Macrophage polarisation plays a role in intermittent hypoxia (IH)-impaired subcutaneous adipogensis in an IH-exposed mouse model

Abstract

INTRODUCTION: Obesity is associated with majority of obstructive sleep apnoea (OSA)–related morbidities including cardiovascular and metabolic disorders. Impairment of adipogenic capacity in pre-adipocytes may promote adipocyte hypertrophy and increase the risks of insulin resistance and type 2 diabetes. Recent research findings have suggested an inhibitory role of macrophage polarisation in adipogenesis. However, the impact of intermittent hypoxia (IH), as a characteristic pathophysiologic feature in OSA, on adipogenesis is unknown. This study investigated the impact of IH on adipogenesis and further explored the role of macrophage polarisation in IH-regulated adipogenic process. METHOD: C57BL/6N male mice were exposed to either IH or room air (RA) for 6 weeks. The profile of IH was cycles of approximately 240 seconds of 10% O2 and 120 seconds of 21% O2, ie 10 cycles/hour. Epididymal adipose tissue (AT) and abdominal AT were isolated to represent visceral (VIS) and subcutaneous (SUB) AT, respectively. Stromal vascular fraction (SVF) from depot-specific AT was isolated, cultured, and underwent two differentiation cycles in vitro. Macrophage-conditioned media from cultured RAW264.7 cell line were also incubated with SUB SVF and VIS SVF during adipogenic differentiation. Oil Red O staining was carried out for identification of oily droplets. Assessments of differentiation-associated markers, adipogenic transcriptional factors, and macrophage markers were achieved by real-time polymerase chain reaction. Morphological change and M1 macrophage infiltration were detected by H&E and immunofluorescence staining on formalin-fixed and paraffin-embedded AT sections (10 μm), respectively. RESULT: After adipogenic differentiation in vitro, oily droplets of SUB SVF from IH-exposed mice were significantly down-regulated in comparison with that derived from RA mice, whilst the inhibitory effect of IH on oily droplets was not observed in VIS SVF. In agreement with the production of oily droplets, differentiation associated markers and adipogenic transcriptional factors were also suppressed in SUB SVF from IH-exposed mice. IH exposure caused hypertrophic adipocytes in SUB AT but not in VIS AT. Furthermore, IH exposure increased infiltration of preferentially M1 macrophages in SUB AT, in line with the reduction of M2 phenotype and increment of M1 phenotype markers in SUB SVF. In support, conditioned-medium from RAW264.7 (M1 macrophage dominant) significantly inhibited the adipogenesis of SUB SVF, suggesting the involvement of macrophage polarisation in IH-inhibited adipogenesis. CONCLUSION: Macrophage polarisation may be a potential mechanism responsible for IH-impaired subcutaneous adipogenesis.