Production Characterization and Function of Pseudoislets from PerinatalCanine Pancreas

摘要

We evaluated the cell composition and function of canine pancreatic pseudoislets (PIs)produced from 42- to 55-day-old fetuses, 1- to 21-day-old pups, and an adult dog pancreas.After mild collagenase treatment, partially digested tissues were cultured for 2–3 weeks.PI production started on culture day 3, was marked for 6 to 9 days, and then stopped. PIproduction was greatest with the neonatal specimens, reaching about 12 million aggregatesper litter (55-day-old fetus) or per pancreas (1-day-old pup). Cell composition at allstages was similar to that in adult pancreatic islets, with predominant β cells, scant αcells and, most importantly, presence of δ cells. Among pancreatic markers assessed byquantitative real-time PCR (qRT-PCR) mRNA assay, insulin showed the highest expressionlevels in PIs from newborn and adult pancreas, although these were more than 1000 timeslower than in adult islets. Pdx1 mRNA expression was high in PIs from 55-day-oldpancreases and was lower at later stages. Consistent with the qRT-PCR results, the insulincontent was far lower than reported in adult dog pancreatic islets. However, insulinrelease by PIs from 1-day-old pups was demonstrated and was stimulated by a high-glucosemedium. PIs were transplanted into euglycemic and diabetic SCID mice. In euglycemicanimals, the transplant cell composition underwent maturation and transplants were stillviable after 6 months. In diabetic mice, the PI transplants produced insulin and partiallycontrolled the hyperglycemia. These data indicate that PIs can be produced ex vivo fromcanine fetal or postnatal pancreases. Although functional PIs can be obtained, theproduction yield is most likely insufficient to meet the requirements for diabetic dogtransplantation without further innovation in cell culture amplification.