Effects of adipocytokines and free fatty acids on viability and apoptosis of rat INS-1E β-cells

R Spinnler; T Gorski; S Schuster; A Garten; S Laue; J Breitfeld; J Kratzsch; A Beck-Sickinger; A Körner; W Kiess

doi:10.1055/s-0031-1277466

Diabetologie und Stoffwechsel, Table of Contents

Effects of adipocytokines and free fatty acids on viability and apoptosis of rat INS-1E β-cells

R Spinnler ¹^,², T Gorski ¹, S Schuster ¹, A Garten ¹, S Laue ¹, J Breitfeld ³, J Kratzsch ⁴, A Beck-Sickinger ⁵, A Körner ¹, W Kiess ¹

¹University Hospital for Children & Adolescents, Leipzig, Germany
²Leipzig University Medical Center, IFB Adiposity Diseases, Leipzig, Germany
³University of Leipzig, Interdisciplinary Centre for Clinical Research, Faculty of Medicine, Leipzig, Germany
⁴University Hospital Leipzig, Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostic, Leipzig, Germany
⁵University of Leipzig, Institute of Biochemistry, Leipzig, Germany

Abstract

Background and aims: The molecular interactions between adipose tissue and β-cells remain to be elucidated. Obesity is associated with a dysregulation of adipocyte secretory function. We asked, whether the adipocytokines, leptin, adiponectin, Nampt and vaspin, or the free fatty acids palmitate and oleate directly influence INS-1E β-cell survival and function.

Methods: Changes in viability of INS-1E cells were detected by WST-1 assay. Adenylate kinase release was determined to assess cytotoxicity. Apoptosis was measured by Annexin V/PI Assay. For further characterization of activated apoptosis signaling pathways, Western blotting of p53, caspase-3 and NF-κB was performed. Insulin secretion was measured by ELISA.

Results: A dose-dependent increase (98±6.4%) in insulin secretion was detected after stimulation with 20mM glucose. Viability was significantly decreased by 91±6.2% after stimulation with IL-1β[10ng/ml], 46±7.8% by TNFα[10ng/ml] and 26.2±6.1% by IFNγ[10ng/ml]. Cytotoxicity was increased by 40% after stimulation with a cytokine combination (IL-1β+IFN-γ[10ng/ml+10ng/ml]). In contrast, adipocytokines showed no effect on viability and cytotoxicity. Palmitate induced cytotoxicity by 70%, oleate by 40% and a palmitate/oleate combination by 20%. Apoptosis was increased after stimulation with a cytokine combination (IL-1β+IFN-γ[10ng/ml+10ng/ml]) by 63±9% and with palmitate[0.5mM] by 20±3.6%. Oleate[0.5mM] and a palmitate/oleat combination[0.25mM+0.25mM] decreased apoptosis by 20±4%. In contrast and most importantly, adipocytokines showed no effects on apoptosis. In addition, adipocytokines did not protect against cytokine-induced apoptosis. Furthermore, no activation of apoptosis signaling pathways were found after stimulation with adipocytokines.

Conclusion: According to our results, leptin, adiponectin, Nampt and vaspin do not induce β-cell death, nor do they influence viability and cytotoxicity in the rat β-cell model INS-1E. However, free fatty acids, particularly palmitate, exert direct effects on β-cell survival.