Polymer particle-based micromolding to fabricate novel microstructures

Conventional micromolding provides rapid and low-cost methods to fabricate polymer microstructures, but has limitations when producing sophisticated designs. To provide more versatile micromolding techniques, we developed methods based on filling micromolds with polymer microparticles, as opposed to polymer melts, to produce microstructures composed of multiple materials, having complex geometries, and made using mild processing conditions. Polymer microparticles of 1 to 30 μm in size were made from PLA, PGA and PLGA using established spray drying and emulsion techniques either with or without encapsulating model drug compounds. These polymer microparticles were filled into PDMS micromolds at room temperature and melted or bonded together to form microstructures according to different protocols. Porous microstructures were fabricated by ultrasonically welding microparticles together in the mold while maintaining the voids inherent in their packing structure. Multi-layered microstructures were fabricated to have different compositions of polymers and encapsulated compounds located in different regions of the microstructures. More complex arrowhead microstructures were fabricated in a two-step process using a single mold. To assess possible applications, microstructures were designed as microneedles for minimally invasive drug delivery. Multi-layer microneedles were shown to insert into cadaver tissue and, according to design, detach from their base substrate and remain embedded in the tissue for controlled release drug delivery over time. We conclude that polymer particle-based micromolding can encapsulate compounds within microstructures composed of multiple materials, having complex geometries, and made using mild processing conditions.     

Microparticles developed by electrohydrodynamic atomization for the local delivery of anticancer drug to treat C6 glioma in vitro

This study aims to fabricate biodegradable polymeric particles by electrohydrodynamic atomization (EHDA) for applications in sustained delivery of anticancer drug-paclitaxel to treat C6 glioma in vitro. Controllable morphologies such as spheres, doughnut shapes and corrugated shapes with sizes from several tens of microns to hundred nanometers of particles were observed by scanning electron microscopy (SEM) and field emission electron microscope (FSEM). The differential scanning calorimetry (DSC) study indicated that paclitaxel could be either in an amorphous or disordered-crystalline phase of a molecular dispersion or a solid solution state in the polymer matrix after fabrication. The X-ray photoelectron spectroscopy (XPS) result suggested that some amount of paclitaxel could exist on the surface layer of the microparticles. The encapsulation efficiency was around 80% and more than 30 days in vitro sustained release profile could be achieved. Cell cycling results suggested that paclitaxel after encapsulation by EHDA could keep its biological function and inhibit C6 glioma cells in G2/M phase. The cytotoxicity of paclitaxel-loaded biodegradable microparticles to C6 glioma cells could be higher than Taxol in the long-term in vitro tests evaluated by MTS assay. The drug delivery devices developed by EHDA in this study could be promising for the local drug delivery to treat malignant glioma.     

Preparation and characterization of starch-poly-ε-caprolactone microparticles incorporating bioactive agents for drug delivery and tissue engineering applications

One limitation associated with the delivery of bioactive agents concerns the short half-life of these molecules when administered intravenously, which results in their loss from the desired site. Incorporation of bioactive agents into depot vehicles provides a means to increase their persistence at the disease site. Major issues are involved in the development of a proper carrier system able to deliver the correct drug, at the desired dose, place and time. In this work, starch-poly-ε-caprolactone (SPCL) microparticles were developed for use in drug delivery and tissue engineering (TE) applications. SPCL microparticles were prepared by using an emulsion solvent extraction/evaporation technique, which was demonstrated to be a successful procedure to obtain particles with a spherical shape (particle size between 5 and 900 μm) and exhibiting different surface morphologies. Their chemical structure was confirmed by Fourier transform infrared spectroscopy. To evaluate the potential of the developed microparticles as a drug delivery system, dexamethasone (DEX) was used as model drug. DEX, a well-known component of osteogenic differentiation media, was entrapped into SPCL microparticles at different percentages up to 93%. The encapsulation efficiency was found to be dependent on the polymer concentration and drug-to-polymer ratio. The initial DEX release seems to be governed mainly by diffusion, and it is expected that the remaining DEX will be released when the polymeric matrix starts to degrade. In this work it was demonstrated that SPCL microparticles containing DEX can be successfully prepared and that these microparticular systems seem to be quite promising for controlled release applications, namely as carriers of important differentiation agents in TE.     

