Name: engineered vascularized muscle flap
Uploaded: 2016-01-11T09:30:00
Description: Watch this Scientific Journal Video about Engineered Vascularized Muscle Flap at JoVE.com

This research was supported by the FP7 European Research Council Grant 281501, ENGVASC.

All animal studies were approved by the Committee of the Ethics of Animal Experiments of the Technion. For this procedure, athymic nude mice were used to avoid immunological rejection. If using another type of mouse, the mice should be shaved prior to the surgical procedure and administration of cyclosporine (or another anti-rejection substitute) is recommended.
1. Scaffold Preparation and Cell Embedding
<ol>
	<li>Prepare scaffolds composed of 1:1 mixture of poly-L-lactic-acid (PLLA) and pol...

<ol>
	<li>Engelsman, A. F., van der Mei, H. C., Ploeg, R. J., Busscher, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+phenomenon+of+infection+with+abdominal+wall+reconstruction.">The phenomenon of infection with abdominal wall reconstruction.</a> Biomaterials. 28 (14), 2314-2327 (2007).</li><li>De Coppi, 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=Myoblast-acellular+skeletal+muscle+matrix+constructs+guarantee+a+long-term+repair+of+experimental+full-thickness+abdominal+wall+defects.">Myoblast-acellular skeletal muscle matrix constructs guarantee a long-term repair of experimental full-thickness abdominal wall defects.</a> Tissue Eng. 12 (7), 1929-1936 (2006).</li><li>Shi, 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=Regeneration+of+full-thickness+abdominal+wall+defects+in+rats+using+collagen+scaffolds+loaded+with+collagen-binding+basic+fibroblast+growth+factor.">Regeneration of full-thickness abdominal wall defects in rats using collagen scaffolds loaded with collagen-binding basic fibroblast growth factor.</a> Biomaterials. 32 (3), 753-759 (2011).</li><li>Yezhelyev, M. V., Deigni, O., Losken, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Management+of+full-thickness+abdominal+wall+defects+following+tumor+resection.">Management of full-thickness abdominal wall defects following tumor resection.</a> Ann Plast Surg. 69 (2), 186-191 (2012).</li><li>Macchiarini, P., Walles, T., Biancosino, C., Mertsching, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=First+human+transplantation+of+a+bioengineered+airway+tissue.">First human transplantation of a bioengineered airway tissue.</a> J Thorac Cardiovasc Surg. 128 (4), 638-641 (2004).</li><li>Macchiarini, 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=Clinical+transplantation+of+a+tissue-engineered+airway.">Clinical transplantation of a tissue-engineered airway.</a> Lancet. 372 (9665), 2023-2030 (2008).</li><li>Atala, A., Bauer, S. B., Soker, S., Yoo, J. J., Retik, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue-engineered+autologous+bladders+for+patients+needing+cystoplasty.">Tissue-engineered autologous bladders for patients needing cystoplasty.</a> Lancet. 367 (9518), 1241-1246 (2006).</li><li>Nishida, K., 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=Corneal+reconstruction+with+tissue-engineered+cell+sheets+composed+of+autologous+oral+mucosal+epithelium.">Corneal reconstruction with tissue-engineered cell sheets composed of autologous oral mucosal epithelium.</a> N Engl J Med. 351 (12), 1187-1196 (2004).</li><li>Petite, H., 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=Tissue-engineered+bone+regeneration.">Tissue-engineered bone regeneration.</a> Nat Biotechnol. 18 (9), 959-963 (2000).</li><li>Banta, M. N., Kirsner, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modulating+diseased+skin+with+tissue+engineering:+actinic+purpura+treated+with+Apligraf.">Modulating diseased skin with tissue engineering: actinic purpura treated with Apligraf.</a> Dermatol Surg. 28 (12), 1103-1106 (2002).</li><li>Vunjak-Novakovic, G., 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=Challenges+in+cardiac+tissue+engineering.">Challenges in cardiac tissue engineering.</a> Tissue engineering. Part B, Reviews. 16 (2), 169-187 (2010).