The molecular mechanisms linking obstructive sleep apnea syndrome (OSA) to obesity and the development of metabolic diseases are still poorly understood. The role of hypoxia (a characteristic feature of OSA) in excessive fat accumulation has been proposed. The present study investigated the possible effects of hypoxia (4% oxygen) on de novo lipogenesis by tracking the major carbon sources in differentiating 3T3-L1 adipocytes. Gas-permeable cultuware was employed to cultivate 3T3-L1 adipocytes in hypoxia (4%) for 7 or 14 days of differentiation. We investigated the contribution of glutamine, glucose or acetate using 13C or 14C labelled carbons to the newly synthesized lipid pool, changes in intracellular lipid content after inhibiting citrate- or acetate-dependent pathways and gene expression of involved key enzymes. The results demonstrate that, in differentiating adipocytes, hypoxia decreased the synthesis of lipids from glucose (44.1 ± 8.8 to 27.5 ± 3.0 pmol/mg of protein, p < 0.01) and partially decreased the contribution of glutamine metabolized through the reverse tricarboxylic acid cycle (4.6% ± 0.2-4.2% ± 0.1%, p < 0.01). Conversely, the contribution of acetate, a citrate- and mitochondria-independent source of carbons, increased upon hypoxia (356.5 ± 71.4 to 649.8 ± 117.5 pmol/mg of protein, p < 0.01). Further, inhibiting the citrate- or acetate-dependent pathways decreased the intracellular lipid content by 58% and 73%, respectively (p < 0.01) showing the importance of de novo lipogenesis in hypoxia-exposed adipocytes. Altogether, hypoxia modified the utilization of carbon sources, leading to alterations in de novo lipogenesis in differentiating adipocytes and increased intracellular lipid content.

Department of Cell Biology Faculty of Science Charles University Prague Czech Republic

Department of Internal Medicine Institute of Medical Biochemistry and Laboratory Diagnostics General University Hospital Prague and 1st Faculty of Medicine Charles University Prague Czech Republic

Department of Internal Medicine Thomayer University Hospital Videnska 800 Prague 140 59 Czech Republic

Department of Pathophysiology 3rd Faculty of Medicine Charles University Ruska 87 Prague 100 00 Czech Republic

Institute of Medical Biochemistry and Laboratory Diagnostics 1st Faculty of Medicine Charles University Prague Czech Republic

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Senaratna, C. V. et al. Prevalence of obstructive sleep apnea in the general population: A systematic review. PubMed DOI

Briancon-Marjollet, A. et al. The impact of sleep disorders on glucose metabolism: Endocrine and molecular mechanisms. PubMed DOI PMC

Kent, B. D., McNicholas, W. T. & Ryan, S. Insulin resistance, glucose intolerance and diabetes mellitus in obstructive sleep apnoea. PubMed PMC

Aurora, R. N. & Punjabi, N. M. Obstructive sleep apnoea and type 2 diabetes mellitus: A bidirectional association. PubMed DOI

Gottlieb, D. J. & Punjabi, N. M. Diagnosis and management of obstructive sleep apnea: A review. PubMed DOI

Jehan, S. et al. Obstructive sleep apnea and obesity: Implications for Public Health. PubMed PMC

Brown, M. A. et al. The impact of sleep-disordered breathing on body Mass Index (BMI): The Sleep Heart Health Study (SHHS). PubMed PMC

Borel, A. L. et al. Sleep apnoea attenuates the effects of a lifestyle intervention programme in men with visceral obesity. PubMed DOI

Zuraikat, F. M. et al. Dimensions of sleep quality are related to objectively measured eating behaviors among children at high familial risk for obesity. PubMed DOI PMC

Shechter, A. Obstructive sleep apnea and energy balance regulation: A systematic review. PubMed DOI PMC

Shechter, A. Effects of continuous positive airway pressure on energy balance regulation: A systematic review. PubMed DOI PMC

Trayhurn, P. Hypoxia and adipose tissue function and dysfunction in obesity. PubMed DOI

Yin, J. et al. Role of hypoxia in obesity-induced disorders of glucose and lipid metabolism in adipose tissue. PubMed DOI PMC

Uchiyama, T., Ota, H., Ohbayashi, C. & Takasawa, S. Effects of intermittent hypoxia on Cytokine expression involved in insulin resistance. PubMed PMC

Lempesis, I. G., van Meijel, R. L. J., Manolopoulos, K. N. & Goossens, G. H. Oxygenation of adipose tissue: A human perspective. PubMed PMC

