Adverse physicochemical properties of tripalmitin in beta cells lead to morphological changes and lipotoxicity in vitro.
OBJECTIVE
Long-term exposure of beta cells to lipids, particularly saturated fatty acids in vitro, results in cellular dysfunction and apoptosis (lipotoxicity); this could contribute to obesity-related diabetes. Our aims were to relate cell death to intracellular triglyceride concentration, composition and localisation following incubation of INS1 cells in saturated and unsaturated NEFA in high and low glucose concentrations.


METHODS
Insulin-producing INS1 cells were cultured (24 h; 3 and 20 mmol/l glucose) with palmitic, oleic or linoleic acids and the resulting intracellular lipids were analysed by gas chromatography and microscopy. Cell death was determined by quantitative microscopy and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and glucose-stimulated insulin secretion by ELISA.


RESULTS
All NEFA (0.5 mmol/l, 0.5% albumin) inhibited glucose-stimulated (20 mmol/l) insulin secretion. Cytotoxicity was evident only with palmitic acid (p<0.05), in which case intracellular triglyceride consisted largely of tripalmitin in angular-shaped dilated endoplasmic reticulum. Cytotoxicity and morphological disruption were reduced by addition of unsaturated NEFA. Triglyceride content (control cells; 14.5 ng/mug protein) increased up to 10-fold following incubation in NEFA (oleic acid 153.2 ng/mug protein; p<0.05) and triglyceride and phospholipid fractions were both enriched with the specific fatty acid added to the medium (p<0.05).


CONCLUSIONS
In INS1 cells, palmitic acid is converted in the endoplasmic reticulum to solid tripalmitin (melting point >65 degrees C), which could induce endoplasmic reticulum stress proteins and signal apoptosis; lipid-induced apoptosis would therefore be a consequence of the physicochemical properties of these triglycerides. Since cellular triglycerides composed of single species of fatty acid are not likely to occur in vivo, destruction of beta cells by saturated fatty acids could be predominantly an in vitro scenario.
Moffitt
J H
JH
Oxford Centre for Diabetes, Endocrinology and Metabolism, Churchill Hospital, Oxford, OX3 7LJ, UK.
Fielding
B A
BA
Evershed
R
R
Berstan
R
R
Currie
J M
JM
Clark
A
A
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D019556:COS Cells; D045744:Cell Line, Tumor; D002470:Cell Survival; D002522:Chlorocebus aethiops; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007340:Insulinoma; D019787:Linoleic Acid / Q000378:metabolism; D051379:Mice; D019301:Oleic Acid / Q000378:metabolism; D019308:Palmitic Acid / Q000378:metabolism; D010190:Pancreatic Neoplasms; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D014280:Triglycerides / Q000737:chemistry* / Q000378:metabolism / Q000633:toxicity*
D005230:Fatty Acids, Nonesterified; D007328:Insulin; D010743:Phospholipids; D014280:Triglycerides; D019301:Oleic Acid; D019308:Palmitic Acid; D019787:Linoleic Acid; C005859:tripalmitin; D005947:Glucose