</li><li>Novosel, E. C., Kleinhans, C., Kluger, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascularization+is+the+key+challenge+in+tissue+engineering.">Vascularization is the key challenge in tissue engineering.</a> Advanced Drug Delivery Reviews. 63 (4-5), 300-311 (2011).</li><li>Lesman, A., Gepstein, L., Levenberg, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascularization+shaping+the+heart.">Vascularization shaping the heart.</a> Ann N Y Acad Sci. 1188, 46-51 (2010).</li><li>Patton Jr, H., Berry, S., Kralovich, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+human+acellular+dermal+matrix+in+complex+and+contaminated+abdominal+wall+reconstructions.">Use of human acellular dermal matrix in complex and contaminated abdominal wall reconstructions.</a> The Am J of Surg. 193 (3), 360-363 (2007).</li><li>Menon, N. G., 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=Revascularization+of+human+acellular+dermis+in+full-thickness+abdominal+wall+reconstruction+in+the+rabbit+model.">Revascularization of human acellular dermis in full-thickness abdominal wall reconstruction in the rabbit model.</a> Ann Plast Surg. 50 (5), 523-527 (2003).</li><li>Buinewicz, B., Rosen, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acellular+cadaveric+dermis+(AlloDerm):+a+new+alternative+for+abdominal+hernia+repair.">Acellular cadaveric dermis (AlloDerm): a new alternative for abdominal hernia repair.</a> Ann Plast Surg. 52 (2), 188-194 (2004).</li><li>Bringman, 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=Hernia+repair:+the+search+for+ideal+meshes.">Hernia repair: the search for ideal meshes.</a> Hernia. 14 (1), 81-87 (2010).</li><li>Meintjes, J., Yan, S., Zhou, L., Zheng, S., Zheng, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthetic+biological+and+composite+scaffolds+for+abdominal+wall+reconstruction.">Synthetic biological and composite scaffolds for abdominal wall reconstruction.</a> Exp rev of med dev. 8 (2), 275-288 (2011).</li><li>Cheng, G., 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=Engineered+blood+vessel+networks+connect+to+host+vasculature+via+wrapping-and-tapping+anastomosis.">Engineered blood vessel networks connect to host vasculature via wrapping-and-tapping anastomosis.</a> Blood. 118 (17), 4740-4749 (2011).</li><li>Shandalov, Y., 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=An+engineered+muscle+flap+for+reconstruction+of+large+soft+tissue+defects.">An engineered muscle flap for reconstruction of large soft tissue defects.</a> PNAS of the USA. 111 (16), 6010-6015 (2014).</li><li>Zhang, T. Y., Suen, C. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+fast+parallel+algorithm+for+thinning+digital+patterns.">A fast parallel algorithm for thinning digital patterns.</a> Commun. ACM. 27 (3), 236-239 (1984).</li><li>Luna, L. G., Luna, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Manual of Histo Stain Meth ; of the Arm Forcs Inst of Path. , (1968).</li><li>Choi, J. H., 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=Adipose+tissue+engineering+for+soft+tissue+regeneration.">Adipose tissue engineering for soft tissue regeneration.</a> Tissue engineering. Part B, Reviews. 16 (4), 413-426 (2010).</li><li>Bellows, C. F., Alder, A., Helton, W. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abdominal+wall+reconstruction+using+biological+tissue+grafts:+present+status+and+future+opportunities.">Abdominal wall reconstruction using biological tissue grafts: present status and future opportunities.</a> Exp rev of med dev. 3 (5), 657-675 (2006).</li><li>Caspi, O., 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=Tissue+engineering+of+vascularized+cardiac+muscle+from+human+embryonic+stem+cells.">Tissue engineering of vascularized cardiac muscle from human embryonic stem cells.</a> Circ Res. 100 (2), 263-272 (2007).</li><li>Kaufman-Francis, K., Koffler, J., Weinberg, N., Dor, Y., Levenberg, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engineered+vascular+beds+provide+key+signals+to+pancreatic+hormone-producing+cells.">Engineered vascular beds provide key signals to pancreatic hormone-producing cells.</a> PloS one. 7 (7), e40741 (2012).