Os autores agradecem a Texas A &amp; M University Engenharia e Estação Experimental (T) para apoio financeiro desta pesquisa. Lindsay prego agradece o apoio do Texas A &amp; M University Louis Stokes Aliança de Participação minoritária (LSAMP) ea National Science Foundation (NSF) Programa de Bolsas de Pós-Graduação de Pesquisa (GRFP). Zhang Dawei, graças a Texas A &amp; M University Dissertação companheirismo.

1. Sintetizando PCL-DA Macrómero <ol><li> Execute a reação acrylation. <ol><li> Pesar 20 g de PCL-diol (M n = ~ 10000 g / mol) em frasco de 250 ml de fundo redondo equipado com uma barra de agitação magnética coberta de teflon. </li><li> Dissolve-se o PCL-diol em DCM. <ol><li> Adicionar 120 mLlof DCM para o frasco (concentração = 0,17 g / ml). </li><li> Coloque um septo de borracha frouxamente dentro do gargalo do frasco de modo a evitar a acumulação de pressão enquanto também previne a evaporação do DCM. </li><li> Solução agitar durante ~ 30 min a ~ 250 rpm, para dissolver completamente o polímero. </li></ol></li><li> Adicionar ~ 6,6 mg de 4-dimetilaminopiridina (DMAP) à solução e dissolver sob agitação. </li><li> Coloque um septo de borracha de forma segura no gargalo do frasco. Permitir que a solução de continuar a agitação. </li><li> Através do septo de borracha, limpar suavemente o frasco com N2 durante ~ 3 min usando uma entrada de N2 positiva agulha e uma agulha de pressão abertacomo uma saída. </li><li> Remover a entrada de N2 e de saída. </li><li> Adicionar 0,56 ml (4,0 mmol) de trietilamina (Et3N) gota a gota através de uma seringa de vidro equipada com uma agulha introduzida através do septo de borracha. </li><li> Adicionar 0,65 ml (8,0 mmol) de cloreto de acriloílo, gota a gota através de uma seringa de vidro equipada com uma agulha introduzida através do septo de borracha. </li><li> E volta a uma entrada de N2 ao balão e permitir que o conteúdo a agitar sob pressão positiva de N2 durante ~ 30 min. </li><li> Pré-aquecer num banho de óleo a 55 ° C. </li><li> Após o atribuído ~ 30 min, remover a entrada de N2 e substituir o septo com um condensador. </li><li> Mergulhe o balão no banho de óleo pré-aquecido. </li><li> Deixar o conteúdo do balão em agitação durante 20 h. </li><li> Após 20 h a alocado, retirar o balão do banho de óleo e permite que o conteúdo arrefecer até à TA. </li><li> Usando um evaporador rotativo, remover o solvente DCM a partir do frasco. </li></ol></li><li> Purify o produto em bruto PCL-DA. <ol><li> Ao balão, adicionar ~ 135 ml de acetato de etilo e dissolve-se o produto em bruto PCL-DA. </li><li> Gravidade filtrar a solução através de papel de filtro para um balão de fundo redondo de 250 ml limpo. (Nota: A solução pode engrossar no papel de filtro, não prontamente passar através Se assim for, cuidadosamente aplicar calor suave com uma pistola de aquecimento.). </li><li> Usando um evaporador rotativo, remover solvente acetato de etilo a partir do frasco. </li><li> Ao balão, adicionar ~ 140 ml de DCM e dissolver o produto em bruto PCL-DA. </li><li> Transferir o conteúdo para um funil de separação de 500 mL. </li><li> Para o funil, adicionar 13,5 mL de carbonato de potássio 2 M (K 2 CO 3). </li><li> Tapar o funil. Misture delicadamente as duas camadas, invertendo o funil e rodando suavemente uma ou duas vezes, tomando cuidado para liberar a pressão através da torneira. Repita 3 vezes. </li><li> Substituir a tampa com uma camada de Parafilm e permitir que a mistura para separar S / N (~ 12 h). </li><li> Colete o fundo, oucamada gânica para um balão de Erlenmeyer de 250 mL. </li><li> Adicionar ~ 5 g de sulfato de magnésio anidro (MgSO4) ao balão e agitar suavemente. </li><li> Gravidade filtrar a mistura através de papel filtro qualitativo e em um ambiente limpo 250 ml balão de fundo redondo. </li><li> Usando um evaporador rotativo, remover o solvente DCM a partir do frasco. </li><li> Seca-se sob alto vácuo para remover o DCM residual. (Nota: PCL-DA devem ser armazenados longe da luz.) </li><li> Confirme acrylation com 1H NMR 22,23. </li></ol></li></ol> 2. Preparar o SMP Andaime (Figura 1) <ol><li> Prepare o modelo de sal fundido. <ol><li> Use de um peneiro de 425 | iM para se obter cloreto de sódio (NaCl) 460 ~ partículas ± 70 um de diâmetro. (Nota: O tamanho médio de partícula pode ser confirmado a partir de microscopia eletrônica de varredura [SEM] imagens com o software ImageJ.) 14 </li><li> Para um frasco de vidro de 3 ml (ID = 12,9 mm), adicionar 1,8 g de NaCl previamente peneirado. </li><li> Adiciona-se lentamente, em quatro porções, 7,5% em peso (com base no peso de sal) água Dl (0,146 g), para o frasco. Misturar com uma espátula de metal, após a adição de cada porção de água. </li><li> Tapar o frasco, enrole em tecido e colocar na vertical num tubo de centrifugação. Centrifugar durante 15 minutos a 3220 x g. </li><li> Retire a tampa e deixe secar ao ar O / N (~ 12 horas). </li></ol></li><li> Em um novo frasco de vidro, preparar uma &quot;solução de macrómero&quot; pela combinação de 0,15 g de PCL-DA por ml de DCM. (Nota:. Para um andaime, ~ 1 ml de solução deve ser preparado) Tampão e misturar a solução a alta velocidade num misturador vortex durante ~ 1 min. </li><li> Em um novo frasco de vidro de 3 ml, preparar uma &quot;solução de foto-iniciador&quot; com base em 10% em peso de 2,2-dimetoxi-2-fenil-acetofenona (DMP) em 1-vinil-2-pirrolidinona (NVP). Combinar 0,115 g de DMP em 1 ml de NVP. (Nota: Para um andaime, ~ 180 ul é necessária.) Cap, embrulhe em papel alumínio (para bloquear a luz) e misturar a solução em alta velocidade em um misturador de vórticepara ~ 1 min. (Nota: Se preparada com antecedência, a solução deve ser refrigerada e protegida da luz). </li><li> Envolver o frasco contendo a solução de macrómero (excluindo a tampa) com folha de alumínio (para bloquear a luz) e adicionar através de uma pipeta de 15% em volume (baseada no volume total da solução de macrómero) da solução de foto-iniciador. Tampão e misturar a solução a alta velocidade num misturador vortex durante ~ 1 min. </li><li> Envolver o frasco que contém o molde sal fundido (excluindo a