Questo studio è stato sostenuto da Innovation Award Farris famiglia e NIH 1R01NR015674-01 di MK. Gli autori ringraziano Josep Nayfach (Qteris, Inc) per la fornitura di un sistema generatore elettromagnetico così come la sua consulenza tecnica. Gli autori ringraziano anche Huan Yan (LCI &amp; Chemical programma interdisciplinare Fisica, Kent State University) per i suoi assistenti tecnici.

Nota: Il processo generale di fabbricazione magnetici microgel gelatina campo-reattivo è illustrato nella Figura 1A. 1. Preparazione soluzioni e sospensioni <ol><li> Preparare una genipina reticolante (1% w / v) soluzione sciogliendo 20 mg di genipina in 2 ml di tampone fosfato salino (PBS 1x; pH 7,4). Vortex la soluzione e mettere in un 50 ° C bagnomaria per 2 ore per sciogliere completamente la soluzione. </li><li> Preparare una soluzione di tensioattivo sciogliendo 20 mg di poli (glicole etilenico) -poly (glicole propilenico) -poly (glicole etilenico) (M w = 2.900 Da, denominato L64) in 200 ml di PBS per essere alla concentrazione di 100 ppm. </li><li> Preparare un 15% (w / v) soluzione di gelatina sciogliendo 64,5 mg di gelatina in 0,43 ml di PBS. Vortex la soluzione e posizionarlo su bagnomaria a 37 ° C fino a raggiungere una fase sol, dove la soluzione diventa fluidico. Poi, vortice la soluzione di gelatina 2 - 3 volte a garantire esimoe l&#39;omogeneità del campione. </li><li> Preparazione di p (NIPAM- co -AAm) / soluzione MNPs con un modello di droga (BSA): <ol><li> Disperse 10,75 mg di idrofila MNPs in 0,43 ml di PBS e quindi sciogliere 12,9 mg di p (NIPAM- co -AAm) nella sospensione MNP per rendere la concentrazione del 3% (w / v). L&#39;aumento della concentrazione di p (NIPAM- co -AAm) può essere utilizzato per ottenere un aumento comportamento deswelling di microgel. </li><li> Utilizzare Texas-Red coniugato albumina sierica bovina (TR-BSA; M w ~ 66 kDa) come modello di droga. Sciogliere 0,5 mg di TR-BSA nella miscela di p (NIPAM- co -AAm) / MNPs. </li></ol></li><li> Preparare miscele di gelatina / p (NIPAM- co -AAm) / / soluzione BSA MNPs (0,86 ml) aggiungendo miscela di p (NIPAM- co -AAm) / MNPs (0,43 ml) nella soluzione di gelatina (0,43 ml) e poi accuratamente vortice loro di fare una miscela omogenea. Così, le concentrazioni di polimeri e MNP diventano metà delle concentrazioni iniziali della miscela finale. </li></ol><ol><li> Versare 15 ml di olio di silicone [polidimetilsilossano (viscosità 350 cSt)] in un becher pulito e sterile. </li><li> Aggiungere immediatamente le miscele acquose pre-preparati di gelatina / p (NIPAM- co -AAm) / / soluzione BSA MNPs (0,86 ml) nel olio di silicone ed emulsionare la miscela acquosa nella fase oleosa mescolando con una barra di agitazione magnetica a 900 rpm a 30 ° C per 30 min. </li></ol> 3. gelificazione e trasferimento di micro-gocce di una soluzione acquosa <ol><li> Trasferire l&#39;emulsione (~ 16 ml) dal bicchiere in una provetta da 50 ml. </li><li> Raffreddare la provetta per 10 min a 4 ° C per gelificazione dei