Os autores gostariam de agradecer ao pessoal das instalações de ressonância magnética na Clínica Semmes-Murphey, Memphis, Tennessee, bem como o departamento de Neurocirurgia da Universidade de Tennessee Health Science Center, em Memphis, Tennessee.

1. Preparação de Gel Phantoms e Dye <ol><li> Prepare a 0,2% de gel de agarose por dissolução de 2 g de 0,1% de agarose em pó, em 1000 ml de água desionizada. Agitar a solução durante cerca de 1 min para garantir a mistura adequada; e microondas imediatamente a solução em 3 intervalos mínimo para 9 min ou até que claro, mexendo entre os intervalos. </li><li> Embora o gel de agarose a ser líquido, vazar a solução em 5 cm x 5 cm x 5 cm recipientes. Permitir que o espaço na parte superior do recipiente para adicionar água e permitir que o gel de agarose para arrefecer e assentar. </li><li> Uma vez que o gel de agarose foi solidificado (cerca de 1-2 horas), adicionar 1 cm de água para a superfície do gel e refrigerar. É melhor utilizar o gel dentro de 24-48 horas de mistura, mas pode ser armazenado durante até uma semana refrigerado 10. </li><li> Preparar um corante radio-contraste em uma seringa de 60 ml consistindo em 50 ml de bromofenol 0,017% de corante azul (BPB) e 2 mM de gadoteridol meios de rádio-contraste. <ol><li> Combine 8,5 mg de corante BPB a 50 ml de água deionizada para criar uma solução BPB 0,017%. </li><li> Adicionar 0,2 ml de estoque 0,5 M gadoteridol à solução 50 ml de BPB 0,017% para criar uma solução gadoteridol 2 mM. </li></ol></li></ol> 2. Preparação do Sistema de Infusão <ol><li> Sistema de bomba de infusão de seringa (método preferido): Para a preparação de uma bomba de seringa, fixar o cateter de infusão directamente para a seringa, através do sensor de pressão, reduzindo o volume morto da linha de perfusão. A função de purga da bomba de seringa podem ser utilizados para limpar a linha de ar usando um bolus maior do que o volume de escorvamento do cateter a uma taxa de 10 mL / min. </li><li> Sistema de bomba de infusão de tubo (método alternativo): ligar a seringa contendo o corante radio-contraste com a bomba de infusão. Ligue o sensor de pressão de saída da bomba, estando o transdutor acoplado ao monitor IV. Anexar um cateter de infusão de 16 L para a extremidade aberta do sensor de pressão. Nota: A ponta do cateter 16 G infusão tem um diâmetro interiormetro de 0,2 mm e um diâmetro externo de 0,35 mm. A ponta é feita de sílica fundida e o comprimento da ponta é de 3 mm. Ele aumenta a aproximadamente 0,75 mm, e continua durante 15 mm, o cateter, depois, passos acima de uma forma cónica para 1,6 mm ou 16 G. </li><li> Preparar para infusão purgando o sistema durante cerca de 15 minutos a 16,667 ul / min para remover quaisquer bolhas de ar. Não exceda a taxa de fluxo de 16,667 mL / min, como a máquina deixará de infusão devido à pressão alta linha. Após a fixação do cateter de infusão para a linha de saída da bomba de infusão, linhas de purga de ar, usando a função &quot;Bolus&quot; sobre a bomba de infusão. </li><li> Fixe o suporte para a infusão do cateter e quadro trajetória ao contêiner fantasma gel (5 cm x 5 cm x 5 cm) e coloque na ressonância magnética. </li></ol> 3. CED Gel Infusion e MR Scanning <ol><li> Zerar o valor da pressão (mmHg) registrado pelo monitor IV antes do início da infusão. </li><li> Inserir o cateter de infusão para o gel de agarose wom a bomba de infusão, que funciona na taxa de fluxo mais baixa possível, neste caso, 1,667 mL / min. </li><li> Começar o de ressonância magnética, usando os parâmetros listados na Tabela 1, e continuar a infusão, a uma taxa de 1,667 mL / min. Infundir o gel a uma taxa constante até que o volume total infundido atinge 60 ul (cerca de 38 min). </li><li> Digitalizar o gel continuamente em 3 min e 50 seg intervalos. Registre as leituras de pressão a cada 60 seg. Uma vez que o volume infundido chega a 60 mL, desligue a bomba de infusão; e digitalização MR completo enquanto continua a gravar leituras de pressão. </li></ol> 4. MR Análise de Dados <ol><li> Para analisar as imagens de RM, use um visualizador DICOM com funcionalidade adequada segmentação ROI. </li><li> Seleccione a estrutura correcta em cada varredura marcado pela secção transversal do cateter, como visto na Figura 1. </li><li> Usando o &quot;ROI - retângulo&quot; ferramenta, selecione a maior parte do gel, que não inclui qualquerporção do local da infusão. A vontade saída do software uma densidade de pixels média com desvio padrão. Encontre o valor que corresponde a três desvios-padrão da média. Este valor é usado como o limiar para determinar quando contraste está presente com uma confiança de 99,7%. </li><li> Usando o &quot;ROI - círculo&quot; ferramenta, cercar o local da infusão com um grande círculo bastante e dar a este um nome único. </li><li> Selecione o círculo e usando o &quot;ROI - definir valores de pixel para&quot; ferramenta, valor limite de entrada encontrados no passo 4.3 em &quot;se o valor atual é maior do que:&quot; e marque apenas nesta linha. Em seguida, em &quot;para este novo valor:&quot;, digite um valor grande (25.000). Repor a densidade de pixels para selecionar a área abrangida pelo limite previamente definido. </li><li> Em seguida, usando o &quot;ROI - crescer região (segmentação 2D/3D)&quot; ferramenta, selecione a região 2D crescendo, o algoritmo de confiança com o parâmetro raio inicial = 2, e escova ROI. Clique dentro do local de infusão para o software para calcular a área total of esta região. </li><li> Supondo que uma perfusão de nuvem esférica, calcular o volume de difusão a partir da área através da seguinte equação: V = 4/3π (√ (Área / π)) 3 </li></ol>

