The Diffusive Properties of Hydrogel Microcapsules for Cell Encapsulation.

Hydrogel microcapsules have been used for decades to encapsulate cells and treat diseases ranging from neurodegenerative disorders to more systemic applications like Type I Diabetes. This cell encapsulation modality has been developed through more cumulative experiments than perhaps any other, owing to the relative ease of accessing the required materials, the commercial availability of droplet-generating instrumentation, and the mild microenvironment and unique permeability properties of hydrogels that are difficult to attain with alternative encapsulation systems employing thermoplastic materials. Because of their size and shape, microcapsules have an inherent advantage over macroencapsulation devices due to the more favorable surface area to volume ratio, which allows for greater efficiency in the amount of cellular cargo that is entrapped and enhanced nutrient exchange and efflux of secreted products. Unfortunately, with this significant positive benefit comes the caveat of difficult or impractical retrievability, highlighting the paradox that is particularly relevant as differentiated stem cell sources become more readily available. This chapter focuses on several techniques that can be used to evaluate the permeability and pore structure of hydrogel microcapsules, including a simplistic model for describing the diffusive behavior of alginate-polycation-alginate (APA) microcapsules with a liquid core, and an ancillary method to evaluate the ultrastructure of the APA membrane including morphometric analysis.

The development of encapsulated cell technologies as therapies for neurological and sensory diseases.

Cell encapsulation therapies involve the implantation of cells that secrete a therapeutic factor to provide clinical benefits. The transplanted cells are protected from immunorejection via encapsulation in a semipermeable membrane. This treatment strategy was originally investigated as a method for protecting pancreatic islets from immunorejection, thus allowing them to secrete insulin as a chronic treatment for diabetes. Since then a significant body of work has been conducted in developing cell encapsulation therapies to treat a variety of different diseases. Many of these conditions involve neurodegeneration, such as Alzheimer's and Parkinson's disease, as cell encapsulation therapies have proven to be particularly suitable for delivering therapeutics to the central nervous system. This is mainly because they offer chronic delivery of a therapeutic and can be implanted proximal to the affected tissue, bypassing the blood brain barrier, which is impermeable to many agents. Whilst these therapies are not yet widely available in the clinic, promising results have been obtained in several advanced clinical trials and further developmental work is currently underway. This review specifically examines the development of encapsulated cell therapies as treatments for neurological and sensory diseases and evaluates the challenges that are yet to be overcome before they can be made available for clinical use.

Functional modulation of choroid plexus epithelial clusters in vitro for tissue repair applications.

One of the primary obstacles in the restoration or repair of damaged tissues is the temporospatial orchestration of biological and physiological events. Cellular transplantation is an important component of tissue repair as grafted cells can serve as replacement cells or as a source of secreted factors. But few, if any, primary cells can perform more than a single tissue repair function. Epithelial cells, derived from the choroid plexus (CP), are an exception to this rule, as transplanted CP is protective and regenerative in animal models as diverse as CNS degeneration and dermal wound repair. They secrete a myriad of proteins with therapeutic potential as well as matrix and adhesion factors, and contain responsive cytoskeletal components potentially capable of precise manipulation of cellular and extracellular niches. Here we isolated CP from neonatal porcine lateral ventricles and cultured the cells under a variety of conditions to specifically modulate tissue morphology (2D vs. 3D) and protein expression. Using qRT-PCR analysis, transmission electron microscopy, and gene microarray studies we demonstrate a fine level of control over CP epithelial cell clusters opening further opportunities for exploration of the therapeutic potential of this unique tissue source.

Secreted products from the porcine choroid plexus accelerate the healing of cutaneous wounds.

The choroid plexus (CP), located at the blood-brain interface, is partially responsible for maintaining the composition of cerebrospinal fluid. Epithelial cell clusters isolated from the CP secrete numerous biologically active molecules, and are neuroprotective when transplanted in animal models of Huntington's disease and stroke. The transcriptomic and proteomic profiles of CP may extend beyond CNS applications due to an abundance of trophic and regenerative factors, including vascular endothelial growth factor, transforming growth factor-beta, and others. We used microarray to investigate the transcriptome of porcine CP epithelium, and then assessed the in vitro and in vivo regenerative capability of secreted CP products in cell monolayers and full-thickness cutaneous wounds. In vitro, CP reduced the void area of fibroblast and keratinocyte scratch cultures by 70% and 33%, respectively, compared to empty capsule controls, which reduced the area by only 35% and 6%, respectively. In vivo, after 10 days of topical application, CP conditioned medium lyophilate dispersed in antibiotic ointment produced a twofold improvement in incision tensile strength compared to ointment containing lyophilized control medium, and an increase in the regeneration of epidermal appendages from roughly 50-150 features per wound. Together, these data identify the CP as a source of secreted regenerative molecules to accelerate and improve the healing of superficial wounds and potentially other similar indications.

