Session: SUN 649-677-Adipocyte Biology
Poster Board SUN-650
Methods: Adipose differentiation was determined by Nile Red lipid staining and cell counting. To determine gene expression quantitative PCR was used. Lipolysis rates were measured by glycerol release after stimulation with isoproterenol. Insulin stimulated glucose uptake was quantified by measuring 14C-labelled 2-deoxyglucose uptake. PI3K (Phosphoinositide-3-kinase) activity was determined by applying FRAP (fluorescence redistribution after photobleaching)/TIRF (total internal reflection fluorescence) analysis.
Results: LipPD1 cells preserved a capacity for adipocyte differentiation of 55.1±4.2% for 29 population doublings. The expression of the adipocyte markers PPARy, FASN, adiponectin and AP2 (FABP4) during differentiation tended to be increased in LipPD1 compared to SGBS. Functional analysis revealed no significant differences between LipPD1 and SGBS in 2-deoxyglucose uptake after insulin stimulation. Lipolysis was activated by isoproterenol and not significantly different in LipPD1 compared to SGBS adipocytes. A fluorescent biomarker, reflecting PI3K activity by redistribution to the plasma membrane, was found membrane-associated to a significantly higher amount in LipPD1 (33.4±2.7%) compared to SGBS adipocytes (20.9±2.3%). A constitutive activation of the kinase AKT was verified by detecting significantly increased Ser473 and Thr308 phosphorylation in LipPD1 cells compared to SGBS. AKT phosphorylation was further enhanced after stimulation with IGF-I.
Conclusion: LipPD1 cells have increased PI3K/AKT activity compared to SGBS. In regard of their adipocyte functions LipPD1 behave like SGBS adipocytes and are a suitable model to investigate molecular mechanisms of adiposity.
Nothing to Disclose: WK, FK, GS, KL, AT, FKW, AAG, AK
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