title: Fluorescence-activated cell sorted rat islet cells and studies of the insulin secretory process.
abstract: Studies of individual cell types in the islets of Langerhans are complicated by the cells' functional coupling by gap junctions and paracrine interaction. Access to purified alpha and beta cells is therefore desirable. We present a simplified and optimized method for fluorescence-activated cell sorting of endocrine pancreatic rat islets. For dispersion of the islets, dispase was superior to trypsin, as the number of vital single cells was higher (1.1 +/- 0.1 x 10(3) vs 0.6 +/- 0.1 x 10(3)/islet, P < 0.05). The purity of the sorted cells was 96.7 +/- 1.2% for the non-beta cells and 97.8 +/- 0.6% for the beta cells (numbers in percentages of endocrine cells). In culture, isolated beta cells, non-beta cells and mixtures of beta and non-beta cells formed aggregates, but not at low temperature (4 degrees C) and not in medium with low serum content (2%). Finally, in pure beta cell aggregates, glucose stimulated changes in cytoplasmic free Ca2+ concentration although both glucose- and arginine-induced insulin secretion was much reduced. We conclude that alpha cells are necessary for insulin secretion but not for glucose sensing.
authors.affiliation: Bartholin Instituttet, Kommunehospitalet, Copenhagen K, Denmark.
authors.forename: K
authors.initials: K
authors.lastname: Josefsen
authors.forename: J P
authors.initials: JP
authors.lastname: Stenvang
authors.forename: H
authors.initials: H
authors.lastname: Kindmark
authors.forename: P O
authors.initials: PO
authors.lastname: Berggren
authors.forename: T
authors.initials: T
authors.lastname: Horn
authors.forename: T
authors.initials: T
authors.lastname: Kjaer
authors.forename: K
authors.initials: K
authors.lastname: Buschard
mesh_terms: D000818:Animals; D002469:Cell Separation; D002478:Cells, Cultured; D005434:Flow Cytometry; D005934:Glucagon / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008854:Microscopy, Electron; D051381:Rats; D011919:Rats, Inbred Strains; D013997:Time Factors
chemical_list: D007328:Insulin; D005934:Glucagon