Targeting mechanisms to better understand the improved lipid status of mild hyperbilirubinemia, an in vitro approach

Autoren:Lang, Claudia; Hirtl, Yvonne; Eder, Paulina; Jasprová, Jana (Charles University Prague); Vítek, Libor (Charles University Prague); Schachner, Daniel; Heiß, Elke; Wagner, Karl-Heinz
Abstrakt:Targeting mechanisms to better understand the improved lipid status of mild hyperbilirubinemia, an in vitro approach Lang CA1*, Hirtl Y1, Eder P1, Jašprová J2, Vitek L2, and Wagner KH1 (1) Department of Nutritional Sciences, University of Vienna, (2) Institute of Medical Biochemistry and Laboratory Medicine, Charles University Prague Gilbert’s Syndrome individuals (GS) exhibit mildly elevated blood levels of the bile pigment bilirubin in its unconjugated form (UCB). Previous phenotypic observations revealed that GS are metabolically healthier than controls based on investigations of the anthropometric, blood lipid and glucose profile [1]. Recently our group provided evidence of increased metabolic activity in PBMCs of GS through phospho-AMPKα1/α2-& PPARα/γ-activation [2]. A highly active metabolism would indeed promote cellular lipid turnover which could lower the risk of chronic diseases. This work explores the impact of UCB in lipid metabolism and characterises whether UCB increases lipid accumulation in vitro. C2C12 mouse muscle cells were treated analogously to physiological concentrations with 17.1µM UCB, 1mM free fatty acids (FFA) and with high (450mg/dL) or no glucose. As positive control 0.5mM AICAR, an AMP analogue was used. Cytotoxicity was analysed via the MTT assay and intracellular UCB levels by HPLC. Lipid accumulation into cells was investigated with the lipophilic fluorescence dye Nile Red by plate reader assay and fluorescence microscopy. Intracellular lipid content in UCB treated muscle cells (+UCB+FFA) compared to negative controls (-UCB+FFA) decreased significantly in glucose-starved culture and by trend at hyperglycemic conditions. However only a minor decline of intracellular lipids was shown highlighting the necessity to investigate fatty acid uptake separately from lipid turnover by more sensitive methods such as GC and C14 palmitate uptake assay. We also showed that UCB and FFA are not toxic to C2C12 cells at given concentrations and that UCB is taken up by cells during incubation. Reference: [1] WAGNER KH, WALLNER M, MÖLZER C, et al, Looking to the horizon: the role of UCB in the development and prevention of age-related chronic diseases. Clinical Science 2015; 129: 1-25 [2] MÖLZER C, WALLNER M, KERN D, et al, Features of an altered AMPK metabolic pathway in Gilbert’s Syndrome, and its role in metabolic health. Scientific reports 2016; 6: 30051 *Lang Claudia Anna,