A doação de sementes da Universidade da Flórida - Faculdade de Medicina é reconhecido agradecimento. Estudos de pós-graduação (Manuel Salinas) foram apoiadas através de oportunidades de minoritários em programas de investigação biomédica - iniciativa de pesquisa para o aprimoramento científico (MBRS-RISE) comunhão: NIH / NIGMS R25 GM061347. O apoio financeiro da Fundação Wallace H. Coulter através Florida International University, Departamento de Engenharia Biomédica é também reconhecido agradecimento. Finalmente, os autores agradecem aos seguintes alunos para a sua assistência durante as várias etapas do processo experimental: Kamau cais, Malaquias Suttle, Kendall Armstrong e Alfonso Abraham.

1. Preparação <ol><li> Projetar e fabricar uma montagem para acomodar uma geometria válvula tri-folheto. Isto, pelo menos, conter um suporte de válvula de sutura-nos folhetos da válvula e de um tubo para alojar o suporte da válvula e acessórios circundantes para fixar o conjunto para o sistema de duplicação de pulso. No nosso caso, utilizamos um sistema comercialmente disponível duplicador de pulso disponíveis a partir ViVitro Labs Inc. (Victoria, BC). Válvula de suporte de cartão, bem como configurações de pré e pós-montagem estão representadas na Figura 1. </li><li> O ciclo inteiro terá de ser preparado antes do uso. Isto envolve dois passos: i) a limpeza do sistema de circuito completo utilizando uma solução de sabão e de água, incluindo a substituição de todos os tubos degradados antes de usar e ii) calibração dos instrumentos conectados ao circuito, ou seja, a bomba que está sendo utilizado, a sonda de fluxo, e os transdutores de pressão (geralmente medido em locais atrial, da aorta e do ventrículo). Calibração pode inicializaçãoei ra me ser realizada utilizando uma solução salina de 1% e deve ser repetido antes de sangue usando uma solução de glicerina-analógico. </li></ol> 2. Nativo da Válvula Aórtica Dissection <ol><li> Obter quatro corações de suínos frescas com a aorta intacta de um matadouro (Institutional Animal Care e do Comitê Use aprovação (IACUC) pode ser necessária) USDA aprovado. No nosso caso, o nosso protocolo de dissecação foi aprovado pelo IACUC na Universidade Internacional da Flórida (Protocolo Número de Aprovação: 11-020). Lavar o coração com água desionizada e colocá-las num recipiente cheio com o fosfato antimicótica / antibiótica e esterilizada de 1% e solução tamponada de transporte salino (PBS) em gelo para o laboratório de ensaio de hidrodinâmica. </li><li> Coloque os corações em uma panela dissecação e remova cuidadosamente o pericárdio. Posicione o coração de tal forma que o lado ventral é voltado para você. Inspecione visualmente e identificar as quatro câmaras do coração e localizar o arco aórtico na aorta intacta. </li><li> Separe o coração em twO metades cortando transversalmente na horizontal a cerca de 0,75 em baixo do anel, ou seja, na junção entre a aorta e o ventrículo esquerdo. Isolar cuidadosamente aorta intacta ainda ligado ao segmento de tecido ventricular esquerda. </li><li> Examinar a válvula aórtica localizado na raiz da aorta, a região entre a aorta ascendente e o anel inferior, garantindo que não há danos ou quaisquer sinais de calcificação. </li><li> Dividir a aorta a ~ 1 em cima do anel e separar o segmento de tecido ventricular esquerda inferior do anel para isolar a válvula da aorta (Figura 2a). </li></ol> 3. Polímero e processo de Sutura valva nativa <ol><li> Coloque a válvula cardíaca interior do suporte da válvula de tal modo que a base de cada válvula está alinhada com a base do suporte do poste. Fixe a válvula no lugar em cada posto temporariamente com um clipe de papel, mas tenha cuidado para não danificar as comissuras ou cúspides. </li><li> Inserir a sutura noagulha. Comece a sutura na parte inferior do suporte da válvula, passando a agulha através do primeiro furo, a partir do exterior para o interior tal que a agulha pode ser facilmente removido a partir do fundo. Em uma forma de looping, comece a sutura da válvula verticalmente até os lugares do suporte da válvula. </li><li> Progresso com a sutura (Figura 2b) ao longo da circunferência do titular e seguro com sutura adicional em torno das pontas das mensagens titular. Os grampos de papel (Figura 2c) pode ser removida quando a válvula é completamente segura por meio de sutura para os três postos de trabalho e na periferia do suporte da válvula (Figuras 2d e 2e). </li></ol> 4. Avaliação hidrodinâmica Nota: O protocolo irá variar dependendo do sistema duplicador de pulso específico que está sendo usado. Todas as informações caontained usado aqui, o ViVitro pulso duplicador Sysytem (ViVitro Labs, Inc., Vancouver, BC). <ol><li> Va Bi-folhetolve <ol><li> Ajustar a taxa de coração do sistema duplicador de pulsos para 70 batimentos / min. </li><li> Seleccionar uma forma de onda de fluxo de propulsão da bomba (no caso do sistema ViVitro o S35 onda foi escolhida para todos os testes hidrodinâmicos). A forma de onda específico utilizado nas nossas experiências é ilustrada por Lim et ai. (2001) 17. </li><li> Ligue o amplificador e bomba de pistão. Aquecer por 15 min. </li><li> Lugar válvula