The long-term potential of biodegradable poly(lactide-co-glycolide) microparticles as the next-generation vaccine adjuvant

Biodegradable polymeric microparticles of poly(lactide-co-glycolide) (PLG) have been extensively evaluated for drug delivery and vaccine applications over the last three decades. Despite a wealth of studies on the use of PLG microparticles in vaccines through controlled release of antigens, there is no commercial PLG-based vaccine as yet. The key challenge that prevented the development of PLG microparticles as commercial vaccines was the instability of encapsulated antigen. Over the years, advancements were made towards maintaining antigen integrity during PLG microparticle preparation and sterilization. In parallel and independently, development of PLG microparticles as therapeutic commercial products established PLG with an excellent safety record in humans, and as a suitable candidate for next-generation vaccines. Through the combination of Toll-like receptor agonist encapsulation and surface adsorption of antigen, PLG microparticles can be used as a vaccine adjuvant to address unmet medical needs, such as vaccines against HIV, malaria and TB. With strategic development of PLG-based vaccines, PLG microparticles can offer advantages over the conventional vaccine adjuvants allowing commercial development of this adjuvant.     

花样结构白蛋白缓释载体的制备及在豚鼠内耳跨膜给药的应用

目的制备花样结构白蛋白(flower-shaped bovine serum albumin,FBSA)微纳材料作为载体进行内耳跨圆窗膜给药研究,为临床寻找新的药物缓释载体奠定基础。方法应用改良的去溶剂法制备FBSA微纳材料,并用荧光显微镜、电子显微镜、粒径分析仪等对其进行系统的表征。通过体外药物释放实验、MTT法来评估其细胞相容性和细胞毒性。通过小动物活体成像观察FBSA在豚鼠听泡内的扩散及在圆窗膜上的附着。结果蛋白基微纳米材料为放射状花样结构,大小约为50⁸0μm。空白FBSA微纳材料的zeta电位是-16.2 mV,其最高的载药量和包封率分别是21.4%和40.0%,具有缓释效果。通过L929细胞的毒性实验测试提示经热变性处理固定后的材料具有更低的毒性和更好的细胞相容性。小动物活体实验可见药物在内耳中扩散及在圆窗膜表面附着。结论成功构建FBSA微纳材料载体,在治疗内耳病的局部给药方面有着良好的应用前景。     

Biodegradable microparticles and fiber fabrics for sustained delivery of cisplatin to treat C6 glioma in vitro

The duration of cisplatin release from most of the drug delivery devices seemed to be shorter than 14 days except large microparticles. The objective of this study was to fabricate and characterize cisplatin-loaded PLA microparticles, PLA/PLGA (30/70) composite microparticles, and fibers as formulations for long-term sustained delivery of cisplatin to treat C6 glioma in vitro by electrospray and electrospinning techniques. Cisplatin-loaded biodegradable microparticles with particle size of around 5 microm and fiber fabrics with diameter of 0.5-1.7 microm were obtained using electrospray and electrospinning techniques. Encapsulation efficiency and in vitro release of formulations were measured by ICP-OES. The encapsulation efficiency for different samples of microparticles was approximately from 33% to 72% and the fiber fabrics had encapsulation efficiency greater than 90%. Cisplatin-loaded microparticles showed typical characteristics of cisplatin release profile: a large initial burst followed by a sustained slow release of 35 days. The composite PLA/PLGA (30/70) microparticles could reduce the initial burst release of cisplatin because of their core-shell structures. In contrast, more than 75 days sustained release could be achieved by fiber fabric formulations without large initial burst. MTT assay was used to quantify the cytotoxicity of different formulations against C6 glioma cells. Microparticle formulations had slightly higher cytotoxicity than free drug. In contrast, the cytotoxicity of fiber fabrics formulation was around 4 times higher than of the free drug based on the actual amount of drug released. The microparticle and fiber fabric formulations presented may be promising for the sustained delivery of cisplatin to eliminate the undesired side effects caused by direct injection of cisplatin solution in systemic administration.     

