Este estudo foi apoiado pelo Prêmio Inovação Farris Família e NIH 1R01NR015674-01 para MK. Os autores agradecem Josep Nayfach (Qteris, Inc) para fornecer um sistema gerador de electro-magnética, bem como a sua consulta técnica. Os autores também agradecem Huan Yan (LCI &amp; Chemical Programa Interdisciplinar de Física, Kent State University) por seus assistentes técnicos.

Nota: O processo global de fabrico de microgéis de gelatina de campo de resposta magnética é ilustrado na Figura 1A. 1. Preparar as soluções e suspensões <ol><li> Prepara-se uma genipina reticulador (1% v / v w) solução por dissolução de 20 mg de genipina em 2 ml de solução salina tamponada com fosfato (PBS 1x; pH 7,4). Vortex a solução e colocar num banho de água a 50 ° C durante 2 h para dissolver completamente a solução. </li><li> Prepara-se uma solução de agente tensioactivo por dissolução de 20 mg de poli (etilenoglicol) -poli (propileno glicol) -poli (etileno glicol) (M w = 2900 Da; referido como L64) em 200 ml de PBS para estar na concentração de 100 ppm. </li><li> Prepara-se uma a 15% (v / W) solução de gelatina dissolvendo 64,5 mg de gelatina em 0,43 ml de PBS. Vortex a solução e colocá-lo num banho de água a 37 ° C até se atingir uma fase de sol, em que a solução se torna fluídica. Em seguida, vortex a solução de gelatina a 2 - 3 vezes para garantir the homogeneidade da amostra. </li><li> Preparação de p (co NIPAM- -AAm) de solução / MNPs com um fármaco modelo (BSA): <ol><li> Dispersa-se 10,75 mg de MNPs hidrófilo em 0,43 ml de PBS e, em seguida, dissolver 12,9 mg de p (co NIPAM- -AAm) na suspensão MNP para tornar a concentração a 3% (w / v). O aumento da concentração de p (co NIPAM- -AAm) pode ser utilizado para atingir um aumento de comportamento deswelling de microgéis. </li><li> Uso de albumina de soro de Texas-Red conjugado bovino (BSA-TR; H w ~ 66 kDa) como uma droga modelo. Dissolve-se 0,5 mg de BSA em TR-se a mistura de p (co NIPAM- -AAm) / MNPs. </li></ol></li><li> Preparar misturas de gelatina / p (co NIPAM- -AAm) / MNPs / solução de BSA (0,86 ml) por adição de mistura de p (co NIPAM- -AAm) / MNPs (0,43 ml) para a solução de gelatina (0,43 ml) e em seguida Agitar bem em vórtice-los para fazer uma mistura homogênea. Assim, as concentrações de polímeros e MNP tornar-se metade das concentrações iniciais na mistura final. </li></ol><ol><li> Despeje 15 ml de óleo de silicone [polidimetilsiloxano (viscosidade 350 cSt)] para um copo limpo e estéril. </li><li> Imediatamente adicionar as misturas aquosas de pré-preparadas de gelatina / p (co NIPAM- -AAm) / MNPs / solução de BSA (0,86 ml) em óleo de silicone a emulsionar e a mistura aquosa na fase de óleo por agitação com uma barra de agitação magnética a 900 rpm a 30 ° C durante 30 min. </li></ol> 3. A gelificação e Transferência de micro-gotas a uma solução aquosa <ol><li> Transferir a emulsão (~ 16 ml) a partir de proveta para um tubo de 50 ml. </li><li> Arrefecer o tubo durante 10 minutos a 4 ° C durante a gelificação das