INSULIN RELEASE AND INSULIN MESSENGER-RNA LEVELS IN RAT ISLETS OF LANGERHANS CULTURED ON EXTRACELLULAR-MATRIX

Primary culture of rat islets of Langerhans lose glucose responsivenes s and eventually die when cultured for a long period of time, In this study we evaluated the effect of matrigel, a basement membrane extract , on (i) islet cell survival, (ii) cell responsiveness following a glu cose challenge, and (iii) mRNA levels for insulin, glucagon, and somat ostatin. Pancreatic islets were isolated by collagenase digestion and plated in culture dishes either coated or not with a matrigel layer. U sing the reverse hemolytic plaque assay, we determined the total numbe r of insulin-secreting cells and the amount of insulin secreted by ind ividual beta cells, After 1 h of exposure to 5 mM glucose, beta cells from 6-month-old rat islets cultured for 6 weeks on matrigel showed an equal number of insulin-secreting cells compared to freshly isolated islets cultured for only 3 days in the absence of matrigel (39.5 +/- 2 .5 vs, 37.1 +/- 2.6%). Furthermore, the release of insulin by cells cu ltured on matrigel for 6 weeks increased in a glucose-dependent manner (p < 0.001) and showed an ED(50) of 7 mM. However, the amount of insu lin released per single beta cell was reduced by 40-60% (p < 0.02) com pared to that released from isolated beta cells derived from a 3-day c ulture of islets, Finally, there was a 35-55% increase (p < 0.05) in t he levels of insulin, glucagon, and somatostatin mRNAs in cells cultur ed for 6 weeks on matrigel. These data suggest a trophic effect of mat rigel on the maintenance of normal beta-cell activity and function aci d may lead the way to the development of a new model for the study of pancreatic islets in long-term culture.