Richard Wood Storrs

Preclinical Development of the Islet Sheet

Abstract: The Islet Sheet is a thin planar bioartificial endocrine pancreas fabricated by gelling highly purified alginate and islets of Langerhans. Acellular alginate layers form a uniform immunoprotective barrier to host rejection of the encapsulated cells, with the tissue nourished by passive diffusion from adjacent host tissue. The overall thickness of the Islet Sheet, 250 μm, is chosen to maximize nutrient diffusion. In this paper we describe the early development of the Islet Sheet, including purification and fractionation of the alginates used, difficulties in maintaining sheet planarity, and preliminary metabolic studies in pancreatectomized dogs. In a key experiment, approximately 75,000 allogeneic islet equivalents in six Islet Sheets were sutured to the omentum of a 7‐kg female beagle dog at the time of pancreatectomy. Fasting euglycemia was maintained for 84 days. Fed blood sugars were usually below 150 mg/dL. A single injection of 2 U insulin was administered on day 9, and antibiotics were administered for two weeks. No other drugs were used. IVGTT post implant was not normal, but seemed to improve between 30 and 60 days. Upon omentectomy and sheet removal the metabolic parameters deteriorated to a frankly diabetic state within seven days. The sheets did not remain flat, but fragments were recovered within hard, mostly acellular capsules. Dithizone staining showed islets within alginate sheets recovered from the interior of these capsules, suggesting that allogeneic islet tissue survived 84 days and was responsible for maintaining fasting euglycemia.

Function and Viability of Human Islets Encapsulated in Alginate Sheets: In Vitro and in Vivo Culture

Islet encapsulation offers an immune system barrier for islet transplantation, and encapsulation within an alginate sheetlike structure offers the ability to be retrievable after transplanted. This study aims to show that human islets encapsulated into islet sheets remain functional and viable after 8 weeks in culture or when transplanted into the subcutaneous space of rats. Human islets were isolated from cadaveric organs. Dissociation and purification were done using enzymatic digestion and a continuous Ficoll-UWD gradient. Purified human islets were encapsulated in alginate sheets. Human Islet sheets were either kept in culture, at 37°C and 5% CO(2), or transplanted subcutaneously into Lewis rats. After 1, 2, 4, and 8 weeks, the human islet sheets were retrieved from the rats and assessed. The viability of the sheets was measured using fluorescein diacetate (FDA)/propidium iodide (PI), and function was measured through glucose-stimulated insulin release, in which the sheets were incubated for an hour in low-glucose concentration (2.8 mmol/L) and then high (28 mmol/L), then high (28 mmol/L) plus 3-isobutyl-1-methylxanthine (50 μm). Human islet sheets remained both viable, above 70%, and functional, with a stimulation index (insulin secretion in high glucose divided by insulin secretion in low glucose) above 1.5, over 8 weeks of culture or subcutaneous transplantation. Islet transplantation continues to make advances in the treatment of type 1 diabetes. These preliminary results suggest that encapsulated islets sheets can survive and maintain islet viability and function in vivo, within the subcutaneous region.