bi-folheto (Figura 2f) em posição aórtica. </li><li> Smear graxa de vácuo em todos os cruzamentos do dispositivo onde poderia ocorrer vazamentos. </li><li> Despeje a glicerina / líquido salino no compartimento fibrilação. Note-se que o sistema de duplicação pulsátil é executado em 2 L de líquido com: 35% / 0,7 L e 65% de glicerina / 1,3 L de solução salina. A solução salina é preparado usando o sal comum bem dissolvido em água desionizada a uma concentração de 9 mg / ml (peso / volume). </li><li> Ligue o transdutor de fluxo que tenha sido colocado na posição aórtica. </li><li> Calibrar ªe bomba. </li><li> Continuar com a calibração do transdutor de fluxo, seguido pelos transdutores de pressão. Da mesma forma que a bomba, basta seguir as instruções dadas pelo software ViVitest (ViVitro Labs Inc.) para cada fluxo e pressão sob a aba calibrar. </li><li> Uma vez que a calibração estiver concluída, inicie a bomba em baixa rpm até que o líquido preenche o compartimento da aorta. Verifique se há vazamentos. Use graxa de vácuo adicional, se necessário. </li><li> Vire as duas torneiras de corte (transdutores aórticas e ventriculares) para a posição aberta. </li><li> Aumentar a rpm da bomba até o volume sistólico chega a 80 ml / batida. </li><li> Permitir que o sistema funcione por 10 minutos até que o fluxo se estabilizou. Fluxo de estabilização pode ser verificada através da observação do fluxo e da pressão de onda exibida na tela. Baixa à variação nenhum entre os ciclos é um bom indicador de estabilização do sistema. </li><li> No software ViVitest selecionar o modo de adquirir. </li><li> Clique em coletar 10 ciclos. </li><li> De modo a analisar, clamber na mesa e salve o arquivo. Também salvar uma imagem da onda usando a opção foto-snap em ViVitest. </li></ol></li><li> Valvas nativas e Polymer <ol><li> Para válvulas de polímeros e animal, siga os mesmos passos 3.1.1 - 3.1.3 das instruções da válvula bi-folheto. </li><li> Coloque o suporte da válvula com a válvula suturado no interior do tubo de vidro da montagem feitos. Sandwich o tubo com as peças superior e inferior e seguras no local com parafusos laterais e nozes. </li><li> Montagem lugar entre a câmara de suporte da válvula aórtica e da aorta de origem. </li><li> Continue com os passos 3.1.5 - 3.1.16 de instruções da válvula de bi-folheto. </li></ol></li></ol> 5. Pós-Processamento <ol><li> Formas de onda de fluxo e pressão <ol><li> Calcule a média dos dados obtidos para cada uma das formas de onda recolhido, ou seja, a pressão da aorta (PA), a pressão do ventrículo esquerdo (VP), e taxa de fluxo (Q). </li><li> Para cada grupo de válvula (polímero, porcino nválvula aórtica ative e bi-folheto), traçar a correspondente AP, VP e Q contra as relações de tempo na mesma trama. </li><li> Para a AP, sobrepor, a válvula aórtica normal, nativo 18, e bi-protética do folheto da válvula 19 parcelas a partir da literatura para fins de validação. </li></ol></li><li> Métricas hidrodinâmicas <ol><li> Para cada válvula testada, as seguintes métricas hidrodinâmicas devem ser computadas: a) queda Encaminhar fluxo de pressão e pressão transvalvar máximo (TVP), b) raiz da aorta mean square (RMS) taxa de fluxo para a frente, c) o fluxo direto da aorta, fechamento, vazamento eo volume regurgitante total d) válvula área final do orifício (EOA), e) transaórtica frente de fluxo, fechamento, vazamentos e perdas totais de energia. <ol><li> Frente a queda de pressão do fluxo é calculado a partir das leituras TVP e podem ser classificados em três intervalos de tempo, P: intervalo que começa e termina com 0 TVP, F: intervalo com fluxo direto e H: intervalo começando com 0 TVP e terminando com 0 fluxo. Máximo TVP é a magradiente de pressão ximum registrado através da válvula a partir das leituras de pressão ventricular e aórtica. </li><li> O atacante vazão RMS (Q rms) fornece uma métrica útil para quantificar a magnitude da taxa de fluxo para a frente como se segue: <img alt="Equação 1" fo:src="/files/ftp_upload/50335/50335eq1highres.jpg" src="/files/ftp_upload/50335/50335eq1.jpg" /> Onde &#39;n&#39; é o número total de pontos no tempo coletado, &#39;Q i&#39; é a medição da taxa de fluxo instantânea coletados em ordem &#39;i&#39;. </li><li> Volumes da aorta para a frente, de fechamento e vazamento são calculados com base nos seguintes intervalos de tempo, Forward: início do fluxo para a frente através da válvula (t o), ao final do fluxo direto (t 1); Encerramento: a partir de t 1 até a instância de fecho de válvula (T 2); vazamento: a partir de t 2 até ao final do ciclo cardíaco (t 3). O volume total de regurgitação é simplesmente a soma de closção e os volumes de vazamento. </li><li> O EOA com base nas propriedades do sangue podem ser calculados para os intervalos de 3, P, F e H do TVP significativo durante cada um destes períodos de 20: <img alt="Equação 1" fo:src="/files/ftp_upload/50335/50335eq2highres.jpg" src="/files/ftp_upload/50335/50335eq2.jpg" /></li><li> As perdas de energia são definidos da seguinte forma 21: <img alt="Equação 1" fo:src="/files/ftp_upload/50335/50335eq3highres.jpg" src="/files/ftp_upload/50335/50335eq3.jpg" /></li></ol></li></ol></li></ol>

