Nous tenons à remercier Jennifer Petrosino et David DiSilvestro de l&#39;aide éditoriale. Cette recherche a été soutenue par numéro 20020728 Prix de l&#39;American Egg Board et le numéro 10040042 Prix de Novo Nordisk Pharmaceuticals ainsi que par le Centre Innovation Alimentaire, Bureau des affaires internationales, Centre for Advanced Functional Foods de recherche, et de l&#39;entrepreneuriat à l&#39;OSU ainsi que la National Science Foundation accorde CEE-0914790 (LJL). Le projet décrit a été soutenue par le Prix Nombre R21OD017244 (OZ) et UL1RR025755 (OSUCCC) du National Center for Research Resources, financé par le Bureau du Directeur de la National Institutes of Health (OD) et soutenu par le NIH Roadmap for Medical Research et NCI P30CA16058. Le contenu est de la seule responsabilité des auteurs et ne représentent pas nécessairement les vues officielles de la National Center for Research Resources ou de la National Institutes of Health.

Le protocole d&#39;étude a été approuvé par l&#39;Université Ohio State comités d&#39;éthique. Les expérimentations animales ont été approuvés par le protocole IACUC. Toutes les procédures ont été effectuées en vertu de la biosécurité au niveau du Cabinet 2 avec flux laminaire. Nous avons suivi toutes les exigences et procédures de sécurité standard. La technique de microencapsulation pour la préparation de microcapsules a été effectuée comme décrit 17, 18. 1. Préparatifs de Matériaux <ol><li> Préparer 10 ml de solution d&#39;alginate de sodium à 2% dans une solution saline physiologique à l&#39;autoclave (0,9% de NaCl dans l&#39;eau). Préparer une solution à 0,05% PLL dans du sérum physiologique. Préparer ces solutions le jour avant et agiter durant la nuit. Filtrer les solutions avec filtre de 0,22 um avant utilisation. </li><li> Préparation du citrate de sodium 50 mM et 100 mM de CaCl 2 dans 0,9% de NaCl et les autoclaver. </li><li> Autoclaver toutes les solutions, les aiguilles, les électrodes, et gobelets. Laver soigneusement les aiguilles propres avec des fils pour éviter le colmatage. </li><li> Déterminer le montant approprié de cellules à utiliser pour les capsules (102 microcapsules sont nécessaires pour chaque cm 2 de bien et il ya environ 500 cellules par capsule 18). Utiliser 1 ml d&#39;alginate de sodium par deux millions de cellules. </li><li> Préparer un «Fibroblast Growth Medium&quot; contenant du sérum de veau à 10%, et 100 U / ml de pénicilline / streptomycine dans une haute teneur en glucose (4500 mg / l de glucose) du milieu de Eagle modifié de Dulbecco (DMEM). <ol><li> Préparer un «milieu de différenciation I &#39;contenant 10% de sérum bovin foetal, 10 ug / ml d&#39;insuline, 1 uM de dexaméthasone, la xanthine mM de 3-isobutyl-1-méthyl 0,5, et 100 U / ml de pénicilline / streptomycine dans du DMEM. </li><li> Préparer contenant 10% de sérum un «milieu de différenciation II &#39;fœtal bovin, 10 pg / ml d&#39;insuline, et 100 U / ml de pénicilline / streptomycine dans DMEM. </li></ol></li><li> Préparer du tampon de lyse contenant une protease comprimé d&#39;inhibiteur par 10 ml de tampon de dosage radio-immunoprécipitation (RITampon PA). </li></ol> 2. Préparation des microbilles d&#39;alginate (figure 1A) <ol><li> Retirer ancien milieu de flacon de culture cellulaire. Rincer les cellules avec 10 ml de PBS. Apportez cellules en suspension avec 0,25% de trypsine-EDTA (2 ml par une confluence flacon T175). </li><li> Compter les cellules en suspension de cellules à l&#39;aide d&#39;un hémocytomètre. Utilisez aliquote de 10 ml de la suspension cellulaire et compter les cellules selon les instructions du fabricant. Centrifuger les cellules restantes dans un milieu de centrifugation à 480 x g à la température ambiante pendant 5 min. </li><li> Suspension du culot cellulaire dans de l&#39;alginate de sodium comme décrit à l&#39;étape 1.4. Transférer la solution d&#39;alginate de sodium-cellule à une seringue de 5 ml. </li><li> Retirer les bulles d&#39;air dans la solution, ajouter une aiguille de calibre 23 et inverser la seringue pour créer un 1 ml poche d&#39;air. </li><li> Placez un petit gobelet (180 ml) contenant 144 ml de solution de CaCl 100 mM 2 sous le bec de l&#39;aiguille de l&#39;encapsulateur. Fixez l&#39;électrode à l&#39;encapsulateur avec la pointe Approximment 2,5 cm au-dessus de la surface de la solution mM CaCl 2 100. </li><li> Passer hermétiquement la seringue contenant