</li><li>Kaully, T., Kaufman-Francis, K., Lesman, A., Levenberg, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascularization--the+conduit+to+viable+engineered+tissues.">Vascularization--the conduit to viable engineered tissues.</a> Tiss eng. Part B, Reviews. 15 (2), 159-169 (2009).</li><li>Koffler, 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=Improved+vascular+organization+enhances+functional+integration+of+engineered+skeletal+muscle+grafts.">Improved vascular organization enhances functional integration of engineered skeletal muscle grafts.</a> PNAS of the USA. 108 (36), 14789-14794 (2011).</li><li>Lesman, 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=Transplantation+of+a+tissue-engineered+human+vascularized+cardiac+muscle.">Transplantation of a tissue-engineered human vascularized cardiac muscle.</a> Tisseng. Part A. 16 (1), 115-125 (2010).</li><li>Levenberg, 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=Engineering+vascularized+skeletal+muscle+tissue.">Engineering vascularized skeletal muscle tissue.</a> Nat Biotechnol. 23 (7), 879-884 (2005).</li><li>Bearzi, 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=PlGF-MMP9-engineered+iPS+cells+supported+on+a+PEG-fibrinogen+hydrogel+scaffold+possess+an+enhanced+capacity+to+repair+damaged+myocardium.">PlGF-MMP9-engineered iPS cells supported on a PEG-fibrinogen hydrogel scaffold possess an enhanced capacity to repair damaged myocardium.</a> Cell death &amp; disease. 5, e1053 (2014).</li><li>Zhang, 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=SDF-1+expression+by+mesenchymal+stem+cells+results+in+trophic+support+of+cardiac+myocytes+after+myocardial+infarction.">SDF-1 expression by mesenchymal stem cells results in trophic support of cardiac myocytes after myocardial infarction.</a> FASEB J : official publication of the .Fed Am Soc Exp Biol. 21 (12), 3197-3207 (2007).</li><li>Dvir, 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=Prevascularization+of+cardiac+patch+on+the+omentum+improves+its+therapeutic+outcome.">Prevascularization of cardiac patch on the omentum improves its therapeutic outcome.</a> PNAS. 106 (35), 14990-14995 (2009).</li><li>Marsano, 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=The+effect+of+controlled+expression+of+VEGF+by+transduced+myoblasts+in+a+cardiac+patch+on+vascularization+in+a+mouse+model+of+myocardial+infarction.">The effect of controlled expression of VEGF by transduced myoblasts in a cardiac patch on vascularization in a mouse model of myocardial infarction.</a> Biomaterials. 34 (2), 393-401 (2013).</li><li>Rufaihah, A. 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=Enhanced+infarct+stabilization+and+neovascularization+mediated+by+VEGF-loaded+PEGylated+fibrinogen+hydrogel+in+a+rodent+myocardial+infarction+model.">Enhanced infarct stabilization and neovascularization mediated by VEGF-loaded PEGylated fibrinogen hydrogel in a rodent myocardial infarction model.</a> Biomaterials. 34 (33), 8195-8202 (2013).</li><li>Nillesen, S. T. 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=Increased+angiogenesis+in+acellular+scaffolds+by+combined+release+of+FGF2+and+VEGF.">Increased angiogenesis in acellular scaffolds by combined release of FGF2 and VEGF.</a> J of Contr Release. 116 (2), e88-e90 (2006).</li><li>Sekine, H., 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=In+vitro+fabrication+of+functional+three-dimensional+tissues+with+perfusable+blood+vessels.">In vitro fabrication of functional three-dimensional tissues with perfusable blood vessels.</a> Nat Commun. 4, 1399 (2013).</li><li>Tee, R., 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=Transplantation+of+engineered+cardiac+muscle+flaps+in+syngeneic+rats.">Transplantation of engineered cardiac muscle flaps in syngeneic rats.</a> Tiss eng. Part A. (19-20), 1992-1999 (2012).</li><li>Morritt, A. 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=Cardiac+tissue+engineering+in+an+in+vivo+vascularized+chamber.">Cardiac tissue engineering in an in vivo vascularized chamber.</a> Circulation. 115 (3), 353-360 (2007).</li></ol>

