Simple or still not discovered TOM complex of the amoeba Acanthamoeba castellanii

Abstract

concentration of FFAs than BTC-6, due to the increased sensitivity of these cells. Mitochondrial function was monitored by measurements of the mitochondrial oxygen consumption (high-resolution respirometry), ATP content (PerkinElmer), mitochondrial membrane potential [flow cytometry (BD)], activity of the caspase-9 (R&D Systems) and ROS generation (DCFH-DA). In preadipocyte cell line: palmitic acid impaired mitochondrial function that was manifested by a decreased intracellular ATP content and an increased ROS generation. Arachidonic acid increased ROS generation and caspase-9 activity related with the mitochondrial apoptosis pathway. Eicosapentaenoic acid decreased mitochondrial respiration and ATP content. In BTC6 line increased the amount of early apoptotic cells after incubation with AA and EPA was observed. There was also a tendency to increased caspase-9 activity after incubation with FFAs. Arachidonic acid slightly increased mitochondrial respiration in beta cells. In conclusion, free fatty acids affect mitochondrial function of selected adipose and pancreatic cell lines which is interesting in the light of a recent data, indicating that regulation of mitochondrial activity may be the target pathway for treating obesity.Systemic energy metabolism requires coordination of different organs to maintain homeostasis and overcome physiological and stress challenges. Skeletal muscle is a major metabolic organ that responds to energy shortage. Here, we show that nutrient deprivation of cultured muscle cells (C2C12) changes their bioenergetic profile. Differentiated myotubes were deprived of glucose and lglutamine for 30 min up to 24 h. Plate-based respirometry showed that the lack of glycolysis was first compensated by an increased mitochondrial ATP turnover. In the sequelae of nutrient deprivation, mitochondrial respiration and ATP production were progressively compromised until reaching levels below reliable detection after 24 h. After 8 h of nutrient deprivation, respiration linked to ATP synthesis was decreased to 36% of non-fasted controls. Fibroblast Growth Factor 21 (FGF21), previously shown to be induced upon a reduced mitochondrial energy output in muscle mediating endocrine crosstalk between organs, increased mRNA levels by 13-fold and the secretion of FGF21 protein by 5-fold. Under these conditions, we assessed the protein secretome unbiased using mass spectrometric analysis. In the complex secretome, we found the highest released protein to be FGF21, confirming the suitability of our analysis to discover novel players mediating fasting induced systemic crosstalk. We identified a total of 1498 proteins; 93 of these were significantly induced by starvation, of which 21 were previously published to be secreted and thus represent potential myokines. While FGF21 regulates carbohydrate and lipid metabolism, other novel proteins may display similar effects to maintain systemic homeostasis and will be explored in vivo, offering novel therapeutics to treat metabolic diseases.