Ota, H. et al. Relationship between intermittent hypoxia and type 2 diabetes in Sleep Apnea Syndrome. PubMed DOI PMC

Oates, E. H. & Antoniewicz, M. R. 13 C-Metabolic flux analysis of 3T3-L1 adipocytes illuminates its core metabolism under hypoxia. PubMed DOI

Weiszenstein, M. et al. Adipogenesis, lipogenesis and lipolysis is stimulated by mild but not severe hypoxia in 3T3-L1 cells. PubMed DOI

Suzuki, T., Shinjo, S., Arai, T., Kanai, M. & Goda, N. Hypoxia and fatty liver. PubMed DOI PMC

Michailidou, Z. et al. Adipocyte pseudohypoxia suppresses lipolysis and facilitates benign adipose tissue expansion. PubMed DOI PMC

Kim, K. H., Song, M. J., Chung, J., Park, H. & Kim, J. B. Hypoxia inhibits adipocyte differentiation in a HDAC-independent manner. PubMed DOI

Musutova, M., Weiszenstein, M., Koc, M. & Polak, J. Intermittent Hypoxia stimulates Lipolysis, but inhibits differentiation and De Novo Lipogenesis in 3T3-L1 cells. PubMed DOI

Shao, M. et al. Pathologic HIF1α signaling drives adipose progenitor dysfunction in obesity. PubMed DOI PMC

Floyd, Z. E., Kilroy, G., Wu, X. & Gimble, J. M. Effects of prolyl hydroxylase inhibitors on adipogenesis and hypoxia inducible factor 1 alpha levels under normoxic conditions. PubMed DOI

Kudo, T. et al. Context-dependent regulation of lipid accumulation in adipocytes by a HIF1α-PPARγ feedback network. PubMed DOI PMC

He, Q. et al. Regulation of HIF-1{alpha} activity in adipose tissue by obesity-associated factors: Adipogenesis, insulin, and hypoxia. PubMed PMC

White, U. & Ravussin, E. Dynamics of adipose tissue turnover in human metabolic health and disease. PubMed DOI PMC

Spalding, K. L. et al. Dynamics of fat cell turnover in humans. PubMed

Yin, J. et al. Role of hypoxia in obesity-induced disorders of glucose and lipid metabolism in adipose tissue. PubMed DOI PMC

Musutova, M. et al. The effect of hypoxia and metformin on fatty acid uptake, storage, and oxidation in L6 differentiated myotubes. PubMed DOI PMC

Felix, J. B., Cox, A. R. & Hartig, S. M. Acetyl-CoA and metabolite fluxes regulate white adipose tissue expansion. PubMed DOI PMC

Holleran, A. L., Briscoe, D. A., Fiskum, G. & Kelleher, J. K. Glutamine metabolism in AS-30D hepatoma cells. Evidence for its conversion into lipids via reductive carboxylation. PubMed DOI

DeBerardinis, R. J. et al. Beyond aerobic glycolysis: Transformed cells can engage in glutamine metabolism that exceeds the requirement for protein and nucleotide synthesis. PubMed DOI PMC

Metallo, C. M. et al. Reductive glutamine metabolism by IDH1 mediates lipogenesis under hypoxia. PubMed DOI PMC

Zhang, G. F. et al. Reductive TCA cycle metabolism fuels glutamine- and glucose-stimulated insulin secretion. PubMed DOI PMC

Xu, H. et al. Acyl-CoA synthetase short-chain family member 2 (ACSS2) is regulated by SREBP-1 and plays a role in fatty acid synthesis in caprine mammary epithelial cells. PubMed DOI

Zhao, S. et al. Dietary fructose feeds hepatic lipogenesis via microbiota-derived acetate. PubMed PMC

Smith, S. B. & Crouse, J. D. Relative contributions of acetate, lactate and glucose to lipogenesis in bovine intramuscular and subcutaneous adipose tissue. PubMed DOI

Gao, X. et al. Acetate functions as an epigenetic metabolite to promote lipid synthesis under hypoxia. PubMed PMC

Kamphorst, J. J., Chung, M. K., Fan, J. & Rabinowitz, J. D. Quantitative analysis of acetyl-CoA production in hypoxic cancer cells reveals substantial contribution from acetate. PubMed PMC

Weiszenstein, M. et al. The Effect of Pericellular Oxygen levels on Proteomic Profile and Lipogenesis in 3T3-L1 differentiated Preadipocytes cultured on gas-permeable cultureware. PubMed DOI PMC

Pavlacky, J. & Polak, J. Technical feasibility and physiological relevance of hypoxic cell culture models. PubMed PMC

Vacek, L. et al. Hypoxia induces saturated fatty acids Accumulation and reduces unsaturated fatty acids independently of reverse tricarboxylic acid cycle in L6 myotubes. PubMed DOI PMC

Ong, C. W., O’Driscoll, D. M., Truby, H., Naughton, M. T. & Hamilton, G. S. The reciprocal interaction between obesity and obstructive sleep apnoea. PubMed DOI

Reinke, C., Bevans-Fonti, S., Drager, L. F., Shin, M. K. & Polotsky, V. Y. Effects of different acute hypoxic regimens on tissue oxygen profiles and metabolic outcomes. PubMed DOI PMC

Lu, H., Gao, Z., Zhao, Z., Weng, J. & Ye, J. Transient hypoxia reprograms differentiating adipocytes for enhanced insulin sensitivity and triglyceride accumulation. PubMed PMC

Shevchenko, A. & Simons, K. Lipidomics: Coming to grips with lipid diversity. PubMed DOI

Smolková, K. & Ježek, P. The role of mitochondrial NADPH-dependent isocitrate dehydrogenase in cancer cells. PubMed DOI PMC

Handy, D. E. & Loscalzo, J. Responses to reductive stress in the cardiovascular system. PubMed DOI PMC

Kampjut, D. & Sazanov, L. A. Structure and mechanism of mitochondrial proton-translocating transhydrogenase. PubMed DOI

Zhang, Z. et al. Serine catabolism generates liver NADPH and supports hepatic lipogenesis. PubMed DOI PMC

Ducluzeau, P. H. et al. Dynamic regulation of mitochondrial network and oxidative functions during 3T3-L1 fat cell differentiation. PubMed DOI

Metallo, C. M. et al. Reductive glutamine metabolism by IDH1 mediates lipogenesis under hypoxia. PubMed PMC

Zhao, S. et al. ATP-Citrate lyase controls a glucose-to-acetate metabolic switch. PubMed DOI PMC

Schug, Z. T. et al. Acetyl-CoA synthetase 2 promotes acetate utilization and maintains Cancer Cell Growth under metabolic stress. PubMed DOI PMC

Wheatley, V. R., Hodgins, L. T., Coon, W. M. & Cutaneous Lipogenesis, I. Evaluation of Model systems and the utilization of acetate, citrate and glucose as compared with other tissues. PubMed DOI

Murphy, A. M. et al. Intermittent hypoxia in obstructive sleep apnoea mediates insulin resistance through adipose tissue inflammation. PubMed DOI

Tang, Y., Wang, J., Cai, W. & Xu, J. RAGE/NF-κB pathway mediates hypoxia-induced insulin resistance in 3T3-L1 adipocytes. PubMed DOI

Magalang, U. J. et al. Intermittent hypoxia suppresses adiponectin secretion by adipocytes. PubMed

Dunlop, M. & Court, J. M. Lipogenesis in developing human adipose tissue. PubMed DOI

Zebisch, K., Voigt, V., Wabitsch, M. & Brandsch, M. Protocol for effective differentiation of 3T3-L1 cells to adipocytes. PubMed DOI

Si, Y., Yoon, J. & Lee, K. Flux profile and modularity analysis of time-dependent metabolic changes of de novo adipocyte formation. PubMed DOI

Dvořák, A., Zelenka, J., Smolková, K., Vítek, L. & Ježek, P. Background levels of neomorphic 2-hydroxyglutarate facilitate proliferation of primary fibroblasts. PubMed DOI

Yoshimoto, M. et al. Characterization of acetate metabolism in tumor cells in relation to cell proliferation: Acetate metabolism in tumor cells. PubMed DOI

Yoshii, Y. et al. Tumor uptake of radiolabeled acetate reflects the expression of cytosolic acetyl-CoA synthetase: Implications for the mechanism of acetate PET. PubMed DOI

Vankoningsloo, S. et al. Mitochondrial dysfunction induces triglyceride accumulation in 3T3-L1 cells: Role of fatty acid β-oxidation and glucose. PubMed DOI

Bligh, E. G. & Dyer, W. J. A rapid method of total lipid extraction and purification. PubMed DOI

Bradford, M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. PubMed DOI

Lu, Y. et al. Inhibition of ACSS2 attenuates alcoholic liver steatosis via epigenetically regulating de novo lipogenesis. PubMed DOI