Este estudio fue apoyado por el Premio a la Innovación de la familia Farris y los NIH 1R01NR015674-01 a MK. Los autores agradecen a Josep Nayfach (Qteris, Inc) para proporcionar un sistema generador electro-magnética, así como su consulta técnica. Los autores también agradecen Huan Yan (Programa Interdisciplinario de Física y Química de LCI, Kent State University) por sus asistentes técnicos.

Nota: El proceso global de fabricación de microgeles de gelatina sensible al campo magnético se ilustra en la Figura 1A. 1. Preparación de soluciones y suspensiones <ol><li> Preparar una genipina reticulante (1% w / v) de solución mediante la disolución de 20 mg de genipina en 2 ml de solución salina tamponada con fosfato (1x PBS; pH 7,4). Vortex la solución y colocar en un 50 ° C baño de agua durante 2 horas para disolver completamente la solución. </li><li> Preparar una solución de tensioactivo disolviendo 20 mg de poli (etilenglicol) -poli (propilenglicol) -poli (etilenglicol) (M w = 2900 Da; referido como L64) en 200 ml de PBS que estar en el concentración de 100 ppm. </li><li> Preparar un 15% (v w /) solución de gelatina disolviendo 64,5 mg de gelatina en 0,43 ml de PBS. Vortex la solución y colocarlo en baño de agua a 37 ° C hasta que se alcanza una fase de sol, donde la solución se vuelve fluido. Entonces, vórtice de la solución de gelatina 2 - 3 veces para asegurar THe homogeneidad de la muestra. </li><li> Preparación de p (NIPAM- co -AAm) solución / MNPs con un fármaco modelo (BSA): <ol><li> Dispersar 10,75 mg de hidrófilo MNPs en 0,43 ml de PBS y luego se disuelven 12,9 mg de p (NIPAM- co -AAm) en la suspensión MNP para hacer la concentración a 3% (w / v). El aumento de la concentración de p (NIPAM- co -AAm) se puede utilizar para lograr un mayor comportamiento de deshinchamiento de microgeles. </li><li> Utilice la albúmina de suero de Texas-Red conjugado bovina (BSA-TR; M w ~ 66 kDa) como fármaco modelo. Disolver 0,5 mg de TR-BSA en la mezcla de p (NIPAM- co -AAm) / MNPs. </li></ol></li><li> Preparar las mezclas de gelatina / p (NIPAM- co -AAm) / solución MNPs / BSA (0,86 ml) mediante la adición de mezcla de p (NIPAM- co -AAm) / MNPs (0,43 ml) en la solución de gelatina (0,43 ml) y luego mezclar bien para hacer una mezcla homogénea. Por lo tanto, las concentraciones de polímeros y MNP se convierten en medio de las concentraciones iniciales de la mezcla final. </li></ol><ol><li> Verter 15 ml de aceite de silicona [polidimetilsiloxano (viscosidad 350 cSt)] en un vaso limpio y estéril. </li><li> Inmediatamente añadir las mezclas acuosas pre-preparadas de gelatina / p (NIPAM- co -AAm) / solución MNPs / BSA (0,86 ml) en el aceite de silicona y emulsionar la mezcla acuosa en la fase oleosa mediante agitación con una barra de agitación magnética a 900 rpm a 30 ° C durante 30 min. </li></ol> 3. La gelificación y transferencia de micro-gotas a una solución acuosa <ol><li> Se transfiere la emulsión (~ 16 ml) de vaso de precipitados en un tubo de 50 ml. </li><li> Enfriar el tubo durante 10 min a 4 ° C para la gelificación de las microgotas en el aceite. </li><li> Llenar el tubo con la solución preparada L64 (a 4 ° C) hasta 50 ml y agitar vigorosamente el tubo. Puede ser posible que una parte de L64 tensioactivos estaría dentro de los microgeles. </li><li> Centrifugar el tubo durante 20 minutos a 2300 xg a 4 ° C. </li><li> Regularly comprobar la presencia de la pastilla de partículas de gel en el lado del tubo. Si las partículas no se ven, centrifugar durante otros 20 minutos a la misma velocidad y temperatura. Proceder para eliminar cuidadosamente el sobrenadante sin perturbar el sedimento formado en la pared interior del tubo. </li><li> Repita los pasos (3.3) a (3.5) una vez más. Cada vez, la transferencia de la muestra a un tubo nuevo para evitar la inclusión de cualquier gotitas de aceite en la suspensión de microgel. Después de este paso, asegurar que los agentes tensioactivos o gotitas de aceite no están presentes en la suspensión de la muestra. Sin embargo, las etapas de separación repetidas pueden conducir a la pérdida de los materiales iniciales. </li></ol> 4. reticulación covalente de los microgeles <ol><li> Añadir 2 ml de solución genipina (preparado en el apartado 1) al sedimento de partículas de gel y se mezcla bien por agitación la solución. </li><li> Transferir rápidamente el tubo de la suspensión en un baño de agua a 23 ° C para iniciar una reacción de reticulación covalente durante una des(. ejemplo, 5 - 120 min) tiempo de reticulación ired. </li><li> Después de la reticulación, retire inmediatamente cualquier reticulantes excesivas descartando la solución genipina, resuspensión de los microgeles en PBS y centrifugando el tubo durante 20 min a 2.300 xg (4 ° C). Si es necesario, se separan con cuidado formada de pellets con una punta de pipeta. Esta etapa de lavado se puede repetir hasta 3 veces si el genipina todavía se queda en la solución. </li><li> Eliminar el sobrenadante y resuspender los microgeles en PBS a una densidad deseada (por ejemplo., 5 × 10 6 microgeles / ml) contando el número con un hemocitómetro. </li><li> Para las observaciones microscópicas, cargar la suspensión de microgel en el espacio entre un portaobjetos y un cubreobjetos y selle la frontera de la hoja de la cubierta con resina epoxi. </li></ol> 5. La aplicación de campo magnético alterno para la activación de Liberación de Fármacos <ol><li> Colocar el tubo con la concentración deseada de microgeles en medios acuosos enla cámara de bobinas magnéticas. Si es necesario, inserte una sonda de temperatura de fibra óptica en el tubo para monitorear los cambios de temperatura de los medios de comunicación durante la aplicación de la AMF. </li><li> Aplicar de alta frecuencia (&gt; 100 kHz) AMF en una intensidad de campo definida (&gt; 5 kA / m) y para una duración especificada. Tras la aplicación de AMF, centrifugar el tubo de muestra durante 20 min a 2.273 xg (4 ° C) y recoger el sobrenadante para cuantificar la cantidad de TR-BSA liberado de microgel a los medios circundantes utilizando espectrofotometría. Las longitudes de onda de excitación y emisión de Texas Red 584 nm y 612 nm, respectivamente. </li></ol>

