Long-term culture of human pancreatic islets in an extracellular matrix: morphological and metabolic effects.
In this experiment, various conditions for embedding cultures of human pancreatic islets in type I collagen gel were studied in an attempt to maintain the highly differentiated functions of islet cells and particularly insulin secretion over a long period of time. The islets isolated by a collagenase digestion technique were plated either on or within the collagen gel and refed with either Eagle's minimum essential medium (5.5 mM D-glucose) or RPMI 1640 medium (11 mM D-glucose) supplemented with 10% FCS and antibiotics. The comparison between the two culture media showed that embedded islets cultured in RPMI had a higher basal insulin secretion rate, survived longer than their MEM counterparts, but exhibited impaired response to an acute glucose test contrasting thus with islets cultured in MEM. The secretory behaviour of islets was also related to the different morphological modifications occurring during culture. Islets directly embedded within the collagen gel more or less maintained their spherical structure and highest secretory capacities. When overlaid with a second layer of collagen, well established monolayers of human islet cells grown on collagen underwent a gradual and complete reorganization into a three-dimensional islet-like structure with a striking reinforcement of their secretory activity. Both cultures were able to survive more than 8 weeks, thus proving the usefulness of such a new model for long-term culture. In contrast, standard cultures on culture treated plastic dishes on which islets cells rapidly established wide monolayers, exhibited a rapid and definitive decline in insulin secretion with a survival not exceeding 14 days. In the light of these different culture conditions, possible mechanisms responsible for disturbance of hormonal release and their implications for in-vitro study of isolated islets functions are discussed. In conclusion, this work is a new example of the permissive effects of collagen matrices on the establishment or maintenance of tissue-like structures in vitro, suggesting the definition of a new model for the study of human pancreatic islets in long-term culture.
Lucas-Clerc
C
C
Laboratoire de Biochimie Médicale A, UER Médicale et Pharmaceutique, Rennes, France.
Massart
C
C
Campion
J P
JP
Launois
B
B
Nicol
M
M
D000328:Adult; D001120:Arginine / Q000494:pharmacology; D002470:Cell Survival; D002478:Cells, Cultured / Q000187:drug effects; D003094:Collagen; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D046508:Culture Techniques / Q000379:methods*; D005109:Extracellular Matrix; D005782:Gels; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D013997:Time Factors
D016895:Culture Media, Serum-Free; D005782:Gels; D007328:Insulin; D003094:Collagen; D001120:Arginine; D005947:Glucose