micro-gotículas no óleo. </li><li> Encha o tubo com a solução L64 preparado (a 4 ° C) até 50 ml e agite o tubo. Pode ser possível que uma porção de L64 tensioactivos estaria dentro dos microgéis. </li><li> Centrifugar o tubo durante 20 minutos a 2.300 xg a 4 o C. </li><li> RegularlY verificar a presença do sedimento de partículas de gel no lado do tubo. Se as partículas não são vistas, centrifuga-se durante mais 20 min à mesma temperatura e velocidade. Avance para remover cuidadosamente o sobrenadante sem perturbar o sedimento formado na parede interior do tubo. </li><li> Repita os passos (3.3) a (3.5) mais uma vez. Cada vez, transferir a amostra para um novo tubo para evitar a inclusão de quaisquer gotículas de óleo na suspensão de microgel. Após este passo, garantir que as gotículas de óleo ou agentes tensioactivos não estão presentes na suspensão de amostra. No entanto, os passos de separação repetidas podem conduzir a perda de materiais iniciais. </li></ol> 4. covalente reticulação dos microgéis <ol><li> Adicionar 2 ml de solução de genipina (preparada na secção 1) para o sedimento de partículas de gel e mistura-los bem com vortex a solução. </li><li> Transferir rapidamente o tubo de a suspensão em banho de água a 23 ° C para iniciar uma reacção de reticulação covalente durante um des(. por exemplo, 5-120 min) Tempo de reticulação Ired. </li><li> Após a reticulação, remover imediatamente qualquer reticuladores excessivas descartando a solução genipina, ressuspendendo os microgéis em PBS e centrifugando o tubo durante 20 minutos a 2300 xg (4 ° C). Se necessário, cautelosamente quebrar pellet formado com uma ponta de pipeta. Este passo de lavagem pode ser repetido até 3 vezes, se a genipina ainda permanece na solução. </li><li> Descartar o sobrenadante e ressuspender as microgéis em PBS a uma densidade desejada (por ex., 5 x 10 6 microgéis / ml) através da contagem do número com um hemocitómetro. </li><li> Para as observações microscópicas, carregar a suspensão microgel no espaço entre uma lâmina de vidro e uma tampa de deslizamento e selar o limite da cobertura de vidro com resina epóxi. </li></ol> 5. A aplicação de campo magnético alternado para desencadear a libertação do fármaco <ol><li> Colocar o tubo com a concentração desejada de microgéis em meios aquosos ema câmara de bobinas magnéticas. Se necessário, inserir uma sonda de temperatura de fibra óptica para dentro do tubo para monitorar a mudança de temperatura dos meios de comunicação durante a aplicação da AMF. </li><li> Aplicar alta frequência (&gt; 100 kHz) AMF a uma intensidade de campo definido (&gt; 5 kA / m) e por um período de tempo especificado. Na sequência da aplicação de AMF, centrifugar os tubos de amostra durante 20 min a 2273 xg (4 ° C) e recolher o sobrenadante para quantificar a quantidade de TR-BSA libertada a partir de microgel nos meios circundantes através de espectrofotometria. Os comprimentos de onda de excitação e emissão de Texas Red são 584 nm e 612 nm, respectivamente. </li></ol>