<ol>
	<li>Langer, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomaterials+in+drug+delivery+and+tissue+engineering:+one+laboratory's+experience.">Biomaterials in drug delivery and tissue engineering: one laboratory's experience.</a> Acc. Chem. Res. 33, 94-101 (2000).</li><li>Rivest, C. M., Morrison, D., Ni, B., Rubib, J., Yadav, V., Mahdavi, A., Karp, J., Khademhosseini, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscale+hydrogels+for+medicine+and+biology:+synthesis,+characteristics+and+applications.">Microscale hydrogels for medicine and biology: synthesis, characteristics and applications.</a> J Mech Mater Struct. 2, 1103-1119 (2007).</li><li>Kawaguchi, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thermoresponsive+microhydrogels:+preparation,+properties+and+applications.">Thermoresponsive microhydrogels: preparation, properties and applications.</a> Polym. Int. 63, 925-932 (2014).</li><li>Vinogradov, S. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colloidal+microgels+in+drug+delivery+applications.">Colloidal microgels in drug delivery applications.</a> Curr. Pharm. Des. 12, 4703-4712 (2006).</li><li>Liechty, W. B., Kryscio, D. R., Slaughter, B. V., Peppas, N. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polymers+for+drug+delivery+systems.">Polymers for drug delivery systems.</a> Annu Rev Chem Biomol Eng. 1, 149-173 (2010).</li><li>Kumari, A., Yadav, S. K., Yadav, S. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biodegradable+polymeric+nanoparticles+based+drug+delivery+systems.">Biodegradable polymeric nanoparticles based drug delivery systems.</a> Colloids Surf. B Biointerfaces. 75, 1-18 (2010).</li><li>Shibayama, M., Tanaka, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volume+Phase-Transition+and+Related+Phenomena+of+Polymer+Gels.">Volume Phase-Transition and Related Phenomena of Polymer Gels.</a> Adv Polym Sci. 109, 1-62 (1993).</li><li>Sung, B., Kim, C., Kim, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biodegradable+colloidal+microgels+with+tunable+thermosensitive+volume+phase+transitions+for+controllable+drug+delivery.">Biodegradable colloidal microgels with tunable thermosensitive volume phase transitions for controllable drug delivery.</a> J Colloid Interface Sci. 450, 26-33 (2015).</li><li>Kumar, C. S., Mohammad, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetic+nanomaterials+for+hyperthermia-based+therapy+and+controlled+drug+delivery.">Magnetic nanomaterials for hyperthermia-based therapy and controlled drug delivery.</a> Adv. Drug Deliv. Rev. 63, 789-808 (2011).</li><li>Mura, S., Nicolas, J., Couvreur, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stimuli-responsive+nanocarriers+for+drug+delivery.">Stimuli-responsive nanocarriers for drug delivery.</a> Nat. Mater. 12, 991-1003 (2013).</li><li>Kong, S. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetic+field+activated+lipid-polymer+hybrid+nanoparticles+for+stimuli-responsive+drug+release.">Magnetic field activated lipid-polymer hybrid nanoparticles for stimuli-responsive drug release.</a> Acta biomaterialia. 9, 5447-5452 (2013).</li><li>Hayashi, 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=Magnetically+responsive+smart+nanoparticles+for+cancer+treatment+with+a+combination+of+magnetic+hyperthermia+and+remote-control+drug+release.">Magnetically responsive smart nanoparticles for cancer treatment with a combination of magnetic hyperthermia and remote-control drug release.</a> Theranostics. 8, 834-844 (2014).</li><li>Suzuki, D., Kawaguchi, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stimuli-sensitive+core/shell+template+particles+for+immobilizing+inorganic+nanoparticles+in+the+core.">Stimuli-sensitive core/shell template particles for immobilizing inorganic nanoparticles in the core.</a> Colloid Polym Sci. 284, 1443-1451 (2006).</li><li>Bhattacharya, S., Eckert, F., Boyko, V., Pich, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature-,+pH-,+and+magnetic-field-sensitive+hybrid+microgels.">Temperature-, pH-, and magnetic-field-sensitive hybrid microgels.</a> Small. 3, 650-657 (2007).</li><li>Wong, J. E., Gaharwar, A. K., Muller-Schulte, D., Bahadur, D., Richtering, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dual-stimuli+responsive+PNiPAM+microgel+achieved+via+layer-by-layer+assembly:+Magnetic+and+thermoresponsive.">Dual-stimuli responsive PNiPAM microgel achieved via layer-by-layer assembly: Magnetic and thermoresponsive.</a> J Colloid Interf Sci. 324, 47-54 (2008).</li><li>Zhao, X., 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=Active+scaffolds+for+on-demand+drug+and+cell+delivery.">Active scaffolds for on-demand drug and cell delivery.