<ol>
	<li>Miranpuri, G. 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=Gene-based+therapy+of+Parkinson's+Disease:+Translation+from+animal+model+to+human+clinical+trial+employing+convection+enhanced+delivery.">Gene-based therapy of Parkinson's Disease: Translation from animal model to human clinical trial employing convection enhanced delivery.</a> Annals of Neurosciences. 19, 133-146 (2012).</li><li>Sillay, K., Hinchman, A., Akture, E., Salamat, S., Miranpuri, G., Williams, J., Berndt, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convection+Enhanced+Delivery+to+the+Brain:+Preparing+for+Gene+Therapy+and+Protein+Delivery+to+the+Brain+for+Functional+and+Restorative+Neurosurgery+by+Understanding+Low-Flow+Neurocatheter+Infusions+Using+the+Alaris&#174;+System+Infusion+Pump.">Convection Enhanced Delivery to the Brain: Preparing for Gene Therapy and Protein Delivery to the Brain for Functional and Restorative Neurosurgery by Understanding Low-Flow Neurocatheter Infusions Using the Alaris&#174; System Infusion Pump.</a> Annals of Neurosciences. 20, (2013).</li><li>Song, D. K., Lonser, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convection-enhanced+delivery+for+the+treatment+of+pediatric+neurologic+disorders.">Convection-enhanced delivery for the treatment of pediatric neurologic disorders.</a> Journal of child neurology. 23, 1231-1237 (2008).</li><li>Bobo, R. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convection-enhanced+delivery+of+macromolecules+in+the+brain.">Convection-enhanced delivery of macromolecules in the brain.</a> Proc Natl Acad Sci USA. 91, 2076-2080 (1994).</li><li>Debinski, W., Tatter, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convection-enhanced+delivery+for+the+treatment+of+brain+tumors.">Convection-enhanced delivery for the treatment of brain tumors.</a> Expert review of neurotherapeutics. 9, 1519-1527 (2009).</li><li>Morrison, P. F., Laske, D. W., Bobo, H., Oldfield, E. H., Dedrick, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-flow+microinfusion:+tissue+penetration+and+pharmacodynamics.">High-flow microinfusion: tissue penetration and pharmacodynamics.</a> The American journal of physiology. 266, 292-305 (1994).</li><li>Nguyen, T. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convective+distribution+of+macromolecules+in+the+primate+brain+demonstrated+using+computerized+tomography+and+magnetic+resonance+imaging.">Convective distribution of macromolecules in the primate brain demonstrated using computerized tomography and magnetic resonance imaging.</a> Journal of neurosurgery. 98, 584-590 (2003).</li><li>Lonser, R. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Successful+and+safe+perfusion+of+the+primate+brainstem:+in+vivo+magnetic+resonance+imaging+of+macromolecular+distribution+during+infusion.">Successful and safe perfusion of the primate brainstem: in vivo magnetic resonance imaging of macromolecular distribution during infusion.</a> Journal of neurosurgery. 97, 905-913 (2002).</li><li>Raghavan, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convection-enhanced+delivery+of+therapeutics+for+brain+disease,+and+its+optimization.">Convection-enhanced delivery of therapeutics for brain disease, and its optimization.</a> Neurosurg Focus. 20, (2006).</li><li>Sillay, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Benchmarking+the+ERG+valve+tip+and+MRI+Interventions+Smart+Flow+neurocatheter+convection-enhanced+delivery+system's+performance+in+a+gel+model+of+the+brain:+employing+infusion+protocols+proposed+for+gene+therapy+for+Parkinson's+disease.">Benchmarking the ERG valve tip and MRI Interventions Smart Flow neurocatheter convection-enhanced delivery system's performance in a gel model of the brain: employing infusion protocols proposed for gene therapy for Parkinson's disease.</a> Journal of neural engineering. 9, (2012).</li><li>Schomberg, D., Wang, A., Marshall, H., Sillay, K., Miranpuri, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ramped-Rate+vs.+continuous+rate+infusions:+An+in+vitro+comparison+of+Convection+Enhanced+Delivery+protocols.">Ramped-Rate vs. continuous rate infusions: An in vitro comparison of Convection Enhanced Delivery protocols.</a> Annals of Neurosciences. 20, (2013).</li><li>Sillay, K. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Perioperative+Brain+Shift+and+Deep+Brain+Stimulating+Electrode+Deformation+Analysis:+Implications+for+rigid+and+non-rigid+devices.">Perioperative Brain Shift and Deep Brain Stimulating Electrode Deformation Analysis: Implications for rigid and non-rigid devices.</a> Ann Biomed Eng. 41, 293-304 (2013).