Peroxisome proliferator activated receptor gamma 2 (PPARg2) is the nutritionally regulated isoform of PPARg. Ablation of PPARg2 in the ob/ob background, PPARg2−/− Lepob/Lepob (POKO mouse), resulted in decreased fat mass, severe insulin resistance, β-cell failure, and dyslipidaemia. Our results indicate that the PPARg2 isoform plays an important role, mediating adipose tissue expansion in response to positive energy balance. Lipidomic analyses suggest that PPARg2 plays an important antilipotoxic role when induced ectopically in liver and muscle by facilitating deposition of fat as relatively harmless triacylglycerol species and thus preventing accumulation of reactive lipid species. Our data also indicate that PPARg2 may be required for the β-cell hypertrophic adaptive response to insulin resistance. In summary, the PPARg2 isoform prevents lipotoxicity by (a) promoting adipose tissue expansion, (b) increasing the lipid-buffering capacity of peripheral organs, and (c) facilitating the adaptive proliferative response of β-cells to insulin resistance.

The transcriptional coactivator peroxisome proliferator-activated receptor-gamma coactivator-1β (PGC-1β) has been implicated in important metabolic processes. A mouse lacking PGC-1β (PGC1βKO) was generated and phenotyped using physiological, molecular, and bioinformatic approaches. PGC1βKO mice are generally viable and metabolically healthy. Using systems biology, we identified a general defect in the expression of genes involved in mitochondrial function and, specifically, the electron transport chain. This defect correlated with reduced mitochondrial volume fraction in soleus muscle and heart, but not brown adipose tissue (BAT). Under ambient temperature conditions, PGC-1β ablation was partially compensated by up-regulation of PGC-1α in BAT and white adipose tissue (WAT) that lead to increased thermogenesis, reduced body weight, and reduced fat mass. Despite their decreased fat mass, PGC1βKO mice had hypertrophic adipocytes in WAT. The thermogenic role of PGC-1β was identified in thermoneutral and cold-adapted conditions by inadequate responses to norepinephrine injection. Furthermore, PGC1βKO hearts showed a blunted chronotropic response to dobutamine stimulation, and isolated soleus muscle fibres from PGC1βKO mice have impaired mitochondrial function. Lack of PGC-1β also impaired hepatic lipid metabolism in response to acute high fat dietary loads, resulting in hepatic steatosis and reduced lipoprotein-associated triglyceride and cholesterol content. Altogether, our data suggest that PGC-1β plays a general role in controlling basal mitochondrial function and also participates in tissue-specific adaptive responses during metabolic stress.

The nuclear receptor peroxisome proliferator-activated receptor-␥ (PPAR␥) is critically required for adipogenesis. PPAR␥ exists as two isoforms, ␥1 and ␥2. PPAR␥2 is the more potent adipogenic isoform in vitro and is normally restricted to adipose tissues, where it is regulated more by nutritional state than PPAR␥1. To elucidate the relevance of the PPAR␥2 in vivo, we generated a mouse model in which the PPAR␥2 isoform was specifically disrupted. Despite similar weight, body composition, food intake, energy expenditure, and adipose tissue morphology, male mice lacking the ␥2 isoform were more insulin resistant than wild-type animals when fed a regular diet. These results indicate that insulin resistance associated with ablation of PPAR␥2 is not the result of lipodystrophy and suggests a specific role for PPAR␥2 in maintaining insulin sensitivity independently of its effects on adipogenesis. Furthermore, PPAR␥2 knockout mice fed a high-fat diet did not become more insulin resistant than those on a normal diet, despite a marked increase in their mean adipocyte cell size. These findings suggest that PPAR␥2 is required for the maintenance of normal insulin sensitivity in mice but also raises the intriguing notion that PPAR␥2 may be necessary for the adverse effects of a high-fat diet on carbohydrate metabolism. Diabetes 54: 1706 -1716, 2005 P eroxisome proliferator-activated receptor-␥ (PPAR␥) plays a central role in adipogenesis and insulin sensitivity. PPAR␥ is expressed as two isoforms, PPAR␥1 and PPAR␥2, which differ only in that PPAR␥2 has 30 extra amino acids at its NH 2 terminus. Under physiological conditions, PPAR␥2 is expressed almost exclusively in white and brown adipocytes, whereas PPAR␥1 is also expressed in colon, macrophages, skeletal muscle, and liver (1). Although there is limited information regarding the functional differences between these two splice variants, PPAR␥2 may be more adipogenic than PPAR␥1 (2,3). PPAR␥2 may play a distinct role in regulating insulin sensitivity as suggested by the strong epidemiological evidence that the PPAR␥2-specific Pro12Ala variant influences diabetes susceptibility in humans (4).We have shown that expression of PPAR␥ isoforms is differentially regulated by nutritional factors (5). Murine studies showed that PPAR␥2 mRNA is markedly downregulated in white adipose tissue (WAT) by fasting and normalized by re-feeding (1). Similarly, PPAR␥2 gene expression is increased in WAT by a high-fat diet (HFD) as well as in mouse models of diet-induced obesity (5). Studies using genetically modified mouse models have addressed the role of PPAR␥ in vivo (6). A proadipogenic role for PPAR␥ in vivo was supported by the global PPAR␥-deficient and the hypomorphic PPAR␥ mouse models (7-9). In addition to a role in promoting adipogenesis, activation of PPAR␥ also improves insulin sensitivity (10). However, the characterization of the heterozygous PPAR␥ knockout mouse provided the paradoxical finding that mice with a 50% reduction in PPAR␥ gene dosage were resistant to HFD-induced ob...