</a> Proc. Natl. Acad. Sci. U.S.A. 108, 67-72 (2011).</li><li>Xu, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Release+of+magnetic+nanoparticles+from+cell-encapsulating+biodegradable+nanobiomaterials.">Release of magnetic nanoparticles from cell-encapsulating biodegradable nanobiomaterials.</a> ACS nano. 6, 6640-6649 (2012).</li><li>Li, Y. 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=Magnetic+Hydrogels+and+Their+Potential+Biomedical+Applications.">Magnetic Hydrogels and Their Potential Biomedical Applications.</a> Adv Funct Mater. 23, 660-672 (2013).</li><li>Cooperstein, M. A., Canavan, H. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+cytotoxicity+of+(N-isopropyl+acrylamide)+and+poly(N-isopropyl+acrylamide)-coated+surfaces.">Assessment of cytotoxicity of (N-isopropyl acrylamide) and poly(N-isopropyl acrylamide)-coated surfaces.</a> Biointerphases. 8, 19 (2013).</li><li>Jorgensen, L., Moeller, E. H., van de Weert, M., Nielsen, H. M., Frokjaer, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparing+and+evaluating+delivery+systems+for+proteins.">Preparing and evaluating delivery systems for proteins.</a> Eur J Pharm Sci. 29, 174-182 (2006).</li><li>Holland, T. A., Tabata, Y., Mikos, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+release+of+transforming+growth+factor-beta+1+from+gelatin+microparticles+encapsulated+in+biodegradable,+injectable+oligo(poly(ethylene+glycol)+fumarate)+hydrogels.">In vitro release of transforming growth factor-beta 1 from gelatin microparticles encapsulated in biodegradable, injectable oligo(poly(ethylene glycol) fumarate) hydrogels.</a> J Control Release. 91, 299-313 (2003).</li><li>Liang, H. C., Chang, W. H., Lin, K. J., Sung, H. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genipin-crosslinked+gelatin+microspheres+as+a+drug+carrier+for+intramuscular+administration:+in+vitro+and+in+vivo+studies.">Genipin-crosslinked gelatin microspheres as a drug carrier for intramuscular administration: in vitro and in vivo studies.</a> J Biomed Mater Res. Part A. 65, 271-282 (2003).</li><li>Solorio, L., Zwolinski, C., Lund, A. W., Farrell, M. J., Stegemann, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gelatin+microspheres+crosslinked+with+genipin+for+local+delivery+of+growth+factors.">Gelatin microspheres crosslinked with genipin for local delivery of growth factors.</a> J Tissue Eng Regen Med. 4, 514-523 (2010).</li><li>Regmi, 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=Hyperthermia+controlled+rapid+drug+release+from+thermosensitive+magnetic+microgels.">Hyperthermia controlled rapid drug release from thermosensitive magnetic microgels.</a> J Mater Chem. 20, 6158-6163 (2010).</li><li>Kim, M. 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=Magnetic+nanoparticle+targeted+hyperthermia+of+cutaneous+Staphylococcus+aureus+infection.">Magnetic nanoparticle targeted hyperthermia of cutaneous Staphylococcus aureus infection.</a> Ann Biomed Eng. 41, 598-609 (2013).</li><li>Ivkov, 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=Application+of+high+amplitude+alternating+magnetic+fields+for+heat+induction+of+nanoparticles+localized+in+cancer.">Application of high amplitude alternating magnetic fields for heat induction of nanoparticles localized in cancer.</a> Clin Cancer Res. 11, 7093s-7103s (2005).</li><li>Huang, S., Fu, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Naturally+derived+materials-based+cell+and+drug+delivery+systems+in+skin+regeneration.">Naturally derived materials-based cell and drug delivery systems in skin regeneration.</a> J Control Release. 142, 149-159 (2010).</li><li>Malafaya, P. B., Silva, G. A., Reis, 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=Natural-origin+polymers+as+carriers+and+scaffolds+for+biomolecules+and+cell+delivery+in+tissue+engineering+applications.">Natural-origin polymers as carriers and scaffolds for biomolecules and cell delivery in tissue engineering applications.</a> Adv. Drug Deliv. Rev. 59, 207-233 (2007).</li><li>Shah, R., Kim, J., Agresti, J., Weitz, D., Chu, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fabrication+of+monodisperse+thermosensitive+microgels+and+gel+capsules+in+microfluidic+devices.">Fabrication of monodisperse thermosensitive microgels and gel capsules in microfluidic devices.</a> Soft Matter. 4, 2303-2309 (2008).</li><li>Hoare, 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=Magnetically+triggered+nanocomposite+membranes:+a+versatile+platform+for+triggered+drug+release.">Magnetically triggered nanocomposite membranes: a versatile platform for triggered drug release.</a> Nano letters. 11, 1395-1400 (2011).</li></ol>

