Alginate has long been the material of choice for immunoprotection of islets due to its low cost and ability to easily form microspheres. Unfortunately, this seaweed-derived material is notoriously prone to fibrotic overgrowth in vivo, resulting in premature graft failure. The purpose of this study was to test an alternative, hyaluronic acid (HA-COL), for in vitro function, viability, and allogeneic islet transplant outcomes in diabetic rats. In vitro studies indicated that the HA-COL gel had diffusion characteristics that would allow small molecules such as glucose and insulin to enter and exit the gel, whereas larger molecules (70 and 500 kDa dextrans) were impeded from diffusing past the gel edge in 24 h. Islets encapsulated in HA-COL hydrogel showed significantly improved in vitro viability over unencapsulated islets and retained their morphology and glucose sensitivity for 28 days. When unencapsulated allogeneic islet transplants were administered to the omentum of outbred rats, they initially were normoglycemic, but by 11 days returned to hyperglycemia. Immunohistological examination of the grafts and surrounding tissue indicated strong graft rejection. By comparison, when using the same outbred strain of rats, allogeneic transplantation of islets within the HA-COL gel reversed long-term diabetes and prevented graft rejection in all animals. Animals were sacrificed at 40, 52, 64, and 80 weeks for evaluation, and all were non-diabetic at sacrifice. Explanted grafts revealed viable islets in the transplant site as well as intact hydrogel, with little or no evidence of fibrotic overgrowth or cellular rejection. The results of these studies demonstrate great potential for HA-COL hydrogel as an alternative to sodium alginate for long-term immunoprotected islet transplantation.

When working with isolated islet preparations, measuring the volume of tissue is not a trivial matter. Islets come in a large range of sizes and are often contaminated with exocrine tissue. Many factors complicate the procedure, and yet knowledge of the islet volume is essential for predicting the success of an islet transplant or comparing experimental groups in the laboratory. In 1990, Ricordi presented the islet equivalency (IEQ), defined as one IEQ equaling a single spherical islet of 150 μm in diameter. The method for estimating IEQ was developed by visualizing islets in a microscope, estimating their diameter in 50 μm categories and calculating a total volume for the preparation. Shortly after its introduction, the IEQ was adopted as the standard method for islet volume measurements. It has helped to advance research in the field by providing a useful tool improving the reproducibility of islet research and eventually the success of clinical islet transplants. However, the accuracy of the IEQ method has been questioned for years and many alternatives have been proposed, but none have been able to replace the widespread use of the IEQ. This article reviews the history of the IEQ, and discusses the benefits and failings of the measurement. A thorough evaluation of alternatives for estimating islet volume is provided along with the steps needed to uniformly move to an improved method of islet volume estimation. The lessons learned from islet researchers may serve as a guide for other fields of regenerative medicine as cell clusters become a more attractive therapeutic option.

Cell microencapsulation is a rapidly expanding field with broad potential for stem cell therapies and tissue engineering research. Traditional alginate microspheres suffer from poor biocompatibility, and microencapsulation of more advanced hydrogels is challenging due to their slower gelation rates. We have developed a novel, noncytotoxic, nonemulsion-based method to produce hydrogel microspheres compatible with a wide variety of materials, called core-shell spherification (CSS). Fabrication of microspheres by CSS derived from two slow-hardening hydrogels, hyaluronic acid (HA) and polyethylene glycol diacrylate (PEGDA), was characterized. HA microspheres were manufactured with two different crosslinking methods: thiolation and methacrylation. Microspheres of methacrylated HA (MeHA) had the greatest swelling ratio, the largest average diameter, and the lowest diffusion barrier. In contrast, PEGDA microspheres had the smallest diameters, the lowest swelling ratio, and the highest diffusion barrier, while microspheres of thiolated HA had characteristics that were in between the other two groups. To test the ability of the hydrogels to protect cells, while promoting function, diabetic NOD mice received intraperitoneal injections of PEGDA or MeHA microencapsulated canine islets. PEGDA microspheres reversed diabetes for the length of the study (up to 16 weeks). In contrast, islets encapsulated in MeHA microspheres at the same dose restored normoglycemia, but only transiently (3-4 weeks). Nonencapsulated canine islet transplanted at the same dose did not restore normoglycemia for any length of time. In conclusion, CSS provides a nontoxic microencapsulation procedure compatible with various hydrogel types.

Cell therapies are hampered by a lack of available delivery systems, resulting in inconsistent outcomes in animal studies and human clinical trials. Hydrogel encapsulants offer a broad range of tunable characteristics in the design of cell delivery vehicles. The focus of the hydrogel field has been on durable encapsulants that provide long-term paracrine function of the cells. However, some cell therapies require cell-to-cell contact in order to elicit their effect. Controlled release microencapsulants would be beneficial in these situations, but appropriate polymers have not been adaptable to microsphere manufacturing because they harden too slowly. We developed and tested a novel microencapsulant formulation (acrylated hyaluronic acid: AHA) with degradation characteristics as a controlled release cell delivery vehicle. The properties of AHA microspheres were evaluated and compared to those of poly­(ethylene glycol) diacrylate (PEGDA), a durable hydrogel. AHA microspheres possessed a higher swelling ratio, lower diffusion barrier, faster degradation rate, a lower storage modulus, and a larger average diameter than microspheres composed of PEGDA. Additionally, in vitro cell viability and release and short-term in vivo biocompatibility in immune competent Sprague–Dawley rats was assessed for each microsphere type. Compared to PEGDA, microspheres composed of AHA resulted in significantly less foreign body response in vivo as measured by a lack of cellularity or fibrotic ring in the surrounding tissue and no cellular infiltration into the microsphere. This study illustrates the potential of AHA microspheres as a degradable cell delivery system with superior encapsulated cell viability and biocompatibility with the surrounding tissue.