title: Hypoxia mimetic induces lipid accumulation through mitochondrial dysfunction and stimulates autophagy in murine preadipocyte cell line.
abstract: Hypoxia occurs within adipose tissue of obese human and mice. However, its role in adipose tissue regulation is still controversial.
  
  
  
  We used murine preadipocyte 3T3-L1 cells and hypoxia was induced by using hypoxia mimetic agents, as CoCl2. To study adipocyte differentiation, we evaluated the adipocyte markers (PPARγ, C/EBPα and aP2), and a preadipocyte marker (pref-1) by qPCR, western blotting and immunofluorescence. Lipid accumulation was evaluated by Oil red-O assay and perilipin levels by western blotting and immunofluorescence. The effect of CoCl2in microRNA, miR-27a and miR-27b, levels was evaluated by qPCR. We also assessed the mitochondrial membrane potential and reactive oxygen species (ROS), superoxide and ATP production. The effect of hypoxia mimetic in autophagy was determined by LC3B and p62 level evaluation by western blotting.
  
  
  
  Our results show that the hypoxia mimetic cobalt chloride increases lipid accumulation with no expression of PPARγ2. Furthermore, using qPCR we observed that the hypoxia mimetic increases microRNAs miR-27a and miR-27b, which are known to block PPARγ2 expression. In contrast, cobalt chloride induces mitochondrial dysfunction, and increases ROS production and autophagy. Moreover, an antioxidant agent, glutathione, prevents lipid accumulation induced by hypoxia mimetic indicating that ROS are responsible for hypoxia-induced lipid accumulation.
  
  
  
  All these results taken together suggest that hypoxia mimetic blocks differentiation and induces autophagy. Hypoxia mimetic also induces lipid accumulation through mitochondrial dysfunction and ROS accumulation.
  
  
  
  This study highlights the importance of adipocyte response to hypoxia, which might impair adipocyte metabolism and compromise adipose tissue function.
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal; PDBEB - Doctoral Program in Experimental Biology and Biomedicine, Interdisciplinary Research Institute (III-UC), University of Coimbra, 3004-504 Coimbra, Portugal.
authors.forename: Ana Patrícia
authors.initials: AP
authors.lastname: Marques
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal; Department of Medical Genetics, Pediatric Unit, Coimbra Hospital and Universitary Center (CHUC), 3000-602 Coimbra, Portugal.
authors.forename: Joana
authors.initials: J
authors.lastname: Rosmaninho-Salgado
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal.
authors.forename: Marta
authors.initials: M
authors.lastname: Estrada
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal.
authors.forename: Vera
authors.initials: V
authors.lastname: Cortez
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal.
authors.forename: Rui Jorge
authors.initials: RJ
authors.lastname: Nobre
authors.affiliation: CNC - Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504 Coimbra, Portugal; Faculty of Pharmacy, University of Coimbra, 3000-548 Coimbra, Portugal. Electronic address: ccavadas@uc.pt.
authors.forename: Cláudia
authors.initials: C
authors.lastname: Cavadas
mesh_terms: D041721:3T3-L1 Cells; D017667:Adipocytes / Q000378:metabolism / Q000473:pathology*; D050156:Adipogenesis / Q000502:physiology; D000273:Adipose Tissue / Q000378:metabolism / Q000473:pathology; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D001343:Autophagy / Q000502:physiology*; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D000860:Hypoxia / Q000378:metabolism / Q000473:pathology*; D050356:Lipid Metabolism / Q000502:physiology*; D008055:Lipids / Q000502:physiology*; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism / Q000473:pathology*; D047495:PPAR gamma / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D022763:CCAAT-Enhancer-Binding Protein-alpha; D008055:Lipids; D035683:MicroRNAs; D047495:PPAR gamma; D017382:Reactive Oxygen Species
keywords: Adipogenesis; Autophagy; Hypoxia; Mitochondrial dysfunction; Reactive oxygen species