The authors have nothing to disclose.

La tecnología descrita aquí demuestra una prueba de concepto en el uso de híbridos de nanopartículas de microgel para la liberación del fármaco magneto-térmico disparada. Esto se logró atrapando físicamente cadenas de MNPs y P (NIPAM- co -AAm) dentro de una red de gelatina tridimensional microescala reticulado por genipina. La plataforma de campo magnético sensible fue suficiente para generar calor dentro del microgel en respuesta a una AMF aplicado de forma remota, que a su vez desencadena la liberac...

Los estímulos que responden a los sistemas de administración de fármacos que permiten una administración de fármacos estrechamente controlada en respuesta a cualquiera de los estímulos endógenos o exógenos (por ejemplo., La temperatura o pH) se han investigado extensamente como nuevos tipos de dispositivos suaves inteligentes para la administración de fármacos. Hidrogeles microescala han sido ampliamente empleado como una plataforma de administración de fármacos en que confieren perfiles controlables y sostenibles de liberación de fármacos, así como química sintonizable y propiedades mecánicas 1-3. En particular, los microgeles coloidales presentan muchas ventajas como un vehículo para la administración de fármacos debido a su rápida capacidad de respuesta a estímulos externos y inyectabilidad adecuado para tejido local de una manera mínimamente invasiva 4. El poli (N-isopropilacrilamida) (PNIPAM) o sus copolímeros han sido ampliamente adoptados en la síntesis de microgeles termo-sensible por injerto de PNIPAM con polímeros biodegradables / biocompatibles incluyendo gelatina, quitosano, ácido alginato, o ácido hialurónico 5,6, En el que una característica de transición de fase de PNIPAM a su temperatura de solución crítica inferior (LCST) se puede utilizar como un disparador de liberación del fármaco 7. Recientemente hemos demostrado una fabricación de microgel termosensible biodegradable, a base de gelatina mediante la incorporación de poli (co N-isopropylacrylamide- acrilamida) [p (NIPAM- co -AAm)] cadenas como un componente menor dentro de las redes de gelatina tridimensionales 8. El / p (NIPAM- co -AAm) microgel gelatina exhibió un deshinchamiento sintonizable de aumento de la temperatura, lo que se correlaciona positivamente con la liberación de albúmina de suero bovino (BSA). Durante los últimos años, ha habido un incremento esfuerzos para desarrollar una plataforma de administración de fármacos magnéticamente sensible que puede desencadenar la liberación de fármaco en un 9,10 moda bajo demanda. El principio básico para la síntesis de la plataforma de administración de fármacos magnéticamente sensible utiliza la característica de nanopartículas superparamagnéticas (NPM) para generar calor cuando reciben una alta frecuencia del campo magnético (AMF), que desencadena una liberación del fármaco sensible a la temperatura alterna. Esta es una promesa para futuras aplicaciones clínicas en que este sistema puede orientar profundamente en el tejido, permite una liberación de fármaco no invasivo y controlado a distancia y se puede combinar con un tratamiento de hipertermia y el sistema de formación de imágenes de resonancia magnética 10-12. Tales plataformas incluyen: (1) las partículas de microgel NPM / PNIPAM híbrido 13-15 y (2) los andamios de hidrogel que incorporan macroscópicas inmovilizan los micronutrientes en polvo 16-18. Las plataformas basadas en microgel PNIPAM demostraron una capacidad de respuesta de transición de fase de volumen finamente sintonizable a los estímulos magnetotérmicos. Sin embargo, todavía se basan en técnicas complejas y sofisticadas en la fabricación y el uso de polímeros PNIPAM con un alto contenido potencialmente pueden ser citotóxicas para las células 19, lo que puede limitar sus aplicaciones in vivo. Los andamios macroscópicas exhiben un parienteLy lenta respuesta a los estímulos externos y requieren un trasplante quirúrgico invasivo en comparación con los microgeles coloidales. La emulsificación de agua-en-aceite ha sido el método estándar para producir submilimétrico o gel micrómetros partículas de tamaño 20. En la interfaz agua-aceite de la emulsión, de partículas de microgel forma una forma esférica debido a la minimización de la energía superficial de la gota de agua en virtud de la fuerza de corte mecánica. Este método permite la producción de una gran cantidad de gotitas de gel esféricas acuosas en un sencillo procedimiento de fabricación y se ha adoptado con éxito para la fabricación de microgeles a base de gelatina para aplicaciones de administración de fármacos 21-23. A continuación, presentamos un método fácil de sintetizar un microgeles a base de gelatina magnetothermally adecuados para hacer posible la aplicación de administración de fármacos empleando el método de emulsión de agua-en-aceite. Esto se consigue incorporando físicamente MNPs de óxido de hierro y p (co NIPAM- -AAm) cadenas como un componente menor dentro de una red de gelatina a microescala esférica que está reticulado covalentemente por un agente de reticulación genipina de origen natural, junto con una alta frecuencia sistema de aplicación de campo magnético (AMF) alterna.