</li><li>Brodsky, E., Block, W., Alexander, A., Emborg, M., Ross, C., Sillay, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraoperative+Device+Targeting+using+Real-Time+MRI.">Intraoperative Device Targeting using Real-Time MRI.</a> Biomedical Sciences and Engineering Conference, BSEC. , (2011).</li><li>Sillay, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Strategies+for+the+delivery+of+multiple+collinear+infusion+clouds+in+convection-enhanced+delivery+in+the+treatment+of+Parkinson's+disease.">Strategies for the delivery of multiple collinear infusion clouds in convection-enhanced delivery in the treatment of Parkinson's disease.</a> Stereotactic and functional neurosurgery. 91, 153-161 (2013).</li><li>Brady, M. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathways+of+infusate+loss+during+convection-enhanced+delivery+into+the+putamen+nucleus.">Pathways of infusate loss during convection-enhanced delivery into the putamen nucleus.</a> Stereotactic and functional neurosurgery. 91, 69-78 (2013).</li><li>White, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+evaluation+of+the+relationships+between+catheter+design+and+tissue+mechanics+in+achieving+high-flow+convection-enhanced+delivery.">An evaluation of the relationships between catheter design and tissue mechanics in achieving high-flow convection-enhanced delivery.</a> J Neurosci Methods. 199, 87-97 (2011).</li><li>Fiandaca, M. S., Forsayeth, J. R., Dickinson, P. J., Bankiewicz, K. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Image-guided+convection-enhanced+delivery+platform+in+the+treatment+of+neurological+diseases.">Image-guided convection-enhanced delivery platform in the treatment of neurological diseases.</a> Neurotherapeutics : the journal of the American Society for Experimental NeuroTherapeutics. 5, 123-127 (2008).</li><li>Jagannathan, J., Walbridge, S., Butman, J. A., Oldfield, E. H., Lonser, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+ependymal+and+pial+surfaces+on+convection-enhanced+delivery.">Effect of ependymal and pial surfaces on convection-enhanced delivery.</a> Journal of neurosurgery. 109, 547-552 (2008).</li><li>Chen, Z. J., Broaddus, W. C., Viswanathan, R. R., Raghavan, R., Gillies, G. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraparenchymal+drug+delivery+via+positive-pressure+infusion:+experimental+and+modeling+studies+of+poroelasticity+in+brain+phantom+gels.">Intraparenchymal drug delivery via positive-pressure infusion: experimental and modeling studies of poroelasticity in brain phantom gels.</a> IEEE transactions on bio-medical engineering. 49, 85-96 (2002).</li><li>Chen, Z. 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=A+realistic+brain+tissue+phantom+for+intraparenchymal+infusion+studies.">A realistic brain tissue phantom for intraparenchymal infusion studies.</a> Journal of neurosurgery. 101, 314-322 (2004).</li><li>Richardson, R. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interventional+MRI-guided+Putaminal+Delivery+of+AAV2-GDNF+for+a+Planned+Clinical+Trial+in+Parkinson's+Disease.">Interventional MRI-guided Putaminal Delivery of AAV2-GDNF for a Planned Clinical Trial in Parkinson's Disease.</a> Mol Ther. 19, 1048-1057 (2011).</li><li>Thorne, R. G., Hrabetova, S., Nicholson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diffusion+of+epidermal+growth+factor+in+rat+brain+extracellular+space+measured+by+integrative+optical+imaging.">Diffusion of epidermal growth factor in rat brain extracellular space measured by integrative optical imaging.</a> Journal of neurophysiology. 92, 3471-3481 (2004).</li><li>Panse, S. J., Fillmore, H. L., Chen, Z. J., Gillies, G. T., Broaddus, W. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+coaxial+tube+catheter+for+central+nervous+system+infusions:+performance+characteristics+in+brain+phantom+gel.">A novel coaxial tube catheter for central nervous system infusions: performance characteristics in brain phantom gel.</a> J Med Eng Technol. 35, 408-414 (2010).</li><li>Linninger, A. A., Somayaji, M. R., Zhang, L., Smitha Hariharan, M., Penn, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rigorous+mathematical+modeling+techniques+for+optimal+delivery+of+macromolecules+to+the+brain.">Rigorous mathematical modeling techniques for optimal delivery of macromolecules to the brain.</a> IEEE transactions on bio-medical engineering. 55, 2303-2313 (2008).</li><li>Sampson, J. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+utility+of+a+patient-specific+algorithm+for+simulating+intracerebral+drug+infusions.">Clinical utility of a patient-specific algorithm for simulating intracerebral drug infusions.</a> Neuro-oncology. 9, 343-353 (2007).</li></ol>