micro-goccioline di olio. </li><li> Riempire il tubo con la soluzione preparata L64 (a 4 ° C) fino a 50 ml e agitare vigorosamente la provetta. Può essere possibile che una porzione di L64 tensioattivi sarebbe nei microgel. </li><li> Centrifugare la provetta per 20 min a 2300 xga 4 ° C. </li><li> Regularly controllare la presenza del pellet di particelle di gel sul lato del tubo. Se le particelle non si vedono, centrifugare per altri 20 minuti alla stessa velocità e la temperatura. Avanti a rimuovere con attenzione il surnatante senza disturbare il pellet formata sulla parete interna del tubo. </li><li> Ripetere i punti (3,3) a (3,5) ancora una volta. Ogni volta, trasferire il campione in una nuova provetta per evitare l&#39;inclusione di eventuali goccioline di olio in sospensione Microgel. Dopo questo passo, assicurano che i tensioattivi o goccioline di olio non sono presenti nella sospensione del campione. Tuttavia, le fasi di separazione ripetute possono portare alla perdita di materiali iniziali. </li></ol> 4. covalente reticolazione del microgel <ol><li> Aggiungere 2 ml di soluzione genipina (preparata nella sezione 1) al pellet di particelle di gel e mescolare bene nel vortex la soluzione. </li><li> Trasferire rapidamente il tubo della sospensione a bagnomaria a 23 ° C per iniziare una reazione di reticolazione covalente durante una des(es., 5 - 120 min) tempo di reticolazione ired. </li><li> Dopo la reticolazione, rimuovere immediatamente eventuali reticolanti eccessivi scartando la soluzione genipina, risospendere le microgel in PBS, e centrifugazione del tubo per 20 minuti a 2300 xg (4 ° C). Se necessario, cautamente si rompono formata pellet con un puntale. Questa fase di lavaggio può essere ripetuto fino a 3 volte se il genipina è ancora presente nella soluzione. </li><li> Eliminare il supernatante e risospendere le microgel in PBS ad una densità desiderata (ad es., 5 × 10 6 microgel / ml) contando il numero con un emocitometro. </li><li> Per osservazioni microscopiche, caricare la sospensione Microgel nello spazio tra un vetrino e coprioggetto e sigillare il confine del vetrino con resina epossidica. </li></ol> 5. Applicazione di campo magnetico alternato per innescare Drug uscita <ol><li> Posizionare il tubo con concentrazione desiderata di microgel in mezzi acquosi inla camera di bobine magnetiche. Se necessario, inserire una sonda di temperatura in fibra ottica nel tubo per monitorare il cambiamento della temperatura del supporto durante l&#39;applicazione del AMF. </li><li> Applicare alta frequenza (&gt; 100 kHz) AMF ad una intensità di campo definito (&gt; 5 kA / m) e per una durata specificata. A seguito dell&#39;applicazione del AMF, centrifugare la provetta per 20 minuti a 2.273 xg (4 ° C) e raccogliere il surnatante per quantificare la quantità di TR-BSA rilasciato dalla microgel ai media circostanti con spettrofotometria. Le lunghezze d&#39;onda di eccitazione ed emissione per Texas Red sono 584 nm e 612 nm, rispettivamente. </li></ol>

