Improved survival of anchorage-dependent cells in core-shell hydrogel microcapsules via co-encapsulation with cell-friendly microspheres.

In this study, we investigated the effect of intracapsular environment on the survival of anchorage-dependent cells (ADCs) encapsulated in alginate microcapsules with three different core structures, i.e. liquid, semi-liquid and microsphere-encapsulating semi-liquid core, using NIH 3T3 fibroblasts as an ADC model. For the latter, we fabricated poly (ɛ-caprolactone) microspheres and co-encapsulated them with the cells, to establish cell-substrate interactions in the capsule. The fibroblast cells co-encapsulated with the microspheres exhibited higher survival and growth than those without. This study provides a "proof of concept" for employing microspheres as a cell-friendly surface to establish intracapsular cell-substrate interactions thus prolonging the survival of encapsulated therapeutic ADCs.

Sustained exenatide delivery via intracapsular microspheres for improved survival and function of microencapsulated porcine islets.

The ability of glucagon-like peptide-1 analogs to enhance glucose-dependent insulin secretion and to inhibit ß cell apoptosis could be of potential benefit for islet transplantation. In this study, we investigated the effect of sustained local delivery of exenatide, a synthetic exendin-4, on the in vitro viability and function of encapsulated porcine islets. Prior to encapsulation, we fabricated exenatide-loaded poly(latic-co-glycolic acid) microspheres, and investigated their release behavior with different initial drug-loading amounts. Exenatide-loaded microspheres, exhibiting a sustained release over 21 days, were subsequently chosen and co-encapsulated with porcine islets in alginate microcapsules. During the 21-day period, the islets co-encapsulated with the exenatide-loaded microspheres exhibited improved survival and glucose-stimulated insulin secretion, compared to those without. This suggested that the intracapsular sustained delivery of exenatide via microspheres could be a promising strategy for improving survival and function of microencapsulated porcine islets for islet xenotransplantation.

Macromolecule release from monodisperse PLG microspheres: control of release rates and investigation of release mechanism.

Novel macromolecular therapeutics such as peptides, proteins, and DNA are advancing rapidly toward the clinic. Because of typically low oral bioavailability, macromolecule delivery requires invasive methods such as frequently repeated injections. Parenteral depots including biodegradable polymer microspheres offer the possibility of reduced dosing frequency but are limited by the inability to adequately control delivery rates. To control release and investigate release mechanisms, we have encapsulated model macromolecules in monodisperse poly(D,L-lactide-co-glycolide) (PLG) microspheres using a double-emulsion method in combination with the precision particle fabrication technique. We encapsulated fluorescein-dextran (F-Dex) and sulforhodamine B-labeled bovine serum albumin (R-BSA) into PLG microspheres of three different sizes: 31, 44, and 80 microm and 34, 47, and 85 microm diameter for F-Dex and R-BSA, respectively. The in vitro release profiles of both compounds showed negligible initial burst. During degradation and release, the microspheres hollowed and swelled at critical time points dependant upon microsphere size. The rate of these events increased with microsphere size resulting in the largest microspheres exhibiting the fastest overall release rate. Monodisperse microspheres may represent a new delivery system for therapeutic proteins and DNA and provide enhanced control of delivery rates using simple injectable depot formulations.

Uniform biodegradable hydrogel microspheres fabricated by a surfactant-free electric-field-assisted method.

Uniform biodegradable hydrogel microspheres (HMS) with precisely controlled size have been fabricated using an electric-field-assisted precision particle fabrication technique. Particle agglomeration was prevented by charging the hydrogel drops and allowing Coulomb repulsion to separate them. As a result, surfactant-free and non-toxic particle fabrication was possible and the resulting microspheres were most suitable for biomedical and food-related applications. Due to the size uniformity, the present HMS may serve as a convenient yet most accurate vehicle for controlled delivery of therapeutic agents and other active ingredients.

Small-molecule release from poly(D,L-lactide)/poly(D,L-lactide-co-glycolide) composite microparticles.

Addition of biodegradable polymer shells surrounding polymeric, drug-loaded microparticles offers the opportunity to control drug release rates. A novel fabrication method was used to produce microparticles with precise control of particle diameter and the thickness of the polymer shell. The effect of shell thickness on release of a model drug, piroxicam, has been clearly shown for 2- to 15-microm thick shells of poly(D,L-lactide) (PDLL) surrounding a poly(D,L-lactide-co-glycolide) (PLG) core and compared to pure PLG microspheres loaded with piroxicam. Furthermore, the core-shell microparticles are compared to microspheres containing blended polymers in the same mass ratios to demonstrate the importance of the core-shell morphology. Combining PDLL(PLG) microcapsules of different shell thicknesses allows nearly constant release rates to be attained for a period of 6 weeks.