Dexamethasone-containing PLGA superparamagnetic microparticles as carriers for the local treatment of arthritis

Superparamagnetic iron oxide nanoparticles (SPIONs) are attractive materials that have been widely used in medicine for diagnostic imaging and therapeutic applications. In our study, SPIONs and the corticosteroid dexamethasone acetate (DXM) are co-encapsulated into PLGA microparticles for the aim of locally treating inflammatory conditions such as arthritis. The magnetic properties conferred by the SPIONs could help to maintain the microparticles in the joint with an external magnet. The aim of this study was to investigate the interaction between magnetic microparticles and human synovial fibroblasts in terms of microparticle uptake (FACS, confocal and optical microscopy), internalization mechanism (Prussian Blue staining, TEM, immunofluorescence), cell toxicity (MTT) and tissue reaction after intra-articular injection (histology). The results show that the microparticles have an excellent biocompatibility with synoviocytes and that they are internalized through a phagocytic process, as demonstrated by fluorescence-activated cell sorting and morphological analyses of cells exposed to microparticles. Histological analysis showed that the prepared microparticles did not induce any inflammatory reaction in the joint. This type of carrier could represent a suitable magnetically retainable intra-articular drug delivery system for treating joint diseases such as arthritis or osteoarthritis.     

Systemic and Mucosal Delivery of Drugs within Polymeric Microparticles Produced by Spray Drying

Encapsulation of therapeutic and diagnostic materials into polymeric particles is a means to protect and control or target the release of active substances such as drugs, vaccines, and genetic material. In terms of mucosal delivery, polymeric encapsulation can be used to promote absorption of the active substance, while particles can improve the half-life of drugs administered systemically. Spray drying is an attractive technology used to produce such microparticles, because it combines both the encapsulation and drying steps in a rapid, single-step operation. Even so, spray drying is not classically associated with processes used for drug and therapeutic material encapsulation, since elevated temperatures could potentially denature the active substance. However, a comprehensive review of the literature revealed a number of studies demonstrating that spray drying can be used to produce microparticulate formulations with labile therapeutics. Polymers commonly employed include synthetics such as methacrylic copolymers and polyesters, and natural materials including chitosan and alginate. Drugs and active substances are diverse and included antibiotics, anti-inflammatory agents, and chemotherapeutics. Regarding the delivery of spray-dried particles, the pulmonary, oral, colonic, and nasal mucosal routes are often investigated because they offer a convenient means of administration, which promotes physician and patient compliance. In addition, spray drying has been widely used to produce polymeric microparticles for systemic delivery in order to control the delivery of drugs, vaccines, or genetic material that may exhibit poor pharmacokinetic profiles or pose toxicity concerns. This review presents a brief introduction to the technology of spray drying and outlines the delivery routes and the applications of spray-dried polymeric microparticles.     

Phagocytotic Competence of Differentiated U937 Cells for Colloidal Drug Delivery Systems in Immune Cells

Drug delivery into immune cells has high potential for the treatment of all kinds of inflammation, allowing a target-oriented transport of active agents. The advantage of this local drug release is the prevention of negative effects of systemic applications and low-dose application. Thereby, the phagocytotic capability of mature phagocytes is essential. Microparticles can be loaded with immune regulatory substances to control and terminate inflammatory processes. In this study, silica microparticles were co-incubated with monocyte/macrophage-like cells in order to determine phagocytotic particle uptake. The phorbol ester-triggered differentiation was proven by the increased expression of surface markers as phosphatidylserine and CD14 and enhanced lysosomal activity. Particle/cell co-incubation results in cell surface attachment followed by phagocytosis. Phagolysosomal ingestion could be determined by co-localization using fluorescence staining techniques. In contrast, no particle interaction with undifferentiated cells could be found. Under phagolysosomal conditions, multilayer degradation within 22 h could be shown, indicating a valuable carrier basis design for the time-controlled delivery of active agents. Subsequently, it can be assumed that a higher differentiation degree allows phagocytosis of microparticles, providing drug delivery into immuno-active cells.     