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Neste estudo, foi demonstrado a utilidade da modificação de um aparelho duplicador pulsátil comercialmente disponível para acomodar geometrias de válvula tri folheto de modo que o teste hidrodinâmico de um polímero e as válvulas nativas porcino pode ser executada. Especificamente, no nosso caso, o sistema foi modificado de um coração esquerdo ViVitro e sistema de simulador sistémica (Figura 1a), controlado através do sistema de aquisição de dados ViViTest (ViVitro Systems, Inc, Victoria, B...

Doença valvular cardíaca muitas vezes resulta de calcificação valvar degenerativa 1, febre reumática 2, endocardite 3,4 ou defeitos congênitos. Quando a lesão de válvula, provocando estenose e / ou prolapso da válvula de refluxo e não pode ser reparado cirurgicamente, a válvula nativa é geralmente substituído por uma válvula protética. Atualmente opções disponíveis incluem válvulas mecânicas (válvulas gaiola de esfera, válvulas de disco basculante, etc.), Homoenxerto e válvulas bioprostéticas (suínos e bovinos válvulas). As válvulas mecânicas são frequentemente recomendadas para pacientes mais jovens com base na sua durabilidade, mas o paciente é obrigado a permanecer em terapia anticoagulante, para a prevenção de complicações trombóticas 5. Próteses valvares homoenxerto e biológicos têm sido escolhas eficazes para evitar a terapia de sangue mais fino, no entanto, estas válvulas têm risco elevado de fibrose, calcificação, degeneração e complicações imunogênicas levando à insuficiência da válvula 6. Válvulas da engenharia de tecidos estão sendo investigados como uma tecnologia emergente 7-9, mas ainda há muito a ser descoberto. Válvulas alternativos duráveis, biocompatíveis, protéticos são necessários para melhorar a qualidade de vida dos pacientes com doença da válvula cardíaca. Mais uma vez, este projeto válvula poderia substituir a prótese biológica utilizados em tecnologia de válvula transcateter, com abordagens transcateter mostrando o potencial de transformar o tratamento de pacientes selecionados com doença valvular cardíaca 10. Como dito pelos padrões atuais, um substituto de válvula cardíaca bem sucedida deve ter as seguintes características de desempenho: &quot;1) permite avançar com fluxo aceitável diferença pequena queda média pressão, 2) impede o fluxo retrógrado com aceitavelmente pequena regurgitação, 3) resiste a embolização; 4) resiste hemólise; 5) resiste à formação de trombos, 6) é biocompatível, 7) é compatível com as técnicas de diagnóstico in vivo e 8) e pode ser entregue no alvo implantávelpopulação; 9) permanece fixo, uma vez colocado, 10) tem um nível de ruído aceitável; 11) tem a função reprodutível; 12) mantém a sua funcionalidade para uma vida razoável, de acordo com sua classe genérica; 13) mantém a sua funcionalidade e esterilidade para uma prateleira razoável vida antes da implantação. &quot;11. Alguns dos defeitos de próteses de válvulas existentes podem, potencialmente, ser superada por uma válvula de polímero. polímeros biocompatíveis, têm sido considerados os melhores candidatos com base em bioestabilidade, anti-hidrólise, anti-oxidação, e propriedades mecânicas vantajosas, tais como alta resistência e viscoelasticidade. Em particular, os polímeros elastoméricos podem prever deformação do material semelhante a dinâmica da válvula nativas. Elastómeros pode ser adaptado para imitar as propriedades dos tecidos moles, e eles podem ser os únicos materiais artificiais disponíveis que são bio-tolerantes e que possa resistir ao acoplamento, in vivo, as tensões de fluido induzida por tensão e de flexão, no entanto, mover-se de maneira semelhante ao saudável,movimento da válvula nativa. Além disso, os elastómeros podem ser produzidos em massa numa variedade de tamanhos, armazenados com facilidade, são esperados para serem dispositivos de baixo custo e pode ser estruturalmente reforçada com reforço fibroso. O conceito de utilização de materiais poliméricos para montar uma válvula de tri-folheto não é novo e tem sido objecto de várias investigações de investigação ao longo dos últimos 50 anos, 12, os quais foram abandonados, em grande parte devido à válvula de durabilidade limitada. Contudo, com o advento de novas metodologias de fabrico 13,14, o reforço de materiais poliméricos 15,16 e integração potencialmente sem costura de substitutos de válvulas de polímero com tecnologia de válvula transcateter, recentemente tem havido um interesse renovado e actividade no desenvolvimento