Transplantation of purified islet cells in diabetic rats. I, Standardization of islet cell grafts

• Published in: Diabetes (New York, N.Y.) Vol. 40; no. 7; pp. 908 - 919
• Main Authors: PIPELEERS, D. G, PIPELEERS-MARICHAL, M, HANNAERT, J.-C, BERGHMANS, M, IN'T WELD, P. A, ROZING, J, VAN DE WINKEL, M, GEPTS, W
• Format: Journal Article
• Language: English
• Published: Alexandria, VA American Diabetes Association 01.07.1991
• Subjects: Animals; Biological and medical sciences; Cell Aggregation; Cell Separation - methods; Cells, Cultured; Diabetes Mellitus, Experimental - surgery; Islets of Langerhans - cytology; Islets of Langerhans - ultrastructure; Islets of Langerhans Transplantation - methods; Islets of Langerhans Transplantation - pathology; Medical sciences; Microscopy, Electron; Rats; Rats, Inbred Strains; Surgery (general aspects). Transplantations, organ and tissue grafts. Graft diseases; Surgery of endocrine glands; Time Factors; Transplantation, Isogeneic; Endocrinopathy; Methodology; Rat; Diabetes mellitus; Rodentia; Langerhans islet; Homograft; Vertebrata; Experimental disease; Mammalia; Animal; Surgery; Technique; Graft surgical; Streptozotocin
• ISSN: 0012-1797; 1939-327X
• DOI: 10.2337/diab.40.7.908

A standardized procedure was developed for the preparation of rat islet cell grafts with selected cell number and composition. After collagenase digestion of pancreases and elutriation of tissue fragments, islets were isolated and dissociated, and cells were purified by autofluorescence-activated cell sorting. Approximately 30% of the initial beta-cell mass was lost during digestion and elimination of small mostly exocrine particles. Fifty percent was recovered in isolated islet preparations and 30% in the purified beta-cell suspensions of greater than 95% purity and viability. Sorting according to cellular flavin adenine dinucleotide content discriminated islet beta-cells from islet endocrine non-beta-cells, fibroblasts, leukocytes, and exocrine cells. Purified endocrine islet cell grafts were prepared by aggregating 10(6) pure beta-cells with or without 8 x 10(5) pure endocrine non-beta-cells. In contrast to intact islets, the purified aggregates were devoid of nonendocrine and damaged cells. Intraportal implantation of a pure beta-cell graft rapidly and permanently normalized the diabetic state of streptozocin-administered animals. The standardized preparation of purified beta-cell grafts allows us to address several metabolic and immunological questions concerning islet cell transplantation in diabetes.