<table border="0" cellpadding="0" cellspacing="0" width="706">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Gelatin</td>
			<td style="width:204px;">Sigma-Aldrich, MO, USA</td>
			<td style="width:119px;">G2500</td>
			<td style="width:167px;">Gelatin type A, porcine skin</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">poly(N-isopropylacrylamide-co-acrylamide)&#160;</td>
			<td style="width:204px;">Sigma-Aldrich, MO, USA</td>
			<td>738727</td>
			<td>MW=20,000, LCST=34-38 oC</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Silicon oil</td>
			<td style="width:204px;">Sigma-Aldrich, MO, USA</td>
			<td>378372</td>
			<td>Viscosity 350 cSt</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Pluoronic L64</td>
			<td style="width:204px;">Sigma-Aldrich, MO, USA</td>
			<td style="width:119px;">435449</td>
			<td>100 ppm poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:216px;">genipin</td>
			<td style="width:204px;">TimTec LLC, DE, USA</td>
			<td>ST080860</td>
			<td>Mw = 226.23;&#160;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Magnetic nanoparticles (MNPs)</td>
			<td style="width:204px;">Micromod Inc, Germany</td>
			<td>79-00-102</td>
			<td>nanomag-D-spio, 100 nm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">TR-BSA</td>
			<td style="width:204px;">Life Technologies, NY USA</td>
			<td>A23017</td>
			<td>Albumin from Bovine Serum (BSA), Texas Red conjugate</td>
		</tr>
	</tbody>
</table>

alternating magnetic field-responsive hybrid gelatin microgels for controlled drug release

nano biomateriales magnéticamente sensible / micro-ingeniería que permiten a, la administración de fármacos estrechamente controlada bajo demanda han sido desarrollados como nuevos tipos de dispositivos suaves inteligentes para aplicaciones biomédicas. Aunque un número de sistemas de administración de fármacos magnéticamente sensibles han demostrado eficacias a través de ya sea la prueba in vitro de estudios de concepto o en aplicaciones preclínicos in vivo, su uso en la práctica clínica es todavía limitada por su biocompatibilidad insuficiente o biodegradabilidad. Además, muchas de las plataformas existentes se basan en técnicas sofisticadas para sus fabricaciones. Recientemente hemos demostrado la fabricación de microgel termosensible biodegradable, a base de gelatina atrapando físicamente poli (N-isopropylacrylamide- acrilamida co) cadenas como un componente menor dentro de una red de gelatina tridimensional. En este estudio, se presenta un método fácil de fabricar una plataforma de liberación de fármaco biodegradable que permite a un magneto-thermally provocado la liberación del fármaco. Esto se logró mediante la incorporación de nanopartículas de óxido de hierro superparamagnético y polímeros termo-sensible dentro de microgeles coloidales a base de gelatina, en conjunción con un sistema de aplicación de campo magnético alterno.

Cuando el protocolo se realiza correctamente, los microgeles fabricadas deben exhibir una morfología esférica bien caracterizado y dispersabilidad coloidal con diámetros en el intervalo entre 5 micras a 20 micras (Figura 1B y C). Cualquiera de MNPs fluorescentes o BSA fluorescente se puede utilizar para confirmar si MNPs o drogas (BSA en este estudio) se encapsulan correctamente dentro del microgel (Figura 1D). Los microgeles fabricados pueden ser est...

Watch this Scientific Journal Video about Alternating Magnetic Field-Responsive Hybrid Gelatin Microgels for Controlled Drug Release at JoVE.com

Alternating Magnetic Field-Responsive Hybrid Gelatin Microgels for Controlled Drug Release

Alternando microgeles de gelatina de campo híbrido-Responsive magnéticos para Controlled Drug Release

Se presenta un método fácil de fabricar una plataforma de liberación del fármaco a base de gelatina biodegradable que es sensible magneto-térmico. Esto se logró mediante la incorporación de nanopartículas de óxido de hierro superparamagnéticas y poli (co N-isopropylacrylamide- acrilamida) dentro de una gelatina esférica micro-red reticulada por genipina, en conjunción con un sistema de aplicación de campo magnético alterno.

Magnetically-responsive nano/micro-engineered biomaterials that enable a tightly controlled, on-demand drug delivery have been developed as new types of ...

alternating-magnetic-field-responsive-hybrid-gelatin-microgels-for

Research

JoVE Journal

Bioengineering

9.7K vistas.  Kent State University.  Se presenta un método fácil de fabricar una plataforma de liberación del fármaco a base de gelatina biodegradable que es sensible magneto-térmico. Esto se logró mediante la incorporación de nanopartículas de óxido de hierro superparamagnéticas y poli (co N-isopropylacrylamide- acrilamida) dentro de una gelatina esférica micro-red reticulada por genipina, en conjunción con un sistema de aplicación de campo magnético alterno. 

Video: Alternating Magnetic Field-Responsive Hybrid Gelatin Microgels for Controlled Drug Release

Magnetically-Assisted Remote Controlled Microcatheter Tip Deflection under Magnetic Resonance Imaging

X-ray fluoroscopy-guided endovascular procedures have several significant limitations, including difficult catheter navigation and use of ionizing radiation, which can potentially be overcome using a magnetically steerable catheter under MR guidance.
	The main goal of this work is to develop a microcatheter whose tip can be remotely controlled using the magnetic field of the MR scanner. This protocol aims to describe the procedures for applying current to the microcoil-tipped microcatheter to produce consistent and controllable deflections.
	A microcoil was fabricated using laser lathe lithography onto a polyimide-tipped endovascular catheter. In vitro testing was performed in a waterbath and vessel phantom under the guidance of a 1.5-T MR system using steady-state free precession (SSFP) sequencing. Various amounts of current were applied to the coils of the microcatheter to produce measureable tip deflections and navigate in vascular phantoms.
	The development of this device provides a platform for future testing and opportunity to revolutionize the endovascular interventional MRI environment.