de válvulas de polímero como um potencialmente alternativa viável para válvulas comerciais actualmente disponíveis. Neste contexto, um protocolo para permitir testes destas válvulas para avaliar a funcionalidade de hidrodinâmica é a primeira etapano processo de avaliação, ainda sistemas de simulador de pulso disponíveis comercialmente geralmente não estão equipados para acomodar os modelos de válvulas de tri-folheto e conter um espaçamento anelar para inserir válvulas cardíacas comercialmente disponíveis (por exemplo, disco de inclinação, as válvulas cardíacas mecânicas bi-folheto). Em segundo lugar, as válvulas de polímero são uma tecnologia emergente cujo hidrodinâmica só pode ser avaliada em um contexto familiar. Apesar da pressão da válvula do coração nativo e dados de fluxo está disponível, é importante para a realização de testes de válvulas nativas aórtica porcina, que são biologicamente compatíveis com válvulas humanos, utilizando o mesmo simulador pulsátil que é utilizado para avaliar as válvulas de polímero de modo a contabilizar diferenças de medição que podem estar dependentes do sistema. Assim, o objetivo deste estudo foi demonstrar como um simulador de pulso disponível comercialmente pode ser equipado com um conjunto de construções para acomodar válvulas tri-folheto e avaliar sistematicamente polímero válvula métricas hidrodinâmicas em um cont relativaext, em comparação com colegas mecânicos e nativo de válvula cardíaca suínos. No nosso caso, as válvulas de polímero de silicone tri-folheto novos anteriormente desenvolvido na Universidade da Flórida 13 formaram o grupo de válvula de polímero.

<table border="1">
	<tbody>
		<tr>
			<td>Name of Reagent/Material</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Pump</td>
			<td>ViVitro Labs</td>
			<td></td>
			<td><a href="http://vivitrolabs.com/products/superpump/" target="_blank">http://vivitrolabs.com/products/superpump/</a></td>
		</tr>
		<tr>
			<td>Flow Meter and Probe</td>
			<td>Carolina Medical</td>
			<td>Model 501D</td>
			<td><a href="http://www.carolinamedicalelectronics.com/documents/FM501.pdf" target="_blank">http://www.carolinamedicalelectronics.com/documents/FM501.pdf</a></td>
		</tr>
		<tr>
			<td>Pressure Transducer</td>
			<td>ViVitro Labs</td>
			<td>HCM018</td>
			<td></td>
		</tr>
		<tr>
			<td>ViVitro Pressure Measuring Assembly</td>
			<td>ViVitro Labs</td>
			<td>6186</td>
			<td></td>
		</tr>
		<tr>
			<td>Valve holder</td>
			<td>WB Engineering</td>
			<td></td>
			<td>Designed by Florida International University. Manufactured by WB Engineering</td>
		</tr>
		<tr>
			<td>Pulse Duplicator</td>
			<td>ViVitro Labs</td>
			<td>PD2010</td>
			<td><a href="http://vivitrolabs.com/wp-content/uploads/Pulse-Duplicator-Accessories1.pdf" target="_blank">http://vivitrolabs.com/wp-content/uploads/Pulse-Duplicator-Accessories1.pdf</a></td>
		</tr>
		<tr>
			<td>Pulse Duplicator Data Acquisition and Control System, including ViViTest Software</td>
			<td>ViVitro Labs</td>
			<td>PDA2010</td>
			<td><a href="http://vivitrolabs.com/products/software-daq" target="_blank">http://vivitrolabs.com/products/software-daq</a></td>
		</tr>
		<tr>
			<td>Porcine Hearts and Native Aortic Valves</td>
			<td>Mary&#39;s Ranch Inc</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bi-leaflet Mechanical Valves</td>
			<td>Saint Jude Medical</td>
			<td></td>
			<td><a href="http://www.sjm.com/" target="_blank">http://www.sjm.com/</a></td>
		</tr>
		<tr>
			<td>High Vacuum Grease</td>
			<td>Dow Corning Corporation</td>
			<td></td>
			<td><a href="http://www1.dowcorning.com/DataFiles/090007b281afed0e.pdf" target="_blank">http://www1.dowcorning.com/DataFiles/090007b281afed0e.pdf</a></td>
		</tr>
		<tr>
			<td>Glycerin</td>
			<td>McMaster-Carr</td>
			<td>3190K293</td>
			<td>99% Natural 5 gal</td>
		</tr>
		<tr>
			<td>Phosphate Buffered Saline (PBS)</td>
			<td>Fisher Scientific</td>
			<td>MT21031CV</td>
			<td>100 ml/heart</td>
		</tr>
		<tr>
			<td>Antimycotic/Antibiotic Solution</td>
			<td>Fisher Scientific</td>
			<td>SV3007901</td>
			<td>1 ml in 100 ml of PBS/heart; 20 ml for ViVitro System</td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>Sigma-Aldrich</td>
			<td>S3014-500G</td>
			<td>9 g/L of deionized water</td>
		</tr>
		<tr>
			<td>Deionized Water</td>
			<td>EMD Millipore Chemicals</td>
			<td></td>
			<td>Millipore Deionized Purification System. 1.3 L for ViVitro System, 200 ml for heart valve dissection process</td>
		</tr>
	</tbody>
</table>