tampa) com folha de alumínio (para bloquear a luz) através de uma pipeta e adicionar a solução de macrómero / fotoiniciador previamente preparada (~ 0,6 ml ou até que o molde está completamente coberta). </li><li> Tapar o frasco, enrole em tecido e colocar na vertical num tubo de centrifugação. Centrifugar durante 10 minutos a 1.260 xg, para distribuir a solução de macrómero ao longo do molde. </li><li> Remover a película de alumínio, uncap o frasco e expor à luz UV (365 nm, 25 W) durante 3 min. O ar seco / N. </li><li> Retire o &quot;scaffol contendo sald &quot;do frasco para injectáveis ​​com uma pinça depois de marcar e fraturando a parte superior do frasco de vidro. </li><li> Em um copo de 400 ml, prepara ~ 200 ml de uma água / solvente de etanol (1: 1 vol: vol). </li><li> Manter o andaime submerso na água / solvente de etanol durante 4 dias, com mudanças de solvente diárias. </li><li> Remover o andaime do solvente e seca ao ar, O / N. </li></ol> 3. Aplicando polydopamine Coating para SMP Andaime (Figura 1) <ol><li> Em 400 ml equipado com uma barra de agitação coberta de teflon, prepara ~ 200 ml de uma solução de cloridrato de dopamina (2 mg / ml em tampão Tris 10 mM, pH = 8,5, 25 ºC). Agita-se a ~ 150 rpm. </li><li> Coloque uma agulha descartável (comprimento = 40 mm; calibre = 20) para o cadafalso, ~ metade da distância através do andaime. Enrole um fio ao redor do centro da agulha. </li><li> Submerge o andaime (com o cubo de agulha acima da superfície da solução) na solução de dopamina agitação por ancoragem do fio até a borda do copo. </li><li> Desgasificar o andaime, colocando uma seringa para dentro do cubo de agulha e utilizando-a para puxar o ar para fora do andaime. (Nota: A desgaseificação é completa quando não mais ar pode ser removido e a solução foi completamente infiltrado o andaime.) </li><li> Manter o andaime submerso na solução de dopamina de se agitar durante 16 h. </li><li> Remover o andaime a partir da solução e remover a agulha. Lavar com água desionizada e seco numa estufa de vácuo à temperatura ambiente durante 24 h. </li><li> Colocar o andaime num forno de 85 ° C durante 1 h. </li><li> Permitir que o andaime arrefecer até à TA. O andaime cilíndrico final será ~ 6 mm de diâmetro x ~ 5 mm de altura. </li></ol> 4. Avaliando o comportamento &quot;auto-montagem&quot; <ol><li> Preparar um &quot;modelo de defeito CMF irregular&quot; utilizando uma folha de plástico rígido cuja espessura é de aproximadamente 5 mm. Utilize uma broca para criar um vácuo dentro da folha de plástico com um diâmetro médio ligeiramente inferior a ~ 6 mm, como demonstrado na Figura 2A. </li><li> Em abEaker, água Dl calor (representando o uso clínico de solução salina) a uma temperatura de ~ 60 ° C. </li><li> Coloque o andaime para o copo de água ~ 60 ºC. Utilize uma pinça para empurrar o andaime abaixo da superfície da água, expondo todos os domínios para a água. Continuar para ~ 2 min ou até que o andaime é visivelmente amoleceu (Figura 2B). </li><li> Remova o cadafalso na taça e pressione imediatamente (com a mão) no defeito modelo. </li><li> Deixa-se arrefecer até à temperatura ambiente (~ 5-10 min) (Figura 2C). </li><li> Retire do defeito para observar o novo, forma fixa temporária e o retorno do estado relativamente mais rígida (Figura 2D). </li></ol> 5. Teste de Comportamento Memória de Forma <ol><li> Utilizando um analisador mecânico dinâmico (DMA; por exemplo, um TA Instruments Q800, tal como aqui utilizado), executar um ensaio de compressão com deformação controlada cíclico térmico-mecânica sobre uma estrutura de suporte ao longo de dois ciclos de (N) para determinar a forma fixidez (R f) emoldar recuperação (R r) (Figura 3). <ol><li> Equilibrar a 60 ° C (T alto) durante 5 min. </li><li> Comprimir a uma tensão máxima (ε m = 50%) a 50% / min. </li><li> Manter a ε m (5 min). </li><li> Arrefecer até 25 ° C (T baixo) e manter durante 10 min para fixar a forma temporária. </li><li> Remover a carga. </li><li> Meça a tensão final no estado livre de estresse (ε u). </li><li> Aquecer de novo para 60 ° C (T alta) e manter durante 10 min para recuperar a forma permanente. </li><li> Meça a tensão recuperado (ε p). </li><li> Enquanto ainda a 60 ° C (T alta), iniciar o ciclo de ND 2 (N = 2), comprimindo o andaime a 50% da altura recuperado após o ciclo de r 1 (n = 1). </li><li> Repita 5.1.3-5.1.8 Para N = 2. </li><li> Calcular R f R e R para N = 1 e 2, utilizando as seguintes equações: R f (n) = [ε L (N) / ε m] r r (n) = [m ε - ε P (N)] / [m ε - ε p (N-1)] </li></ol></li></ol> 6. A visualização Pore Tamanho e Pore Interconectividade <ol><li> Microscopia eletrônica de varredura (SEM; por exemplo, uma FEI Quanta SEM como aqui utilizado), observar o tamanho dos poros e poros interconectividade. <ol><li> Com uma pinça para segurar o andaime SMP, submergir em N2 líquido durante 1 min. </li><li> Remover do N2 líquido e fractura ao longo do meio do andaime com uma lâmina de barbear limpo. </li><li> Usando fita de carbono, apor uma das metades do SMP andaime sobre a amostrapalco com a superfície fraturada voltado para cima. </li><li> Por pulverização catódica, revestimento com Au-Pt (~ 4 nm). </li><li> Capturar a imagem SEM a uma voltagem de aceleração de 10-15 kV recomendado (Figura 4A). </li></ol></li></ol> 7. Testes in vitro de bioatividade <ol><li> Num tubo de centrífuga de 50 ml, adicionar 30 ml de ~ 1X SBF 24. </li><li> Obter um andaime em sua forma original é permanente, cilindricamente-moldado. Corte o andaime ao meio (em toda a borda circular) usando uma lâmina limpo. </li><li> Coloque um andaime metade indivíduo no tubo de centrífuga preparada e cap. </li><li> Manter o tubo a 37 ° C num banho de água em condições estáticas sem alterações SBF. </li><li> Após 14 dias, remover o andaime de SBF e secar ao ar durante 24 horas. </li><li> Usando a fita de carbono, apor o andaime para o palco amostra com a superfície fraturada voltado para cima. </li><li> Por pulverização catódica, revestimento com Au-Pt (~ 4 nm). </li><li> Capturar a imagem SEM em um rerecomendou tensão de aceleração de 10-15 kV (Figura 4B). </li></ol>

