Abstract
IntroductionHypoxia regulates adipocyte metabolism. Hexosamine biosynthesis is implicated in murine 3T3L1 adipocyte differentiation and is a possible underlying mechanism for hypoxia’s effects on adipocyte metabolism.MethodsLipid metabolism was studied in human visceral and subcutaneous adipocytes in in vitro hypoxic culture with adipophilic staining, glycerol release, and palmitate oxidation assays. Gene expression and hexosamine biosynthesis activation was studied with QRTPCR, immunofluorescence microscopy, and Western blotting.ResultsHypoxia inhibits lipogenesis and induces basal lipolysis in visceral and subcutaneous human adipocytes. Hypoxia induces fatty acid oxidation in visceral adipocytes but had no effect on fatty acid oxidation in subcutaneous adipocytes. Hypoxia inhibits hexosamine biosynthesis in adipocytes. Inhibition of hexosamine biosynthesis with azaserine attenuates lipogenesis and induces lipolysis in adipocytes in normoxic conditions, while promotion of hexosamine biosynthesis with glucosamine in hypoxic conditions slightly increases lipogenesis.ConclusionsHypoxia’s net effect on human adipocyte lipid metabolism would be expected to impair adipocyte buffering capacity and contribute to systemic lipotoxicity. Our data suggest that hypoxia may mediate its effects on lipogenesis and lipolysis through inhibition of hexosamine biosynthesis. Hexosamine biosynthesis represents a target for manipulation of adipocyte metabolism.
Highlights
Differentiated in adipogenic medium over 14 days exhibit progressive accumulation of cytoplasmic lipid and increased transcript levels of genes associated with adipocyte metabolism, including PPAR-c, fatty acid synthase (FAS), ATGL, and SREBP1c, as well as GFAT, the gene that encodes the ratelimiting enzyme involved in Hexosamine biosynthesis (HBS)
HBS Increases during Human Adipocyte Differentiation Given the observed increase in GFAT transcript levels with differentiation, we studied HBS over the course of adipocyte differentiation
When compared to undifferentiated SVF, mature differentiated human visceral adipocytes demonstrated increased expression on Western blot analysis of adipocyte protein lysates with antibody specific for OGlcNAc, the glycosylation moiety that is covalently linked to multiple cellular proteins when HBS is activated, indicative of increased HBS over the course of adipocyte differentiation (Figure 2A)
Summary
Most data suggest that increased HBS is associated with increased insulin resistance in in vitro and in vivo systems [4,6,7,8,9], some studies demonstrate that HBS improves or has no effect on insulin resistance [10,11,12]. These data are primarily derived from murine systems and involve study of glucose homeostasis. The role of HBS in regulating lipid metabolism in adipocytes and its relationship to hypoxia are unknown, and data from human systems is sparse
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