Os autores declaram que não têm interesses financeiros concorrentes.

<html> Os passos críticos para assegurar o êxito da infusão são: a linha de purga de ar de infusão, a mistura do gel de agarose, a análise dos dados de RM, utilizando pequenos diâmetros do cateter interno, utilizando um passo desenhos de cateter para minimizar o refluxo, e minimizando a pressão sentida pela gel ou tecido no qual a droga está a ser infundido. Como afirmado anteriormente, a principal detrimento para o sucesso da infusão é de perfusão de ar. Corretamente e completamente purgar a linha de infusã...

Administração realçada por convecção (CED) tem sido proposto como uma opção de tratamento para um largo espectro de desordens neurológicas, incluindo tumores malignos do cérebro, epilepsia, distúrbios metabólicos, doenças neurodegenerativas (tais como a doença de Parkinson) 1, acidente vascular cerebral e trauma 2. CED emprega fluxo maior pressão positiva para a distribuição de uma droga ou outra infusato. CED prevê a entrega segura, confiável e homogêneo de compostos de massa molecular, variando de baixa a alta, em volumes clinicamente relevantes 3. Entrega da droga tradicional para o tecido cerebral é severamente limitada pela barreira sangue-cérebro 4. Formada pelas junções apertadas entre as células endoteliais que formam os capilares do cérebro, os blocos de barreira sangue-cérebro e moléculas polares de elevado peso molecular a partir de entrar no parênquima do cérebro. Infusão cérebro intraparenquimal direto via CED pode superar as limitações das modalidades de entrega de drogas terapêuticas anteriorese permite a utilização de agentes terapêuticos que não atravessam a barreira sangue-cérebro, e portanto, tem sido previamente disponível como opções de tratamento viáveis ​​5. Pesquisadores de os EUA National Institutes of Health (NIH) descreveu CED no início de 1990 como um meio de alcançar maiores concentrações de drogas terapêuticas que por difusão sozinho 6-8. Os primeiros métodos de CED envolvido implantar um ou mais cateteres no cérebro, ligando uma bomba de infusão para o cateter, e bombeando os agentes terapêuticos directamente na região alvo. O aumento da fracção de distribuição e concentração relativamente estável é relatado para ocorrer como a pressão positiva criada pela bomba de infusão faz com que os tecidos para dilatar e permitem a permeação do fármaco 9. A técnica fundamental para CED permanece praticamente a mesma que foi descrita pela primeira vez. Avanços no desenho de cateter 10, técnica de infusão <sup&gt; 11, monitoramento de pressão na linha 2, e monitoramento em tempo real de ressonância magnética para corrigir a mudança cérebro 12, 13, otimizar infusões múltiplas colineares 14, e monitorar a perda infusato 15 aumentaram a segurança e eficácia do tratamento 10. Importância adicional foi colocada no cateter de design e estratégia de infusão incluindo a taxa de fluxo. Successful CED, com refluxo cateter limitado e danos nos tecidos, tem sido correlacionada com design cateter e taxa de infusão. O uso de um cateter com um diâmetro estreito e uma taxa de infusão de baixo para limitar o refluxo ao longo da interface cérebro-cateter, bem como limitar os danos na ponta do cateter 16. RM fornece confirmação visual do local correto para a colocação do cateter de infusão e, assim, a entrega da droga, ao mesmo tempo, permitindo a correção de refluxo infusão ou entrega aberrante 17. As imagens de RM também pode ser usada para aproximar e controlar os volumes de distribuição (Vd) Do fármaco infundido. O Vd é calculada usando um valor de intensidade de sinal ressonância magnética superior a três desvios padrão acima da média da envolvente gel não-infundido como um limite para a segmentação 18. O Vd é uma medição útil para CED pois representa o volume do fármaco distribuído no cérebro. Juntamente com o volume infundido (VI), um rácio pode ser gerado (Vd / Vi) quantificar o volume coberto pelo fármaco infundido. Fantasmas em gel de agarose imitar várias propriedades mecânicas cruciais do cérebro humano importantes para a compreensão CED, tais como: Vd, interações gel-cateter, propriedades poroelásticos e morfologia infusão nuvem 10. As misturas de 0,2% de gel de agarose foram mostrados para imitar in vivo alterações na fracção de poros locais provocadas pela dilatação de gel devido ao DCE. Uma fração de poros semelhante ao cérebro humano promove interações semelhantes e medições precisas de Vd 19. Adicionalmente, concentrações semelhantes de umgéis garose tais como 0,6% e 0,8% mostraram perfis similares de pressão de perfusão para o cérebro 20. Além disso, os géis de agarose translúcidos têm a vantagem de visualização em tempo real de colocação do cateter e infusão de refluxo. Fantasmas em gel de agarose são relativamente baratos de produzir. O custo dos fantasmas de gel de agarose pode ser a chave para o futuro da formação generalizada em toda a cirurgia neurológica. Devido a estas propriedades, os géis de agarose a fornecer um substituto útil, replicar muitos dos atributos chave da infusão de cérebro humano, sem a utilização de tecido do cérebro. Como dito acima, imagem-guiada CED em modelos de gel de agarose fornece um benéfico método in vitro para teste, pesquisa e treinamento. O objetivo deste artigo é descrever como recriar fantasmas gel de agarose, para delinear os protocolos de ensaio e análise CED apropriados, e para resolver os erros mais comuns enfrentados durante infusões de DEC para o tratamento de doenças neurológicas.

<table border="0" cellpadding="0" cellspacing="0" width="590">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:205px;">Prohance</td>
			<td style="width:71px;">Bracco</td>
			<td style="width:119px;"></td>
			<td style="width:195px;">Gadoteridol radio contrast media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:205px;">Bromophenol Blue Dye</td>
			<td style="width:71px;">Biorad</td>
			<td style="width:119px;">161-0404</td>
			<td>Dye for infusate visualization</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:205px;">Agarose Gel Powder</td>
			<td style="width:71px;">Biorad</td>
			<td style="width:119px;">161-3101EDU</td>
			<td>Agarose powder for creating gels</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:205px;">Medrad Veris MR Vital Signs Monitor</td>
			<td style="width:71px;">Medrad</td>
			<td style="width:119px;"></td>
			<td>MR safe infusion pressure monitor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:205px;">16 Gauge SmartFlow Catheter</td>
			<td style="width:71px;">SurgiVision</td>
			<td style="width:119px;"></td>
			<td>Infusion catheter</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:205px;">Medrad Continuum MR Infusion System</td>
			<td style="width:71px;">Medrad</td>
			<td style="width:119px;"></td>
			<td>MR safe infusion pump</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:205px;">SMART Frame MRI Guided Trajectory Frame</td>
			<td style="width:71px;">ClearPoint</td>
			<td style="width:119px;"></td>
			<td>Infusion catheter frame</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:205px;">Osirix Imaging Software and DICOM Viewer</td>
			<td style="width:71px;">Osirix Imaging Software</td>
			<td style="width:119px;"></td>
			<td>OsiriX 32-bit DICOM Viewer</td>
		</tr>
	</tbody>
</table>