la solution d&#39;alginate de sodium-cellule dans la pompe à seringue. Fixer le tube de caoutchouc à l&#39;ouverture de la seringue. Poussez le piston jusqu&#39;à ce que la solution d&#39;alginate de sodium-cellule entre mi-chemin à travers le tube. Régler la tension à 5,4 kV. Régler le diamètre de 12,06 mm sur la pompe à seringue. Réglez la vitesse de 3 ml / h. Démarrer la pompe. Allumez l&#39;encapsulateur et maintenir la tension à 5,4 kV. </li><li> Fermer la fenêtre de la hotte et éviter toute vibration inutile jusqu&#39;à la fin de la formation des microbilles d&#39;alginate. Après tout solution passe à travers l&#39;aiguille, les microbilles se solidifier en forme de billes d&#39;alginate dans la solution mM CaCl 2 pour cent supplémentaire de 20 min avant le revêtement avec PLL. </li></ol> 3. microbilles de revêtement avec PLL (figure 1B) <ol><li> Retirer le bécher contenant les microbilles en forme de billes d&#39;alginate de sodium et de cellules transférer ces MicrobEADS dans un tube de centrifugation de 50 ml. Retirer la solution de CaCl2 à partir de la pastille de microbilles. </li><li> Laver les microbilles sphériques d&#39;alginate de cellules par addition de 30 ml de NaCl à 0,9%. Agiter le tube à la main en douceur. Retirer NaCl à 0,9% avec 25 ml pipette après les microbilles ont précipité par la gravitation. Répéter deux fois de plus pour un total de 3 lavages. </li><li> Utiliser 10 ml de solution de PLL à 0,05% pour chaque 1 ml de solution d&#39;alginate de sodium. Ajouter 0,05% PLL et le vortex à 1000 rotations par minute pendant 10 minutes. Remarque: Habituellement, 10 min est suffisante pour le revêtement PLL. </li><li> Une fois la couche PLL est formé, retirez la solution PLL et laver les capsules 3 fois comme décrit au paragraphe 3.2. </li></ol> 4. Suppression de l&#39;alginate de base (figure 1C) <ol><li> Ajouter 30 ml de solution de citrate de sodium à 50 mM. Attendre 5 minutes, ou jusqu&#39;à ce que tout l&#39;alginate de sodium soit dissous. Laver capsules trois fois comme décrit à l&#39;étape 3.1.1. </li><li> Enlever le NaCl à 0,9%, ajouter 20 ml de milieu de culture dans le tube de 50 mlet transférer tous les capsules contenant les cellules dans un ballon de culture de cellules. Poignée cellules encapsulées dans des conditions de culture cellulaire standard 18. </li></ol> 5. In Vitro applications à l&#39;étude de xénogreffes et cellule hôte Interactions ou Cinétique de métabolite Afflux / Efflux entre cellules (Figure 2) <ol><li> La culture de cellules hôtes sur plaque à 24 puits jusqu&#39;à la confluence des co-cultures. Utilisez le &#39;moyen de croissance des fibroblastes »pour préadipocytes de culture. </li><li> microcapsules de transfert 24 dans des cellules hôtes confluentes de plaque contenant aussi. Ajouter microcapsules pour parvenir à une monocouche (102 microcapsules / cm 2 de bien). </li><li> Induire la différenciation pré-adipocytes en milieu de différenciation I. Chaque 48 heures, les médias de changement au milieu de différenciation II pendant six jours. Lyse des cellules dans du tampon RIPA. Utilisez 50 pg de protéine par condition pour analyser l&#39;expression de protéines en Western blot. </li></ol> 6. En Vivo Demande de Traitemeent de l&#39;obésité (Figure 3) <ol><li> Mélanger 4% d&#39;isoflurane avec de l&#39;oxygène pour l&#39;induction de l&#39;anesthésie et 2% d&#39;isoflurane avec de l&#39;oxygène pour l&#39;entretien de l&#39;anesthésie. Confirmer la profondeur de l&#39;anesthésie adéquate par pincement de l&#39;orteil. <ol><li> Appliquer une pommade anti-démangeaison sur les yeux pour protéger les cornées de se dessécher. </li></ol></li><li> Utilisation des cellules encapsulées qui sont marqués de façon permanente avec la protéine de fluorescence artificiel, tels que la protéine de fluorescence verte (GFP). </li><li> Utiliser une seringue de trois ml et une aiguille de calibre 20 pour injecter des cellules encapsulées. <ol><li> Injecter 0,5 x 10 6 cellules en suspension dans 0,2 ml de PBS dans chaque dépôt de graisse IAB qui se trouve dans la zone intrapéritonéale entre une gonade et les reins comme le montre la figure 3. Utilisez ce volume de PBS et le nombre de cellules pour des souris avec un poids moyen de 40 g. Ajuster le nombre de cellules et le volume chez des souris qui ont un poids différent ou pour la détermination des effets dépendants de la dose. </li></ol></li></ol>

<ol>
	<li>Vogeli, 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=Multiple+chronic+conditions:+prevalence,+health+consequences,+and+implications+for+quality,+care+management,+and+costs.">Multiple chronic conditions: prevalence, health consequences, and implications for quality, care management, and costs.</a> Journal of general internal medicine. 22, 391-395 (2007).</li><li>Vija, 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=Mesenchymal+stem+cells:+Stem+cell+therapy+perspectives+for+type+1+diabetes.">Mesenchymal stem cells: Stem cell therapy perspectives for type 1 diabetes.</a> Diabetes &amp; metabolism. 35, 85-93 (2009).</li><li>Acarregui, A., Orive, G., Pedraz, J. L., Hernandez, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Therapeutic+applications+of+encapsulated+cells.">Therapeutic applications of encapsulated cells.</a> Methods in molecular biology. 1051, 349-364 (2013).</li><li>Chang, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Semipermeable+Microcapsules.">Semipermeable Microcapsules.</a> Science. 146, 524-525 (1964).</li><li>Elliott, R. B., 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=Live+encapsulated+porcine+islets+from+a+type+1+diabetic+patient+9.5+yr+after+xenotransplantation.">Live encapsulated porcine islets from a type 1 diabetic patient 9.5 yr after xenotransplantation.</a> Xenotransplantation. 14, 157-161 (2007).</li><li>Lim, F., Sun, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microencapsulated+islets+as+bioartificial+endocrine+pancreas.">Microencapsulated islets as bioartificial endocrine pancreas.</a> Science. 210, 908-910 (1980).</li><li>Vos, P., Spasojevic, M., Faas, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment+of+diabetes+with+encapsulated+islets.">Treatment of diabetes with encapsulated islets.</a> Advances in experimental medicine and biology. 670, 38-53 (2010).</li><li>Cotton, 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=Engineering+challenges+in+cell-encapsulation+technology.">Engineering challenges in cell-encapsulation technology.</a> Trends in biotechnology. 14, 158-162 (1996).</li><li>Yach, D., Stuckler, D., Brownell, K. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiologic+and+economic+consequences+of+the+global+epidemics+of+obesity+and+diabetes.">Epidemiologic and economic consequences of the global epidemics of obesity and diabetes.</a> Nature medicine. 12, 62-66 (2006).</li><li>Ng, 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=Global,+regional,+and+national+prevalence+of+overweight+and+obesity+in+children+and+adults+during+1980-2013:+a+systematic+analysis+for+the+Global+Burden+of+Disease+Study+2013.">Global, regional, and national prevalence of overweight and obesity in children and adults during 1980-2013: a systematic analysis for the Global Burden of Disease Study 2013.</a> Lancet. 384, 766-781 (2014).</li><li>Kissebah, A. 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=Relation+of+body+fat+distribution+to+metabolic+complications+of+obesity.">Relation of body fat distribution to metabolic complications of obesity.</a> The Journal of clinical endocrinology and metabolism. 54, 254-260 (1982).</li><li>Bray, G. 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=Relation+of+central+adiposity+and+body+mass+index+to+the+development+of+diabetes+in+the+Diabetes+Prevention+Program.">Relation of central adiposity and body mass index to the development of diabetes in the Diabetes Prevention Program.</a> The American journal of clinical nutrition. 87, 1212-1218 (2008).</li><li>Klein, 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=What+are+subcutaneous+adipocytes+really+good+for.">What are subcutaneous adipocytes really good for.</a> Experimental dermatology. 16, 45-70 (2007).</li><li>Tran, T. T., Yamamoto, Y., Gesta, S., Kahn, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beneficial+effects+of+subcutaneous+fat+transplantation+on+metabolism.">Beneficial effects of subcutaneous fat transplantation on metabolism.</a> Cell metabolism. 7, 410-420 (2008).</li><li>Seale, P., Kajimura, S., Spiegelman, B. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcriptional+control+of+brown+adipocyte+development+and+physiological+function--of+mice+and+men.">Transcriptional control of brown adipocyte development and physiological function--of mice and men.</a> Genes &amp; development. 23, 788-797 (2009).</li><li>Kozak, L. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+variation+in+brown+fat+activity+and+body+weight+regulation+in+mice:+lessons+for+human+studies.">Genetic variation in brown fat activity and body weight regulation in mice: lessons for human studies.</a> Biochimica et biophysica acta. 1842, 370-376 (2014).</li><li>Zhang, X., He, H., Yen, C., Ho, W., Lee, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+biodegradable,+immunoprotective,+dual+nanoporous+capsule+for+cell-based+therapies.">A biodegradable, immunoprotective, dual nanoporous capsule for cell-based therapies.</a> Biomaterials. 29, 4253-4259 (2008).</li><li>Yang, 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=The+prolonged+survival+of+fibroblasts+with+forced+lipid+catabolism+in+visceral+fat+following+encapsulation+in+alginate-poly-L-lysine.">The prolonged survival of fibroblasts with forced lipid catabolism in visceral fat following encapsulation in alginate-poly-L-lysine.</a> Biomaterials. 33, 5638-5649 (2012).</li><li>Chang, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Artificial+cells+with+emphasis+on+bioencapsulation+in+biotechnology.">Artificial cells with emphasis on bioencapsulation in biotechnology.</a> Biotechnology annual review. 1, 267-295 (1995).</li><li>Chang, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hybrid+artificial+cells:+microencapsulation+of+living+cells.">Hybrid artificial cells: microencapsulation of living cells.</a> ASAIO journal. 38, 128-130 (1992).</li><li>Koo, J., Chang, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Secretion+of+erythropoietin+from+microencapsulated+rat+kidney+cells:+preliminary+results.">Secretion of erythropoietin from microencapsulated rat kidney cells: preliminary results.</a> The International journal of artificial organs. 16, 557-560 (1993).</li><li>Weidenauer, U., Bodmer, D., Kissel, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microencapsulation+of+hydrophilic+drug+substances+using+biodegradable+polyesters.+Part+I:+evaluation+of+different+techniques+for+the+encapsulation+of+pamidronate+di-sodium+salt.">Microencapsulation of hydrophilic drug substances using biodegradable polyesters. Part I: evaluation of different techniques for the encapsulation of pamidronate di-sodium salt.</a> Journal of microencapsulation. 20, 509-524 (2003).</li><li>Smidsrod, O., Skjak-Braek, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alginate+as+immobilization+matrix+for+cells.">Alginate as immobilization matrix for cells.</a> Trends in biotechnology. 8, 71-78 (1990).</li><li>Lewinska, D., Rosinski, S., Werynski, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+process+conditions+during+impulsed+electrostatic+droplet+formation+on+size+distribution+of+hydrogel+beads.">Influence of process conditions during impulsed electrostatic droplet formation on size distribution of hydrogel beads.</a> Artificial cells, blood substitutes, and immobilization biotechnology. 32, 41-53 (2004).</li><li>Chan, E. S., Lee, B. B., Ravindra, P., Poncelet, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prediction+models+for+shape+and+size+of+ca-alginate+macrobeads+produced+through+extrusion-dripping+method.">Prediction models for shape and size of ca-alginate macrobeads produced through extrusion-dripping method.</a> Journal of colloid and interface science. 338, 62-72 (2009).</li><li>Bhujbal, S. V., Paredes-Juarez, G. A., Niclou, S. P., de Vos, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+influencing+the+mechanical+stability+of+alginate+beads+applicable+for+immunoisolation+of+mammalian+cells.">Factors influencing the mechanical stability of alginate beads applicable for immunoisolation of mammalian cells.</a> Journal of the behavior of biomedical materials. 37, 196-208 (2014).</li><li>Gushchina, L. V., Yasmeen, R., Ziouzenkova, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Moderate+vitamin+A+supplementation+in+obese+mice+regulates+tissue+factor+and+cytokine+production+in+a+sex-specific+manner.">Moderate vitamin A supplementation in obese mice regulates tissue factor and cytokine production in a sex-specific manner.</a> Archives of biochemistry and biophysics. 539, 239-247 (2013).</li><li>Ziouzenkova, 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=Retinaldehyde+represses+adipogenesis+and+diet-induced+obesity.">Retinaldehyde represses adipogenesis and diet-induced obesity.</a> Nature. 13, 695-702 (2007).</li><li>Yasmeen, R., Jeyakumar, S. M., Reichert, B., Yang, F., Ziouzenkova, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+contribution+of+vitamin+A+to+autocrine+regulation+of+fat+depots.">The contribution of vitamin A to autocrine regulation of fat depots.</a> Biochimica et biophysica acta. 1821, 190-197 (2012).</li><li>Liu, 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=miR-133a+regulates+adipocyte+browning+in+vivo.">miR-133a regulates adipocyte browning in vivo.</a> PLoS genetics. 9, e1003626 (2013).</li><li>Rao, R. 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=Meteorin-like+is+a+hormone+that+regulates+immune-adipose+interactions+to+increase+beige+fat+thermogenesis.">Meteorin-like is a hormone that regulates immune-adipose interactions to increase beige fat thermogenesis.</a> Cell. 157, 1279-1291 (2014).</li><li>Kir, 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=Tumour-derived+PTH-related+protein+triggers+adipose+tissue+browning+and+cancer+cachexia.">Tumour-derived PTH-related protein triggers adipose tissue browning and cancer cachexia.</a> Nature. 513, 100-104 (2014).</li></ol>

No conflicts of interest declared.

Diverses méthodes ont été utilisées pour encapsuler des cellules, y compris le séchage, l&#39;extrusion, et 19 l&#39;émulsion. Dans ce procédé, les billes d&#39;alginate sont extrudes à travers une aiguille, puis revêtue d&#39;PLL et le noyau alginate sera dissoute pour terminer l&#39;encapsulation. Bien que cette méthode a été utilisée pendant des années, la formation des perles avec la taille désirée et forme sphérique est toujours difficile. La taille des capsules est fortement dépendant...

L&#39;incidence croissante des maladies chroniques 1 a stimulé les études sur la transplantation de populations de cellules thérapeutiques 2. Cellules souches syngéniques ou allogéniques sont les types de cellules les plus couramment utilisés pour ces applications 2. Cependant, ces traitements ne permettent pas le contrôle de la différenciation et la migration de cellules souches après l&#39;implantation et ne sont pas économiques efficace. La transplantation de cellules génétiquement modifiées avec les fonctions bénéfiques prévoit améliorer le traitement de nombreuses maladies. Cependant, les modifications génétiques de cellules sont reconnus par le système immunitaire de l&#39;hôte, par conséquent, ces traitements nécessitent trois immunosuppression. L&#39;encapsulation de cellules produisant de l&#39;insuline a été développé par le Dr Chang 4. La technique est basée sur l&#39;encapsulation des cellules dans des gouttelettes d&#39;alginate qui sont immergés dans une solution de chlorure de calcium. Les molécules d&#39;alginate sont constitués d&#39;acide mannuronique (M) et guluronique (G) et peuvent être connectés par Ca 2+. Après la gélification, les billes sont mises en suspension une poly-L-lysine (PLL) solution. Au cours de cette étape, PLL se lie à G et M dans les molécules d&#39;alginate qui établit la membrane de la capsule. La porosité de la membrane de la capsule peut être modulée en faisant varier les concentrations de S PLL, et le temps d&#39;incubation et de la température. La liaison de PLL dépend aussi du type et de la concentration d&#39;alginate. Des matrices d&#39;alginate réticulé par des ions Ca 2+, sont instables dans le milieu physiologique ou dans des solutions tampons à forte concentration communs de phosphate et les ions citrate. Ces tampons peuvent extraire Ca 2+ de l&#39;alginate et liquéfier le noyau. La liquéfaction du noyau alginate offre un espace à l&#39;intérieur des capsules pour le mouvement cellulaire et la croissance. Les cellules encapsulées dans de l&#39;alginate polyanionique avec polycationique poly-L-lysine (APL) sont imperméables pour des immunoglobulines mais ont afflux d&#39;éléments nutritifs et l&#39;efflux de toxines. Les propriétés de ceux-ci APL permettent long terme survival des cellules encapsulées après transplantation dans génétiquement différents hôtes. Elliott et al. A rapporté la survie de fonctionnement des cellules du pancréas de porc encapsulés dans un patient humain neuf ans après l&#39;implantation 5. techniques d&#39;encapsulation peuvent être classés en microencapsulation (3-800 um) et macroencapsulation (supérieure à 1000 um). Microcapsules sont plus durables que macrocapsules 6. Depuis sa découverte par le Dr Chang et ses collègues en 1964, microencapsulation a été largement utilisé pour l&#39;encapsulation de cellules produisant l&#39;insuline anabolisants, hormones et autres molécules bioactives 7. Ces traitements confrontés à plusieurs défis dans le tissu hôte, y compris la fibrose et la réponse immunitaire 8. Dans un premier temps, les effets secondaires liés à la qualité de biopolymères ont été résolus. Cependant, la transplantation de cellules anabolisants déclenche toujours des effets secondaires, tels que la fibrose, à la suite de l&#39;hormone overproduction à l&#39;extérieur d&#39;une glande spécialisé. Au cours des dernières décennies, l&#39;obésité et le diabète de type 2 a atteint des proportions épidémiques 9. Plus de 30% des personnes adultes dans le monde sont en surpoids et obèses 10. Intra-abdominale (IAB) formation accrue de graisse augmente l&#39;incidence de l&#39;inflammation chronique et favorise le diabète de type 2, les maladies cardiovasculaires, de certains cancers et d&#39;autres pathologies 11-13. Plusieurs éléments de preuve suggèrent que la pathogenèse associée avec de la graisse IAB peut être évitée par les adipocytes spécifiques. Des études récentes ont montré que la transplantation d&#39;adipocytes sous-cutanés dans la région IAB peut améliorer le métabolisme et diminuer la résistance de l&#39;obésité et de l&#39;insuline chez les rongeurs in vivo 14. Une réduction efficace de la résistance de l&#39;obésité et de l&#39;insuline a été associé à adipocytes thermogénique capables de dissiper l&#39;énergie sous forme de chaleur de 15,16. Modification thermogénique des adipocytes peut être obtenue par transfection stabledes gènes participant à la désolidarisation de protons mitochondrial, comme la protéine de découplage 1 (Ucp1) ou de gènes régulant l&#39;expression de gènes et d&#39;autres Ucp1 thermogénique 15,16. Nos études récentes ont montré que la déficience en aldéhyde déshydrogénase 1 a1 (aldh1a1) conduit à la rénovation de Iab graisse thermogénique qui réduit l&#39;obésité et la résistance à l&#39;insuline chez ces souris 17,18. Notamment, l&#39;encapsulation des thermogénique aldh1a1 déficient (aldh1a1 - / -) préadipocytes médiatise même effet thérapeutique dans IAB graisse chez les souris de type sauvage obèses, ce qui suggère de nouvelles possibilités thérapeutiques pour le traitement de l&#39;IAB graisses 18. Dans les paramètres expérimentaux, des cellules encapsulées permettent aux chercheurs d&#39;étudier les effets des populations de cellules spécifiques dans une manière rentable 19. Ici, nous discutons du procédé d&#39;encapsulation d&#39;une lignée cellulaire catabolique thermogénique et son laboratoire et l&#39;application thérapeutique dans un modèle de souris de l&#39;obésité. Le protocole décrit tphases ROIS pour la production de microcapsules (figure 1): la formation des microbilles d&#39;alginate (Figure 1A), la formation de la polycationique poly-L-lysine (PLL) de membranes à la surface des microbilles (figure 1B), et l&#39;enlèvement de la alginate noyaux (Figure 1C).

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Encapsulation device (VAR V1)</td>
			<td>Nisco</td>
			<td>LIN-0042</td>
			<td>None</td>
		</tr>
		<tr>
			<td>KD scientific syringe pump</td>
			<td>KD scientific</td>
			<td>780100Y</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Olympus microscope&#160;</td>
			<td>Olympus Optical</td>
			<td>IX70-S8F2</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Sodium alginate</td>
			<td>Sigma</td>
			<td>MKBP8122V</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Poly-l-lysine hydrobromide (PLL)</td>
			<td>Sigma</td>
			<td>020M5006V</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Calcium chloride</td>
			<td>Sigma</td>
			<td>SLBJ2662V</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Sodium citrate tribasic dihydrate</td>
			<td>Sigma</td>
			<td>030M0200</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Sodium chloride</td>
			<td>Sigma</td>
			<td>SLBD2595V</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Mini-PROTEAN TGX Gels</td>
			<td>Bio-Rad</td>
			<td>456-1093</td>
			<td>None</td>
		</tr>
		<tr>
			<td>ATGL primary antibody (from rabbit)</td>
			<td>Cell Signaling</td>
			<td>2138S</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Secondary anti body (anti rabbit)</td>
			<td>LI-COR</td>
			<td>926-68071</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Radio-Immunoprecipitation Assay (RIPA) buffer</td>
			<td>Boston BioProducts</td>
			<td>D25Y6Z</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Phosphate buffered saline (PBS)</td>
			<td>Sigma</td>
			<td>RNBD2893</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Trypsin</td>
			<td>Gibco</td>
			<td>25200-056</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Cortizone 10 anti-itch ointment</td>
			<td>Cortizone 10</td>
			<td>C4029138</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s Modified Eagle Medium (DMEM)</td>
			<td>Gibco</td>
			<td>11965-092</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Newborn calf serum (CS)</td>
			<td>Sigma</td>
			<td>N4762</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Fetal bovine serum (FBS)</td>
			<td>Sigma</td>
			<td>F4135</td>
			<td>None</td>
		</tr>
		<tr>
			<td>3-Isobutyl-1-methylxanthine (IBMX)</td>
			<td>Sigma</td>
			<td>I0516</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Dexamethasone</td>
			<td>Sigma</td>
			<td>D4902</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Insulin (bovine)</td>
			<td>Sigma</td>
			<td>I5879</td>
			<td>None</td>
		</tr>
		<tr>
			<td>Protease inhibitor cocktail tablets</td>
			<td>Roche</td>
			<td>4693159001</td>
			<td>None</td>
		</tr>
	</tbody>
</table>

encapsulation thermogenic preadipocytes for transplantation into adipose tissue depots

Cellule encapsulation est développé pour piéger des cellules viables dans des membranes semi-perméables. Les cellules encapsulées greffées peuvent échanger des metabolites de faible poids moléculaire dans les tissus de l&#39;hôte traité pour obtenir une survie à long terme. La membrane semi-perméable permet greffé cellules encapsulées pour éviter le rejet par le système immunitaire. Le procédé d&#39;encapsulation a été conçu pour permettre une libération contrôlée de composés bioactifs, tels que l&#39;insuline, des hormones et d&#39;autres cytokines. Nous décrivons ici un procédé pour l&#39;encapsulation de cellules cataboliques, qui consomment des lipides pour la production de chaleur et de dissipation d&#39;énergie (thermogenèse) dans le tissu adipeux intra-abdominale de souris obèses. Encapsulation de cellules cataboliques thermogénique peut être potentiellement applicable à la prévention et au traitement de l&#39;obésité et le diabète de type 2. Une autre application potentielle de cellules cataboliques peut inclure la désintoxication d&#39;alcools ou d&#39;autres métabolites toxiques et des polluants environnementaux.

La figure 1 montre que chaque étape de la production de microbilles peut être contrôlé au microscope. La figure 2A montre comment la co-culture des adipocytes avec une monocouche de cellules encapsulées. La figure 2B est un exemple représentatif d&#39;une étude quantitative utilisant des adipocytes / microcapsules co-cultures que ont été décrits dans la section 5. lysats des adipocytes ont été analysés en Western blot. Des cellules encapsulées sont analys?...

Watch this Scientific Journal Video about Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots at JoVE.com

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

Encapsulation thermogénique préadipocytes pour la transplantation dans dépôts de tissu adipeux

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

encapsulation-thermogenic-preadipocytes-for-transplantation-into

Research

JoVE Journal

Medicine

9.2K vues.  The Ohio State University. Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

Vidéo: Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Subretinal Transplantation of MACS Purified Photoreceptor Precursor Cells into the Adult Mouse Retina

Vision impairment and blindness due to the loss of the light-sensing cells of the retina, i.e. photoreceptors, represents the main reason for disability in industrialized countries. Replacement of degenerated photoreceptors by cell transplantation represents a possible treatment option in future clinical applications. Indeed, recent preclinical studies demonstrated that immature photoreceptors, isolated from the neonatal mouse retina at postnatal day 4, have the potential to integrate into the adult mouse retina following subretinal transplantation. Donor cells generated a mature photoreceptor morphology including inner and outer segments, a round cell body located at the outer nuclear layer, and synaptic terminals in close proximity to endogenous bipolar cells. Indeed, recent reports demonstrated that donor photoreceptors functionally integrate into the neural circuitry of host mice. For a future clinical application of such cell replacement approach, purified suspensions of the cells of choice have to be generated and placed at the correct position for proper integration into the eye. For the enrichment of photoreceptor precursors, sorting should be based on specific cell surface antigens to avoid genetic reporter modification of donor cells. Here we show magnetic-associated cell sorting (MACS) - enrichment of transplantable rod photoreceptor precursors isolated from the neonatal retina of photoreceptor-specific reporter mice based on the cell surface marker CD73. Incubation with anti-CD73 antibodies followed by micro-bead conjugated secondary antibodies allowed the enrichment of rod photoreceptor precursors by MACS to approximately 90%. In comparison to flow cytometry, MACS has the advantage that it can be easier applied to GMP standards and that high amounts of cells can be sorted in relative short time periods. Injection of enriched cell suspensions into the subretinal space of adult wild-type mice resulted in a 3-fold higher integration rate compared to unsorted cell suspensions.

Cell transplantation represents a strategy for the treatment of retinal degeneration characterized by photoreceptor loss. Here we describe a method for enrichment of transplantable photoreceptors and their subretinal grafting into adult mice.

Transplantation sous-rétinienne de cellules précurseurs de photorécepteurs purifiées par MACS dans la rétine adulte de la souris

Vision impairment and blindness due to the loss of the light-sensing cells of the retina, i.e. photoreceptors, represents the main reason for disability ...

Recherche

Médecine

Transplantation of Pulmonary Valve Using a Mouse Model of Heterotopic Heart Transplantation

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed results. However, the cellular and molecular mechanisms of the neotissue development, valve thickening, and stenosis development are not researched extensively. To answer the above questions, we developed a murine heterotopic heart valve transplantation model. A heart valve was harvested from a valve donor mouse and transplanted to a heart donor mouse. The heart with a new valve was transplanted heterotopically to a recipient mouse. The transplanted heart showed its own heartbeat, independent of the recipient&#8217;s heartbeat. The blood flow was quantified using a high frequency ultrasound system with a pulsed wave Doppler. The flow through the implanted pulmonary valve showed forward flow with minimal regurgitation and the peak flow was close to 100 mm/sec. This murine model of heart valve transplantation is highly versatile, so it can be modified and adapted to provide different hemodynamic environments and/or can be used with various transgenic mice to study neotissue development in a tissue engineered heart valve.

In order to understand the cellular and molecular mechanisms underlying neotissue formation and stenosis development in tissue engineered heart valves, a murine model of heterotopic heart valve transplantation was developed. A pulmonary heart valve was transplanted to recipient using the heterotopic heart transplantation technique.

Transplantation de valve pulmonaire en utilisant un modèle de souris de transplantation cardiaque hétérotopique

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed ...

Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely related to metabolism and energy expenditure, and its dysfunction is one important contributor for obesity and diabetes. Contrary to previous belief, recent PET/CT imaging studies indicated the BAT depots are still present in human adults. PET imaging clearly shows that BAT has considerably high uptake of 18F-FDG under certain conditions. In this video report, we demonstrate that Cerenkov luminescence imaging (CLI) with 18F-FDG can be used to optically image BAT in small animals. BAT activation is observed after intraperitoneal injection of norepinephrine (NE) and cold treatment, and depression of BAT is induced by long anesthesia. Using multiple-filter Cerenkov luminescence imaging, spectral unmixing and 3D imaging reconstruction are demonstrated. Our results suggest that CLI with 18F-FDG is a practical technique for imaging BAT in small animals, and this technique can be used as a cheap, fast, and alternative imaging tool for BAT research.

In this video report, we show the application of Cerenkov Luminescence Imaging (CLI) for interscapular brown adipose tissue in mice under activated and depressed conditions.

Cerenkov Luminescence imagerie de Interscapular le tissu adipeux brun

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely ...

Localization, Identification, and Excision of Murine Adipose Depots

Obesity has increased dramatically in the last few decades and affects over one third of the adult US population. The economic effect of obesity in 2005 reached a staggering sum of $190.2 billion in direct medical costs alone. Obesity is a major risk factor for a wide host of diseases. Historically, little was known regarding adipose and its major and essential functions in the body. Brown and white adipose are the two main types of adipose but current literature has identified a new type of fat called brite or beige adipose. Research has shown that adipose depots have specific metabolic profiles and certain depots allow for a propensity for obesity and other related disorders. The goal of this protocol is to provide researchers the capacity to identify and excise adipose depots that will allow for the analysis of different factorial effects on adipose; as well as the beneficial or detrimental role adipose plays in disease and overall health. Isolation and excision of adipose depots allows investigators to look at gross morphological changes as well as histological changes. The adipose isolated can also be used for molecular studies to evaluate transcriptional and translational change or for in vitro experimentation to discover targets of interest and mechanisms of action. This technique is superior to other published techniques due to the design allowing for isolation of multiple depots with simplicity and minimal contamination.

Due to the drastic and negative connection between obesity and other comorbidities, research on the role adipose plays in disease and overall health is warranted. We present a protocol for the isolation and excision of adipose depots allowing for the study of adipose using in situ and in vitro methods.

Localisation, identification, et l&#39;excision des dépôts adipeux murin

Obesity has increased dramatically in the last few decades and affects over one third of the adult US population. The economic effect of obesity in 2005 ...

Human Brown Adipose Tissue Depots Automatically Segmented by Positron Emission Tomography/Computed Tomography and Registered Magnetic Resonance Images

Reliably differentiating brown adipose tissue (BAT) from other tissues using a non-invasive imaging method is an important step toward studying BAT in humans. Detecting BAT is typically confirmed by the uptake of the injected radioactive tracer 18F-Fluorodeoxyglucose (18F-FDG) into adipose tissue depots, as measured by positron emission tomography/computed tomography (PET-CT) scans after exposing the subject to cold stimulus. Fat-water separated magnetic resonance imaging (MRI) has the ability to distinguish BAT without the use of a radioactive tracer. To date, MRI of BAT in adult humans has not been co-registered with cold-activated PET-CT. Therefore, this protocol uses 18F-FDG PET-CT scans to automatically generate a BAT mask, which is then applied to co-registered MRI scans of the same subject. This approach enables measurement of quantitative MRI properties of BAT without manual segmentation. BAT masks are created from two PET-CT scans: after exposure for 2 hr to either thermoneutral (TN) (24 &#176;C) or cold-activated (CA) (17 &#176;C) conditions. The TN and CA PET-CT scans are registered, and the PET standardized uptake and CT Hounsfield values are used to create a mask containing only BAT. CA and TN MRI scans are also acquired on the same subject and registered to the PET-CT scans in order to establish quantitative MRI properties within the automatically defined BAT mask. An advantage of this approach is that the segmentation is completely automated and is based on widely accepted methods for identification of activated BAT (PET-CT). The quantitative MRI properties of BAT established using this protocol can serve as the basis for an MRI-only BAT examination that avoids the radiation associated with PET-CT.

The method presented here uses 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography/computed tomography (PET-CT) and fat-water separated magnetic resonance imaging (MRI), each scanned following 2 hr exposure to thermoneutral (24 &#176;C) and cold conditions (17 &#176;C) in order to map brown adipose tissue (BAT) in adult human subjects.

Dépôts de tissus humains adipeux brun segmentés automatiquement par tomographie par émission de positons / informatisée et la résonance magnétique images enregistrées

Reliably differentiating brown adipose tissue (BAT) from other tissues using a non-invasive imaging method is an important step toward studying BAT in ...

Transplantation Into the Mouse Ovarian Fat Pad

Orthotopic transplantation assays in mice are invaluable for studies of cell regeneration and neoplastic transformation. Common approaches for orthotopic transplantation of ovarian surface and tubal epithelia include intraperitoneal and intrabursal administration of cells. The respective limitations of these methods include poorly defined location of injected cells and limited space volume. Furthermore, they are poorly suited for long-term structural preservation of transplanted organs. To address these challenges, we have developed an alternative approach, which is based on the introduction of cells and tissue fragments into the mouse fat pad. The mouse ovarian fat pad is located in the immediate vicinity of the ovary and uterine tube (aka oviduct, fallopian tube), and provides a familiar microenvironment for cells and tissues of these organs. In our approach fluorescence-labeled mouse and human cells, and fragments of the uterine tube are engrafted by using minimally traumatic dorsal incision surgery. Transplanted cells and their outgrowths are easily located in the ovarian fat pad for over 40 days. Long-term transplantation of the entire uterine tube allows correct preservation of all principle tissue components, and does not result in adverse side effects, such as fibrosis and inflammation. Our approach should be uniquely applicable for answering important biological questions such as differentiation, regenerative and neoplastic potential of specific cell populations. Furthermore, it should be suitable for studies of microenvironmental factors in normal development and cancer.

We describe an ovarian fad pad transplantation assay suitable for studies of normal and transformed epithelia of the female reproductive tract. The mouse fat pad allows transplantation of large tissue fragments, is easily accessible for surgery and imaging, and provides the most favorable native environment for tissues of M&#252;llerian origin.

Transplantation Dans le tapis de souris ovarien Fat

Orthotopic transplantation assays in mice are invaluable for studies of cell regeneration and neoplastic transformation. Common approaches for orthotopic ...

A Technique for Subcutaneous Abdominal Adipose Tissue Biopsy via a Non-diathermy Method

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To obtain subcutaneous adipose tissue biopsies in the abdominal area, researchers and physicians use either a surgical or a needle-based technique. However, surgical subcutaneous fat biopsies can offer tissue samples that may provide a more comprehensive overview of the complexities of biological indices in white adipose tissue. Usually, a surgical adipose tissue biopsy includes a diathermy treatment for cauterizing blood vessels to prevent excessive bleeding. Nevertheless, side effects, such as flash fires and skin lesions in the tissue, have been reported after diathermy. Therefore, we aimed to standardize a surgical abdominal adipose tissue biopsy performed under local anesthesia using a non-diathermy method. We conducted 115 subcutaneous adipose tissue biopsies in healthy men using a non-diathermy abdominal surgical biopsy method. Our results showed three cases of excessive post-operation bleeding out of 115 operations (2.61%).In conclusion, our standardized subcutaneous abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men at the bedside, with minimal side effects.

We standardized an abdominal adipose tissue biopsy using a non-diathermy method performed under local anesthesia. Three cases of excessive post-operation bleeding out of 115 operations (2.61%) occurred.We conclude that an abdominal adipose tissue surgical biopsy using a non-diathermy method can be safely applied to healthy men.

Une Technique de biopsie du tissu adipeux sous-cutané Abdominal via une méthode Non-diathermie

Adipose tissue biopsies offer tissue samples that, upon analysis, may provide insightful overviews of mechanisms relating to metabolism and disease. To ...

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 &#181;m internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Videomicroscopy systems are used to examine functional properties of isolated adipose tissue arterioles in response to physiological and pharmacological stimuli. This technique can be used to examine microvascular phenotypes in different adipose tissue domains in obese humans.

Évaluation de la fonction microvasculaire tissu adipeux humain en utilisant la vidéomicroscopie

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. ...

Transplantation of Adipose Tissue-Derived Stem Cell Sheet to Reduce Leakage After Partial Colectomy in A Rat Model

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of clinical efficacy, they are until now imperfect solutions. Stem cell therapy using ASC sheets could provide a solution to this problem. ASCs are considered as promising candidates for promoting tissue healing because of their trophic and immunomodulatory properties. Here, we provide methods to produce high-density ASC sheets, that are transplanted onto a colorectal anastomosis in a rat model to reduce the leakage. ASCs formed cell sheets in thermo-responsive culture dishes that could be easily detached. On the day of the transplantation, a partial colectomy with a 5-suture colorectal anastomosis was performed. Animals were immediately transplanted with 1 ASC sheet per rat. ASC sheets adhered spontaneously to the anastomosis without any glue, suture, or any biomaterial. Animal groups were sacrificed 3 and 7 days postoperatively. Compared to transplanted animals, the incidence of anastomotic abscesses and leakage was higher in control animals. In our model, the transplantation of ASC sheets after colorectal anastomosis was successful and associated with a lower leakage rate.

The preparation and transplantation of adipose tissue derived stem cell (ASC) sheets onto insufficiently sutured colorectal anastomoses in a rat model is presented. This novel application shows that ASC sheets can reduce colorectal anastomosis leakage.

Transplantation de la feuille les cellules souches dérivées du tissu adipeux pour réduire les fuites après colectomie partielle dans un modèle de Rat

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of ...

Clinical Protocol of Producing Adipose Tissue-Derived Stromal Vascular Fraction for Potential Cartilage Regeneration

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the knees by using different forms of mesenchymal stem cells (MSCs). In Korea, bone marrow concentrates and cord blood-derived stem cells have been approved by the Korean Food and Drug Administration (KFDA) for cartilage regeneration. In addition, an adipose tissue-derived stromal vascular fraction (SVF) has been allowed by the KFDA for joint injections in human patients. Autologous adipose tissue-derived SVF contains extracellular matrix (ECM) in addition to mesenchymal stem cells. ECM excretes various cytokines that, along with hyaluronic acid (HA) and platelet-rich plasma (PRP) activated by calcium chloride, may help MSCs to regenerate cartilage and improve knee functions. In this article, we presented a protocol to improve knee functions by regenerating cartilage-like tissue in human patients with OA. The result of the protocol was first reported in 2011 followed by a few additional publications. The protocol involves liposuction to obtain autologous lipoaspirates that are mixed with collagenase. This lipoaspirates-collagenase mixture is then cut and homogenized to remove large fibrous tissue that may clog up the needle during the injection. Afterwards, the mixture is incubated to obtain adipose tissue-derived SVF. The resulting adipose tissue-derived SVF, containing both adipose tissue-derived MSCs and remnants of ECM, is injected into knees of patients, combined with HA and calcium chloride activated PRP. Included are three cases of patients who were treated with our protocol resulting in improvement of knee pain, swelling, and range of motion along with MRI evidence of hyaline cartilage-like tissue.

Here, we present a protocol to produce an adipose tissue-derived stromal vascular fraction and its application to improve knee functions by regenerating cartilage-like tissue in human patients with osteoarthritis.

Protocole clinique de produire des dérivés du tissu adipeux stroma vasculaire Fraction pour la régénération du Cartilage potentiels

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the ...