A novel preparation method of paclitaxcel-loaded folate-modified chitosan microparticles and in vitro evaluation

A new chitosan microparticles loading paclitaxel (PTX) for application as an oral delivery system were developed using a novel double emulsion crosslinking method. To improve the targeted effect, folic acid (FA) was introduced onto the surface of microparticles using chemical method. The method was based on Schiff reaction between amino group of chitosan and carboxyl group of FA, and folate-chitosan (FA-CS) conjugate was characterized using infrared spectrum analysis (FT-IR), and the microparticles were named as FA-CS-PTX/MPs. FA-CS-PTX/MPs had larger size of average diameter 223.6 nm, while PTX-loaded chitosan microparticles (CS-PTX/MPs) had 179.1 nm average diameter. The zeta potential of CS-PTX/MPs and FA-CS-PTX/MPs was 22.3 and 33.1 mV, respectively. SEM and TEM showed both the two microparticles had well-defined spherical structure. The in vitro drug release was studied under different pH conditions, and a two-phase kinetics model was found to be the most adequate kinetic model. Furthermore, the cytotoxicity activities of drug-carriers against L929 cells and the cellular uptake of PTX-loaded microparticles against HepG2 cells were investigated. Results demonstrated that FA-CS-PTX/MPs might be a promising drug carrier for promoting PTX cellular uptake and could be used as a potential tumor-targeted drug vector.     

Bioadhesive Microdevices for Drug Delivery: A Feasibility Study

A variety of delivery systems have been devised to improve the oral bioavailability of drugs including enterically coated tablets, capsules, particles, liposomes, and others. Microfabrication technology, however, may offer some potential advantages over conventional drug delivery technologies. This technology, combined with appropriate surface chemistry, may permit the highly localized and unidirectional release of drugs, permeation enhancers, and/or promoters. In this study, we demonstrate the fabrication of prototype reservoir containing microparticles and a surface chemistry protocol that can be used to bind lectin via avidin-biotin interactions to silicon microparticles. In vitro studies show enhanced bioadhesion of these lectin conjugated microparticles. Such an approach can be used to improve the absorption of pharmacologically active biopolymers such as peptides, proteins and oligonucleotides into circulation at targeted sights in the GI system. Moreover, the use of microfabrication allows one to tailor the size, shape, reservoir volume, and surface characteristics of the drug delivery vehicle.     

Microfabricated Drug Delivery Systems: Concepts to Improve Clinical Benefit

Important classes of drugs have yet to benefit from advances in drug delivery technology. Strategies to provide reasonable oral bioavailability of peptide and proteins drugs remain elusive, for example. Systemic cancer drugs produce dose-limiting toxicities largely due to their lack of selectivity. Although delivery systems such as immunotoxins and liposomes improve selectivity of a few cancer drugs, current technology is not suitable for the vast majority of such molecules. Systems able to mimic the body's natural feedback mechanisms for secretion of hormones such as insulin represents yet another unmet medical need. Microfabrication techniques may permit the creation of drug delivery systems that possess a combination of structural, mechanical, and perhaps electronic features which may surmount some of these challenges. In this review, drug delivery concepts are presented which capitalize on the strengths of microfabrication. Possible applications include micromachined silicon membranes to create implantable biocapsules for the immunoisolation of pancreatic islet cells—as a possible treatment for diabetes—and sustained release of injectable drugs needed over long time periods. Asymmetrical, drug-loaded microfabricated particles with specific ligands linked to the surface are proposed for improving oral bioavailability of peptide (and perhaps protein) drugs. Similarly designed particles with sizes in the 2–10 m range may be safe to administer intravenously and a clinical strategy is suggested for using such microparticles for treating solid tumors. Although hypothetical now, work is in progress to prove the concepts presented here and to validate the intuitive belief that there is an important place for microfabricated systems in drug delivery.     

Polymeric particulate technologies for oral drug delivery and targeting: A pathophysiological perspective

The oral route for delivery of pharmaceuticals is the most widely used and accepted. Nanoparticles and microparticles are increasingly being applied within this arena to optimize drug targeting and bioavailability. Frequently the carrier systems used are either constructed from or contain polymeric materials. Examples of these nanocarriers include polymeric nanoparticles, solid lipid nanocarriers, self-nanoemulsifying drug delivery systems and nanocrystals. It is the purpose of this review to describe these cutting edge technologies and specifically focus on the interaction and fate of these polymers within the gastrointestinal system.     

