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Autotaxin and its product lysophosphatidic acid suppress brown adipose differentiation and promote diet-induced obesity in mice

ABSTRACT: Brown adipose tissue (BAT) is a thermogenic organ that dissipates stored energy as heat to maintain body temperature in infants and small mammals. This process may also provide protection from development of diet-induced obesity. We found that the bioactive lipid mediator lysophosphatidic acid (LPA) markedly decreases differentiation of cultured primary brown adipocyte precursors, while potent selective inhibitors of the LPA-generating enzyme autotaxin (ATX) promote differentiation. Transgenic mice overexpressing ATX exhibited reduced expression of BAT-related genes in peripheral white adipose tissue and accumulated significantly more fat than wild-type controls when fed a high fat diet. Our results indicate that ATX and its product LPA are physiologically relevant negative regulators of brown fat adipogenesis and suggest that a decrease in peripheral brown adipose tissue results in increased susceptibility to diet-induced obesity in mice.

ORGANISM(S): Mus musculus

PROVIDER: GSE26442 | GEO | 2011/06/01

SECONDARY ACCESSION(S): PRJNA136785

REPOSITORIES: GEO

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Project description:Brown adipose tissue (BAT) is a thermogenic organ that dissipates stored energy as heat to maintain body temperature in infants and small mammals. This process may also provide protection from development of diet-induced obesity. We found that the bioactive lipid mediator lysophosphatidic acid (LPA) markedly decreases differentiation of cultured primary brown adipocyte precursors, while potent selective inhibitors of the LPA-generating enzyme autotaxin (ATX) promote differentiation. Transgenic mice overexpressing ATX exhibited reduced expression of BAT-related genes in peripheral white adipose tissue and accumulated significantly more fat than wild-type controls when fed a high fat diet. Our results indicate that ATX and its product LPA are physiologically relevant negative regulators of brown fat adipogenesis and suggest that a decrease in peripheral brown adipose tissue results in increased susceptibility to diet-induced obesity in mice. Primary BAT cell cultures were established as previously reported with some minor modifications (Cannon B., 2001; NÃ©chad M., 1983). Four- to 6-week-old mice were euthanized under aseptic conditions and interscapular BAT (IBAT) dissected, finely minced in digestion buffer containing 123 mM NaCl, 5mM HCl, 1.3 mM CaCl2, 5mM glucose, 1.5% (w/v) crude bovine serum albumin fraction V, 100 mM HEPES, 0.2% collagenase type II (Sigma-Aldrich Corp., St. Louis, MO), pH 7.4, and digested in 10 ml of the same buffer for 30 min at 37Â°C. The supernatant obtained was then filtered using a 250 Âµm nylon mesh and kept in ice for 15 minutes to allow fat droplets and mature cells to float. The resulting infranatant was filtered through a 30 Âµm nylon mesh to enrich the fraction with precursor cells and centrifuged at 700g for 10 min. The pellet obtained was washed once in warm DMEM and resuspended in DMEM medium containing 10% newborn calf serum, 10 mM HEPES, 4mM glutamine, 25 Âµg/ml sodium ascorbate (Sigma-Aldrich Corp., St. Louis, MO)., 100U/ml penicillin and 100mg/ml streptomycin. Cells were seeded onto 35mm dish plates. Insulin treatment was started on day 1 at 50 nM and the concentration increased to 100 nM at day 5 and 200 nM at day 7. Differentiated/confluent cells were collected for biochemical analysis (~day 9). To identify genes regulated by ATX and LPA signaling, 9 DIV primary BAT cultures were treated with the vehicle (DMSO), specific ATX inhibitor HA155 (2.5ÂµM), or LPA (1-Oleoly-LPA, 5ÂµM) (Avanti Polar Lipids, AL, USA) during differentiation. Treatment of the BAT cultures did not affect confluence or markers of proliferation. RNA was extracted from three cultures per condition using TRIzol reagent (Invitrogen, Carlsbad, CA) as described previously (Blalock EM., 2003).

Project description:Thoracic perivascular adipose tissue (PVAT) is a unique adipose depot that likely influences vascular function and susceptibility to pathogenesis in obesity and metabolic syndrome. Surprisingly, PVAT has been reported to share characteristics of both brown and white adipose, but a detailed direct comparison to interscapular brown adipose tissue (BAT) has not been performed. Here we show by full genome DNA microarray analysis that global gene expression profiles of PVAT are virtually identical to BAT, with equally high expression of Ucp-1, Cidea and other genes known to be uniquely or very highly expressed in BAT. PVAT and BAT also displayed nearly identical phenotypes upon immunohistochemical analysis, and electron microscopy confirmed that PVAT contained multilocular lipid droplets and abundant mitochondria. Compared to white adipose tissue (WAT), PVAT and BAT from C57BL/6 mice fed a high fat diet for 13 weeks had markedly lower expression of immune cell-enriched mRNAs, suggesting resistance to obesity-induced inflammation. Indeed, staining of BAT and PVAT for macrophage markers (F4/80, CD68) in obese mice showed virtually no macrophage infiltration, and FACS analysis of BAT confirmed the presence of very few CD11b+/CD11c+ macrophages in BAT (1.0%) in comparison to WAT (31%). In summary, murine PVAT from the thoracic aorta is virtually identical to interscapular BAT, is resistant to diet-induced macrophage infiltration, and thus may play an important role in protecting the vascular bed from thermal and inflammatory stress. 8-week-old male C57BL6/J mice were fed a normal (ND) or high fat diet (HFD) (Research Diets 12451, 45 kcal% fat) for 13 weeks. Mice were then euthanized and four different adipose depots were harvested for RNA analysis: perivascular fat from the lesser curvature of the aortic arch (PVAT), interscapular brown adipose (BAT), inguinal adipose tissue (SAT), and epididymal adipose tissue (VAT). 250 ng total RNA pooled from two mice was used for cDNA synthesis; 3 biological replicates per tissue and diet were performed for a total of 24 hybridizations.

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Autotaxin and its product lysophosphatidic acid suppress brown adipose differentiation and promote diet-induced obesity in mice

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