Function of microdissected pancreatic islets cultured in a chemically defined medium. I. Insulin content and release

Abstract

Summary. Microdissected pancreatic islets from non-inbred ob/ ob-mice were cultured for 6 or 7 days in serum-free tissue culture medium 199. The insulin content of the islets decreased 60% during culture in 17 mM or 28 mM glucose and about 70% in the presence of 3.3 mM or 5.6 mM glucose. At the end of a culture period in high glucose, the sum of the insulin in the islet plus that in the culture medium was almost twice as high as the insulin content of fresh islets, indicating an active insulin biosynthesis. The maximal insulin response to glucose after culture in 17 mM or 28 mM glucose was about 40% of that in fresh islets; after culture in 3.3 mM glucose it was 10%. Half-maximal stimulation was observed at a glucose concentration of 5 mM for islets cultured with high glucose as compared to 9 mM for fresh islets. Like glucose, glibenclamide was a more effective insulin stimulator after culture with a high glucose concentration than with a low one. However, leucine-induced insulin release was not affected by the glucose concentration in the preceding culture medium. Whereas potentiation of glucose-stimulated release by arginine or dibutyryl-cAMP was independent of glucose concentration during the culture, theophylline released three times more insulin when the islets had been cultured with high glucose.

Maintenance of insulin release from pancreatic islets stored in the cold for up to 5 weeks.

“…The present studies confirm the findings of other authors (1,14) that storage in high-glucose media for 1 wk at 37WC is associated with synthesis and release of 1 1 insulin during this period, and with a fall of islet insulin content. We also confirm (1) that storage at 370C in a high-glucose medium maintains a greater insulin response to glucose than storage in a low-glucose medium.…”

“…The relatively complex TCM 199, rather than only Hanks' balanced salt solution, may have provided some necessary nutritional requirement. In addition, hand-dissection instead of collagenase isolation, which may damage the islets during long-term storage in the absence of serum (1), or the use of 8°instead of 4VC, may have been of importance.…”

“…Culture of islets at 370C has been shown to result in a decreased ability to release insulin in response to glucose stimulation (1,2). The use of low temperature in the preservation of kidneys (3), as well as other tissues (4)(5)(6), for transplantation has been extensively studied and found to maintain tissue function for a longer time than storage at warmer temperatures.…”

Cyclic Adenosine-3′,5′ -monophosphate Stimulates Islet B Cell Replication in Neonatal Rat Pancreatic Monolayer Cultures

“…Measurement of cAMP levels in the cultures revealed dissociations between effects on B cell replication and insulin release. Thus, addition of 0.1 mM IBMX, or 0.1 nM cholera toxin, to 5.6 mM glucose produced slightly greater increases in cAMP levels and B cell replication than did 16.7 mM glucose, whereas insulin release was increased significantly more with 16 (15-20 mM) has been shown to increase glucose metabolism (1, 2), insulin release and content (3)(4)(5)(6), and insulin biosynthesis (7), and also to improve the survival (6) and to stimulate the replication (8)(9)(10)(11) of islet B cells. Glucose has also been reported to stimulate adenylate cyclase (5,12) and to increase cyclic adenosine-3',5'-monophosphate (cAMP) levels (13) Green ancd Taylor (26).…”

Effect of nutrients, hormones and serum on survival of rat islet beta cells in culture

“…Various media have been used to maintain islet beta-cell preparations in culture. Previous papers have reported how the medium content in serum, glucose and hormones may influence the function of islets and islet cell lines that are recovered from culture [1][2][3][4][5]. Information is scarce on the extent of cell losses under different culture conditions.…”