The effect of starch and starch-bioactive glass composite microparticles on the adhesion and expression of the osteoblastic phenotype of a bone cell line

There is a clear need for the development of microparticles that can be used simultaneously as carriers of stem/progenitor cells and as release systems for bioactive agents, such as growth factors or differentiation agents. In addition, when thinking on bone-tissue-engineering applications, it would be very useful if these microparticles are biodegradable and could be made to be bioactive. Microparticles with all those characteristics could be cultured together with adherent cells in appropriate bioreactors to form in vitro constructs that can then be used in tissue-engineering therapies. In this work, we have characterized the response of MC3T3-E1 pre-osteoblast cells to starch-based microparticles. We evaluated the adhesion, proliferation, expression of osteoblastic markers and mineralization of cells cultured at their surface. The results clearly show that MC3T3-E1 pre-osteoblast cells adhere to the surface of both polymeric and composite starch-based microparticles and express the typical osteoblastic marker genes. Furthermore, the cells were found to mineralize the extracellular matrix (ECM) during the culture period. The obtained results indicate that starch-based microparticles, known already to be biodegradable, bioactive and able to be used as carriers for controlled release applications, can simultaneously be used as carriers for cells. Consequently, they can be used as templates for forming hybrid constructs aiming to be applied in bone-tissue-engineering applications.     

Chitosan-modified porous silicon microparticles for enhanced permeability of insulin across intestinal cell monolayers

Porous silicon (PSi) based particulate systems are emerging as an important drug delivery system due to its advantageous properties such as biocompatibility, biodegradability and ability to tailor the particles' physicochemical properties. Here, annealed thermally hydrocarbonized PSi (AnnTHCPSi) and undecylenic acid modified AnnTHCPSi (AnnUnTHCPSi) microparticles were developed as a PSi-based platform for oral delivery of insulin. Chitosan (CS) was used to modify the AnnUnTHCPSi microparticles to enhance the intestinal permeation of insulin. Surface modification with CS led to significant increase in the interaction of PSi microparticles with Caco-2/HT-29 cell co-culture monolayers. Compared to pure insulin, the CS-conjugated microparticles significantly improved the permeation of insulin across the Caco-2/HT-29 cell monolayers, with ca. 20-fold increase in the amount of insulin permeated and ca. 7-fold increase in the apparent permeability (P_app) value. Moreover, among all the investigated particles, the CS-conjugated microparticles also showed the highest amount of insulin associated with the mucus layer and the intestinal Caco-2 cells and mucus secreting HT-29 cells. Our results demonstrate that CS-conjugated AnnUnTHCPSi microparticles can efficiently enhance the insulin absorption across intestinal cells, and thus, they are promising microsystems for the oral delivery of proteins and peptides across the intestinal cell membrane.     

Recent developments of collagen-based materials for medical applications and drug delivery systems

In this review, an attempt was made to summarize some of the recent developments in the application of collagen as a biomaterial and in drug delivery systems. The main applications covered include: collagen for burn/wound cover dressings; osteogenic and bone filling materials; antithrombogenic surfaces; and immobilization of therapeutic enzymes. Recently, collagen used as a carrier for drug delivery has attracted many researchers throughout the world. The use of collagen for various drug delivery systems has also been reviewed in this article. Collagen-based drug delivery systems include: injectable microspheres based on gelatin (degraded form of collagen); implantable collagen-synthetic polymer hydrogels; interpenetrating networks of collagen; and synthetic polymers collagen membranes for ophthalmic delivery. Recent efforts to use collagen-liposomal composites for controlled drug delivery, as well as collagen as controlling membranes for transdermal delivery, were also reviewed. In this review, the main emphasis was on the work done in our laboratory.     

Localized delivery of paclitaxel in solid tumors from biodegradable chitin microparticle formulations

Paclitaxel (Taxol)-containing chitin and chitin-Pluronic F-108 microparticles were formulated as biodegradable systems for localized administration in solid tumors. The microparticles were characterized by Fourier transform infrared (FT-IR) spectroscopy, scanning electron microscopy (SEM), and swelling studies in phosphate-buffered saline (PBS, pH 7.4). Lysozyme-induced degradation and in vitro release of paclitaxel was examined in PBS at 37 degrees C. The percent change in tumor volume was used to assess efficacy of the Formulations after local administration in murine Lewis lung carcinoma model. FT-IR confirmed higher degree of acetylation in chitin microparticles from the starting chitosan sample and the SEM showed that the chitin-Pluronic F-108 microparticles were significantly more porous than chitin microparticles. Due to higher porosity, chitin-Pluronic microparticles were able to imbibe higher swelling medium and degraded much faster in the presence of lysozyme than chitin microparticles. After 48 h. 51% of incorporated paclitaxel was released from chitin-Pluronic microparticles as compared to 28% from chitin microparticles. In vivo studies in Lewis lung carcinoma-bearing mice showed that the tumor volumes after 6 days using paclitaxel-loaded chitin and chitin-Pluronic F-108 microparticles was 458 and 307 mm3, respectively. In contrast, the tumor volume was 997 mm3 for the untreated control. The results of this study show that chitin and chitin-Pluronic F-108 microparticles are biodegradable drug delivery systems that can be useful for localized delivery of paclitaxel in solid tumors.     