Current applied to an endovascular microcatheter with microcoil tip made by laser lathe lithography can achieve controllable deflections under magnetic resonance (MR) guidance, which may improve speed and efficacy of navigation of vasculature during various endovascular procedures.

Magnéticamente Asistencia Remota Controlada microcatéter desviación de la punta bajo la resonancia magnética

X-ray fluoroscopy-guided  endovascular procedures have several significant limitations, including  difficult catheter navigation and use of ionizing ...

Investigación

Bioingeniería

Characteristics of Precipitation-formed Polyethylene Glycol Microgels Are Controlled by Molecular Weight of Reactants

This work describes the formation of poly(ethylene glycol) (PEG) microgels via a photopolymerized precipitation reaction. Precipitation reactions offer several advantages over traditional microsphere fabrication techniques. Contrary to emulsion, suspension, and dispersion techniques, microgels formed by precipitation are of uniform shape and size, i.e.&#160;low polydispersity index, without the use of organic solvents or stabilizers. The mild conditions of the precipitation reaction, customizable properties of the microgels, and low viscosity for injections make them applicable for in vivo purposes. Unlike other fabrication techniques, microgel characteristics can be modified by changing the starting polymer molecular weight. Increasing the starting PEG molecular weight increased microgel diameter and swelling ratio. Further modifications are suggested such as encapsulating molecules during microgel crosslinking. Simple adaptations to the PEG microgel building blocks are explored for future applications of microgels as drug delivery vehicles and tissue engineering scaffolds.

This work describes the formation of poly(ethylene glycol) (PEG) microgels via a photopolymerized precipitation reaction. Increasing the PEG molecular weight increased microgel diameter and swelling ratio. Simple adaptations to the PEG microgel precipitation reaction are explored for future applications of microgels as drug delivery vehicles and tissue engineering scaffolds.

Características de los formados Precipitación microgeles polietilenglicol son controlados por Peso molecular de los reactivos

This work describes the formation of poly(ethylene glycol) (PEG) microgels via a photopolymerized precipitation reaction. Precipitation reactions offer ...

Hybrid &#181;CT-FMT imaging and image analysis

Fluorescence-mediated tomography (FMT) enables longitudinal and quantitative determination of the fluorescence distribution in vivo and can be used to assess the biodistribution of novel probes and to assess disease progression using established molecular probes or reporter genes. The combination with an anatomical modality, e.g., micro computed tomography (&#181;CT), is beneficial for image analysis and for fluorescence reconstruction. We describe a protocol for multimodal &#181;CT-FMT imaging including the image processing steps necessary to extract quantitative measurements. After preparing the mice and performing the imaging, the multimodal data sets are registered. Subsequently, an improved fluorescence reconstruction is performed, which takes into account the shape of the mouse. For quantitative analysis, organ segmentations are generated based on the anatomical data using our interactive segmentation tool. Finally, the biodistribution curves are generated using a batch-processing feature. We show the applicability of the method by assessing the biodistribution of a well-known probe that binds to bones and joints.

Hybrid  &#181;CT-FMT imaging and image analysis

We describe a protocol for hybrid imaging, combining fluorescence-mediated tomography (FMT) with micro computed tomography (&#181;CT). After fusion and reconstruction, we perform interactive organ segmentation to extract quantitative measurements of the fluorescence distribution.

Híbrido de imágenes μCT-FMT y análisis de imágenes

Fluorescence-mediated tomography (FMT) enables longitudinal and quantitative determination of the fluorescence distribution in vivo and can be used to ...

Custom-designed Laser-based Heating Apparatus for Triggered Release of Cisplatin from Thermosensitive Liposomes with Magnetic Resonance Image Guidance

Liposomes have been employed as drug delivery systems to target solid tumors through exploitation of the enhanced permeability and retention (EPR) effect resulting in significant reductions in systemic toxicity. Nonetheless, insufficient release of encapsulated drug from liposomes has limited their clinical efficacy. Temperature-sensitive liposomes have been engineered to provide site-specific release of drug in order to overcome the problem of limited tumor drug bioavailability. Our lab has designed and developed a heat-activated thermosensitive liposome formulation of cisplatin (CDDP), known as HTLC, to provide triggered release of CDDP at solid tumors. Heat-activated delivery in vivo was achieved in murine models using a custom-built laser-based heating apparatus that provides a conformal heating pattern at the tumor site as confirmed by MR thermometry (MRT). A fiber optic temperature monitoring device was used to measure the temperature in real-time during the entire heating period with online adjustment of heat delivery by alternating the laser power. Drug delivery was optimized under magnetic resonance (MR) image guidance by co-encapsulation of an MR contrast agent (i.e.,&#160;gadoteridol) along with CDDP into the thermosensitive liposomes as a means to validate the heating protocol and to assess tumor accumulation. The heating protocol consisted of a preheating period of 5 min prior to administration of HTLC and 20 min heating post-injection. This heating protocol resulted in effective release of the encapsulated agents with the highest MR signal change observed in the heated tumor in comparison to the unheated tumor and muscle. This study demonstrated the successful application of the laser-based heating apparatus for preclinical thermosensitive liposome development and the importance of MR-guided validation of the heating protocol for optimization of drug delivery.