image-guided convection-enhanced delivery into agarose gel models of the brain

Administração realçada por convecção (CED) tem sido proposto como uma opção de tratamento para uma grande variedade de doenças neurológicas. Neuroinfusion cateter CED permite o fluxo em massa de pressão positiva para entregar maiores quantidades de terapêutica para um alvo intracraniano do que os métodos tradicionais de distribuição de drogas. A utilidade clínica de ressonância magnética tempo real guiada CED (RCED) reside na capacidade para alvejar com precisão, monitorizar a terapia, e identificar complicações. Com o treinamento, RCED é eficiente e as complicações podem ser minimizadas. O modelo em gel de agarose do cérebro fornece uma ferramenta acessível para testes CED, pesquisa e treinamento. Cérebro simulado RCED permite a prática da cirurgia simulada ao mesmo tempo proporcionar feedback visual da infusão. Análise de infusão permite calcular a fracção de distribuição (Vd / Vi) permitindo que o aluno para verificar a semelhança do modelo, em comparação com tecido cerebral humano. Este artigo descreve o nosso fantasma cérebro em gel de agarose e descreve-me importantetrics durante uma infusão de DEC e análise de protocolos, enquanto que abordam problemas comuns enfrentados durante a infusão CED para o tratamento de doenças neurológicas.

Interpretação e análise de infusões de DEC envolvem vários fatores importantes, como a fração de distribuição e refluxo infusato. O cálculo de fração de distribuição depende muito do cálculo do Vd. Portanto interpretação exata das imagens de RM é fundamental. Propomos um método semi-automatizado para reproduzir com fiabilidade destas medições, listados acima. Estes métodos objectivamente determinar a área de corte transversal da nuvem de solução de infusão e um raio aproximado. Enquanto variáv...

Watch this Scientific Journal Video about Image-guided Convection-enhanced Delivery into Agarose Gel Models of the Brain at JoVE.com

Image-guided Convection-enhanced Delivery into Agarose Gel Models of the Brain

Administração realçada por convecção (CED) tem sido proposto como uma opção de tratamento para uma grande variedade de doenças neurológicas. A fim de preparar profissionais de saúde para a adoção de CED, são necessários modelos de formação acessíveis. Descreve-se o uso de gel de agarose como tal modelo do cérebro humano para o teste, pesquisa e treinamento.

Imagem-guiado Entrega reforçada-Convecção em Modelos gel de agarose do Cérebro

Convection-enhanced delivery (CED) has been proposed as a treatment option for a wide range of neurological diseases. Neuroinfusion catheter CED allows ...

image-guided-convection-enhanced-delivery-into-agarose-gel-models

Research

JoVE Journal

Medicine

11.3K visualizações.  University of Tennessee Health Science Center.  Administração realçada por convecção (CED) tem sido proposto como uma opção de tratamento para uma grande variedade de doenças neurológicas. A fim de preparar profissionais de saúde para a adoção de CED, são necessários modelos de formação acessíveis. Descreve-se o uso de gel de agarose como tal modelo do cérebro humano para o teste, pesquisa e treinamento. 

Vídeo: Image-guided Convection-enhanced Delivery into Agarose Gel Models of the Brain

MRI and PET in Mouse Models of Myocardial Infarction

Myocardial infarction is one of the leading causes of death in the Western world. The similarity of the mouse heart to the human heart has made it an ideal model for testing novel therapeutic strategies.
In vivo magnetic resonance imaging (MRI) gives excellent views of the heart noninvasively with clear anatomical detail, which can be used for accurate functional assessment. Contrast agents can provide basic measures of tissue viability but these are nonspecific. Positron emission tomography (PET) is a complementary technique that is highly specific for molecular imaging, but lacks the anatomical detail of MRI. Used together, these techniques offer a sensitive, specific and quantitative tool for the assessment of the heart in disease and recovery following treatment.
In this paper we explain how these methods are carried out in mouse models of acute myocardial infarction. The procedures described here were designed for the assessment of putative protective drug treatments. We used MRI to measure systolic function and infarct size with late gadolinium enhancement, and PET with fluorodeoxyglucose (FDG) to assess metabolic function in the infarcted region. The paper focuses on practical aspects such as slice planning, accurate gating, drug delivery, segmentation of images, and multimodal coregistration. The methods presented here achieve good repeatability and accuracy maintaining a high throughput.

We describe how to perform MRI and PET imaging of the mouse heart. The protocol is tailored to assess treatment efficacy in models of myocardial infarction and heart failure.

Ressonância magnética e PET no rato modelos de Infarto do Miocárdio

Myocardial infarction is one of the leading causes of death in the Western world. The similarity of the mouse heart to the human heart has made it an ...