The advances in tissue engineering have been met with a growing demand for substitute tissues for reconstruction of various tissue types. A variety of synthetic 1,17,18 and biological 14-16 materials as well as fabrication methods have been assessed for their capacity to address these demands. However, despite the progress in clinical care and in tissue engineering, the restoration of full-thickness abdominal wall defects remains a challenge. A tissue adequate for reconstruction of such massive defe...

Abdominal wall defects often arise following severe trauma, cancer treatment, burns and removal of infected mesh. These defects often involve significant tissue loss, requiring complicated surgical procedures and presenting a major challenge for plastic reconstruction surgeons 1-4. Tissue engineering researchers seeking new sources for artificial tissues have explored different materials, cell sources and growth factors. Successful restorations of various tissues, such as trachea 5,6, bladder 7, cornea 8, bone 9and skin 10, by implantation of engineered tissues were previously reported. However, fabrication of a thick vascularized engineered tissue, particularly for reconstruction of large defects, remains a significant challenge in tissue engineering.
One of the main factors limiting the thickness of a viable tissue construct is the control of oxygen supply to its constituent cells. When relying on diffusion, construct thickness is limited to that of a very thin layer. The maximum distance between oxygen- and nutrient-supplying capillaries in vivo is approximately 200&#160;&#181;m, which correlates with the diffusion limit of oxygen11,12. Insufficient vascularization can result in tissue ischemia and escalate to tissue resorption or necrosis 13.
In addition, the ideal material used for tissue reconstruction must be biocompatible and non-immunogenic. It must also be capable of promoting further integration of host cells with the biomaterial, and maintaining structural integrity. Various biological 14-16 and synthetic 1,17,18 matrices have been previously explored for tissue reconstruction, however their use remain limited due to lack of effective blood supply, infections or insufficient tissue strength.
In this study, a biocompatible, cell-embedded scaffold comprised of Food and Drug Administration (FDA)-approved poly L-lactic acid (PLLA)/poly lactic-co-glycolic acid (PLGA), was implanted around the femoral artery and vein (AV) vessels of a nude mouse and separated from the surrounding tissue, ensuring vascularization from the AV vessels only. One week post-implantation, the graft was viable, thick and well vascularized. This thick vascularized tissue with the AV vessels, was then transferred as a pedicled flap to an abdominal full-thickness defect in the same mouse. One week post-transfer, the flap was viable, vascularized and well integrated with the surrounding tissue, bearing sufficient strength to support abdominal viscera. Thus, the engineered thick, vascularized tissue flap, bearing an autologous pedicle, presents a novel method for repairing full-thickness abdominal wall defects.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>small fine straight scissors</td>
			<td>Fine Science Tools (FST)</td>