<ol>
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The authors have nothing to disclose.

La tecnologia qui descritta illustra un prototipo sull&#39;uso di ibridi nanoparticelle Microgel per magneto-termico innescata rilascio del farmaco. Questo è stato ottenuto intrappolando fisicamente catene MNPs e P (NIPAM- co -AAm) all&#39;interno di una rete di microscala gelatina tridimensionale reticolato per genipina. La piattaforma di campo-responsive magnetico era sufficiente a generare calore all&#39;interno del Microgel in risposta ad una AMF applicata remoto, che a sua volta innescato il rilascio di u...

Gli stimoli-reattivi sistemi di drug delivery che consentono una consegna di droga strettamente controllato in risposta a stimoli sia endogeni o esogeni (ad es., Temperatura o pH) sono stati ampiamente studiati come nuovi tipi di dispositivi intelligenti morbide per la consegna della droga. Idrogel MicroScale sono stati ampiamente utilizzati come piattaforma di consegna della droga, nel senso che conferiscono profili di rilascio di droga controllabili e sostenibili, nonché chimiche sintonizzabile e le proprietà meccaniche 1-3. In particolare, i microgel colloidali presentano molti vantaggi come veicolo di somministrazione di farmaci per la loro rapida risposta a stimoli esterni e adatto iniettabilità al tessuto locale in una maniera minimamente invasiva 4. Il poli (N-isopropilacrilammide) (pNIPAM) o suoi copolimeri sono state ampiamente adottate nella sintesi microgel termo-responsive mediante innesto pNIPAM con / polimeri biocompatibili biodegradabili compresi gelatina, chitosano, acido alginato, o acido ialuronico 5,6, In cui una caratteristica transizione di fase pNIPAM alla sua temperatura critica più bassa soluzione (LCST) può essere utilizzato come un trigger di rilascio del farmaco 7. Abbiamo recentemente dimostrato una fabbricazione di biodegradabile, Microgel termo-sensibile di gelatina a base incorporando poli (N-isopropylacrylamide- co -acrylamide) [p (NIPAM- co -AAm)] catene come un componente minore all&#39;interno di reti di gelatina tridimensionali 8. Il / p (NIPAM- co -AAm) microgel gelatina esibito una deswelling sintonizzabile all&#39;aumento di temperatura, che positivamente correlata al rilascio di albumina sierica bovina (BSA). Nel corso degli ultimi anni, ci sono stati aumentando gli sforzi per sviluppare una piattaforma di consegna della droga magneticamente sensibile che può innescare il rilascio di farmaco in un 9,10 di moda on-demand. Il principio di base per la sintesi della piattaforma di drug delivery magneticamente sensibile utilizza la caratteristica di nanoparticelle superparamagnetiche (MNPs) per generare calore quando ricevono una alta frequenza campo magnetico alternato (AMF), che innesca un rilascio del farmaco sensibile alla temperatura. Questo promettente per future applicazioni cliniche in che questo sistema può indirizzare in profondità nel tessuto, permette un non-invasivo e telecomandata rilascio del farmaco e può essere combinato con il trattamento di ipertermia e risonanza magnetica sistema di imaging 10-12. Tali piattaforme includono: (1) le particelle MNPs / pNIPAM ibrido MicroGEL 13-15 e (2) ponteggi idrogel macroscopici che incorporano immobilizzato MNPs 16-18. Le piattaforme MicroGel pNIPAM basate dimostrato una transizione risposta finemente accordabile fase del volume agli stimoli magnetotermici. Tuttavia, ancora si basano su tecniche complesse e sofisticate nella fabbricazione e l&#39;uso di polimeri pNIPAM ad alto contenuto può essere potenzialmente citotossici per le cellule 19, che possono limitare le loro applicazioni in vivo. I ponteggi macroscopiche esibiscono un parentely lenta risposta a stimoli esterni e richiedono un trapianto chirurgico invasivo rispetto a microgel colloidali. L&#39;emulsione acqua-in-olio è stato il metodo standard per la produzione di submillimetrica o gel di dimensioni micrometriche particelle 20. All&#39;interfaccia acqua-olio dell&#39;emulsione, Microgel particelle costituisce una forma sferica dovuta alla minimizzazione dell&#39;energia di superficie della gocciolina di acqua sotto la forza di taglio meccanica. Questo metodo permette la produzione di una grande quantità di goccioline acquose gel sferiche in una semplice procedura di fabbricazione ed è stato adottato con successo per la fabbricazione microgel a base di gelatina per applicazioni drug delivery 21-23. Qui, vi presentiamo un metodo facile per sintetizzare un magnetothermally reattivo microgel a base di gelatina per l&#39;applicazione somministrazione di farmaci utilizzando il metodo di emulsione acqua in olio. Questo è stato ottenuto incorporando fisicamente MNPs ossido di ferro e p (NIPAM- co -AAm) catene come componente minore all&#39;interno di una rete di gelatina microscala sferica che è covalentemente reticolata da un genipina reticolante di derivazione naturale, in combinazione con un alta frequenza alternata magnetico sistema di applicazione di campo (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-responsive / biomateriali micro-ingegneria nano che consentono a un strettamente controllato, on-demand somministrazione di farmaci sono stati sviluppati nuovi tipi di dispositivi morbide intelligenti per applicazioni biomediche. Sebbene un certo numero di sistemi di rilascio di farmaci magneticamente rispondenti hanno dimostrato efficacies attraverso sia la prova in vitro di studi concettuali o in applicazioni preclinici in vivo, il loro uso in ambito clinico è ancora limitato dalla loro biocompatibilità e biodegradabilità insufficiente. Inoltre, molte delle piattaforme esistenti si basano su tecniche sofisticate per loro invenzioni. Abbiamo recentemente dimostrato la fabbricazione di biodegradabili, microgel termo-reattivo a base di gelatina intrappolando fisicamente poli (N-isopropylacrylamide- co -acrylamide) catene come componente minore all&#39;interno di una rete di gelatina tridimensionale. In questo studio, presentiamo un metodo facile per fabbricare una piattaforma di rilascio del farmaco biodegradabile che consente a un magneto-thermally innescato rilascio del farmaco. Ciò è stato ottenuto incorporando nanoparticelle di ossido di ferro superparamagnetico e polimeri termo-reattivo in microgel colloidali a base di gelatina, in combinazione con un sistema di applicazione di campo magnetico alternato.

