Long-term hypoxia exposure impairs metabolic pathways of de novo lipogenesis in young mouse adipocytes
Authors: Slovakova Lucie , Pospisilova Katerina , Vavra Jiri , Polak Jan 
Published: May 2023

Introduction: Obstructive sleep apnea syndrome, manifested by tissue hypoxia, is causally linked with the development of metabolic diseases such as Type 2 diabetes. Although there is convincing epidemiological evidence, molecular mechanisms mediating the adverse metabolic effects remain unclear. Hypoxia may play an important role and modulate metabolic pathways in young adipocytes including the utilization of glucose and acetate or the activation of the reverse tricarboxylic acid cycle (rTCA) leading to increase de novo lipogenesis and excessive fat accumulation. The aim of this study was to investigate the impact of long-term hypoxia on important pathways of de novo lipogenesis in mouse early and late adipocytes. Hypothesis: Hypoxia affects the utilization of carbon sources for de novo lipogenesis differently in early and late adipocytes. Methods: Mouse 3T3-L1 cells were differentiated into adipocytes for 7 days - early adipocytes (EA) or 14 days - late adipocytes (LA). The cells were cultured in gas-permeable cultureware allowing exposure to sustained oxygen levels of 21% O2 or 4% O2. [13C-5]-glutamine (2.5mM) was added to culture media for 7 and 14 days to assess 13C incorporation to newly synthesized lipids via rTCA and [13C-1]-glutamine (2.5mM) was added to media for 24h to assess 13C incorporation to rTCA metabolites. The 13C incorporation studies were analyzed via gas chromatography-mass spectrometry. The contribution of glucose and acetate to de novo lipogenesis was assessed using [14C]-glucose and [14C]-acetate (both 0.2 μCi) provided in culture media for 2h at the end of experiments. Subsequently, radioactivity in extracted lipids was measured via scintillation counter. Statistical significance was assessed by 2-way ANOVA followed by Tukey's test. Data are presented as means ± SD, N=9. Data Hypoxia had no effect on lipid accumulation in EA, lipid accumulation in LA was increased by 134,41% vs control. The 13C incorporation to lipids was increased in EA by 42,2% vs control and by 13,33% vs LA. The 13C incorporation in citrate was increased by 8,71%, in malate by 9,58% in EA vs control, in LA the incorporation remained unchanged. The [14C]-glucose incorporation to lipids was elevated in EA vs control by 209,98% and EA vs LA by 60,02%. The [14C]-acetate incorporation to lipids was increased in EA by 198,43% vs control and decreased by 45,13% vs LA. Differences are statistically significant (p <0.05). Summary: The effect of hypoxia on lipid accumulation was observed at the late stages of adipocyte maturation. However, in EA hypoxia initially stimulated the activation of rTCA to partially contribute to de novo lipogenesis and significantly increased 14C-glucose incorporation. In LA the rTCA and 14C-glucose incorporation were reduced to the control level and the contribution of acetate was increased. Conclusion: Hypoxia modified metabolic pathways in maturating adipocytes leading to increased de novo lipogenesis and thus hypertrophic mature adipocytes. Project was supported by Grant Agency of the Charles University project GAUK 294822 and by grant AZV project NU21-01-00259 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.