Low-intensity intermittent hypoxia promotes subcutaneous adipogenic differentiation

Abstract

Introduction: Obstructive sleep apnoea (OSA), characterised by intermittent hypoxia (IH), is highly associated with obesity. Depot-specific adipogenic differentiation, an important physiological mechanism in maintaining adipose tissue homeostasis, could be regulated by intracellular transcriptional factors, extracellular signalling pathways, and inflammation. However, the impact of IH on adipogenesis is unclear. This study aimed to investigate the role of IH in adipogenic differentiation in vivo and in vitro. Methods: The Sprague Dawley rats were subjected to intermittent normoxia (IN) or IH (each cycle: 240 seconds for 10% O2 and 120 seconds for 21% O2) for 6 weeks. Human subcutaneous preadipocytes (HPAs) underwent six differentiation cycles into mature adipocytes. Each differentiation cycle consisted of two sequential procedures of differentiation (for 3 days) and maintenance (for 2 days). During each 3-day differentiation, HPAs were subjected to IH (IH; 1% for 10 min and 21% O2 for 5 min per cycle; 5% CO2) or IN treatment. The degree of adipogenic differentiation was evaluated by the measurements of adipogenic transcriptional factors (CEBPα, PPARγ CEBPδ, and CHOP) and adipocyte-specific proteins (FABP4 and GLUT4) using Q-PCR and/or Western blots. The production of oily droplets in HPAs was detected by Oil Red O staining. The analysis of insulin-like growth factor 1 receptor (IGF-1R)/Akt pathway in HPAs was achieved by incubation of IGF-1R selective kinase inhibitor NVP-AEW541 (0.1 μmol/L) using Western blots. ELISA was applied to detect adiponectin in tissue-conditioned media and pro-inflammatory markers (interleukin [IL]-6 and monocyte chemoattractant protein [MCP]-1) in HPAs-conditioned media. Results: The up-regulation of pro-adipogenic markers (CEBPα, PPARγ, FABP4) and down-regulation of antiadipogenic markers CHOP were found in IH-exposed subcutaneous adipose tissue (SAT) but not visceral adipose tissue (VAT). In addition, IH exposure facilitated the release of adiponectin in SAT- but not VATconditioned media. In line with in-vivo results, IH accelerated the accumulation of oil droplets in HPAs. During differentiation, IH caused elevation of adipogenesis-associated markers (FABP4, GLUT4, CEBPα, andPPARγ) compared to cells exposed to IN. Moreover, the reduction in expression of CEBPδ was prevented by IH exposure with adipogenic development of HPAs. The pro-adipogenic role of IGF-1R/Akt activation in IHexposed HPAs was significantly attenuated by IGF-1R kinase inhibitor NVP-AEW541. In addition, IH also induced elevated levels of IL-6 and MCP-1 in the conditioned media during the process of HPAs differentiation. Conclusion: Low-frequency IH exposure could promote the adipogenic differentiation of SAT and subcutaneous preadipocytes (HPAs) via regulating transcriptional factors, IGF-1/Akt signalling pathway and inflammation.