Organ Culture of Fetal Mouse Pancreas: The Effect of Culture Conditions on Insulin and Glucagon Secretion

Abstract

Seventeen-day-old mouse pancreas was grown in organ culture under various culture conditions and the insulin released into the medium, either chronically or during acute incubations, was measured. Three media, DME, RPMI 1640, and TC199, were tested, each at glucose concentrations of 1, 2, 3, and 4 g/L. Fetal pancreata maintained in RPM11640 at 4 g/L glucose secreted the most insulin over a 25-day period. Tissue grown in RPMI 1640 and DME secreted more insulin in higher glucose concentrations, but explants grown in TC199 secreted low levels of insulin regardless of glucose concentration. Fetal pancreata grown in medium supplemented with 5% FCS secreted less insulin in acute incubations than tissues grown in 10%, 15%, 20%, or 30% FCS, which secreted equivalent amounts. There was no difference in the amount of insulin released by islets grown in media supplemented with 15% FCS from four different batches.Although fetal islets grown in high glucose concentrations generally secreted more insulin (and less glucagon) than "normoglycemic" controls, their ability to release insulin in subsequent acute incubations was impaired. Explants, which had been maintained for 2 wk in 2.5 or 4 g/L glucose and a further 3 days in normoglycemic medium, secreted less insulin and glucagon in acute release exeriments than tissue maintained in 1 g/L glucose throughout.Light microscopy of cultured fetal pancreas showed that tissue remained viable at all glucose concentrations tested and differences in response could not be attributed to selective necrosis. Tranplantation of cultured islets indicated that tissue grown under normoglycemic conditions functioned better than tissue grown in hyperglycemic media. DIABETES 30:804-812, October 1981.

A Comparison of Portal Versus Systemic Venous Drainage in Murine Foetal Pancreatic Islet Transplantation

“…Four age, sex and strain matched mice served as controls. Three weeks after STZ injection the diabetic animals were grafted with one half of one 17-day gestation foetal pancreas which had been in organ culture for 14 days (Collier et al, 1981). The grafts were placed either under the left renal capsule (systemic drainage) or on to the hilar surface of the spleen (portal drainage) (Mandel, Hoffman and Carter, 1981).…”

Effects of Stimulation of Adenylate Cyclase and Protein Kinase-C on Cultured Fetal Rat B-Cells*

“…Cultures of fetal islets have at least one advantage over these in vivo experiments. They make it possible to study separately the putative factors that may be involved in the maturation of B-cells (9)(10)(11)(12)(13)(14). A high concentration of glucose was found to be a necessary but insufficient factor to achieve full secretory maturation of fetal Bcells in some (13,14), but not all, studies (15).…”

Detrimental Effect of High Medium Glucose Concentration on Subsequent Endocrine Function of Transplanted Organ‐cultured Foetal Mouse Pancreas

“…In previous studies we showed that foetal islets grown in high concentrations of glucose secreted more insulin into the medium than islets grown in physiological concentrations, but they responded poorly to an acute hyperglycaemic stimulus (Collier et al, 1981). In a separate preliminary study (Mandel et al, 1980) we suggested that islets grown in "hyperglycaemic' media functioned poorly as grafts in diabetic hosts when compared with islets cultured Fig.…”

“…Pancreases were dissected from 17-day CKA/H/WEHI foetuses and each was explanted on to a piece of millipore filter (0 45 jum, Millipore Corporation, Bedford, Massachusetts) which was placed in a tube containing 1ml of medium supplemented with 15% foetal calf serum (FCS) so that the explants were at tlie cas-medium inlerface, as previously described (Collier et al, 1981). There were 4 culture groups: Dulbecco's modified Eagles (DME) medium with I g/1 glucose; DME with 4 g/1 glucose: RPMI 1640 wilh 1 g/1 glucose and RPMI 1640 with 4 g/1 glucose, with 9 replicates in each group.…”