AN MRI-compatible&#160;custom-designed laser-based heating apparatus has been developed to provide local heating of subcutaneous tumors in order to activate release of agents from thermosensitive liposomes specifically at the tumor region.

Diseño personalizado basado en láser máquinas de calentamiento para lanzamiento disparada de cisplatino de termosensible liposomas con Resonancia Magnética Orientación Imagen

Liposomes have been employed as drug delivery systems to target solid tumors through exploitation of the enhanced permeability and retention (EPR) effect ...

Composite Scaffolds of Interfacial Polyelectrolyte Fibers for Temporally Controlled Release of Biomolecules

Various scaffolds used in tissue engineering require a controlled biochemical environment to mimic the physiological cell niche. Interfacial polyelectrolyte complexation (IPC) fibers can be used for controlled delivery of various biological agents such as small molecule drugs, cells, proteins and growth factors. The simplicity of the methodology in making IPC fibers gives flexibility in its application for controlled biomolecule delivery. Here, we describe a method of incorporating IPC fibers into two different polymeric scaffolds, hydrophilic polysaccharide and hydrophobic polycaprolactone, to create a multi-component composite scaffold. We showed that IPC fibers can be easily embedded into these polymeric structures, enhancing the capability for sustained release and improved preservation of biomolecules. We also created a composite polymeric scaffold with topographical cues and sustained biochemical release that can have synergistic effects on cell behavior. Composite polymeric scaffolds with IPC fibers represent a novel and simple method of recreating the cellular niche.

Scaffolds for tissue engineering need to recapitulate the complex biochemical and biophysical microenvironment of the cellular niche. Here, we show the use of interfacial polyelectrolyte complexation fibers as a platform to create composite, multi-component polymeric scaffolds with sustained biochemical release.

Los andamios compuestos de fibras interfacial Polyelectrolyte para lanzamiento Temporalmente controlada de biomoléculas

Various scaffolds used in tissue engineering require a controlled biochemical environment to mimic the physiological cell niche. Interfacial ...

Magnetic and Thermal-sensitive Poly(N-isopropylacrylamide)-based Microgels for Magnetically Triggered Controlled Release

Magnetically and thermally sensitive poly(N-isopropylacrylamide) (PNIPAAm)/Fe3O4-NH2 microgels with the encapsulated anti-cancer drug curcumin (Cur) were designed and fabricated for magnetically triggered release. PNIPAAm-based magnetic microgels with a spherical structure were produced via a temperature-induced emulsion followed with physical-crosslinking by mixing PNIPAAm, polyethylenimine (PEI), and Fe3O4-NH2 magnetic nanoparticles. Because of their dispersity, the Fe3O4-NH2 nanoparticles were embedded inside the polymer matrix. The amine groups exposed on the Fe3O4-NH2 and PEI surface supported the spherical structure by physically crosslinking with the amide groups of the PNIPAAm. The hydrophobic anti-cancer drug curcumin can be dispersed in water after encapsulation into the microgels. The microgels were characterized by transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), and UV-Vis spectral analysis. Furthermore, magnetically triggered release was studied under an external high frequency magnetic field (HFMF). A significant &#34;burst release&#34; of curcumin was observed after applying the HFMF to the microgels due to the magnetic inductive heating (hyperthermia) effect. This manuscript describes the magnetically triggered controlled release of Cur-PNIPAAm/Fe3O4-NH2 encapsulated curcumin, which can be potentially applied for tumor therapy.

Magnetic and Thermal-sensitive PolyN-isopropylacrylamide-based Microgels for Magnetically Triggered Controlled Release

This manuscript describes the preparation of magnetic and thermal-sensitive microgels via a temperature-induced emulsion without chemical reaction. These sensitive microgels were synthesized by mixing poly(N-isopropylacrylamide) (PNIPAAm), polyethylenimine (PEI) and Fe3O4-NH2 nanoparticles for the potential use in magnetically and thermally triggered drug-release.

Poliuretano Magnético y Térmico<emN</em&gt; -isopropilacrilamida) para liberación controlada magnéticamente

Magnetically and thermally sensitive poly(N-isopropylacrylamide) (PNIPAAm)/Fe3O4-NH2 microgels with the encapsulated anti-cancer drug curcumin (Cur) were ...

Repressing Gene Transcription by Redirecting Cellular Machinery with Chemical Epigenetic Modifiers

Regulation of chromatin compaction is an important process that governs gene expression in higher eukaryotes. Although chromatin compaction and gene expression regulation are commonly disrupted in many diseases, a locus-specific, endogenous, and reversible method to study and control these mechanisms of action has been lacking. To address this issue, we have developed and characterized novel gene-regulating bifunctional molecules. One component of the bifunctional molecule binds to a DNA-protein anchor so that it will be recruited to an allele-specific locus. The other component engages endogenous cellular chromatin-modifying machinery, recruiting these proteins to a gene of interest. These small molecules, called chemical epigenetic modifiers (CEMs), are capable of controlling gene expression and the chromatin environment in a dose-dependent and reversible manner. Here, we detail a CEM approach and its application to decrease gene expression and histone tail acetylation at a Green Fluorescent Protein (GFP) reporter located at the Oct4 locus in mouse embryonic stem cells (mESCs). We characterize the lead CEM (CEM23) using fluorescent microscopy, flow cytometry, and chromatin immunoprecipitation (ChIP), followed by a quantitative polymerase chain reaction (qPCR). While the power of this system is demonstrated at the Oct4 locus, conceptually, the CEM technology is modular and can be applied in other cell types and at other genomic loci.