In vitro degradation of polyanhydride/polyester core-shell double-wall microspheres.

Double-wall microspheres (DWMS), comprising distinct polymer core and shell phases, are useful and interesting for controlled-release drug delivery. In particular, the presence of a surface-eroding polymer core may be expected to limit water penetration and, therefore, delay degradation of the core phase and drug release. In this study, solid microspheres and DWMS were fabricated using a surface-eroding polymer (poly[1,6-bis(p-carboxyphenoxy)hexane]; PCPH) and a bulk-eroding polymer (poly(D,L-lactide-co-glycolide); PLG). Erosion of the particles was observed by optical and electron microscopy, while polymer degradation was followed by gel permeation chromatography, during incubation in buffer at 37 degrees C. Degradation and erosion were very different depending on which polymer formed the particle shell. Nevertheless, the relatively thin (approximately 5 microm) PCPH shells could not prevent water penetration, and the PLG cores completely eroded by 6 weeks of incubation.

Controlling surface nano-structure using flow-limited field-injection electrostatic spraying (FFESS) of poly(D,L-lactide-co-glycolide).

Improved control of surface micro- and nano-structure may lead to enhanced performance of degradable biomedical devices such as surgical dressings, vascular grafts, tissue engineering scaffolds, sutures, and structures for guided tissue regeneration. An electrohydrodynamic method called flow-limited field-injection electrostatic spraying (FFESS) has been developed as an improved technique for the controlled deposition of polymeric material. Injecting charge using a nano-sharpened tungsten needle in a process called field ionization can efficiently induce an ionic state in a solution of poly(D,L-lactide-co-glycolide) increasing its capacity to carry charge. As a result, sprays have been produced that are finer and more precisely controlled than sprays produced by conventional electrospraying techniques, which employ hypodermic needles as the spray nozzle. Here, the effect of FFESS variables including applied voltage, polymer solution flow rate, and solvent properties (surface tension, viscosity, vapor pressure) on spray performance have been qualitatively evaluated. Under certain conditions, increasing the applied voltage produced an increasingly rough surface morphology. Similarly, by reducing solvent surface tension and increasing solvent vapor pressure, more distinct surface structures could be formed including uniform nanoparticles. Working ranges of the important parameters for the production of specific structure types such as smooth or porous surfaces, non-woven or melded fibers, and distinct or melded nanoparticles have been defined. FFESS technology provides a simple yet powerful technique for fabricating biomedical devices with a precisely defined nano-structure potentially capable of utilizing a broad range of biocompatible polymeric materials.

Microsphere size, precipitation kinetics and drug distribution control drug release from biodegradable polyanhydride microspheres.

A thorough understanding of the factors affecting drug release mechanisms from surface-erodible polymer devices is critical to the design of optimal delivery systems. Poly(sebacic anhydride) (PSA) microspheres were loaded with three model drug compounds (rhodamine B, p-nitroaniline and piroxicam) with a range of polarities (water solubilities). The drug release profiles from monodisperse particles of three different sizes were compared to release from polydisperse microspheres. Each of the model drugs exhibited different release mechanisms. Drug distribution within the polymer was investigated by laser scanning confocal microscopy and scanning electron microscopy. Rhodamine, the most hydrophilic compound investigated, was localized strongly toward the microsphere surface, while the much more hydrophobic compound, piroxicam, distributed more evenly. Furthermore, all three compounds were most uniformly distributed in the smallest microspheres, most likely due to the competing effects of drug diffusion out of the nascent polymer droplets and the precipitation of polymer upon solvent extraction, which effectively "traps" the drug in the polymer matrix. The differing drug distributions were manifested in the drug release profiles. Rhodamine was released very quickly independent of microsphere size. Thus, extended release profiles may not be obtainable if the drug strongly redistributes in the microspheres. The release of p-nitroaniline was more prolonged, but still showed little dependence on microsphere size. Hence, when water-soluble drugs are encapsulated with hydrophobic polymers, it may be difficult to tailor release profiles by controlling microsphere size. The piroxicam-loaded microspheres exhibit the most interesting release profiles, showing that release duration can be increased by decreasing microsphere size, resulting in a more uniform drug distribution.

Biodegradable gelatin microspheres enhance the neuroprotective potency of osteopontin via quick and sustained release in the post-ischemic brain.