Medicina

Heterotopic Mucosal Engrafting Procedure for Direct Drug Delivery to the Brain in Mice

Delivery of therapeutics into the brain is impeded by the presence of the blood-brain barrier (BBB) which restricts the passage of polar and high molecular weight compounds from the bloodstream and into brain tissue. Some direct delivery success in humans has been achieved via implantation of transcranial catheters; however this method is highly invasive and associated with numerous complications. A less invasive alternative would be to dose the brain through a surgically implanted, semipermeable membrane such as the nasal mucosa that is used to repair skull base defects following endoscopic transnasal tumor removal surgery in humans. Drug transfer though this membrane would effectively bypass the BBB and diffuse directly into the brain and cerebrospinal fluid. Inspired by this approach, a surgical approach in mice was developed that uses a donor septal mucosal membrane engrafted over an extracranial surgical BBB defect. This model has been shown to effectively allow the passage of high molecular weight compounds into the brain. Since numerous drug candidates are incapable of crossing the BBB, this model is valuable for performing preclinical testing of novel therapies for neurological and psychiatric diseases.

A mouse model of human endoscopic skull base reconstruction has been developed that creates a semipermeable interface between the brain and nose using nasal mucosal grafts. This method allows researchers to study delivery to the central nervous system of high molecular weight therapeutics which are otherwise excluded by the blood-brain barrier when administered systemically.

Heterotópico mucosa enxertando Procedimento para entrega da droga direto para o cérebro em ratos

Delivery of therapeutics into the brain is impeded by the presence of the blood-brain barrier (BBB) which restricts the passage of polar and high ...

Mouse Kidney Transplantation: Models of Allograft Rejection

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely replicates both the technical and pathological processes that occur in human renal transplantation. Although mouse models of allogeneic rejection in organs other than the kidney exist, and are more technically feasible, there is evidence that different organs elicit disparate rejection modes and dynamics, for instance the time course of rejection in cardiac and renal allograft differs significantly in certain strain combinations. This model is an attractive tool for many reasons despite its technical challenges. As inbred mouse strain haplotypes are well characterized it is possible to choose donor and recipient combinations to model acute allograft rejection by transplanting across MHC class I and II loci. Conversely by transplanting between strains with similar haplotypes a chronic process can be elicited were the allograft kidney develops interstitial fibrosis and tubular atrophy. We have modified the surgical technique to reduce operating time and improve ease of surgery, however a learning curve still needs to be overcome in order to faithfully replicate the model. This study will provide key points in the surgical procedure and aid the process of establishing this technique.

Here, we present a protocol to study the immunology of rejection. The surgical model presented reports a short operating time and a concise technique. Depending on the donor-recipient strain combination, the transplanted kidney may develop acute cellular rejection or chronic allograft damage, defined by interstitial fibrosis and tubular atrophy.

Rato Transplante Renal: Modelos de rejeição de enxertos

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely ...

Transposon Mediated Integration of Plasmid DNA into the Subventricular Zone of Neonatal Mice to Generate Novel Models of Glioblastoma

An urgent need exists to test the contribution of new genes to the pathogenesis and progression of human glioblastomas (GBM), the most common primary brain tumor in adults with dismal prognosis. New potential therapies are rapidly emerging from the bench and require systematic testing in experimental models which closely reproduce the salient features of the human disease. Herein we describe in detail a method to induce new models of GBM with transposon-mediated integration of plasmid DNA into cells of the subventricular zone of neonatal mice. We present a simple way to clone new transposons amenable for genomic integration using the Sleeping Beauty transposon system and illustrate how to monitor plasmid uptake and disease progression using bioluminescence, histology and immuno-histochemistry. We also describe a method to create new primary GBM cell lines. Ideally, this report will allow further dissemination of the Sleeping Beauty transposon system among brain tumor researchers, leading to an in depth understanding of GBM pathogenesis and progression and to the timely design and testing of effective therapies for patients.

Here we describe an efficient and versatile protocol to induce, monitor and analyze novel glioblastomas (GBM) using transposon DNA injected into the ventricles of neonatal mice. Cells of the subventricular zone, which take up the plasmid, transform, proliferate and generate tumors with histo-pathological characteristics of human GBM.

Transposon Mediated Integração de DNA plasmídeo na zona subventricular de Neonatal Ratos para gerar modelos novela de Glioblastoma

An urgent need exists to test the contribution of new genes to the pathogenesis and progression of human glioblastomas (GBM), the most common primary ...

Improved Method for the Establishment of an In Vitro Blood-Brain Barrier Model Based on Porcine Brain Endothelial Cells

The aim of this protocol presents an optimized procedure for the purification and cultivation of pBECs and to establish in vitro blood-brain barrier (BBB) models based on pBECs in mono-culture (MC), MC with astrocyte-conditioned medium (ACM), and non-contact co-culture (NCC) with astrocytes of porcine or rat origin. pBECs were isolated and cultured from fragments of capillaries from the brain cortices of domestic pigs 5-6 months old. These fragments were purified by careful removal of meninges, isolation and homogenization of grey matter, filtration, enzymatic digestion, and centrifugation. To further eliminate contaminating cells, the capillary fragments were cultured with puromycin-containing medium. When 60-95% confluent, pBECs growing from the capillary fragments were passaged to permeable membrane filter inserts and established in the models. To increase barrier tightness and BBB characteristic phenotype of pBECs, the cells were treated with the following differentiation factors: membrane permeant 8-CPT-cAMP (here abbreviated cAMP), hydrocortisone, and a phosphodiesterase inhibitor, RO-20-1724 (RO). The procedure was carried out over a period of 9-11 days, and when establishing the NCC model, the astrocytes were cultured 2-8 weeks in advance. Adherence to the described procedures in the protocol has allowed the establishment of endothelial layers with highly restricted paracellular permeability, with the NCC model showing an average transendothelial electrical resistance (TEER) of 1249 &#177; 80 &#937; cm2, and paracellular permeability (Papp) for Lucifer Yellow of 0.90 10-6 &#177; 0.13 10-6 cm sec-1 (mean &#177; SEM, n=55). Further evaluation of this pBEC phenotype showed good expression of the tight junctional proteins claudin 5, ZO-1, occludin and adherens junction protein p120 catenin. The model presented can be used for a range of studies of the BBB in health and disease and, with the highly restrictive paracellular permeability, this model is suitable for studies of transport and intracellular trafficking.