Regulation of the chromatin environment is an essential process required for proper gene expression. Here, we describe a method for controlling gene expression through the recruitment of chromatin-modifying machinery in a gene-specific and reversible manner.

Reprimiendo la transcripción genética redirigiendo la maquinaria celular con modificadores químicos de epigenéticas

Regulation of chromatin compaction is an important process that governs gene expression in higher eukaryotes. Although chromatin compaction and gene ...

Use of High-Throughput Automated Microbioreactor System for Production of Model IgG1 in CHO Cells

Automated microscale bioreactors (15 mL) can be a useful tool for cell culture engineers. They facilitate the simultaneous execution of a wide variety of experimental conditions while minimizing potential process variability. Applications of this approach include: clone screening, temperature and pH shifts,&#160;media and supplement optimization. Furthermore, the small reactor volumes are conducive to large Design of Experiments that investigate a wide range of conditions. This allows upstream processes to be significantly optimized before scale-up where experimentation is more limited in scope due to time and economic constraints. Automated microscale bioreactor systems offer various advantages over traditional small scale cell culture units, such as shake flasks or spinner flasks. However, during pilot scale process development&#160;significant care must be taken to ensure that these advantages are realized. When run with care, the system can enable high level automation, can be programmed to run DOE&#39;s with a higher number of variables&#160;and can reduce sampling time when integrated with a nutrient analyzer or cell counter. Integration of the expert-derived heuristics presented here, with current automated microscale bioreactor experiments can minimize common pitfalls that hinder meaningful results. In the extreme, failure to adhere to the principles laid out here can lead to equipment damage that requires expensive repairs. Furthermore, the microbioreactor systems have small culture volumes&#160;making characterization of cell culture conditions difficult. The number and amount of samples taken in-process in batch mode culture is limited as operating volumes cannot fall below 10 mL. This method will discuss the benefits and drawbacks of microscale bioreactor systems.

A detailed protocol for the concurrent operation of 48 parallel cell cultures under varied conditions in a microbioreactor system is presented. Cell culture process, harvest and subsequent antibody titer analysis are described.

Microbioreactor uso de alto rendimiento automático para la producción de IgG1 modelo en células CHO

Automated microscale bioreactors (15 mL) can be a useful tool for cell culture engineers. They facilitate the simultaneous execution of a wide variety of ...

Wet-spinning-based Molding Process of Gelatin for Tissue Regeneration

This article presents an inexpensive method to fabricate gelatin, as a natural polymer, into monofilament fibers or other appropriate forms. Through the wet spinning method, gelatin fibers are produced by smooth extrusion in a suitable coagulation medium. To increase the functional surface of these gelatin fibers and their ability to mimic the features of tissues, gelatin can be molded into a tube form by referring to this concept. Examined by in vitro and in vivo tests, the gelatin tubes demonstrate a great potential for application in tissue engineering. Acting as a suitable filling gap material, gelatin tubes can be used to substitute the tissue in the damaged area (e.g., in the nervous or cardiovascular system), as well as to promote regeneration by providing a direct replacement of stem cells and neural circuitry. This protocol provides a detailed procedure for creating a biomaterial based on a natural polymer, and its implementation is expected to greatly benefit the development of correlative natural polymers, which help to realize tissue regeneration strategies.

We developed and describe a protocol based on the wet spinning concept, for the construction of gelatin-based biomaterials used for the application of tissue engineering.

Proceso de moldeo basada en mojado-spinning de gelatina para la regeneración de los tejidos

This article presents an inexpensive method to fabricate gelatin, as a natural polymer, into monofilament fibers or other appropriate forms. Through the ...

Primary Clarification of CHO Harvested Cell Culture Fluid using an Acoustic Separator

Primary clarification is an essential step in a biomanufacturing process for the initial removal of cells from therapeutic products within the harvested cell culture fluid. While traditional methods like centrifugation or filtration are widely implemented for cell removal, the equipment for these processes have large footprints and operation can involve contamination risks and filter fouling. Additionally, traditional methods may not be ideal for continuous bioprocessing schemes for primary clarification. Thus, an alternate application using acoustic (sound) waves was investigated to continuously separate cells from the cell culture fluid. Presented in this study is a detailed protocol for using a bench-scale acoustic wave separator (AWS) for the primary separation of culture fluid containing a monoclonal IgG1 antibody from a CHO cell bioreactor harvest. Representative data are presented from the AWS and demonstrate how to achieve effective cell clarification and product recovery. Finally, potential applications for AWS in continuous bioprocessing are discussed. Overall, this study provides a practical and general protocol for the implementation of AWS in primary clarification for CHO cell cultures and further describes its application potential in continuous bioprocessing.

Presented here is a protocol for the primary clarification of CHO cell culture using an acoustic separator. This protocol can be used for the primary clarification of shake flask cultures or bioreactor harvests and has the potential application for continuous clarification of the cell bleed material during perfusion bioreactor operations.

Clarificación primaria del fluido de cultivo celular cosechado con CHO utilizando un separador acústico

Primary clarification is an essential step in a biomanufacturing process for the initial removal of cells from therapeutic products within the harvested ...

Alternando microgeles de gelatina de campo híbrido-Responsive magnéticos para Controlled Drug Release

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