Quando il protocollo è eseguita correttamente, i microgel fabbricati devono mostrare una morfologia sferica ben caratterizzato e disperdibilità colloidale con diametri nel campo tra 5 micron a 20 micron (Figura 1B e C). Sia MNPs fluorescenti o BSA fluorescente può essere utilizzata per confermare se MNPs o droghe (BSA in questo studio) sono correttamente incapsulati all&#39;interno del Microgel (Figura 1D). I microgel prefabbricati possono essere stab...

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 microgel ibrida Gelatina di campo magnetico-Responsive per Controlled Drug uscita

Vi presentiamo un metodo facile per fabbricare una piattaforma di rilascio del farmaco a base di gelatina biodegradabili che è magneto-termico sensibile. Ciò è stato ottenuto incorporando nanoparticelle di ossido di ferro superparamagnetiche e poli (N-isopropylacrylamide- co -acrylamide) all&#39;interno di una gelatina sferica micro-rete reticolato da genipina, in combinazione con un sistema di applicazione di campo magnetico alternato.

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 Views.  Kent State University.  Vi presentiamo un metodo facile per fabbricare una piattaforma di rilascio del farmaco a base di gelatina biodegradabili che è magneto-termico sensibile. Ciò è stato ottenuto incorporando nanoparticelle di ossido di ferro superparamagnetiche e poli (N-isopropylacrylamide- co -acrylamide) all'interno di una gelatina sferica micro-rete reticolato da genipina, in combinazione con un sistema di applicazione di campo magnetico alternato. 

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.

Magneticamente-Assisted Remote Controlled Suggerimento Flessione Microcatetere in Risonanza Magnetica

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

Ricerca

Bioingegneria

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.

Hybrid Imaging μCT-FMT e analisi delle immagini

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.

Appositamente studiata apparecchi di riscaldamento a base laser per la Release Innescata di Cisplatino da termosensibile liposomi con Risonanza Magnetica Guidance Immagine

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.

Ponteggi composto di fibre di interfacce polielettrolita per rilascio controllato di Biomolecole temporalmente

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 sensibile magnetico e termico (<em&gt; N</em&gt; -isopropilacrilammide) per il rilascio controllato magneticamente attivato

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.

Reprimere la trascrizione genica reindirizzando macchinario cellulare con modificatori epigenetici chimici

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 di High-Throughput Microbioreactor sistema automatizzato per la produzione del modello IgG1 in cellule 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 di stampaggio basati su bagnato-filatura di gelatina per la rigenerazione tissutale

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.

Chiarificazione primaria del fluido di coltura cellulare raccolto da CHO utilizzando un separatore acustico

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

Alternando microgel ibrida Gelatina di campo magnetico-Responsive per Controlled Drug uscita

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