Improved Method for the Establishment of an In Vitro Blood-Brain Barrier Model Based on Porcine Brain Endothelial Cells

The aim of the protocol is to present an optimized procedure for the establishment of an in vitro blood-brain barrier (BBB) model based on primary porcine brain endothelial cells (pBECs). The model shows high reproducibility, high tightness, and is suitable for studies of transport and intracellular trafficking in drug discovery.

Melhor método para o estabelecimento de uma In Vitro barreira hemato - encefálica modelo baseado em células endoteliais do cérebro de suínos

The aim of this protocol presents an optimized procedure for the purification and cultivation of pBECs and to establish in vitro blood-brain barrier (BBB) ...

Studying the Hypothalamic Insulin Signal to Peripheral Glucose Intolerance with a Continuous Drug Infusion System into the Mouse Brain

Insulin regulates systematic metabolism in the hypothalamus and the peripheral insulin response. An inflammatory reaction in peripheral adipose tissues contributes to type 2 diabetes mellitus (T2DM) development and appetite regulation in the hypothalamus. Chemokine CCL5 and C-C chemokine receptor type 5 (CCR5) levels have been suggested to mediate arteriosclerosis and glucose intolerance in type 2 diabetes mellitus (T2DM). In addition, CCL5 plays a neuroendocrine role in the hypothalamus by regulating food intake and body temperature, thus, prompting us to investigate its function in hypothalamic insulin signaling and the regulation of peripheral glucose metabolism.
The micro-osmotic pump brain infusion system is a quick and precise way to manipulate CCL5 function and study its effect in the brain. It also provides a convenient alternative approach to generating a transgenic knockout animal. In this system, CCL5 signaling was blocked by intracerebroventricular (ICV) infusion of its antagonist, MetCCL5, using a micro-osmotic pump. The peripheral glucose metabolism and insulin responsiveness was detected by the Oral Glucose Tolerance Test (OGTT) and Insulin Tolerance Test (ITT). Insulin signaling activity was then analyzed by protein blot from tissue samples derived from the animals.
After 7-14 days of MetCCL5 infusion, the glucose metabolism and insulin responsiveness was impaired in mice, as seen in the results of the OGTT and ITT. The IRS-1 serine302 phosphorylation was increased and the Akt activity was reduced in mice hypothalamic neurons following CCL5 inhibition. Altogether, our data suggest that blocking CCL5 in the mouse brain increases the phosphorylation of IRS-1 S302 and interrupts hypothalamic insulin signaling, leading to a decrease in insulin function in peripheral tissues as well as the impairment of glucose metabolism.

This protocol studies the role of chemokine (C-C motif) ligand 5 (CCL5) in the hypothalamus by delivering an antagonist, MetCCL5, into the mouse brain using a micro-osmotic pump brain infusion system. This transient inhibition of CCL5 activity interrupted hypothalamic insulin signaling, leading to glucose intolerance and peripheral systemic insulin sensitivity.

Estudando o sinal de insulina hipotalâmico a intolerância de glicose periférica com um sistema de infusão contínua de drogas dentro do cérebro de rato

Insulin regulates systematic metabolism in the hypothalamus and the peripheral insulin response. An inflammatory reaction in peripheral adipose tissues ...

Image-Guided Resection of Glioblastoma and Intracranial Implantation of Therapeutic Stem Cell-seeded Scaffolds

Glioblastoma (GBM), the most common and aggressive primary brain cancer, carries a life expectancy of 12-15 months. The short life expectancy is due in part to the inability of the current treatment, consisting of surgical resection followed by radiation and chemotherapy, to eliminate invasive tumor foci. Treatment of these foci may be improved with tumoricidal human mesenchymal stem cells (MSCs). MSCs exhibit potent tumor tropism and can be engineered to express therapeutic proteins that kill tumor cells. Advancements in preclinical models indicate that surgical resection induces premature MSC loss and reduces therapeutic efficacy. Efficacy of MSC treatment can be improved by seeding MSCs on a biodegradable poly(lactic acid) (PLA) scaffold. MSC delivery into the surgical resection cavity on a PLA scaffold restores cell retention, persistence, and tumor killing. To study the effects of MSC-seeded PLA implantation on GBM, an accurate preclinical model is needed. Here we provide a preclinical surgical protocol for image-guided tumor resection of GBM in immune-deficient mice followed by MSC-seeded scaffold implantation. MSCs are engineered with lentiviral constructs to constitutively express and secrete therapeutic TNF&#945;-related apoptosis-inducing ligand (TRAIL) as well as green fluorescent protein (GFP) to allow fluorescent tracking. Similarly, the U87 tumor cells are engineered to express mCherry and firefly luciferase, providing dual fluorescent/luminescent tracking. While currently used for investigating stem cell mediated delivery of therapeutics, this protocol could be modified to investigate the impact of surgical resection on other GBM interventions.

Tumor-seeking therapeutic mesenchymal stem cells (MSCs) show promise as a treatment for invasive glioblastoma. Optimal transplantation involves delivery of MSCs into the tumor resection cavity on scaffolds. Here, preclinical techniques to study MSC treatment of glioblastoma are provided including: image-guided tumor resection; implantation of MSC-seeded scaffolds; and postoperative therapy tracking.

