Background: Exposure to intermittent hypoxia (IH) may play a role in the development of metabolic impairments in the context of obstructive sleep apnea syndrome, probably by elevated plasma levels of free fatty acids. Employing gas-permeable cultureware to grow differentiated human and mouse adipocytes in vitro, we directly studied the effects of pericellular oxygen fluctuations on key adipocyte metabolic functions-spontaneous lipolytic rates, triglyceride accumulation, de novo lipogenesis, and expression of adipocyte-specific marker genes. Materials and Methods: 3T3-L1 fibroblasts and human subcutaneous preadipocytes were differentiated under conditions that induced repetitive pericellular-oxygen cycles IH between 1% O2 (5 min) and 16% O2 (5 min), continuously for 14 days or under control conditions. Chemicals were used to inhibit the flux of acetyl-CoA from glycolysis (alfa-cyano-4-hydroxy cinnamate) or the tricarboxylic acid cycle (SB204990), or to stimulate the flux of acetyl-CoA from pyruvate to the lipogenic pool. Lipolytic rate, intracellular lipids, and expression of adipocyte differentiation markers were assessed and t-test or ANOVA were used to find significant differences. Results: The rate of lipolysis increased by 211% in 3T3-L1 cells and by 39% in obese human adipocytes. Exposure to IH reduced intracellular lipid stores by 37% and reduced the expression of adipocyte differentiation markers. Pharmacological stimulation or inhibition of de novo lipogenesis did not modify the intracellular lipid content under IH. Conclusions: Pericellular oxygen fluctuations directly stimulated lipolysis, but did not increase de novo lipogenesis from endogenous substrates. Similarly, IH hampered adipocyte differentiation from precursors.
- MeSH
- acetylkoenzym A metabolismus MeSH
- buněčná diferenciace genetika fyziologie MeSH
- buňky 3T3-L1 MeSH
- citrátový cyklus MeSH
- glykolýza MeSH
- hypoxie buňky genetika fyziologie MeSH
- kinetika MeSH
- lidé MeSH
- lipogeneze genetika fyziologie MeSH
- lipolýza genetika fyziologie MeSH
- myši MeSH
- spotřeba kyslíku genetika MeSH
- stanovení celkové genové exprese MeSH
- triglyceridy metabolismus MeSH
- tukové buňky metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Obstructive sleep apnea (OSA) is associated with insulin resistance, glucose intolerance, and type 2 diabetes. Causal mechanisms mediating this association are not well defined; however, augmented lipolysis in adipose might be involved. Here, we investigated the effect of acipimox treatment (lipolysis inhibitor) on glucose tolerance and insulin sensitivity in mice exposed to intermittent hypoxia (IH). C57BL6/J mice were exposed for 14 days to IH or control conditions. IH was created by decreasing the fraction of inspired oxygen from 20.9 to 6.5%, 60 times/h. Control exposure was air (fraction of inspired oxygen, 20.9%) delivered at an identical flow rate. Acipimox was provided in drinking water (0.5 g/ml) during exposures. After exposures, intraperitoneal insulin (0.5 IU/kg) and glucose (1 g/kg) tolerance tests were performed, and primary adipocytes were isolated for lipolysis experiments. IH elevated fasting glucose by 51% and worsened glucose tolerance and insulin sensitivity by 33 and 102%, respectively. In parallel, IH increased spontaneous lipolysis by 264%, and reduced epididymal fat mass by 15% and adipocyte size by 8%. Acipimox treatment prevented IH-induced lipolysis and increased epididymal fat mass and adipocyte size by 19 and 10%, respectively. Acipimox fully prevented IH-induced impairments in fasting glycemia, glucose tolerance, and insulin sensitivity. For all reported results, P less than 0.05 was considered significant. Augmented lipolysis contributes to insulin resistance and glucose intolerance observed in mice exposed to IH. Acipimox treatment ameliorated the metabolic consequences of IH and might represent a novel treatment option for patients with obstructive sleep apnea.