protocol for relative hydrodynamic assessment of tri-leaflet polymer valves

Limitações de válvulas atualmente disponíveis protéticos, xenotransplantes e homoenxertos levaram um ressurgimento recente da evolução da área de tri-folheto próteses valvares polímero. No entanto, a identificação de um protocolo para a avaliação inicial da funcionalidade da válvula hidrodinâmica polímero é importante, durante as fases iniciais do processo de concepção. Características em sistemas in vitro duplicador de pulsos não são configurados para acomodar materiais tri folheto flexíveis e, além disso, a avaliação da funcionalidade da válvula de polímero tem de ser feito num contexto em relação a válvulas cardíacas e próteses nativos sob condições de teste idênticas, para que a variabilidade nas medições de diferentes instrumentos podem ser evitados. Deste modo, foi realizada a avaliação da hidrodinâmica i) nativa (n = 4, o diâmetro médio, D = 20 mm), ii) bi-folheto mecânica (n = 2, D = 23 mm) e iii) polímero de válvulas (n = 5, D = 22 mm) através da utilização de um sistema de duplicação de impulso disponível comercialmente (ViVitro LabsInc, Victoria, BC) que foi modificado para acomodar geometrias de válvula tri folheto. Válvulas de silicone Tri-folheto desenvolvidos na Universidade da Flórida, constituíram o grupo válvula de polímero. Uma mistura na proporção de 35:65 de glicerina para a água foi utilizada para imitar as propriedades físicas do sangue. Caudal instantâneo foi medido na interface do ventrículo esquerdo, e as unidades da aorta, enquanto a pressão foi registada nas posições ventriculares e da aorta. Foi utilizado Bi-folheto e dados de válvulas nativas da literatura para validar o fluxo e leituras de pressão. As seguintes métricas hidrodinâmicas foram relatados: frente a queda de pressão do fluxo, da raiz da aorta taxa média de praça em frente do fluxo, o fechamento da aorta, o vazamento eo volume regurgitante, fechamento transaórtica, vazamento, e as perdas totais de energia. Os resultados representativos indicam que as métricas hidrodinâmicos dos três grupos de válvulas pode ser obtido com sucesso pela incorporação de um conjunto de custom-built em um sistema duplicador de pulsos disponíveis comercialmente e subsequently, em comparação objetiva fornecer insights sobre aspectos funcionais do projeto da válvula de polímero.

Fluxo representativo e ondas de pressão são mostrados nas Figuras 3, 4 e 5. As parcelas foram em média sobre o tamanho da amostra de válvulas para cada grupo testado, que foi, n = 5, 4 e 2 válvulas de polímero, porcina nativa e grupos bi-folheto, respectivamente. As métricas hidrodinâmicas médios e o erro padrão da média para estas amostras são apresentados na Tabela 1. <img alt="Figura 1" src...