			<td>14090-09</td>
			<td></td>
		</tr>
		<tr>
			<td>spring scissors</td>
			<td>Fine Science Tools (FST)</td>
			<td>15003-08</td>
			<td></td>
		</tr>
		<tr>
			<td>straight forceps with fine tip</td>
			<td>Fine Science Tools (FST)</td>
			<td>11251-20</td>
			<td></td>
		</tr>
		<tr>
			<td>serrated forceps&#160;</td>
			<td>Fine Science Tools (FST)</td>
			<td>11050-10</td>
			<td></td>
		</tr>
		<tr>
			<td>needle holder</td>
			<td>Fine Science Tools (FST)</td>
			<td>12500-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Small vessel cauterizer&#160;</td>
			<td>Fine Science Tools (FST)</td>
			<td>18000-00</td>
			<td></td>
		</tr>
		<tr>
			<td>Duratears</td>
			<td>Alcon</td>
			<td align="right">5686</td>
			<td></td>
		</tr>
		<tr>
			<td>Sedaxylan</td>
			<td>Euravet</td>
			<td>DJ03</td>
			<td></td>
		</tr>
		<tr>
			<td>Clorketam 1000</td>
			<td>Vetoquinol</td>
			<td>4A0726B</td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine</td>
			<td>vetmarket</td>
			<td>B15100</td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 silk sutures</td>
			<td>Assut sutures</td>
			<td align="right">647</td>
			<td></td>
		</tr>
		<tr>
			<td>6-0 polypropylene sutures</td>
			<td>Assut sutures</td>
			<td>9351F</td>
			<td></td>
		</tr>
		<tr>
			<td>8-0 silk sutures</td>
			<td>Assut sutures</td>
			<td align="right">684568</td>
			<td></td>
		</tr>
		<tr>
			<td>Insulin syringe (6mm needle)</td>
			<td>BD</td>
			<td align="right">324911</td>
			<td></td>
		</tr>
		<tr>
			<td>Vevo 2100 high-resolution ultrasound system</td>
			<td>VisualSonics inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>MS250 non-linear transducer</td>
			<td>VisualSonics inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Micromarker non-targeted contrast agent</td>
			<td>VisualSonics inc.</td>
			<td>VS-11694</td>
			<td></td>
		</tr>
		<tr>
			<td>tail vein catheter</td>
			<td>VisualSonics inc.</td>
			<td>VS-11912</td>
			<td></td>
		</tr>
		<tr>
			<td>Vevo 2100 software</td>
			<td>VisualSonics inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>fluorescein isothiocyanate-conjugated dextran</td>
			<td>Sigma</td>
			<td>FD500S</td>
			<td></td>
		</tr>
		<tr>
			<td>Matlab</td>
			<td>Mathworks, MA, USA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Kimwipes</td>
			<td>Kimtech</td>
			<td align="right">34120</td>
			<td></td>
		</tr>
		<tr>
			<td>antigen unmasking solution</td>
			<td>Vector laboratories</td>
			<td>H-3300</td>
			<td></td>
		</tr>
		<tr>
			<td>anti-CD31 antibody</td>
			<td>Abcam&#160;</td>
			<td>ab28364</td>
			<td></td>
		</tr>
		<tr>
			<td>biotinylated goat anti-rabbit (secondary) antibody</td>
			<td>Vector laboratories</td>
			<td>BA-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>streptavidin-peroxidase</td>
			<td>Jackson&#160;</td>
			<td>016-030-084</td>
			<td></td>
		</tr>
		<tr>
			<td>Mayer&#39;s hamatoxylin solution</td>
			<td>Sigma-Aldrich</td>
			<td>MHS-16</td>
			<td></td>
		</tr>
		<tr>
			<td>aminoethylcarbazole (AEC) substrate kit</td>
			<td>Life technologies, Invitrogen&#160;</td>
			<td>00-2007</td>
			<td></td>
		</tr>
		<tr>
			<td>Vectamount</td>
			<td>Vector laboratories</td>
			<td>H-5501</td>
			<td></td>
		</tr>
	</tbody>
</table>