The intracellular uptake of CD95 modified paclitaxel-loaded poly(lactic-co-glycolic acid) microparticles

The CD95/CD95L receptor-ligand system is mainly recognised in the induction of apoptosis. However, it has also been shown that CD95L is over-expressed in many cancer types where it modulates immune-evasion and together with its receptor CD95 promotes tumour growth. Here, we show that CD95 surface modification of relatively large microparticles >0.5 μm in diameter, including those made from biodegradable polylactic-co-glycolic acid (PLGA), enhances intracellular uptake by a range of CD95L expressing cells in a process akin to phagocytosis. Using this approach we describe the intracellular uptake of microparticles and agent delivery in neurons, medulloblastoma, breast and ovarian cancer cells in vitro. CD95 modified paclitaxel-loaded PLGA microparticles are shown to be significantly more effective compared to conventional paclitaxel therapy (Taxol) at the same dose in subcutaneous medulloblastoma (???P < 0.0001) and orthotopic ovarian cancer xenograft models where a >65-fold reduction in tumour bioluminescence was measured after treatment (?P = 0.012). This drug delivery platform represents a new way of manipulating the normally advantageous tumour CD95L over-expression towards a therapeutic strategy. CD95 functionalised drug carriers could contribute to the improved function of cytotoxics in cancer, potentially increasing drug targeting and efficacy whilst reducing toxicity.     

Directed transport of bacteria-based drug delivery vehicles: bacterial chemotaxis dominates particle shape

Several attenuated and non-pathogenic bacterial species have been demonstrated to actively target diseased sites and successfully deliver plasmid DNA, proteins and other therapeutic agents into mammalian cells. These disease-targeting bacteria can be employed for targeted delivery of therapeutic and imaging cargos in the form of a bio-hybrid system. The bio-hybrid drug delivery system constructed here is comprised of motile Escherichia coli MG1655 bacteria and elliptical disk-shaped polymeric microparticles. The transport direction for these vehicles can be controlled through biased random walk of the attached bacteria in presence of chemoattractant gradients in a process known as chemotaxis. In this work, we utilize a diffusion-based microfluidic platform to establish steady linear concentration gradients of a chemoattractant and investigate the roles of chemotaxis and geometry in transport of bio-hybrid drug delivery vehicles. Our experimental results demonstrate for the first time that bacterial chemotactic response dominates the effect of body shape in extravascular transport; thus, the non-spherical system could be more favorable for drug delivery applications owing to the known benefits of using non-spherical particles for vascular transport (e.g. relatively long circulation time).     

Dynamics of magnetic particles in cylindrical Halbach array: implications for magnetic cell separation and drug targeting

Magnetic nanoparticles for therapy and diagnosis are at the leading edge of the rapidly developing field of bionanotechnology. In this study, we have theoretically studied motion of magnetic nano- as well as micro-particles in the field of cylindrical Halbach array of permanent magnets. Magnetic flux density was modeled as magnetostatic problem by finite element method and particle motion was described using system of ordinary differential equations—Newton law. Computations were done for nanoparticles Nanomag^®-D with radius 65 nm, which are often used in magnetic drug targeting, as well as microparticles DynaBeads-M280 with radius 1.4 µm, which can be used for magnetic separation. Analyzing snapshots of trajectories of hundred magnetite particles of each size in the water as well as in the air, we have found that optimally designed magnetic circuits of permanent magnets in quadrupolar Halbach array have substantially shorter capture time than simple blocks of permanent magnets commonly used in experiments, therefore, such a Halbach array may be useful as a potential source of magnetic field for magnetic separation and targeting of magnetic nanoparticles as well as microparticles for delivery of drugs, genes, and cells in various biomedical applications.