Watch this Scientific Journal Video about Protocol for Relative Hydrodynamic Assessment of Tri-leaflet Polymer Valves at JoVE.com

Protocol for Relative Hydrodynamic Assessment of Tri-leaflet Polymer Valves

Tem havido um interesse renovado no desenvolvimento de válvulas de polímeros. Aqui, os objetivos são demonstrar a viabilidade de modificação de um duplicador de pulso comercial para acomodar geometrias tri-folheto e definir um protocolo para apresentar polímero dados hidrodinâmicos de válvulas em relação aos dados de válvulas nativas e protéticas, recolhidos em condições quase idênticas.

Protocolo Relativo Avaliação hidrodinâmica de Tri-folheto polímero Válvulas

Limitations of currently available prosthetic valves, xenografts, and homografts have prompted a recent resurgence of developments in the area of ...

protocol-for-relative-hydrodynamic-assessment-tri-leaflet-polymer

Research

JoVE Journal

Bioengineering

13.7K visualizações.  Florida International University. Tem havido um interesse renovado no desenvolvimento de válvulas de polímeros. Aqui, os objetivos são demonstrar a viabilidade de modificação de um duplicador de pulso comercial para acomodar geometrias tri-folheto e definir um protocolo para apresentar polímero dados hidrodinâmicos de válvulas em relação aos dados de válvulas nativas e protéticas, recolhidos em condições quase idênticas.

Vídeo: Protocol for Relative Hydrodynamic Assessment of Tri-leaflet Polymer Valves

A Microfluidic-based Hydrodynamic Trap for Single Particles

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science. Methods for particle trapping based on optical, magnetic, electrokinetic, and acoustic techniques have led to major advancements in physics and biology ranging from the molecular to cellular level. In this article, we introduce a new microfluidic-based technique for particle trapping and manipulation based solely on hydrodynamic fluid flow. Using this method, we demonstrate trapping of micro- and nano-scale particles in aqueous solutions for long time scales. The hydrodynamic trap consists of an integrated microfluidic device with a cross-slot channel geometry where two opposing laminar streams converge, thereby generating a planar extensional flow with a fluid stagnation point (zero-velocity point). In this device, particles are confined at the trap center by active control of the flow field to maintain particle position at the fluid stagnation point. In this manner, particles are effectively trapped in free solution using a feedback control algorithm implemented with a custom-built LabVIEW code. The control algorithm consists of image acquisition for a particle in the microfluidic device, followed by particle tracking, determination of particle centroid position, and active adjustment of fluid flow by regulating the pressure applied to an on-chip pneumatic valve using a pressure regulator. In this way, the on-chip dynamic metering valve functions to regulate the relative flow rates in the outlet channels, thereby enabling fine-scale control of stagnation point position and particle trapping. The microfluidic-based hydrodynamic trap exhibits several advantages as a method for particle trapping. Hydrodynamic trapping is possible for any arbitrary particle without specific requirements on the physical or chemical properties of the trapped object. In addition, hydrodynamic trapping enables confinement of a "single" target object in concentrated or crowded particle suspensions, which is difficult using alternative force field-based trapping methods. The hydrodynamic trap is user-friendly, straightforward to implement and may be added to existing microfluidic devices to facilitate trapping and long-time analysis of particles. Overall, the hydrodynamic trap is a new platform for confinement, micromanipulation, and observation of particles without surface immobilization and eliminates the need for potentially perturbative optical, magnetic, and electric fields in the free-solution trapping of small particles.

In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice.

A Armadilha hidrodinâmica microfluídicos baseados em partículas individuais

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science.  Methods for ...

Bioengenharia

Design of a Cyclic Pressure Bioreactor for the Ex Vivo Study of Aortic Heart Valves

The aortic valve, located between the left ventricle and the aorta, allows for unidirectional blood flow, preventing backflow into the ventricle. Aortic valve leaflets are composed of interstitial cells suspended within an extracellular matrix (ECM) and are lined with an endothelial cell monolayer. The valve withstands a harsh, dynamic environment and is constantly exposed to shear, flexion, tension, and compression. Research has shown calcific lesions in diseased valves occur in areas of high mechanical stress as a result of endothelial disruption or interstitial matrix damage1-3. Hence, it is not surprising that epidemiological studies have shown high blood pressure to be a leading risk factor in the onset of aortic valve disease4. 
The only treatment option currently available for valve disease is surgical replacement of the diseased valve with a bioprosthetic or mechanical valve5. Improved understanding of valve biology in response to physical stresses would help elucidate the mechanisms of valve pathogenesis. In turn, this could help in the development of non-invasive therapies such as pharmaceutical intervention or prevention. Several bioreactors have been previously developed to study the mechanobiology of native or engineered heart valves6-9. Pulsatile bioreactors have also been developed to study a range of tissues including cartilage10, bone11 and bladder12. The aim of this work was to develop a cyclic pressure system that could be used to elucidate the biological response of aortic valve leaflets to increased pressure loads. 
The system consisted of an acrylic chamber in which to place samples and produce cyclic pressure, viton diaphragm solenoid valves to control the timing of the pressure cycle, and a computer to control electrical devices. The pressure was monitored using a pressure transducer, and the signal was conditioned using a load cell conditioner. A LabVIEW program regulated the pressure using an analog device to pump compressed air into the system at the appropriate rate. The system mimicked the dynamic transvalvular pressure levels associated with the aortic valve; a saw tooth wave produced a gradual increase in pressure, typical of the transvalvular pressure gradient that is present across the valve during diastole, followed by a sharp pressure drop depicting valve opening in systole. The LabVIEW program allowed users to control the magnitude and frequency of cyclic pressure. The system was able to subject tissue samples to physiological and pathological pressure conditions. This device can be used to increase our understanding of how heart valves respond to changes in the local mechanical environment.