engineered vascularized muscle flap

One of the main factors limiting the thickness of a tissue construct and its consequential viability and applicability in vivo, is the control of oxygen supply to the cell microenvironment, as passive diffusion is limited to a very thin layer. Although various materials have been described to restore the integrity of full-thickness defects of the abdominal wall, no material has yet proved to be optimal, due to low graft vascularization, tissue rejection, infection, or inadequate mechanical properties. This protocol describes a means of engineering a fully vascularized flap, with a thickness relevant for muscle tissue reconstruction. Cell-embedded poly L-lactic acid/poly lactic-co-glycolic acid constructs are implanted around the mouse femoral artery and vein and maintained in vivo for a period of one or two weeks. The vascularized graft is then transferred as a flap towards a full thickness defect made in the abdomen. This technique replaces the need for autologous tissue sacrifications and may enable the use of in vitro engineered vascularized flaps in many surgical applications.

Graft vascularization and perfusion in vivo
The grafts were implanted one or two weeks prior to their transfer as axial flaps. At one and two weeks post-implantation, gross observation of the graft area revealed viable and vascularized tissue grafts. These grafts proved to be highly vascularized, as determined by positive CD31 immunostaining (Figure 1A), and highly perfused, as evidenced by FITC-dextran tail vein injection and ultrasound measurements. Many vessels wer...

Watch this Scientific Journal Video about Engineered Vascularized Muscle Flap at JoVE.com

Engineered Vascularized Muscle Flap

To date, thick tissue defects are typically reconstructed by applying autologous tissue flaps or engineered tissues. In this protocol, we present a new method for engineering vascularized tissue flap bearing an autologous pedicle, to serve as a substitute to autologous flaps.

One of the main factors limiting the thickness of a tissue construct and its consequential viability and applicability in vivo, is the control of oxygen ...

Research

JoVE Journal

Bioengineering

Engineered 3D Silk-collagen-based Model of Polarized Neural Tissue

Despite huge efforts to decipher the anatomy, composition and function of the brain, it remains the least understood organ of the human body. To gain a ...

Applications of In Vivo Functional Testing of the Rat Tibialis Anterior for Evaluating Tissue Engineered Skeletal Muscle Repair

Applications of In Vivo Functional Testing of the Rat Tibialis Anterior for Evaluating Tissue Engineered Skeletal Muscle Repair

Despite the regenerative capacity of skeletal muscle, permanent functional and/or cosmetic deficits (e.g., volumetric muscle loss (VML) resulting from ...

Microfluidic Bioprinting for Engineering Vascularized Tissues and Organoids

Engineering vascularized tissue constructs and organoids has been historically challenging. Here we describe a novel method based on microfluidic ...

Tissue-Engineered Graft for Circumferential Esophageal Reconstruction in Rats

The use of biocompatible materials for circumferential esophageal reconstruction is a technically challenging task in rats and requires an optimal implant ...

Fabrication of Zero Mode Waveguides for High Concentration Single Molecule Microscopy

In single molecule fluorescence enzymology, background fluorescence from labeled substrates in solution often limits fluorophore concentration to pico- to ...

Generation of Self-assembled Vascularized Human Skin Equivalents

Human skin equivalents (HSEs) are tissue engineered constructs that model epidermal and dermal components of human skin. These models have been used to ...

Assessing Functional Metrics of Skeletal Muscle Health in Human Skeletal Muscle Microtissues

Three-dimensional (3D) in vitro models of skeletal muscle are a valuable advancement in biomedical research as they afford the opportunity to study ...

Fibroblast Derived Human Engineered Connective Tissue for Screening Applications

Fibroblasts are phenotypically highly dynamic cells, which quickly transdifferentiate into myofibroblasts in response to biochemical and biomechanical ...

Engineered Lung Tissues Prepared from Decellularized Lung Slices

There is a need for improved 3-dimensional (3D) lung models that recapitulate the architectural and cellular complexity of the native lung alveolus ex ...

A Reliable Porcine Fascio-Cutaneous Flap Model for Vascularized Composite Allografts Bioengineering Studies

Vascularized Composite Allografts (VCA) such as hand, face, or penile transplant represents the cutting-edge treatment for devastating skin defects, ...