title: Glycated fetal calf serum affects the viability of an insulin-secreting cell line in vitro.
abstract: The purpose of the present study was to evaluate the direct effects of advanced glycation end products (AGEs) on beta-cells by their exposure to a glycated serum to estimate the cellular viability and the related insulin secretion. Glycation of fetal calf serum was obtained by incubation with 50 mol/L ribose at 37 degrees C for 7 days; at the end of this incubation period, the pentosidine content ranged between 15 and 16 x 10(5) pmol/L. HIT-T15 cells, a pancreatic islet cell line, were grown and cultured for 5 days in Roswell Park Memorial Institute (RPMI) medium containing either not glycated (NGS) or glycated (GS) fetal calf serum. Cellular oxidative stress (ie, thiobarbituric acid-reactive substances) was assessed by high-performance liquid chromatography. Cellular viability was evaluated by detection of proliferation, cell necrosis, and cell apoptosis rate. The insulin secretion and the related intracellular content were evaluated by enzyme-linked immunosorbent assay. The present study reported, after 5 days of exposure to the glycation environment, a moderately reduced cellular proliferation (-20.44% +/- 2.92%) with a corresponding increase of cell necrosis (+67.7% +/- 1.56%) and cell apoptosis (+39.83% +/- 2.92%) rate in comparison with the untreated cells. Oxidative intracellular stress was higher in GS conditions compared with the NGS ones (+293.3% +/- 87.53%). Insulin release from GS-treated HIT-T15 cells was lower than that of NGS-treated cells both when cells were stimulated with low glucose concentration (2.8 mmol/L, -30.3% +/- 4.91%) or when they were challenged with high glucose concentration (16.7 mmol/L, -29.2% +/- 5.82%). Incubation of HIT-T15 cells with glycated serum also caused a significant decrease of insulin intracellular content (-44.47% +/- 9.98%). Thus, AGEs were shown to exert toxic effects on insulin-secreting cells. Chronically high intracellular oxidative stress, due to accumulation of AGEs, affects the insulin secretion machinery. The present data suggest a pivotal role of the non-enzymatic glycation process in the onset and progression of diabetes during aging and a direct adverse effect of a glycated environment on the pancreatic islet cells.
authors.affiliation: Department of Internal Medicine and Medical Specialties, Gerontology and Geriatrics, 16132 Genova, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Luciano Viviani
authors.forename: Alessandra
authors.initials: A
authors.lastname: Puddu
authors.forename: Gianni
authors.initials: G
authors.lastname: Sacchi
authors.forename: Anna
authors.initials: A
authors.lastname: Garuti
authors.forename: Daniela
authors.initials: D
authors.lastname: Storace
authors.forename: Arianna
authors.initials: A
authors.lastname: Durante
authors.forename: Fiammetta
authors.initials: F
authors.lastname: Monacelli
authors.forename: Patrizio
authors.initials: P
authors.lastname: Odetti
mesh_terms: D000367:Age Factors; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D002417:Cattle; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D017127:Glycation End Products, Advanced / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives / Q000494:pharmacology; D018384:Oxidative Stress / Q000502:physiology; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism
chemical_list: D017127:Glycation End Products, Advanced; D007328:Insulin; D017392:Thiobarbituric Acid Reactive Substances; D001120:Arginine; C062187:pentosidine; D008239:Lysine