- MeSH
- adipozita účinky léků MeSH
- experimentální diabetes mellitus krev komplikace genetika patologie MeSH
- fenotyp MeSH
- glukosa metabolismus MeSH
- hypoxie komplikace metabolismus MeSH
- inzulin metabolismus MeSH
- lipolýza * účinky léků MeSH
- mastné kyseliny krev MeSH
- modely nemocí na zvířatech MeSH
- myši inbrední C57BL MeSH
- obstrukční spánková apnoe krev komplikace genetika patologie MeSH
- pyraziny farmakologie MeSH
- regulace genové exprese účinky léků MeSH
- signální transdukce účinky léků MeSH
- tukové buňky účinky léků patologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Methyltriphenylphosphonium (TPMP) salts have been widely used to measure the mitochondrial membrane potential and the triphenylphosphonium (TPP+) moiety has been attached to many bioactive compounds including antioxidants to target them into mitochondria thanks to their high affinity to accumulate in the mitochondrial matrix. The adverse effects of these compounds on cellular metabolism have been insufficiently studied and are still poorly understood. Micromolar concentrations of TPMP cause a progressive inhibition of cellular respiration in adherent cells without a marked effect on mitochondrial coupling. In permeabilized cells the inhibition was limited to NADH-linked respiration. We found a mixed inhibition of the Krebs cycle enzyme 2-oxoglutarate dehydrogenase complex (OGDHC) with an estimated IC50 3.93 [3.70-4.17] mM, which is pharmacologically plausible since it corresponds to micromolar extracellular concentrations. Increasing the lipophilic character of the used TPP+ compound further potentiates the inhibition of OGDHC activity. This effect of TPMP on the Krebs cycle ought to be taken into account when interpreting observations on cells and mitochondria in the presence of TPP+ derivatives. Compounds based on or similar to TPP+ derivatives may also be used to alter OGDHC activity for experimental or therapeutic purposes.
- MeSH
- buněčné linie MeSH
- citrátový cyklus účinky léků MeSH
- citrátsynthasa účinky léků metabolismus MeSH
- glutamátdehydrogenasa účinky léků metabolismus MeSH
- isocitrátdehydrogenasa účinky léků metabolismus MeSH
- ketoglutarátdehydrogenasový komplex antagonisté a inhibitory metabolismus MeSH
- kosterní svaly enzymologie MeSH
- krysa rodu rattus MeSH
- malátdehydrogenasa účinky léků metabolismus MeSH
- membránový potenciál mitochondrií účinky léků MeSH
- oniové sloučeniny farmakologie MeSH
- potkani Wistar MeSH
- pyruvátdehydrogenasový komplex účinky léků metabolismus MeSH
- svalové mitochondrie účinky léků enzymologie MeSH
- tritylové sloučeniny farmakologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
In-vitro investigation of the effects of hypoxia is limited by physical laws of gas diffusion and cellular O2 consumption, making prolonged exposures to stable O2 concentrations impossible. Using a gas-permeable cultureware, chronic effects of mild and severe hypoxia on triglyceride accumulation, lipid droplet size distribution, spontaneous lipolysis and gene expression of adipocyte-specific markers were assessed. 3T3-L1 cells were differentiated under 20%, 4% or 1% O2 using a gas-permeable cultureware. Triglyceride accumulation, expression of genes characteristic for advanced adipocyte differentiation and involvement of key lipogenesis enzymes were assessed after exposures. Lipogenesis increased by 375% under mild hypoxia, but dropped by 43% in severe hypoxia. Mild, but not severe, hypoxia increased formation of large lipid droplets 6.4 fold and strongly induced gene expression of adipocyte-specific markers. Spontaneous lipolysis increased by 488% in mild, but only by 135% in severe hypoxia. Inhibition of ATP-dependent citrate lyase suppressed hypoxia-induced lipogenesis by 81% and 85%. Activation of HIF inhibited lipogenesis by 59%. Mild, but not severe, hypoxia stimulates lipolysis and promotes adipocyte differentiation, probably through excess of acetyl-CoA originating from tricarboxylic acid cycle independently of HIF activation.