Encapsulated porcine islet transplantation: an evolving therapy for the treatment of type I diabetes.

BACKGROUND: Allogeneic tissue-based therapies for Type I diabetes have demonstrated efficacy but are limited due to tissue-sourcing constraints, as the number of patients exceeds that of tissue donors. Porcine islets derived from designated pathogen-free sources could be an alternative, particularly if delivered in a way that evades the host immune system's rejection. METHODS: This review focuses on approaches designed to protect xenogeneic islets from immune rejection by provision of perm-selective barriers. RESULTS: Designated pathogen-free herds could provide a supply of wild-type porcine islets that are well tolerated when administered in a suitable protective delivery vehicle. Such barrier systems have enabled amelioration of diabetes in a variety of animal models and preliminary evidence suggests that similar results could be attained in humans. CONCLUSION: With advances in biomaterial design, source tissue selection, and the evolution of critical cell processing techniques, contemporary encapsulated porcine islet therapies offer a new level of clinical promise.

Formulating the alginate-polyornithine biocapsule for prolonged stability: evaluation of composition and manufacturing technique.

Alginate encapsulation is one of the most widely used techniques for introducing cell-based therapeutics into the body. Numerous encapsulation methodologies exist, utilizing a variety of alginates, purification technologies, and unique polycationic membrane components. The stability of a conventional alginate formulation encapsulated using a commercially available technique and apparatus has been characterized extensively. The current study employs an encapsulation protocol and ultra-pure alginate pioneered at the University of Perugia. The enhanced microcapsules were produced, characterized, and implanted into the brain, peritoneal cavity, and subcutaneous space of Long-Evans rats. After 14, 28, 60, 90, 120, and 180 or 215 days, capsules were explanted and the surface was analyzed using Fourier-transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Image analysis was carried out to measure changes in diameter and wall thickness. FTIR peak analysis and surface morphology from SEM indicated that the enhanced encapsulation technique and formulation produced a stable biocapsule capable of survival in all sites, including the harsh peritoneal environment, for at least 215 days. Preimplant analysis showed a marked increase in the structural integrity of the enhanced formulation with improved elasticity and burst strength compared with the baseline formulation, which remained stable for less than 60 days. The enhanced microcapsule composition showed advantages in physical strength and longevity, indicating that small changes in encapsulation methodologies and materials selection can dramatically impact the stability and longevity of alginate microcapsules and their contents.

Stability of alginate-polyornithine microcapsules is profoundly dependent on the site of transplantation.

Alginate encapsulation is a form of cell-based therapy with numerous preclinical successes but recalcitrant complications related to stability and reproducibility. Understanding how alginate stability varies across different transplant sites will help identify indications that might benefit most from this approach. Alginate stability has been quantified in the peritoneum, but there are no systematic studies comparing its relative stability across transplant sites. This study compares the stability of alginate-polycation microcapsules implanted in the peritoneum to those implanted in the brain and subcutaneous space at 14, 28, 60, 90, 120, and 180 days in-life. Using Fourier-Transform Infrared Spectroscopy (FTIR), the surface of explanted capsules was analyzed for the relative proportion of alginate (outer coat) and the polycationic polyornithine (middle coat). Using a mathematic relationship between FTIR peaks related to these two material components, an index was generated to compare the stability of four different alginates. A notable difference was observed with rapid breakdown in the peritoneum. Conversely, identical alginate capsules transplanted into the brain or subcutaneous space were stable for the 6 month study. These data suggest that (1) successful intraperitoneal transplantation requires modifications of the capsule configuration, the host environment, or both and (2) that sites such as the brain and subcutaneous space are inherently less hostile to conventional alginate capsule configurations.

Improving relative bioavailability of dicumarol by reducing particle size and adding the adhesive poly(fumaric-co-sebacic) anhydride.

PURPOSE: This study was carried out to show the effect of particle size reduction and bioadhesion on the dissolution and relative bioavailability of dicumarol. METHODS: Formulations were produced by a variety of methods including a novel technique to reduce particle size as well as phase inversion with poly(fumaric-co-sebacic)anhydride p(FA:SA) to create nanospheres. Drug was administered to groups of pigs and rats via oral gavage of a suspension, and dicumarol concentration in the blood was measured using a double extraction technique. RESULTS: In vitro results showed improved dissolution in both the micronized formulation and the encapsulated p(FA:SA) nanospheres. In vivo, relative bioavailability of a spray-dried formulation was increased by 17% in the rat and 72% in the pig by further reduction in particle size. The bioadhesive p(FA:SA) formulation also improved relative bioavailability over the spray-dried drug, increasing it by 55% in the rat and 96% in the pig. Additionally, the p(FA:SA) formulation prolonged Tmax and decreased Cmax in both species. CONCLUSION: This work demonstrates the importance of particle size and bioadhesion to improve oral bioavailability of ducumarol.