without damaging the graft (Figure 1B),...

Watch this Scientific Journal Video about Vein Interposition Model: A Suitable Model to Study Bypass Graft Patency at JoVE.com

Vein Interposition Model: A Suitable Model to Study Bypass Graft Patency

This video demonstrates a model to study the development of myointimal hyperplasia after venous interposition surgery in rats.

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. ...

The overall goal of this procedure is to study bypass graft patency in a rat vein interposition model. This method can help answer key questions in the bypass surgery field, such as underlying path of biological and physiological processes during bypass occlusion. The main advantage of this technique is that the surgical procedure is easy and fast, which can be used to study novel therapeutics and tracts in vivo.Demonstrating the technique will be Grigol Tediashvili, a micro-surgeon from our laboratory. Before the surgery, check for a sufficient depth of anesthesia by pinching the rat's hind feet and verifying an absence of reflexes. Then, apply some vet ointment to the eyes to prevent dryness while under anesthesia.Next, spread the hind legs and fix their position using tape. Then, shave the inguinal hair with a hair trimmer, followed by disinfecting the area widely using povidone-iodine followed by 80%alcohol. Repeat these alternating scrubs a total of three times.Now under a microscope, perform a vertical incision along the linea inguinalis. Next, use two forceps to gently separate the subcutaneous tissues, and expose the superficial epigastric vein up to its origin on the femoral vein. Do this very carefully, as the vein is fragile.Next, stop the blood flow through the superficial epigastric vein using two clamps. Now, harvest a segment of the vein. Carefully lift the isolated vein and cut through the vessel using micro-scissors.Then, place the vein segment on sterile gauze. There, carefully glide a 30-gauge needle into one end and flush the vein with heparin. Then, transfer the vein into 1%lidocaine, and keep it on ice to prevent spasms.Now, euthanize the donor rat by increasing the anesthesia to 5%isoflurane. Next, in preparation for the anastamoses, prepare the recipient rat in the same way as the donor rat up to the shaving step. After two to three minutes of 5%isoflurane, open the donor rat's abdomen along the linea alba, cut through the diaphragm, and remove the heart to stop circulation.Continue preparing the recipient rat for surgery. Shave the medial side of the legs with a hair trimmer and disinfect the skin with three alternating scrubs of povidone-iodine and 80%ethanol. Before proceeding further, perform another toe pinch to monitor the anesthesia.Now, perform a median femoral incision from the knee to the inguinal fold. Under the microscope, use two forceps to separate the femoral artery from its surroundings. Then, use two micro clamps to stop the blood flow, placing the proximal clamp first.Next, cut out a short segment of the clamped femoral artery and discard it. Shorten the remaining arterial stump to leave a gap that is one to two millimeters larger than the vein graft. Then, flush the arterial stump with heparin from a 30-gauge needle.If the adventitia protrudes slightly beyond the vessel stump, use forceps to pull the adventitia slightly over the end of the vessel, and then, remove a piece of adventitia. Now, place the harvested vein segment between the arterial stumps in the correct orientation, and adjust the gap size for a good fit. Now, perform the proximal anastomosis first using 10-0 prolene suture.Start with suturing each lateral side, then add three more sutures to the ventral side. Finish with three stitches on the dorsal side. Next, connect the distal vessels using the same technique, starting with a suture on each lateral side, followed by three sutures on the ventral side, and then finishing on the dorsal side.After completing the anastomoses, load two one-milliliter syringes with the fibrin glue components. Then, carefully lift the graft with forceps, and apply approximately 100 microliters of component one under the graft, followed by 100 microliters of component two. The two components must always be added in the same volumes.Now, place the graft back in its position, and apply 100 microliters of each component over the graft to completely cover the graft and the anastomosis. This prevents anastomotic insufficiency and overdistention of the vein graft. Now, carefully open the clamps starting with the distal clamp.Then, confirm a successful surgery by checking for a visible pulse in the transplanted vein and distal artery of the graft. Before closing the skin, remove any excessive glue that could impede closure. Then, close the skin layers using 5-0 prolene sutures.Before the rat wakes up, inject four to five milligrams of carprofen per kilogram subcutaneously. Keep an eye on the animal until it has regained sufficient consciousness to maintain sternal recumbency. Then, give the rat a home cage without other animals.For the next three days, provide metamizole in the drinking water, and check the rat's recovery daily. Once fully recovered, the rat can be housed with other rats. Animals recover well from the surgery and showed excellent physical condition post-operation.Successful integration of the vein into the femoral artery and graft patency after transplantation was confirmed noninvasively using duplex sonography. By transplanting the vein of a luke-positive rat to a luke-negative rat, bioluminescence imaging was used to monitor the graft's presence. It was found that myointimal hyperplasia develops progressively in the graft over time.Histological staining with Masson's trichrome demonstrates myointimal formation inside the internal elastic lamina. To confirm that the model reproduces the dynamics of myointimal hyperplasia, luminal obliteration was measured. A gradual loss of graft patency was noted from seven days to 28 days post-surgery, which fits the model nicely.Once mastered, this technique can be done in 20 minutes if it is performed properly. Thank you for watching, and good luck with your experiments.

vein-interposition-model-suitable-model-to-study-bypass-graft

Research

JoVE Journal

Medicine

8.6K visualizações.  University Heart Center Hamburg. The overall goal of this procedure is to study bypass graft patency in a rat vein interposition model. This method can help answer key questions in the bypass surgery field, such as underlying path of biological and physiological processes during bypass occlusion. The main advantage of this technique is that the surgical procedure is easy and fast, which can be used to study novel therapeutics and tracts in vivo.Demonstrating the technique will be Grigol Tediashvili, a micro-surgeon fr...