- MeSH
- acetylkoenzym A metabolismus MeSH
- adipogeneze účinky léků genetika MeSH
- ATP-citrát-(pro-S)-lyasa genetika metabolismus MeSH
- buněčná diferenciace účinky léků MeSH
- buňky 3T3-L1 MeSH
- citrátový cyklus účinky léků genetika MeSH
- diacylglycerol-O-acyltransferasa genetika metabolismus MeSH
- faktor 1 indukovatelný hypoxií - podjednotka alfa genetika metabolismus MeSH
- hypoxie buňky MeSH
- kyslík farmakologie MeSH
- lipidová tělíska chemie účinky léků MeSH
- lipogeneze účinky léků genetika MeSH
- lipolýza účinky léků genetika MeSH
- myši MeSH
- perilipin 1 genetika metabolismus MeSH
- proteiny vázající mastné kyseliny genetika metabolismus MeSH
- regulace genové exprese účinky léků MeSH
- signální transdukce MeSH
- sterolesterasa genetika metabolismus MeSH
- triglyceridy metabolismus MeSH
- tukové buňky cytologie účinky léků metabolismus MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Publikační typ
- abstrakt z konference MeSH
- Publikační typ
- abstrakt z konference MeSH
Pericellular oxygen concentration represents an important factor in the regulation of cell functions, including cell differentiation, growth and mitochondrial energy metabolism. Hypoxia in adipose tissue has been associated with altered adipokine secretion profile and suggested as a possible factor in the development of type 2 diabetes. In vitro experiments provide an indispensable tool in metabolic research, however, physical laws of gas diffusion make prolonged exposure of adherent cells to desired pericellular O2 concentrations questionable. The aim of this study was to investigate the direct effect of various O2 levels (1%, 4% and 20% O2) on the proteomic profile and triglyceride accumulation in 3T3-L1 differentiated preadipocytes using gas-permeable cultureware. Following differentiation of cells under desired pericellular O2 concentrations, cell lysates were subjected to two-dimensional gel electrophoresis and protein visualization using Coomassie blue staining. Spots showing differential expression under hypoxia were analyzed using matrix-assisted laser desorption/ionization mass spectrometry. All identified proteins were subjected to pathway analysis. We observed that protein expression of 26 spots was reproducibly affected by 4% and 1% O2 (17 upregulated and 9 downregulated). Pathway analysis showed that mitochondrial energy metabolism and triglyceride synthesis were significantly upregulated by hypoxia. In conclusion, this study demonstrated the direct effects of pericellular O2 levels on adipocyte energy metabolism and triglyceride synthesis, probably mediated through the reversed tricarboxylic acid cycle flux.
- MeSH
- 2D gelová elektroforéza MeSH
- buněčná diferenciace účinky léků MeSH
- buňky 3T3-L1 MeSH
- citrátový cyklus účinky léků MeSH
- down regulace účinky léků MeSH
- kultivované buňky MeSH
- kyslík farmakologie MeSH
- lipidy biosyntéza MeSH
- lipogeneze účinky léků MeSH
- myši MeSH
- permeabilita MeSH
- plyny chemie MeSH
- počet buněk MeSH
- proteomika * MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- triglyceridy metabolismus MeSH
- tukové buňky cytologie účinky léků metabolismus MeSH
- upregulace účinky léků MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Modern lifestyle has profoundly modified human sleep habits. Sleep duration has shortened over recent decades from 8 to 6.5 hours resulting in chronic sleep deprivation. Additionally, irregular sleep, shift work and travelling across time zones lead to disruption of circadian rhythms and asynchrony between the master hypothalamic clock and pacemakers in peripheral tissues. Furthermore, obstructive sleep apnea syndrome (OSA), which affects 4 - 15% of the population, is not only characterized by impaired sleep architecture but also by repetitive hemoglobin desaturations during sleep. Epidemiological studies have identified impaired sleep as an independent risk factor for all cause of-, as well as for cardiovascular, mortality/morbidity. More recently, sleep abnormalities were causally linked to impairments in glucose homeostasis, metabolic syndrome and Type 2 Diabetes Mellitus (T2DM). This review summarized current knowledge on the metabolic alterations associated with the most prevalent sleep disturbances, i.e. short sleep duration, shift work and OSA. We have focused on various endocrine and molecular mechanisms underlying the associations between inadequate sleep quality, quantity and timing with impaired glucose tolerance, insulin resistance and pancreatic β-cell dysfunction. Of these mechanisms, the role of the hypothalamic-pituitary-adrenal axis, circadian pacemakers in peripheral tissues, adipose tissue metabolism, sympathetic nervous system activation, oxidative stress and whole-body inflammation are discussed. Additionally, the impact of intermittent hypoxia and sleep fragmentation (key components of OSA) on intracellular signaling and metabolism in muscle, liver, fat and pancreas are also examined. In summary, this review provides endocrine and molecular explanations for the associations between common sleep disturbances and the pathogenesis of T2DM.
- Publikační typ
- časopisecké články MeSH
- Publikační typ
- abstrakt z konference MeSH
- Publikační typ
- abstrakt z konference MeSH
