Insulin release and insulin mRNA levels in rat islets of Langerhans cultured on extracellular matrix

• Published in: Pancreas Vol. 13; no. 1; p. 47
• Main Authors: Perfetti, R, Henderson, T E, Wang, Y, Montrose-Rafizadeh, C, Egan, J M
• Format: Journal Article
• Language: English
• Published: United States 01.07.1996
• Subjects: Animals; Base Sequence; Collagen; Culture Techniques - methods; DNA, Complementary - genetics; Drug Combinations; Extracellular Matrix; Glucagon - genetics; Insulin - genetics; Insulin - metabolism; Insulin Secretion; Islets of Langerhans - metabolism; Laminin; Male; Proteoglycans; Rats; Rats, Wistar; RNA, Messenger - genetics; RNA, Messenger - metabolism; Somatostatin - genetics
• ISSN: 0885-3177
• DOI: 10.1097/00006676-199607000-00006

Primary culture of rat islets of Langerhans lose glucose responsiveness 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 glucose challenge, and (iii) mRNA levels for insulin, glucagon, and somatostatin. Pancreatic islets were isolated by collagenase digestion and plated in culture dishes either coated or not with a matrigel layer. Using the reverse hemolytic plaque assay, we determined the total number of insulin-secreting cells and the amount of insulin secreted by individual 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 cultured on matrigel for 6 weeks increased in a glucose-dependent manner (p < 0.001) and showed an ED50 of 7 mM. However, the amount of insulin released per single beta cell was reduced by 40-60% (p < 0.02) compared to that released from isolated beta cells derived from a 3-day culture of islets. Finally, there was a 35-55% increase (p < 0.05) in the levels of insulin, glucagon, and somatostatin mRNAs in cells cultured for 6 weeks on matrigel. These data suggest a trophic effect of matrigel on the maintenance of normal beta-cell activity and function and may lead the way to the development of a new model for the study of pancreatic islets in long-term culture.