Ressecção guiada por imagem de Glioblastoma e intracraniana implantação de andaimes terapêuticos de células-tronco-semeado

Glioblastoma (GBM), the most common and aggressive primary brain cancer, carries a life expectancy of 12-15 months. The short life expectancy is due in ...

Predicting Treatment Response to Image-Guided Therapies Using Machine Learning: An Example for Trans-Arterial Treatment of Hepatocellular Carcinoma

Intra-arterial therapies are the standard of care for patients with hepatocellular carcinoma who cannot undergo surgical resection. The objective of this study was to develop a method to predict response to intra-arterial treatment prior to intervention.
The method provides a general framework for predicting outcomes prior to intra-arterial therapy. It involves pooling clinical, demographic and imaging data across a cohort of patients and using these data to train a machine learning model. The trained model is applied to new patients in order to predict their likelihood of response to intra-arterial therapy.
The method entails the acquisition and parsing of clinical, demographic and imaging data from N patients who have already undergone trans-arterial therapies. These data are parsed into discrete features (age, sex, cirrhosis, degree of tumor enhancement, etc.) and binarized into true/false values (e.g., age over 60, male gender, tumor enhancement beyond a set threshold, etc.). Low-variance features and features with low univariate associations with the outcome are removed. Each treated patient is labeled according to whether they responded or did not respond to treatment. Each training patient is thus represented by a set of binary features and an outcome label. Machine learning models are trained using N - 1 patients with testing on the left-out patient. This process is repeated for each of the N patients. The N models are averaged to arrive at a final model.
The technique is extensible and enables inclusion of additional features in the future. It is also a generalizable process that may be applied to clinical research questions outside of interventional radiology. The main limitation is the need to derive features manually from each patient. A popular modern form of machine learning called deep learning does not suffer from this limitation, but requires larger datasets.

Intra-arterial therapies are the standard of care for patients with hepatocellular carcinoma who cannot undergo surgical resection. A method for predicting response to these therapies is proposed. The technique uses pre-procedural clinical, demographic, and imaging information to train machine learning models capable of predicting response prior to treatment.

Prevendo a resposta do tratamento para terapias guiada por imagem utilizando aprendizado de máquina: um exemplo para o Trans-Arterial tratamento do Carcinoma hepatocelular

Intra-arterial therapies are the standard of care for patients with hepatocellular carcinoma who cannot undergo surgical resection. The objective of this ...

An Image Guided Transapical Mitral Valve Leaflet Puncture Model of Controlled Volume Overload from Mitral Regurgitation in the Rat

Mitral regurgitation (MR) is a widely prevalent heart valve lesion, which causes cardiac remodeling and leads to congestive heart failure. Though the risks of uncorrected MR and its poor prognosis are known, the longitudinal changes in cardiac function, structure and remodeling are incompletely understood. This knowledge gap has limited our understanding of the optimal timing for MR correction, and the benefit that early versus late MR correction may have on the left ventricle. To investigate the molecular mechanisms that underlie left ventricular remodeling in the setting of MR, animal models are necessary. Traditionally, the aorto-caval fistula model has been used to induce volume overload, which differs from clinically relevant lesions such as MR. MR represents a low-pressure volume overload hemodynamic stressor, which requires animal models that mimic this condition. Herein, we describe a rodent model of severe MR in which the anterior leaflet of the rat mitral valve is perforated with a 23G needle, in a beating heart, with echocardiographic image guidance. The severity of MR is assessed and confirmed with echocardiography, and the reproducibility of the model is reported.&#160;&#160;

A rodent model of left heart volume overload from mitral regurgitation is reported. Mitral regurgitation of controlled severity is induced by advancing a needle of defined dimensions into the anterior leaflet of the mitral valve, in a beating heart, with ultrasound guidance.&#160;

Um modelo de punção de folheto de válvula mitral transapical guiada por imagem de sobrecarga de volume controlada da regurgitação mitral no rato

Mitral regurgitation (MR) is a widely prevalent heart valve lesion, which causes cardiac remodeling and leads to congestive heart failure. Though the ...

Intratracheal Administration of Dry Powder Formulation in Mice

In the development of inhalable dry powder formulations, it is essential to evaluate their biological activities in preclinical animal models. This paper introduces a noninvasive method of intratracheal delivery of dry powder formulation in mice. A dry powder loading device that consists of a 200 &#181;L gel loading pipette tip connected to an 1 mL syringe via a three-way stopcock is presented. A small amount of dry powder (1-2 mg) is loaded into the pipette tip and dispersed by 0.6 mL of air in the syringe. Because pipette tips are disposable and inexpensive, different dry powder formulations can be loaded into different tips in advance. Various formulations can be evaluated in the same animal experiment without device cleaning and dose refilling, thereby saving time and eliminating the risk of cross-contamination from residual powder. The extent of powder dispersion can be inspected by the amount of powder remaining in the pipette tip. A protocol of intubation in mouse with a custom-made light source and a guiding cannula is included. Proper intubation is one of the key factors that influences the intratracheal delivery of dry powder formulation to the deep lung region of the mouse.

Dry powder formulations for inhalation have great potential in treating respiratory diseases. Before entering human studies, it is necessary to evaluate the efficacy of the dry powder formulation in preclinical studies. A simple and noninvasive method of the administration of dry powder in mice through the intratracheal route is presented.

Administração Intratraqueal de Formulação de Pó Seco em Camundongos

In the development of inhalable dry powder formulations, it is essential to evaluate their biological activities in preclinical animal models. This paper ...