<ol>
	<li>Neovius, E., Engstrand, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Craniofacial+reconstruction+with+bone+and+biomaterials:+review+over+the+last+11+years.">Craniofacial reconstruction with bone and biomaterials: review over the last 11 years.</a> J Plast Reconstr Aesthet Surg. 63, 1615-1623 (2010).</li><li>Elsalanty, M. E., Genecov, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bone+grafts+in+craniofacial+surgery.">Bone grafts in craniofacial surgery.</a> Craniomaxillofac Trauma Reconstr. 2, 125-134 (2009).</li><li>Hollister, S. 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=Engineering+craniofacial+scaffolds.">Engineering craniofacial scaffolds.</a> Orthod Craniofacial Res. 8, 162-173 (2005).</li><li>Albrektsson, T., Johansson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteoinduction,+osteoconduction+and+osseointegration.">Osteoinduction, osteoconduction and osseointegration.</a> Eur Spine J. 10, S96-S101 (2001).</li><li>Blokhuis, T. J., Arts, J. 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=Bioactive+and+osteoinductive+bone+graft+substitutes:+Definitions,+facts+and+myths.">Bioactive and osteoinductive bone graft substitutes: Definitions, facts and myths.</a> Injury. 42, S26-S29 (1016).</li><li>Zhang, D., 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=A+bioactive+&#8220;self-fitting&#8221;+shape+memory+polymer+scaffold+with+potential+to+treat+cranio-maxillo+facial+bone+defects.">A bioactive &#8220;self-fitting&#8221; shape memory polymer scaffold with potential to treat cranio-maxillo facial bone defects.</a> Acta Biomater. 10, 4597-4605 (2014).</li><li>Lendlein, A., Kelch, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shape-memory+polymers.">Shape-memory polymers.</a> Angew. Chem. Int. Ed. 41, 2034-2057 (2002).</li><li>Hu, J., Zhu, Y., Huang, H., Lu, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+advances+in+shape-memory+polymers:+Structure,+mechanism,+functionality,+modeling+and+applications.">Recent advances in shape-memory polymers: Structure, mechanism, functionality, modeling and applications.</a> Prog Polym Sci. 37, 1720-1763 (2012).</li><li>Middleton, J. C., Tipton, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthetic+biodegradable+polymers+as+orthopedic+devices.">Synthetic biodegradable polymers as orthopedic devices.</a> Biomaterials. 21, 2335-2346 (2000).</li><li>Sun, H., Mei, L., Song, C., Cui, X., Wang, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+in+vivo+degradation,+absorption+and+excretion+of+PCL-based+implant.">The in vivo degradation, absorption and excretion of PCL-based implant.</a> Biomaterials. 27, 1735-1740 (2006).</li><li>Woodruff, M. A., Hutmacher, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+return+of+a+forgotten+polymer-Polycaprolactone+in+the+21st+century.">The return of a forgotten polymer-Polycaprolactone in the 21st century.</a> Prog Polym Sci. 35, 1217-1256 (2010).</li><li>Wang, S., Lu, L., Gruetzmacher, J. A., Currier, B. L., Yaszemski, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synthesis+and+characterizations+of+biodegradable+and+crosslinkable+poly(&#949;-caprolactone+fumarate),+poly(ethylene+glycol+fumarate),+and+their+amphiphilic+copolymer.">Synthesis and characterizations of biodegradable and crosslinkable poly(&#949;-caprolactone fumarate), poly(ethylene glycol fumarate), and their amphiphilic copolymer.</a> Biomaterials. 27, 832-841 (2006).</li><li>Zhang, D., Petersen, K. M., Grunlan, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inorganic-organic+shape+memory+polymer+(SMP)+foams+with+highly+tunable+properties.">Inorganic-organic shape memory polymer (SMP) foams with highly tunable properties.</a> ACS Appl Mater Interfaces. 5, 186-191 (2012).</li><li>Zhang, D., Burkes, W. L., Schoener, C. A., Grunlan, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porous+inorganic-organic+shape+memory+polymers.">Porous inorganic-organic shape memory polymers.</a> Polymer. 53, 2935-2941 (2012).</li><li>Van der Stok, J., Van Lieshout, E. M., El-Massoudi, Y., Van Kralingen, G. H., Patka, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bone+substitutes+in+the+Netherlands-a+systematic+literature+review.">Bone substitutes in the Netherlands-a systematic literature review.</a> Acta Biomater. 7, 739-750 (2011).</li><li>Lee, H., Dellatore, S. M., Miller, W. M., Messersmith, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mussel-inspired+surface+chemistry+for+multifunctional+coatings.">Mussel-inspired surface chemistry for multifunctional coatings.</a> Science. 318, 426-430 (2007).</li><li>Hong, 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=Non-covalent+self-assembly+and+covalent+polymerization+co-contribute+to+polydopamine+formation.">Non-covalent self-assembly and covalent polymerization co-contribute to polydopamine formation.</a> Adv Funct Mater. 22, 4711-4717 (2012).</li><li>Ryu, J., Ku, S. H., Lee, H., Park, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mussel-inspired+polydopamine+coating+as+a+universal+route+to+hydroxyapatite+crystallization.">Mussel-inspired polydopamine coating as a universal route to hydroxyapatite crystallization.</a> Adv Funct Mater. 20, 2132-2139 (2010).</li><li>Lee, Y. 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=Polydopamine-mediated+immobilization+of+multiple+bioactive+molecules+for+the+development+of+functional+vascular+graft+materials.">Polydopamine-mediated immobilization of multiple bioactive molecules for the development of functional vascular graft materials.</a> Biomaterials. 33, 8343-8352 (2012).</li><li>Wu, C., Fan, W., Chang, J., Xiao, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mussel-inspired+porous+SiO+2+scaffolds+with+improved+mineralization+and+cytocompatibility+for+drug+delivery+and+bone+tissue+engineering.">Mussel-inspired porous SiO 2 scaffolds with improved mineralization and cytocompatibility for drug delivery and bone tissue engineering.</a> J Mater Chem. 21, 18300-18307 (2011).</li><li>Ku, S. H., Ryu, J., Hong, S. K., Lee, H., Park, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=General+functionalization+route+for+cell+adhesion+on+non-wetting+surfaces.">General functionalization route for cell adhesion on non-wetting surfaces.</a> Biomaterials. 31, 2535-2541 (2010).</li><li>Schoener, C. A., Weyand, C. B., Murthy, R., Grunlan, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shape+memory+polymers+with+silicon-containing+segments.">Shape memory polymers with silicon-containing segments.</a> J Mater Chem. 20, 1787-1793 (2010).</li><li>Zhang, D., Giese, M. L., Prukop, S. L., Grunlan, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Poly(&#949;-caprolactone)-based+shape+memory+polymers+with+variable+polydimethylsiloxane+soft+segment+lengths.">Poly(&#949;-caprolactone)-based shape memory polymers with variable polydimethylsiloxane soft segment lengths.</a> J Polym Sci Pol Chem. 49, 754-761 (2011).</li><li>Kokubo, T., Takadama, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+useful+is+SBF+in+predicting+in+vivo+bone+bioactivity.">How useful is SBF in predicting in vivo bone bioactivity.</a> Biomaterials. 27, 2907-2915 (2006).</li><li>Murphy, W. L., Dennis, R. G., Kileny, J. L., Mooney, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Salt+fusion:+an+approach+to+improve+pore+interconnectivity+within+tissue+engineering+scaffolds.">Salt fusion: an approach to improve pore interconnectivity within tissue engineering scaffolds.</a> Tissue Eng. 8, 43-52 (2002).</li><li>Yang, Q., Chen, L., Shen, X., Tan, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+of+polycaprolactone+tissue+engineering+scaffolds+by+improved+solvent+casting/particulate+leaching+method.">Preparation of polycaprolactone tissue engineering scaffolds by improved solvent casting/particulate leaching method.</a> J Macromol Sci Phys. 45, 1171-1181 (2006).</li><li>Madbouly, S. A., Kratz, K., Klein, F., L&#252;zow, K., Lendlein, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thermomechanical+behaviour+of+biodegradable+shape-memory+polymer+foams.">Thermomechanical behaviour of biodegradable shape-memory polymer foams.</a> 1190, (2009).</li><li>Luo, X., Mather, P. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparation+and+characterization+of+shape+memory+elastomeric+composites.">Preparation and characterization of shape memory elastomeric composites.</a> Macromolecules. 42, 7251-7253 (2009).</li><li>Lendlein, A., Schmidt, A. M., Langer, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=AB-polymer+networks+based+on+oligo(&#603;-caprolactone)+segments+showing+shape-memory+properties.">AB-polymer networks based on oligo(&#603;-caprolactone) segments showing shape-memory properties.</a> Proc Natl Acad Sci. 98, 842-847 (2001).</li></ol>

The authors have nothing to disclose.

Este protocolo descreve a preparação de, uma estrutura de suporte com base em PCL revestidos por polydopamine cujo comportamento de auto-montagem, bem como osteoindutividade e bioactividade, faz com que seja de interesse para o tratamento de defeitos ósseos irregulares CMF. Aspectos do protocolo pode ser alterado para alterar várias características de andaime. O protocolo começa com um acrylation de PCL-diol para permitir a cura por UV. No exemplo relatados, o PCL-diol H n é</sub...

Atualmente considerada o padrão ouro do crânio-maxilo-facial (CMF) Tratamentos de defeito ósseo, o transplante de enxerto autólogo colhido é prejudicado por procedimentos de enxerto complexos, área doadora morbidade e disponibilidade limitada 1. Uma dificuldade particular está a moldar e que fixa o auto-enxerto rígida firmemente no defeito de modo a obter a osteointegração e para evitar a reabsorção do enxerto. A engenharia de tecidos tem sido investigado como uma estratégia alternativa a auto-enxertia e substitutos ósseos artificiais (por exemplo, cimento ósseo) 2,3. Crítica para o sucesso de uma abordagem de engenharia de tecidos é um andaime com um conjunto específico de propriedades. Em primeiro lugar, a fim de alcançar a osteointegração, o andaime deve formar um contacto estreito com o tecido ósseo adjacente 4. O andaime também deve ser osteocondutor, permitindo a migração celular, difusão de nutrientes e neotissue 4,5 deposição. Este comportamento é geralmente conseguida com sca biodegradávelffolds exibindo uma morfologia de poro altamente interligada. Por último, o andaime deve ser bioativa, de modo a promover a integração e ligação com áreas de tecido ósseo 5. Aqui, apresentamos um protocolo para preparar um andaime engenharia de tecidos com estas propriedades. Importante, este andaime exibe a capacidade de &quot;auto-ajuste&quot; em CMF irregulares defeitos, devido ao seu comportamento com memória de forma 6. Polímeros com memória de forma Thermoresponsive (SMPS) são conhecidos por sofrerem alteração da forma, por exposição ao calor 7,8. SMPs são compostas de &quot;pontos de Netpoint&quot; (isto é ligações cruzadas físicas ou químicas) que determinam a forma permanente e &quot;segmentos de comutação&quot; que mantêm a forma temporária e recuperar a forma permanente. Os segmentos de comutação exibem uma temperatura de transição térmica (t trans) correspondente, quer ao de transição vítrea (Tg) de transição ou fusão (Tm) do polímero. Comoresultado, SMPs pode ser deformado sequencialmente em uma forma temporária no T&gt; T trans, fixado na forma temporária no T &lt;T trans, e recuperou-se à forma permanente no T&gt; T trans. Assim, um andaime SMP poderia conseguir &quot;auto-ajuste&quot; dentro de um defeito CMF da seguinte 6. Após a exposição a aquecer salina (T&gt; T trans), um andaime SMP iria se tornar maleável, permitindo uma cilíndrico andaime genericamente preparado para ser em um defeito irregular mão-pressionado, com recuperação de forma promovendo a expansão do andaime para o limite defeito. Com o arrefecimento (T &lt;T trans), o andaime voltaria ao seu estado relativamente mais rígida, com forma fixidez manter sua nova forma temporária dentro do defeito. Neste protocolo, um andaime SMP é preparado a partir de policaprolactona (PCL), um polímero biodegradável estudado extensivamente para a regeneração de tecidos e outras aplicações biomédicas 9-11. Para memória de forma, thE T M de PCL serve como a trans t e varia entre 43 e 60 ° C, dependendo do peso molecular do PCL 12. Neste protocolo, o T trans (ou seja, T m) do andaime é 56,6 ± 0,3 ºC 6. A fim de alcançar osteocondutividade, foi desenvolvido um protocolo para fazer andaimes SMP baseados em PCL com poros altamente interligados com base num método de fundição 6,13,14-solvente partículas de lixiviação (SCPL). Diacrilato de policaprolactona (PCL-DA) (M = N ~ 10000 g / mol) foi utilizado para permitir a rápida, reticulação fotoquímica e foi dissolvido em diclorometano (DCM) para permitir-solvente sobre o molde de fundição de sal. Após cura fotoquímica e evaporação do solvente, o molde sal foi removido por lixiviação em água. O tamanho médio de sal regula o tamanho de poro andaime. Mais importante, o modelo de sal foi fundido com água antes de o solvente de fundição para alcançar poro interconnectivity. A bioactividade foi conferida ao andaime SMP pela formação in situ de um revestimento nas paredes dos poros polydopamine 6. A bioactividade é frequentemente introduzido no andaimes pela inclusão de vidro ou de vidro cerâmico de enchimento 15. Contudo, estes podem dar origem a propriedades mecânicas indesejadas quebradiças. A dopamina tem sido mostrado para formar, uma camada aderente fina polydopamine em uma variedade de substratos 16-19. Neste protocolo, o andaime PMS foi submetido a uma solução ligeiramente básica (pH = 8,5) de dopamina para formar um revestimento de polydopamine nanothick em todas as superfícies de parede do poro 6. Além de aumentar a hidrofilicidade da superfície para melhorar a adesão e espalhamento, polydopamine tem sido mostrado para ser bioactivo em termos de formação de hidroxiapatite (HAp) após exposição ao fluido corporal simulado (SBF) 18,20,21. Num último passo, o andaime revestido é exposto a um tratamento térmico a 85 ° C (T&gt; T trans) WHich leva a scaffold densificação. O tratamento térmico foi observado anteriormente a ser essencial para o comportamento de memória de forma andaime, talvez devido a domínios cristalinos PCL reorganizando a maior proximidade 14. Nós descrevemos adicionalmente os métodos para caracterizar o comportamento de auto-montagem dentro de um defeito modelo irregular, moldar o comportamento de memória em termos de ensaios de compressão mecânica cíclico térmicos com deformação controlada (isto é, recuperação da forma e moldar fixidez), morfologia de poros, e de bioactividade in vitro. São também apresentadas estratégias para adequar propriedades de andaime.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Polycaprolactone-diol (Mn ~ 10,000 g/mol)</td>
			<td>Sigma-Aldrich</td>
			<td>440752</td>
			<td></td>
		</tr>
		<tr>
			<td>Dichloromethane (DCM)</td>
			<td>Sigma-Aldrich</td>
			<td>D65100</td>
			<td>Dried over 4A molecular sieves</td>
		</tr>
		<tr>
			<td>4-dimethylaminopyridine (DMAP)</td>
			<td>Sigma-Aldrich</td>
			<td>D5640</td>
			<td></td>
		</tr>
		<tr>
			<td>Triethylamine (Et3N)</td>
			<td>Sigma-Aldrich</td>
			<td>T0886</td>
			<td></td>
		</tr>
		<tr>
			<td>Acryloyl chloride</td>
			<td>Sigma-Aldrich</td>
			<td>A24109</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethyl Acetate</td>
			<td>Sigma-Aldrich</td>
			<td>319902</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium Carbonate (K2CO3)</td>
			<td>Sigma-Aldrich</td>
			<td>209619</td>
			<td></td>
		</tr>
		<tr>
			<td>Anhydrous magnesium sulfate (MgSO4)</td>
			<td>Fisher</td>
			<td>M65</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium chloride (NaCl)</td>
			<td>Sigma-Aldrich</td>
			<td>S9888</td>
			<td></td>
		</tr>
		<tr>
			<td>2,2-dimethoxy-2-phenyl acetophenone (DMP)</td>
			<td>Sigma-Aldrich</td>
			<td>196118</td>
			<td></td>
		</tr>
		<tr>
			<td>1-vinyl-2-pyrrolidinone (NVP)</td>
			<td>Sigma-Aldrich</td>
			<td>V3409</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol</td>
			<td>Sigma-Aldrich</td>
			<td>459844</td>
			<td></td>
		</tr>
		<tr>
			<td>Dopamine Hydrochloride</td>
			<td>Sigma-Aldrich</td>
			<td>H8502</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris buffer (2mol/L)</td>
			<td>Fisher</td>
			<td>BP1759</td>
			<td>Used at 10 mM concentration, pH = 8.5</td>
		</tr>
		<tr>
			<td>Sieve</td>
			<td>VWR</td>
			<td>47729-972</td>
			<td></td>
		</tr>
		<tr>
			<td>UV-Transilluminator (365 nm, 25 W)</td>
			<td>UVP</td>
			<td>95-0426-02</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Eppendorf</td>
			<td>5810 R</td>
			<td></td>
		</tr>
		<tr>
			<td>Dynamic Mechanical Analyzer (DMA)</td>
			<td>TA Instruments</td>
			<td>Q800</td>
			<td></td>
		</tr>
		<tr>
			<td>High Resolution Sputter Coater</td>
			<td>Cressington</td>
			<td>208HR</td>
			<td></td>
		</tr>
		<tr>
			<td>Scanning Electron Microscope (SEM)</td>
			<td>FEI</td>
			<td>Quanta 600</td>
			<td></td>
		</tr>
	</tbody>
</table>

fabrication of a bioactive, pcl-based "self-fitting" shape memory polymer scaffold

Tissue engineering has been explored as an alternative strategy for the treatment of critical-sized cranio-maxillofacial (CMF) bone defects. Essential to the success of this approach is a scaffold that is able to conformally fit within an irregular defect while also having the requisite biodegradability, pore interconnectivity and bioactivity. By nature of their shape recovery and fixity properties, shape memory polymer (SMP) scaffolds could achieve defect &#8220;self-fitting.&#8221; In this way, following exposure to warm saline (~60 &#186;C), the SMP scaffold would become malleable, permitting it to be hand-pressed into an irregular defect. Subsequent cooling (~37 &#186;C) would return the scaffold to its relatively rigid state within the defect. To meet these requirements, this protocol describes the preparation of SMP scaffolds prepared via the photochemical cure of biodegradable polycaprolactone diacrylate (PCL-DA) using a solvent-casting particulate-leaching (SCPL) method. A fused salt template is utilized to achieve pore interconnectivity. To realize bioactivity, a polydopamine coating is applied to the surface of the scaffold pore walls. Characterization of self-fitting and shape memory behaviors, pore interconnectivity and in vitro bioactivity are also described.

O resultante PMS andaime baseado no PCL é capaz de auto-montagem em um modelo de defeito de CMF (Figura 2). Depois de uma breve exposição a aquecer salina (~ 60 ° C), o andaime cilíndrica amolece permitindo que a escora seja pressionada manualmente para dentro e para expandir dentro do defeito modelo. Após arrefecimento até à TA, o andaime é fixada na sua nova forma temporária, que é mantida após a remoção do defeito. O comportamento de memória de forma de um ...

Watch this Scientific Journal Video about Fabrication of a Bioactive, PCL-based "Self-fitting" Shape Memory Polymer Scaffold at JoVE.com

Fabrication of a Bioactive, PCL-based &quot;Self-fitting&quot; Shape Memory Polymer Scaffold

Scaffolds capable of fitting within cranio-maxillofacial (CMF) bone defects while exhibiting osteoconductivity and bioactivity are of interest. This protocol describes the preparation of a shape memory scaffold based on polycaprolactone diacrylate (PCL-DA) using a solvent-casting particulate-leaching (SCPL) method employing a fused salt template and application of a bioactive polydopamine coating.

Fabricação de uma baseada em PCL Bioativos, &quot;Self-fitting&quot; Memória de Forma Polymer Andaime

Tissue engineering has been explored as an alternative strategy for the treatment of critical-sized cranio-maxillofacial (CMF) bone defects. Essential to ...

fabrication-bioactive-pcl-based-self-fitting-shape-memory-polymer

Research

JoVE Journal

Bioengineering

Fabrication of a Bioactive, PCL-based "Self-fitting" Shape Memory Polymer Scaffold

12.5K visualizações.  Texas A&M University. Scaffolds capable of fitting within cranio-maxillofacial (CMF) bone defects while exhibiting osteoconductivity and bioactivity are of interest. This protocol describes the preparation of a shape memory scaffold based on polycaprolactone diacrylate (PCL-DA) using a solvent-casting particulate-leaching (SCPL) method employing a fused salt template and application of a bioactive polydopamine coating.

Vídeo: Fabrication of a Bioactive, PCL-based "Self-fitting" Shape Memory Polymer Scaffold

Shape Memory Polymers for Active Cell Culture

Shape memory polymers (SMPs) are a class of "smart" materials that have the ability to change from a fixed, temporary shape to a pre-determined permanent shape upon the application of a stimulus such as heat1-5. In a typical shape memory cycle, the SMP is first deformed at an elevated temperature that is higher than its transition temperature, Ttrans [either the melting temperature (Tm) or the glass transition temperature (Tg)]. The deformation is elastic in nature and mainly leads to a reduction in conformational entropy of the constituent network chains (following the rubber elasticity theory). The deformed SMP is then cooled to a temperature below its Ttrans while maintaining the external strain or stress constant. During cooling, the material transitions to a more rigid state (semi-crystalline or glassy), which kinetically traps or "freezes" the material in this low-entropy state leading to macroscopic shape fixing. Shape recovery is triggered by continuously heating the material through Ttrans under a stress-free (unconstrained) condition. By allowing the network chains (with regained mobility) to relax to their thermodynamically favored, maximal-entropy state, the material changes from the temporary shape to the permanent shape.
Cells are capable of surveying the mechanical properties of their surrounding environment6. The mechanisms through which mechanical interactions between cells and their physical environment control cell behavior are areas of active research. Substrates of defined topography have emerged as powerful tools in the investigation of these mechanisms. Mesoscale, microscale, and nanoscale patterns of substrate topography have been shown to direct cell alignment, cell adhesion, and cell traction forces7-14. These findings have underscored the potential for substrate topography to control and assay the mechanical interactions between cells and their physical environment during cell culture, but the substrates used to date have generally been passive and could not be programmed to change significantly during culture. This physical stasis has limited the potential of topographic substrates to control cells in culture.
Here, active cell culture (ACC) SMP substrates are introduced that employ surface shape memory to provide programmed control of substrate topography and deformation. These substrates demonstrate the ability to transition from a temporary grooved topography to a second, nearly flat memorized topography. This change in topography can be used to control cell behavior under standard cell culture conditions.

A method for developing cell culture substrates with the ability to change topography during culture is described. The method makes use of smart materials known as shape memory polymers that have the ability to memorize a permanent shape. This concept is adaptable to a wide range of materials and applications.

Forma Polímeros Memória da Cultura célula ativa

Shape memory polymers (SMPs) are a class of "smart" materials that have the ability to change from a fixed, temporary shape to a pre-determined permanent ...

Bioengenharia

A Chitosan Based, Laser Activated Thin Film Surgical Adhesive, 'SurgiLux': Preparation and Demonstration

Sutures are a 4,000 year old technology that remain the 'gold-standard' for wound closure by virtue of their repair strength (~100 KPa). However, sutures can act as a nidus for infection and in many procedures are unable to effect wound repair or interfere with functional tissue regeneration.1 Surgical glues and adhesives, such as those based on fibrin and cyanoacrylates, have been developed as alternatives to sutures for the repair of such wounds. However, current commercial adhesives also have significant disadvantages, ranging from viral and prion transfer and a lack of repair strength as with the fibrin glues, to tissue toxicity and a lack of biocompatibility for the cyanoacrylate based adhesives. Furthermore, currently available surgical adhesives tend to be gel-based and can have extended curing times which limit their application.2 Similarly, the use of UV lasers to facilitate cross-linking mechanisms in protein-based or albumin 'solders' can lead to DNA damage while laser tissue welding (LTW) predisposes thermal damage to tissues.3 Despite their disadvantages, adhesives and LTW have captured approximately 30% of the wound closure market reported to be in excess of US $5 billion per annum, a significant testament to the need for sutureless technology.4
In the pursuit of sutureless technology we have utilized chitosan as a biomaterial for the development of a flexible, thin film, laser-activated surgical adhesive termed 'SurgiLux'. This novel bioadhesive uses a unique combination of biomaterials and photonics that are FDA approved and successfully used in a variety of biomedical applications and products. SurgiLux overcomes all the disadvantages associated with sutures and current surgical adhesives (see Table 1).
In this presentation we report the relatively simple protocol for the fabrication of SurgiLux and demonstrate its laser activation and tissue weld strength. SurgiLux films adhere to collagenous tissue without chemical modification such as cross-linking and through irradiation using a comparatively low-powered (120 mW) infrared laser instead of UV light. Chitosan films have a natural but weak adhesive attraction to collagen (~3 KPa), laser activation of the chitosan based SurgiLux films emphasizes the strength of this adhesion through polymer chain interactions as a consequence of transient thermal expansion.5 Without this 'activation' process, SurgiLux films are readily removed.6-9 SurgiLux has been tested both in vitro and in vivo on a variety of tissues including nerve, intestine, dura mater and cornea. In all cases it demonstrated good biocompatibility and negligible thermal damage as a consequence of irradiation.6-10

A Chitosan Based, Laser Activated Thin Film Surgical Adhesive, &#039;SurgiLux&#039;: Preparation and Demonstration

The fabrication of a novel, flexible thin film surgical adhesive from FDA approved ingredients, chitosan and indocyanine green is described. Bonding of this adhesive to collagenous tissue through a simple activation process with a low-powered infra-red laser is demonstrated.

A base de quitosana, Laser Activated adesiva Thin Film cirúrgica &quot;, SurgiLux &#39;: Preparação e demonstração

Sutures are a 4,000 year old technology that remain the 'gold-standard' for wound closure by virtue of their repair strength (~100 KPa). However, sutures ...

Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization

Implantable microdevices are gaining significant attention for several biomedical applications1-4. Such devices have been made from a range of materials, each offering its own advantages and shortcomings5,6. Most prominently, due to the microscale device dimensions, a high modulus is required to facilitate implantation into living tissue. Conversely, the stiffness of the device should match the surrounding tissue to minimize induced local strain7-9. Therefore, we recently developed a new class of bio-inspired materials to meet these requirements by responding to environmental stimuli with a change in mechanical properties10-14. Specifically, our poly(vinyl acetate)-based nanocomposite (PVAc-NC) displays a reduction in stiffness when exposed to water and elevated temperatures (e.g. body temperature). Unfortunately, few methods exist to quantify the stiffness of materials in vivo15, and mechanical testing outside of the physiological environment often requires large samples inappropriate for implantation. Further, stimuli-responsive materials may quickly recover their initial stiffness after explantation. Therefore, we have developed a method by which the mechanical properties of implanted microsamples can be measured ex vivo, with simulated physiological conditions maintained using moisture and temperature control13,16,17. 
To this end, a custom microtensile tester was designed to accommodate microscale samples13,17 with widely-varying Young's moduli (range of 10 MPa to 5 GPa). As our interests are in the application of PVAc-NC as a biologically-adaptable neural probe substrate, a tool capable of mechanical characterization of samples at the microscale was necessary. This tool was adapted to provide humidity and temperature control, which minimized sample drying and cooling17. As a result, the mechanical characteristics of the explanted sample closely reflect those of the sample just prior to explantation.
The overall goal of this method is to quantitatively assess the in vivo mechanical properties, specifically the Young's modulus, of stimuli-responsive, mechanically-adaptive polymer-based materials. This is accomplished by first establishing the environmental conditions that will minimize a change in sample mechanical properties after explantation without contributing to a reduction in stiffness independent of that resulting from implantation. Samples are then prepared for implantation, handling, and testing (Figure 1A). Each sample is implanted into the cerebral cortex of rats, which is represented here as an explanted rat brain, for a specified duration (Figure 1B). At this point, the sample is explanted and immediately loaded into the microtensile tester, and then subjected to tensile testing (Figure 1C). Subsequent data analysis provides insight into the mechanical behavior of these innovative materials in the environment of the cerebral cortex.

Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization

A method is discussed by which the in vivo mechanical behavior of stimuli-responsive materials is monitored as a function of time. Samples are tested ex vivo using a microtensile tester with environmental controls to simulate the physiological environment. This work further promotes understanding the in vivo behavior of our material.

Teste de microtração ambientalmente controlado mecanicamente adaptativas nanocompósitos poliméricos para<em&gt; Ex vivo</em&gt; Caracterização

Implantable microdevices are gaining significant attention for several biomedical applications1-4. Such devices have been made from a range of materials, ...

Microfluidic Fabrication of Polymeric and Biohybrid Fibers with Predesigned Size and Shape

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and used to dictate the shape as well as the diameter of a core stream.&#160;Grooves in the top and bottom of a microfluidic channel were designed to direct the sheath fluid and shape the core fluid.&#160;By matching the viscosity and hydrophilicity of the sheath and core fluids, the interfacial effects are minimized and complex fluid shapes can be formed.&#160;Controlling the relative flow rates of the sheath and core fluids determines the cross-sectional area of the core fluid.&#160;Fibers have been produced with sizes ranging from 300 nm to ~1 mm, and fiber cross-sections can be round, flat, square, or complex as in the case with double anchor fibers. Polymerization of the core fluid downstream from the shaping region solidifies the fibers.&#160;Photoinitiated click chemistries are well suited for rapid polymerization of the core fluid by irradiation with ultraviolet light.&#160;Fibers with a wide variety of shapes have been produced from a list of polymers including liquid crystals, poly(methylmethacrylate), thiol-ene and thiol-yne resins, polyethylene glycol, and hydrogel derivatives. Minimal shear during the shaping process and mild polymerization conditions also makes the fabrication process well suited for encapsulation of cells and other biological components.

Two adjacent fluids passing through a grooved microfluidic channel can be directed to form a sheath around a prepolymer core; thereby&#160;determining both shape and cross-section. Photoinitiated polymerization, such as thiol click chemistry, is well suited for rapidly solidifying the core fluid into a microfiber with predetermined size and shape.

Fabricação microfluida de fibras poliméricas e biohibridas com tamanho e forma predesigned

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and ...

Manufacturing Of Robust Natural Fiber Preforms Utilizing Bacterial Cellulose as Binder

A novel method of manufacturing rigid and robust natural fiber preforms is presented here. This method is based on a papermaking process, whereby loose and short sisal fibers are dispersed into a water suspension containing bacterial cellulose. The fiber and nanocellulose suspension is then filtered (using vacuum or gravity) and the wet filter cake pressed to squeeze out any excess water, followed by a drying step. This will result in the hornification of the bacterial cellulose network, holding the loose natural fibers together.
Our method is specially suited for the manufacturing of rigid and robust preforms of hydrophilic fibers. The porous and hydrophilic nature of such fibers results in significant water uptake, drawing in the bacterial cellulose dispersed in the suspension. The bacterial cellulose will then be filtered against the surface of these fibers, forming a bacterial cellulose coating. When the loose fiber-bacterial cellulose suspension is filtered and dried, the adjacent bacterial cellulose forms a network and hornified to hold the otherwise loose fibers together.
The introduction of bacterial cellulose into the preform resulted in a significant increase of the mechanical properties of the fiber preforms. This can be attributed to the high stiffness and strength of the bacterial cellulose network. With this preform, renewable high performance hierarchical composites can also be manufactured by using conventional composite production methods, such as resin film infusion (RFI) or resin transfer molding (RTM). Here, we also describe the manufacturing of renewable hierarchical composites using double bag vacuum assisted resin infusion.

We present a novel method of manufacturing rigid and robust short natural fiber preforms using a papermaking process. Bacterial cellulose acts simultaneously as the binder for the loose fibers and provides rigidity to the fiber preforms. These preforms can be infused with a resin to produce truly green hierarchical composites.

Fabricação de fibras naturais robustos Preformas Utilizando bacteriana Celulose como Binder

A novel method of manufacturing rigid and robust natural fiber preforms is presented here. This method is based on a papermaking process, whereby loose ...

Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness and low bioactivity of metals. In this study, we have developed a new method towards fabricating a new type of bioactive and mechanically reliable porous metal scaffolds-densified porous Ti scaffolds. The method consists of two fabrication processes, 1) the fabrication of porous Ti scaffolds by dynamic freeze casting, and 2) coating and densification of the porous scaffolds. The dynamic freeze casting method to fabricate porous Ti scaffolds allowed the densification of porous scaffolds by minimizing the chemical contamination and structural defects. The densification process is distinctive for three reasons. First, the densification process is simple, because it requires a control of only one parameter (degree of densification). Second, it is effective, as it achieves mechanical enhancement and sustainable release of biomolecules from porous scaffolds. Third, it has broad applications, as it is also applicable to the fabrication of functionally graded porous scaffolds by spatially varied strain during densification.

Bioactive and mechanically reliable metal scaffolds have been fabricated through a method which consists of two processes, dynamic freeze casting for the fabrication of porous Ti, and coating and densification of the Ti scaffolds. The densification process is simple, effective and applicable to the fabrication of functionally graded scaffolds.

Fabricação de Mecanicamente Tunable e Bioativos metal andaimes para aplicações biomédicas

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness ...

Fabrication of Inverted Colloidal Crystal Poly(ethylene glycol) Scaffold: A Three-dimensional Cell Culture Platform for Liver Tissue Engineering

The ability to maintain hepatocyte function in vitro, for the purpose of testing xenobiotics&#39; cytotoxicity, studying virus infection and developing drugs targeted at the liver, requires a platform in which cells receive proper biochemical and mechanical cues. Recent liver tissue engineering systems have employed three-dimensional (3D) scaffolds composed of synthetic or natural hydrogels, given their high water retention and their ability to provide the mechanical stimuli needed by the cells. There has been growing interest in the inverted colloidal crystal (ICC) scaffold, a recent development, which allows high spatial organization, homotypic and heterotypic cell interaction, as well as cell-extracellular matrix (ECM) interaction. Herein, we describe a protocol to fabricate the ICC scaffold using poly (ethylene glycol) diacrylate (PEGDA) and the particle leaching method. Briefly, a lattice is made from microsphere particles, after which a pre-polymer solution is added, properly polymerized, and the particles are then removed, or leached, using an organic solvent (e.g., tetrahydrofuran). The dissolution of the lattice results in a highly porous scaffold with controlled pore sizes and interconnectivities that allow media to reach cells more easily. This unique structure allows high surface area for the cells to adhere to as well as easy communication between pores, and the ability to coat the PEGDA ICC scaffold with proteins also shows a marked effect on cell performance. We analyze the morphology of the scaffold as well as the hepatocarcinoma cell (Huh-7.5) behavior in terms of viability and function to explore the effect of ICC structure and ECM coatings. Overall, this paper provides a detailed protocol of an emerging scaffold that has wide applications in tissue engineering, especially liver tissue engineering.

Fabrication of Inverted Colloidal Crystal Polyethylene glycol Scaffold: A Three-dimensional Cell Culture Platform for Liver Tissue Engineering

This manuscript presents a detailed protocol for the fabrication of an emerging three-dimensional hepatocyte culture platform, the inverted colloidal crystal scaffold, and the concomitant techniques to assess hepatocyte behavior. The size-controllable pores, interconnectivity and ability to conjugate extracellular matrix proteins to the poly(ethylene glycol) (PEG) scaffold enhance Huh-7.5 cell performance.

Fabricação de Inverted coloidal de cristal poli (etileno-glicol) Andaime: Uma Plataforma Cultura celular tridimensional para Liver Engenharia de Tecidos

The ability to maintain hepatocyte function in vitro, for the purpose of testing xenobiotics&#39; cytotoxicity, studying virus infection and developing ...

Fabrication and Characterization of Optical Tissue Phantoms Containing Macrostructure

The rapid development of new optical imaging techniques is dependent on the availability of low-cost, customizable, and easily reproducible standards. By replicating the imaging environment, costly animal experiments to validate a technique may be circumvented. Predicting and optimizing the performance of in vivo and ex vivo imaging techniques requires testing on samples that are optically similar to tissues of interest. Tissue-mimicking optical phantoms provide a standard for evaluation, characterization, or calibration of an optical system. Homogenous polymer optical tissue phantoms are widely used to mimic the optical properties of a specific tissue type within a narrow spectral range. Layered tissues, such as the epidermis and dermis, can be mimicked by simply stacking these homogenous slab phantoms. However, many in vivo imaging techniques are applied to more spatially complex tissue where three dimensional structures, such as blood vessels, airways, or tissue defects, can affect the performance of the imaging system. 
This protocol describes the fabrication of a tissue-mimicking phantom that incorporates three-dimensional structural complexity using material with optical properties of tissue. Look-up tables provide India ink and titanium dioxide recipes for optical absorption and scattering targets. Methods to characterize and tune the material optical properties are described. The phantom fabrication detailed in this article has an internal branching mock airway void; however, the technique can be broadly applied to other tissue or organ structures.

Optical tissue phantoms are essential tools for calibration and characterization of optical imaging systems and validation of theoretical models. This article details a method for phantom fabrication that includes replication of tissue optical properties and three-dimensional tissue structure.

Fabricação e caracterização de espectros ópticos de tecido contendo Macrostructure

The rapid development of new optical imaging techniques is dependent on the availability of low-cost, customizable, and easily reproducible standards. By ...

Microdissection of Primary Renal Tissue Segments and Incorporation with Novel Scaffold-free Construct Technology

Kidney transplantation is now a mainstream therapy for end-stage renal disease. However, with approximately 96,000 people on the waiting list and only one-fourth of these patients achieving transplantation, there is a dire need for alternatives for those with failing organs. In order to decrease the harmful consequences of dialysis along with the overall healthcare costs it incurs, active investigation is ongoing in search of alternative solutions to organ transplantation. Implantable tissue-engineered renal cellular constructs are one such feasible approach to replacing lost renal functionality. Here, described for the first time, is the microdissection of murine kidneys for isolation of living corticomedullary renal segments. These segments are capable of rapid incorporation within scaffold-free endothelial-fibroblast constructs which may enable rapid connection with host vasculature once implanted. Adult mouse kidneys were procured from living donors, followed by stereoscope microdissection to obtain renal segments 200 - 300 &#181;m in diameter. Multiple renal constructs were fabricated using primary renal segments harvested from only one kidney. This method demonstrates a procedure which could salvage functional renal tissue from organs that would otherwise be discarded.

Tissue-engineered renal constructs provide a solution for the organ shortage and deleterious effects of dialysis. Here, we describe a protocol to micro dissect murine kidneys for isolation of cortico-medullary segments. These segments are implanted into scaffold-free cellular constructs, forming renal organoids.

Microdissection de segmentos de tecido Renal primária e incorporação com tecnologia de construção romance livre de andaime

Kidney transplantation is now a mainstream therapy for end-stage renal disease. However, with approximately 96,000 people on the waiting list and only ...

Scalable Fabrication of Stretchable, Dual Channel, Microfluidic Organ Chips

A significant number of lead compounds fail in the pharmaceutical pipeline because animal studies often fail to predict clinical responses in human patients. Human Organ-on-a-Chip (Organ Chip) microfluidic cell culture devices, which provide an experimental in vitro platform to assess efficacy, toxicity, and pharmacokinetic (PK) profiles in humans, may be better predictors of therapeutic efficacy and safety in the clinic compared to animal studies. These devices may be used to model the function of virtually any organ type and can be fluidically linked through common endothelium-lined microchannels to perform in vitro studies on human organ-level and whole body-level physiology without having to conduct experiments on people. These Organ Chips consist of two perfused microfluidic channels separated by a permeable elastomeric membrane with organ-specific parenchymal cells on one side and microvascular endothelium on the other, which can be cyclically stretched to provide organ-specific mechanical cues (e.g., breathing motions in lung). This protocol details the fabrication of flexible, dual channel, Organ Chips through casting of parts using 3D printed molds, enabling combination of multiple casting and post-processing steps. Porous poly (dimethyl siloxane) (PDMS) membranes are cast with micrometer sized through-holes using silicon pillar arrays under compression. Fabrication and assembly of Organ Chips involves equipment and steps that can be implemented outside of a traditional cleanroom. This protocol provides researchers with access to Organ Chip technology for in vitro organ- and body-level studies in drug discovery, safety and efficacy testing, as well as mechanistic studies of fundamental biological processes.

Here, we present a protocol that describes the fabrication of stretchable, dual channel, organ chip microfluidic cell culture devices for recapitulating organ-level functionality in vitro.

Fabricação escalável de Stretchable, Dual Channel, órgão Microfluidic Chips

A significant number of lead compounds fail in the pharmaceutical pipeline because animal studies often fail to predict clinical responses in human ...