Gelatin microspheres (GMSs) are widely used as drug carriers owing to their excellent biocompatibilities and toxicologically safe degradation products. The drug release profile is easily tailored by controlling the cross-linking density and surface-to-volume ratio, i.e. size, of the GMS. In this study, we employed GMSs which are 25 µm in diameter and cross-linked with 0.03125% glutaraldehyde, to enable rapid initial and a subsequent sustained release. Therapeutic potency of human recombinant osteopontin (rhOPN) with or without encapsulation into GMSs was investigated after administrating them to rat stroke model (Sprague-Dawley; middle cerebral artery occlusion, MCAO). The administration of rhOPN/GMS (100 ng/100 µg) at 1h post-MCAO reduced the mean infarct volume by 81.8% of that of the untreated MCAO control and extended the therapeutic window at least to 12h post-MCAO, demonstrating a markedly enhanced therapeutic potency for the use of OPN in the post-ischemic brain. Scanning electron microscopy micrographs revealed that GMSs maintained the three-dimensional shape for more than 5 days in normal brain but were degraded rapidly in the post-ischemic brain, presumably due to high levels of gelatinase induction. After encapsulation with GMS, the duration of OPN release was markedly extended; from the period of 2 days to 5 days in normal brain, and from 2 days to 4 days in the post-ischemic brain; these encompass the critical period for recovery processes, such as vascularization, and controlling inflammation. Together, these results indicate that GMS-mediated drug delivery has huge potential when it was used in the hyperacute period in the post-ischemic brain.

The effect of biodegradable gelatin microspheres on the neuroprotective effects of high mobility group box 1 A box in the postischemic brain.

High mobility group box 1 (HMGB1) is a family of endogenous molecules that is released by necrotic cells and causes neuronal damages by triggering inflammatory processes. In the cerebral ischemic brain, sustained and regulated suppression of HMGB1 has been emerged as a therapeutic means to grant neuroprotection. HMGB1 consists of two HMG boxes (A and B) and an acidic C-terminal tail, and the A box peptide antagonistically competes with HMGB1 for its receptors. In the middle cerebral artery occlusion (MCAO) in rats, a murine model of transient cerebral ischemia, administration of HMGB1 A box intraparenchymally, after encapsulated in biodegradable gelatin microspheres (GMS), which enhances the stability of peptide inside and allows its sustained delivery, at 1 h, 3 h, or 6 h after MCAO, reduced mean infarct volumes by, respectively, 81.3%, 42.6% and 30.7% of the untreated MCAO-brain, along with remarkable improvement of neurological deficits. Furthermore, the administration of HMGB1 A box/GMS suppressed proinflammatory cytokine inductions more strongly than the injection of non-encapsulated HMGB1 A box. Given that insulted brains-like ischemia have enhanced gelatinase activity than the normal brain, our results suggest that GMS-mediated delivery of therapeutic peptides is a promising means to provide efficient neuroprotection in the postischemic brain.

Monodisperse liquid-filled biodegradable microcapsules.

PURPOSE: Encapsulation of liquids into biodegradable polymer microcapsules has been a challenging task due to production limitations stemming from solution viscosity, phase stabilization, molecular localization, and scalable production. We report an extension of Precision Particle Fabrication (PPF) technology for the production of monodisperse liquid-filled microcapsules containing an oil or aqueous core and contrast these results to double-walled microspheres. MATERIALS AND METHODS: PPF technology utilizes a coaxial nozzle to produce a liquid core jet surrounded by a polymer annular jet, which is further encompassed by a non-solvent carrier stream, typically 0.5% wt/vol polyvinyl alcohol in water. Jet diameters are controlled by the volumetric flow rate of each phase. The compound jet is then disrupted into uniform core/shell droplets via a controllable acoustic wave and shell material is hardened by solvent extraction. RESULTS: Monodisperse polymeric microcapsules demonstrated a narrow size distribution and the formation of a continuous shell leading to efficient encapsulation of various liquid cores. The intermingling of core and shell phases and the localization of different molecular probes (fluorescent dyes and fluorescently labeled proteins) to the core or shell phase provided additional evidence of phase separation and molecular partitioning, respectively. We also demonstrate the pulsatile release of bovine serum albumin encapsulated in an aqueous core. CONCLUSIONS: PPF technology provided exceptional control of the overall size and shell thickness of microcapsules filled with various types of oil or water. This technique may enable advanced delivery profiles of pharmaceuticals or nutraceuticals.