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The authors have nothing to disclose.

A tecnologia aqui descrita demonstra uma prova de conceito de utilização de híbridos nanopartícula-microgel para a libertação do fármaco magneto-desencadeada termicamente. Isto foi conseguido por aprisionamento fisicamente cadeias MnPs e P (co NIPAM- -AAm) dentro de uma rede tridimensional de gelatina microescala reticulado por genipina. A plataforma de campo de resposta magnética foi suficiente para gerar calor no interior do microgel em resposta a um AMF remotamente aplicada, que por sua vez, provocou...

Estímulos responsiva sistemas de administração de fármacos que permitem uma entrega de drogas rigorosamente controlado em resposta a estímulos endógenos ou quer exógenas (por exemplo., Temperatura ou pH) têm sido investigados extensivamente como novos tipos de dispositivos inteligentes suaves para a entrega de drogas. Hidrogeles microscópicos têm sido largamente utilizados como uma plataforma de entrega de droga na medida em que conferem perfis controlável e sustentável de libertação de fármaco, bem como sintonizável química e propriedades mecânicas 1-3. Em particular, os microgéis coloidais apresentam muitas vantagens como um veículo para a entrega de drogas devido à sua capacidade de resposta rápida a estímulos externos e injectabilidade adequado para o tecido local de forma minimamente invasiva 4. O poli (N-isopropilacrilamida) (pNIPAM) ou seus copolímeros têm sido amplamente adotados em sintetizar microgï¿½s termo-sensível por enxertia pNIPAM com polímeros biodegradáveis ​​/ biocompatíveis, incluindo gelatina, a quitosana, ácido alginato, ou ácido hialurônico 5,6, No qual uma característica de transição de fase de pNIPAM à sua temperatura crítica inferior de solução (TCIS) pode ser utilizada como um gatilho de libertação de medicamento 7. Recentemente, demonstrou uma fabricação de biodegradável microgel de resposta térmica, com base em gelatina por incorporação de poli (co-N isopropylacrylamide- -acrilamida) [P (co NIPAM- -AAm)] cadeias como um componente menor de gelatina dentro das redes tridimensionais 8. O / P (co NIPAM- -AAm) microgel de gelatina exibiu um deswelling sintonizável ao aumento de temperatura, o que positivamente correlacionada com a libertação de albumina de soro bovino (BSA). Durante os últimos anos, tem vindo a aumentar os esforços para desenvolver uma plataforma de entrega de drogas magneticamente sensível que pode desencadear a liberação de drogas em um 9,10 de moda on-demand. O princípio básico para a síntese de plataforma de entrega de droga que responde magneticamente utiliza a característica de nanopartículas superparamagnéticas (MNPs) para gerar calor quando recebem uma alta frequência de campo magnético alternado (AMF), o que desencadeia uma liberação da droga sensível à temperatura. Esta é uma promessa para futuras aplicações clínicas em que este sistema pode atingir profundamente no tecido, permite uma libertação de fármaco não-invasivo e remotamente controlada e pode ser combinada com tratamento de hipertermia e sistema de ressonância magnética 10-12. Tais plataformas incluem: (1) Partículas híbridas MNPs / pNIPAM microgel 13-15 e (2) andaimes hidrogel macroscópicas incorporando imobilizada MNPs 16-18. As plataformas de microgel baseados em pNIPAM demonstraram uma resposta de transição de fase de volume finamente sintonizável a estímulos magneto-térmico. No entanto, eles ainda dependem de técnicas complexas e sofisticadas na fabricação e a utilização de polímeros pNIPAM com um teor elevado pode ser potencialmente citotóxico para as células 19, o que pode limitar as suas aplicações in vivo. Os andaimes macroscópicas exibem um parenteLY lenta resposta a estímulos externos e exigem um transplante cirúrgico invasivo em comparação com os microgéis de coloidais. A emulsão água-em-óleo tem sido o método padrão para produzir submillimeter ou gel de partículas de tamanho micrométrico 20. Na interface óleo-água da emulsão, microgel de partícula tem uma forma esférica, devido à minimização da energia de superfície da gotícula de água, sob a força de cisalhamento mecânico. Este método permite a produção de uma grande quantidade de gotículas esféricas de gel aquoso num processo de fabrico simples e tem sido adoptada com sucesso para a fabricação de microgéis à base de gelatina, para aplicações de entrega de drogas 21-23. Aqui, nós apresentamos um método fácil para sintetizar uma microgéis magnetothermally responsivos à base de gelatina para aplicação de entrega de drogas através da utilização do método de emulsificação de água-em-óleo. Isto foi conseguido por incorporando fisicamente MNPs óxido de ferro e P (co NIPAM- -AAm) cadeias como um componente menor dentro de uma rede de gelatina microescala esférica que está covalentemente reticulados por um agente reticulante genipina de origem natural, em conjunto com uma elevada frequência de alternância de um sistema de aplicação do campo magnético (AMF).