Design of a Cyclic Pressure Bioreactor for the Ex Vivo Study of Aortic Heart Valves

A cyclic pressure bioreactor capable of subjecting heart valve tissue to physiological and pathological pressure conditions has been designed. A LabVIEW program allows users to control pressure magnitude, amplitude and frequency. This device can be used to study the mechanobiology of heart valve tissue or isolated cells.

Projeto de um biorreator de pressão cíclica para o Ex Vivo Estudo de válvulas cardíacas Aórtica

The aortic valve, located between the left ventricle and the aorta, allows for unidirectional blood flow, preventing backflow into the ventricle. Aortic ...

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 ...

Polymer Microarrays for High Throughput Discovery of Biomaterials

The discovery of novel biomaterials that are optimized for a specific biological application is readily achieved using polymer microarrays, which allows a combinatorial library of materials to be screened in a parallel, high throughput format1. Herein is described the formation and characterization of a polymer microarray using an on-chip photopolymerization technique 2. This involves mixing monomers at varied ratios to produce a library of monomer solutions, transferring the solution to a glass slide format using a robotic printing device and curing with UV irradiation. This format is readily amenable to many biological assays, including stem cell attachment and proliferation, cell sorting and low bacterial adhesion, allowing the ready identification of 'hit' materials that fulfill a specific biological criterion3-5. Furthermore, the use of high throughput surface characterization (HTSC) allows the biological performance to be correlated with physio-chemical properties, hence elucidating the biological-material interaction6. HTSC makes use of water contact angle (WCA) measurements, atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). In particular, ToF-SIMS provides a chemically rich analysis of the sample that can be used to correlate the cell response with a molecular moiety. In some cases, the biological performance can be predicted from the ToF-SIMS spectra, demonstrating the chemical dependence of a biological-material interaction, and informing the development of hit materials5,3.

A description of the formation of a polymer microarray using an on-chip photopolymerization technique. The high throughput surface characterization using atomic force microscopy, water contact angle measurements, X-ray photoelectron spectroscopy and time of flight secondary ion mass spectrometry and a cell attachment assay is also described.

Microarrays de polímeros para Discovery High Throughput de Biomateriais

The discovery of novel biomaterials that are optimized for a specific biological application is readily achieved using polymer microarrays, which allows a ...

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, ...

Intra-lymph Node Injection of Biodegradable Polymer Particles

Generation of adaptive immune response relies on efficient drainage or trafficking of antigen to lymph nodes for processing and presentation of these foreign molecules to T and B lymphocytes. Lymph nodes have thus become critical targets for new vaccines and immunotherapies. A recent strategy for targeting these tissues is direct lymph node injection of soluble vaccine components, and clinical trials involving this technique have been promising. Several biomaterial strategies have also been investigated to improve lymph node targeting, for example, tuning particle size for optimal drainage of biomaterial vaccine particles. In this paper we present a new method that combines direct lymph node injection with biodegradable polymer particles that can be laden with antigen, adjuvant, or other vaccine components. In this method polymeric microparticles or nanoparticles are synthesized by a modified double emulsion protocol incorporating lipid stabilizers. Particle properties (e.g.&#160;size, cargo loading) are confirmed by laser diffraction and fluorescent microscopy, respectively. Mouse lymph nodes are then identified by peripheral injection of a nontoxic tracer dye that allows visualization of the target injection site and subsequent deposition of polymer particles in lymph nodes. This technique allows direct control over the doses and combinations of biomaterials and vaccine components delivered to lymph nodes and could be harnessed in the development of new biomaterial-based vaccines.

Lymph nodes are the immunological tissues that orchestrate immune response and are a critical target for vaccines. Biomaterials have been employed to better target lymph nodes and to control delivery of antigens or adjuvants. This paper describes a technique combining these ideas to inject biocompatible polymer particles into lymph nodes.

Injeção de linfonodo intra-linfático de partículas de polímeros biodegradáveis

Generation of adaptive immune response relies on efficient drainage or trafficking of antigen to lymph nodes for processing and presentation of these ...