Coenzyme A (CoA) is an obligatory cofactor in all branches of life. CoA and its derivatives are involved in major metabolic pathways, allosteric interactions and the regulation of gene expression. Abnormal biosynthesis and homeostasis of CoA and its derivatives have been associated with various human pathologies, including cancer, diabetes and neurodegeneration. Using an anti-CoA monoclonal antibody and mass spectrometry, we identified a wide range of cellular proteins which are modified by covalent attachment of CoA to cysteine thiols (CoAlation). We show that protein CoAlation is a reversible post-translational modification that is induced in mammalian cells and tissues by oxidising agents and metabolic stress. Many key cellular enzymes were found to be CoAlated in vitro and in vivo in ways that modified their activities. Our study reveals that protein CoAlation is a widespread post-translational modification which may play an important role in redox regulation under physiological and pathophysiological conditions.

Aims/hypothesis Circulating lipopolysaccharide-binding protein (LBP) is an acute-phase reactant known to be increased in obesity. We hypothesised that LBP is produced by adipose tissue (AT) in association with obesity. Methods LBP mRNA and LBP protein levels were analysed in AT from three cross-sectional (n=210, n=144 and n=28) and three longitudinal (n=8, n=25, n=20) human cohorts; in AT from genetically manipulated mice; in isolated adipocytes; and in human and murine cell lines. The effects of a high-fat diet and exposure to lipopolysaccharide (LPS) and peroxisome proliferator-activated receptor (PPAR)γ agonist were explored. Functional in vitro and ex vivo experiments were also performed. Results LBP synthesis and release was demonstrated to increase with adipocyte differentiation in human and mouse AT, Electronic supplementary material The online version of this article

BackgroundAlthough quite challenging, neuroprotective therapies in ischemic stroke remain an interesting strategy to counter mechanisms of ischemic injury and reduce brain tissue damage. Among potential neuroprotective drug, cyclin-dependent kinases (CDK) inhibitors represent interesting therapeutic candidates. Increasing evidence indisputably links cell cycle CDKs and CDK5 to the pathogenesis of stroke. Although recent studies have demonstrated promising neuroprotective efficacies of pharmacological CDK inhibitors in related animal models, none of them were however clinically relevant to human treatment.Methodology/Principal FindingsIn the present study, we report that systemic delivery of (S)-roscovitine, a well known inhibitor of mitotic CDKs and CDK5, was neuroprotective in a dose-dependent manner in two models of focal ischemia, as recommended by STAIR guidelines. We show that (S)-roscovitine was able to cross the blood brain barrier. (S)-roscovitine significant in vivo positive effect remained when the compound was systemically administered 2 hrs after the insult. Moreover, we validate one of (S)-roscovitine in vivo target after ischemia. Cerebral increase of CDK5/p25 activity was observed 3 hrs after the insult and prevented by systemic (S)-roscovitine administration. Our results show therefore that roscovitine protects in vivo neurons possibly through CDK5 dependent mechanisms.Conclusions/SignificanceAltogether, our data bring new evidences for the further development of pharmacological CDK inhibitors in stroke therapy.

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Pyruvate carboxylase (PC) plays a crucial role in various metabolic pathways, including gluconeogenesis, lipogenesis, and glucose-induced insulin secretion. Here we showed for the first time that the PC gene is transcriptionally regulated by peroxisome proliferator-activated receptor-␥ (PPAR␥) in vitro and in vivo in white and brown adipose tissue. PC mRNA and protein are markedly increased during differentiation of 3T3-L1 cells and HIB-1B, in parallel with the expression of the adipogenic transcription factors, CCAAT-enhancer binding protein ␣, PPAR␥1, and PPAR␥2. Tumor necrosis factor-␣, a cytokine that blocks differentiation of 3T3-L1 cells, suppressed PC expression. Co-transfection studies in 3T3-L1 preadipocytes or HEK293T cells with a 2.3-kb promoter fragment of mouse PC gene linked to a luciferase reporter construct and with plasmids overexpressing retinoid X receptor ␣/PPAR␥1 or retinoid X receptor ␣/PPAR␥2 showed a 6 -8-fold increase above the basal promoter activity. Furthermore, treatment of these transfected cells with the PPAR␥ agonist doubled the promoter activity. Mutation of the putative PPARresponse element-(؊386/؊374) of this 2.3-kb PC promoter fragment abolished the PPAR␥ response. Gel shift and chromatin immunoprecipitation assays demonstrated that endogenous PPAR␥ binds to this functional PPAR-response element of the PC promoter. Mice with targeted disruption of the PPAR␥2 gene displayed ϳ50 -60% reduction of PC mRNA and protein in white adipose tissue. Similarly, in brown adipose tissue of PPAR␥2-deficient mice subjected to cold exposure, PC mRNA was 40% lower than that of wild type mice. Impaired in vitro differentiation of white adipocytes of PPAR␥2 knock-out mice was also associated with a marked reduction of PC mRNA. Our findings identified PC as a PPAR␥-regulated gene and suggested a role for PPAR␥ regulating intermediary metabolism.