<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

Magneticamente reactivo nano / micro-biomateriais engenharia que permitem uma entrega de drogas rigorosamente controlada, a pedido têm sido desenvolvidos como novos tipos de dispositivos moles inteligentes para aplicações biomédicas. Embora um certo número de sistemas de entrega de drogas magneticamente reactivo demonstraram eficácias, quer através de provas in vitro de estudos de concepção ou em aplicações de pré-clínicos in vivo, a sua utilização na prática clínica é ainda limitada pela sua biocompatibilidade ou a biodegradabilidade insuficiente. Além disso, muitas das plataformas existentes dependem de técnicas sofisticadas para sua fabricação. Recentemente, demonstrou a fabricação de biodegradável microgel de resposta térmica, com base em gelatina por aprisionamento fisicamente poli (co-acrilamida N isopropylacrylamide-) cadeias como um componente menor dentro de uma rede tridimensional de gelatina. Neste estudo, são apresentados um método fácil de fabricar uma plataforma de libertação de fármaco biodegradáveis ​​que permite uma magneto-thermally desencadeada a libertação do fármaco. Isto foi conseguido através da incorporação de nanopartículas de óxido de ferro superparamagnético e polímeros de resposta térmica dentro de microgéis coloidais à base de gelatina, em conjunto com um sistema de aplicação do campo magnético alternado.

Quando o protocolo é realizado correctamente, os microgéis fabricadas deverão exibir uma morfologia esférica bem caracterizada e dispersibilidade coloidal com diâmetros na gama entre 5 um a 20 um (Figura 1B e C). Qualquer um MNPs fluorescentes ou BSA fluorescente pode ser utilizado para confirmar se MNPs ou drogas (BSA neste estudo) são adequadamente encapsulado dentro do microgel (Figura 1D). Os microgéis fabricadas podem ser estáveis ​​e ar...

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 microgéis de gelatina híbrido Campo-Responsive magnética para controlada de fármacos de libertação

Nós apresentamos um método fácil de fabricar uma plataforma de liberação do fármaco à base de gelatina biodegradável que é magneto-termicamente responsivo. Isto foi conseguido através da incorporação de nanopartículas de óxido de ferro superparamagnético e poli (N-co isopropylacrylamide- -acrilamida) dentro de uma rede de micro-reticulada de gelatina esférica por genipina, em conjunto com um sistema de aplicação do campo magnético alternado.

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 visualizações.  Kent State University.  Nós apresentamos um método fácil de fabricar uma plataforma de liberação do fármaco à base de gelatina biodegradável que é magneto-termicamente responsivo. Isto foi conseguido através da incorporação de nanopartículas de óxido de ferro superparamagnético e poli (N-co isopropylacrylamide- -acrilamida) dentro de uma rede de micro-reticulada de gelatina esférica por genipina, em conjunto com um sistema de aplicação do campo magnético alternado. 

Vídeo: 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.

Magneticamente Assistida deflexão da ponta Remote Controlled microcateter em Imagem por Ressonância Magnética

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

Bioengenharia

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.

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 imagem μCT-FMT e análise de imagem

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.

Customizado aparelho de aquecimento à base de laser para lançamento acionado de cisplatina de termossensível lipossomas com Ressonância Magnética Orientação Imagem

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.

Andaimes de mistura de fibras Interfacial polieletrólito para lançamento 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.

Poli (Magnetic and Thermal-sensitive Poly)<em&gt; N</em&gt; Microgels com base em isopropilacrilamida) para liberação controlada com disparo magnético

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.

Reprimir a transcrição do Gene redirecionando maquinaria celular com modificadores epigenéticas químicas

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.

Uso do elevado-Throughput Microbioreactor sistema automatizado para produção de IgG1 modelo em 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.

Processo de moldagem baseados em molhado-fiação de gelatina para regeneração de tecidos

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.

Esclarecimento Primário do Fluido de Cultura Celular Colhida cho usando um 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 microgéis de gelatina híbrido Campo-Responsive magnética para controlada de fármacos de libertação

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