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 ...

Culturing Mouse Cardiac Valves in the Miniature Tissue Culture System

Heart valve disease is a major burden in the Western world and no effective treatment is available. This is mainly due to a lack of knowledge of the molecular, cellular and mechanical mechanisms underlying the maintenance and/or loss of the valvular structure. 
Current models used to study valvular biology include in vitro cultures of valvular endothelial and interstitial cells. Although, in vitro culturing models provide both cellular and molecular mechanisms, the mechanisms involved in the 3D-organization of the valve remain unclear. While in vivo models have provided insight into the molecular mechanisms underlying valvular development, insight into adult valvular biology is still elusive. 
In order to be able to study the regulation of the valvular 3D-organization on tissue, cellular and molecular levels, we have developed the Miniature Tissue Culture System. In this ex vivo flow model the mitral or the aortic valve is cultured in its natural position in the heart. The natural configuration and composition of the leaflet are maintained allowing the most natural response of the valvular cells to stimuli. The valves remain viable and are responsive to changing environmental conditions. This MTCS may provide advantages on studying questions including but not limited to, how does the 3D organization affect valvular biology, what factors affect 3D organization of the valve, and which network of signaling pathways regulates the 3D organization of the valve.

Here, we present an ex vivo flow model in which murine cardiac valves can be cultured allowing the study of the biology of the valve.

Cultura Rato Cardiac Valves no Sistema de Cultura de Tecidos Miniature

Heart valve disease is a major burden in the Western world and no effective treatment is available. This is mainly due to a lack of knowledge of the ...

Film Extrusion of Crambe abyssinica/Wheat Gluten Blends

Crambe abyssinica is a plant with potential for use in industrial (non-food) plant oil production. The side stream from this oil production is a high-protein crambe meal that has limited value, as it is not fit for food or feed use. However, it contains proteins that could potentially make it a suitable raw material for higher-value products. The purpose of this study was to find methods of making this side stream into extruded films, showing that products with a higher value can be produced. The study mainly considered the development of material compositions and methods of preparing and extruding the material. Wheat gluten was added as a supportive protein matrix material, together with glycerol as a plasticizer and urea as a denaturant. The extrudate was evaluated with respect to mechanical (tensile testing) and oxygen barrier properties, and the extrudate structure was revealed visually and by scanning electron microscopy. A denser, more homogeneous material had a lower oxygen transmission rate, higher strength, and higher extensibility. The most homogeneous films were made at an extruder die temperature of 125-130 &#176;C. It is shown here that a film can be extruded with promising mechanical and oxygen barrier properties, the latter especially after a final compression molding step.

Film Extrusion of Crambe abyssinica/Wheat Gluten Blends

The side stream from plant oil production of Crambe abyssinica has limited value. The purpose of this study was to find methods for extruding materials based on this side stream, demonstrating that products with a higher value can be produced. The extrudates were found to have promising properties.

Extrusão de filme de<em&gt; Abyssinica Crambe</em&gt; / Glúten de trigo Blends

Crambe abyssinica is a plant with potential for use in industrial (non-food) plant oil production. The side stream from this oil production is a ...

Synthesis of Functionalized 10-nm Polymer-coated Gold Particles for Endothelium Targeting and Drug Delivery

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting and drug delivery are some of the applications of these AuNPs, but endothelial extracellular matrices&#39; defensive properties hamper particle uptake. To address this issue, we describe a synthesis method for ultrasmall gold nanoparticles to improve vascular delivery, with customizable functional groups and polymer lengths for further adjustments. The protocol yields 2.5 nm AuNPs that are capped with tetrakis(hydroxymethyl)phosphonium chloride (THPC). The replacement of THPC with hetero-functional polyethylene glycol (PEG) on the surface of the AuNP increases the hydrodynamic radius to 10.5 nm while providing various functional groups on the surface. The last part of the protocol includes an optional addition of a fluorophore to allow the AuNPs to be visualized under fluorescence to track nanoparticle uptake. Dialysis and lyophilization were used to purify and isolate the AuNPs. These fluorescent nanoparticles can be visualized in both in vitro and in vivo experiments due to the biocompatible PEG coating and fluorescent probes. Additionally, the size range of these nanoparticles render them an ideal candidate for probing the glycocalyx without disrupting normal vasculature function, which may lead to improved delivery and therapeutics.

We describe a method of synthesizing biocompatible 10-nm gold nanoparticles, functionalized by coating poly-ethylene glycol onto the surface. These particles can be used in vitro and in vivo for delivering therapeutics to nanoscale cellular and extracellular spaces that are difficult to access with conventional nanoparticle sizes.

Síntese de funcionalizados 10 nm polímero-revestida com partículas de ouro para o direcionamento do endotélio e entrega da droga

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting ...