Acyl-Coenzyme A Thioesterase 9 Traffics Mitochondrial Short-Chain Fatty Acids Toward De Novo Lipogenesis and Glucose Production in the Liver.

BACKGROUND AND AIMS: Obesity-induced pathogenesis of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) is associated with increased de novo lipogenesis (DNL) and hepatic glucose production (HGP) that is due to excess fatty acids. Acyl-coenzyme A (CoA) thioesterase (Acot) family members control the cellular utilization of fatty acids by hydrolyzing (deactivating) acyl-CoA into nonesterified fatty acids and CoASH. APPROACH AND RESULTS: Using Caenorhabditis elegans, we identified Acot9 as the strongest regulator of lipid accumulation within the Acot family. Indicative of a maladaptive function, hepatic Acot9 expression was higher in patients with obesity who had NAFLD and NASH compared with healthy controls with obesity. In the setting of excessive nutrition, global ablation of Acot9 protected mice against increases in weight gain, HGP, steatosis, and steatohepatitis. Supportive of a hepatic function, the liver-specific deletion of Acot9 inhibited HGP and steatosis in mice without affecting diet-induced weight gain. By contrast, the rescue of Acot9 expression only in the livers of Acot9 knockout mice was sufficient to promote HGP and steatosis. Mechanistically, hepatic Acot9 localized to the inner mitochondrial membrane, where it deactivated short-chain but not long-chain fatty acyl-CoA. This unique localization and activity of Acot9 directed acetyl-CoA away from protein lysine acetylation and toward the citric acid (TCA) cycle. Acot9-mediated exacerbation of triglyceride and glucose biosynthesis was attributable at least in part to increased TCA cycle activity, which provided substrates for HGP and DNL. ß-oxidation and ketone body production, which depend on long-chain fatty acyl-CoA, were not regulated by Acot9. CONCLUSIONS: Taken together, our findings indicate that Acot9 channels hepatic acyl-CoAs toward increased HGP and DNL under the pathophysiology of obesity. Therefore, Acot9 represents a target for the management of NAFLD.

Submitochondrial Protein Translocation Upon Stress Inhibits Thermogenic Energy Expenditure.

Mitochondria-rich brown adipocytes dissipate cellular fuel as heat by thermogenic energy expenditure (TEE). Prolonged nutrient excess or cold exposure impair TEE and contribute to the pathogenesis of obesity, but the mechanisms remain incompletely understood. Here we report that stress-induced proton leak into the matrix interface of mitochondrial innermembrane (IM) mobilizes a group of proteins from IM into matrix, which in turn alter mitochondrial bioenergetics. We further determine a smaller subset that correlates with obesity in human subcutaneous adipose tissue. We go on to show that the top factor on this short list, acyl-CoA thioesterase 9 (ACOT9), migrates from the IM into the matrix upon stress where it enzymatically deactivates and prevents the utilization of acetyl-CoA in TEE. The loss of ACOT9 protects mice against the complications of obesity by maintaining unobstructed TEE. Overall, our results introduce aberrant protein translocation as a strategy to identify pathogenic factors. One-Sentence Summary: Thermogenic stress impairs mitochondrial energy utilization by forcing translocation of IM-bound proteins into the matrix.

Human hepatocyte PNPLA3-148M exacerbates rapid non-alcoholic fatty liver disease development in chimeric mice.

Advanced non-alcoholic fatty liver disease (NAFLD) is a rapidly emerging global health problem associated with pre-disposing genetic polymorphisms, most strikingly an isoleucine to methionine substitution in patatin-like phospholipase domain-containing protein 3 (PNPLA3-I148M). Here, we study how human hepatocytes with PNPLA3 148I and 148M variants engrafted in the livers of broadly immunodeficient chimeric mice respond to hypercaloric diets. As early as four weeks, mice developed dyslipidemia, impaired glucose tolerance, and steatosis with ballooning degeneration selectively in the human graft, followed by pericellular fibrosis after eight weeks of hypercaloric feeding. Hepatocytes with the PNPLA3-148M variant, either from a homozygous 148M donor or overexpressed in a 148I donor background, developed microvesicular and severe steatosis with frequent ballooning degeneration, resulting in more active steatohepatitis than 148I hepatocytes. We conclude that PNPLA3-148M in human hepatocytes exacerbates NAFLD. These models will facilitate mechanistic studies into human genetic variant contributions to advanced fatty liver diseases.

Transcriptional Regulation in Non-Alcoholic Fatty Liver Disease.

Obesity is the primary risk factor for the pathogenesis of non-alcoholic fatty liver disease (NAFLD), the worldwide prevalence of which continues to increase dramatically. The liver plays a pivotal role in the maintenance of whole-body lipid and glucose homeostasis. This is mainly mediated by the transcriptional activation of hepatic pathways that promote glucose and lipid production or utilization in response to the nutritional state of the body. However, in the setting of chronic excessive nutrition, the dysregulation of hepatic transcriptional machinery promotes lipid accumulation, inflammation, metabolic stress, and fibrosis, which culminate in NAFLD. In this review, we provide our current understanding of the transcription factors that have been linked to the pathogenesis and progression of NAFLD. Using publicly available transcriptomic data, we outline the altered activity of transcription factors among humans with NAFLD. By expanding this analysis to common experimental mouse models of NAFLD, we outline the relevance of mouse models to the human pathophysiology at the transcriptional level.

Fatty acid activation in thermogenic adipose tissue.

Channeling carbohydrates and fatty acids to thermogenic tissues, including brown and beige adipocytes, have garnered interest as an approach for the management of obesity-related metabolic disorders. Mitochondrial fatty acid oxidation (ß-oxidation) is crucial for the maintenance of thermogenesis. Upon cellular fatty acid uptake or following lipolysis from triglycerides (TG), fatty acids are esterified to coenzyme A (CoA) to form active acyl-CoA molecules. This enzymatic reaction is essential for their utilization in ß-oxidation and thermogenesis. The activation and deactivation of fatty acids are regulated by two sets of enzymes called acyl-CoA synthetases (ACS) and acyl-CoA thioesterases (ACOT), respectively. The expression levels of ACS and ACOT family members in thermogenic tissues will determine the substrate availability for ß-oxidation, and consequently the thermogenic capacity. Although the role of the majority of ACS and ACOT family members in thermogenesis remains unclear, recent proceedings link the enzymatic activities of ACS and ACOT family members to metabolic disorders and thermogenesis. Elucidating the contributions of specific ACS and ACOT family members to trafficking of fatty acids towards thermogenesis may reveal novel targets for modulating thermogenic capacity and treating metabolic disorders.

Obesity Impairs Oligopeptide/Amino Acid-Induced Ghrelin Release and Smooth Muscle Contractions in the Human Proximal Stomach.

SCOPE: The satiation properties of proteins involve effects on gut peptide release and gastrointestinal motility which may be altered during obesity. This study compares the in vitro response and role of amino acid (AA) taste receptors (TASR) in the effect of AAs and a casein hydrolysate on ghrelin release and smooth muscle (SM) contractions in the proximal gut of lean and obese patients. METHODS AND RESULTS: Basal ghrelin release, measured from mucosal segments, is maximal in the fundus and decreased distally. Obesity selectively impaires the stimulatory effect of a casein hydrolyaste on ghrelin release in the fundus but does not affect its inhibitory effect in the small intestine (SI). The SM contractions induced by a casein hydrolysate and AAs are stronger in strips from the SI than from the fundus but are reduced in the stomach of obese patients. The region-dependent expression of AA-TASRs in the mucosa and SM layer is affected by obesity. Most of the AA-induced responses are reduced by the umami antagonist, lactisole. l-Met-induced responses involve bitter taste receptors. CONCLUSION: Region-specific targeting of AA taste receptors on both enteroendocrine and SM cells with specific AA-enriched diets might be a useful strategy to combat obesity as well as hypomotility disorders.

The role of nutrient sensing in the metabolic changes after gastric bypass surgery.

Taste receptors coupled to the gustatory G-protein, gustducin, on enteroendocrine cells sense nutrients to regulate gut hormone release. During Roux-en-Y gastric bypass (RYGB) surgery, the altered nutrient flow to more distal regions can affect gustducin-mediated gut hormone release and hence energy and glucose homeostasis. We studied the role of gustducin-mediated signaling in the metabolic improvements and intestinal adaptations along the gut after RYGB surgery in wild-type (WT) and α-gustducin-/- (α-gust-/-) mice. RYGB surgery decreased body weight in WT and α-gust-/- mice, whereas food intake was only decreased in WT mice. Pair-feeding to the RYGB group improved glucose homeostasis to a similar extent in WT mice. GLP1 levels were increased in both genotypes, PYY levels in α-gust-/- mice and octanoyl ghrelin levels were not affected after RYGB surgery. In WT mice, nutrients act via α-gustducin to increase L-cell differentiation (foregut) and L-cell number (foregut and hindgut) in a region-dependent manner. In α-gust-/- mice, the effect on gut hormone levels is probably tuned via increased peptide sensor and glucose transporter expression in the Roux limb and increased caecal butyrate and propionate levels in the hindgut that activate free fatty acid receptors. Finally, signaling via α-gustducin plays a role in the increased ion transport of the foregut but not in the improvement in colonic barrier function. In conclusion, RYGB surgery decreased body weight in both WT and α-gust-/- mice. Elevated plasma GLP1 and PYY levels might mediate this effect, although α-gustducin differentially affects several regulatory systems in the foregut and hindgut, tuning gut hormone release.

Supplementation of oligofructose, but not sucralose, decreases high-fat diet induced body weight gain in mice independent of gustducin-mediated gut hormone release.

SCOPE: Enteroendocrine cells sense nutrients through taste receptors similar to those on the tongue. Sweet and fatty acid taste receptors (FFAR) coupled to the gustatory G-protein, gustducin, on enteroendocrine cells play a role in gut hormone release. We studied if supplementation of artificial (sucralose) or prebiotic (oligofructose; OFS) sweeteners target gustducin-mediated signaling pathways to alter gut hormone release and reduce obesity-associated disorders. METHODS AND RESULTS: Wild-type (WT) and α-gustducin knockout (α-gust-/- ) mice were fed a high-fat diet and gavaged once daily (8 wk) with water or equisweet concentrations of sweeteners. OFS but not sucralose decreased body weight gain (-19 ± 3%, p < 0.01), fat pad mass (-55 ± 6%, p < 0.001), and insulin resistance (-39 ± 5%, p < 0.001) independent of α-gustducin. Neither sweetener improved glucose intolerance, while solely OFS improved the disturbed colonic permeability. OFS decreased (-65 ± 8%, p < 0.001) plasma glucagon-like peptide 1 (GLP-1) but not ghrelin and peptide YY (PYY) levels in WT mice. Cecal acetate and butyrate levels were reduced by OFS in both genotypes suggesting enhanced uptake of SCFAs that may target FFAR2 (upregulated expression) in adipose tissue. CONCLUSION: OFS, but not sucralose, reduced body weight gain and decreased intestinal permeability, but not glucose intolerance. Effects were not mediated by altered gut hormone levels or gustducin-mediated signaling. Artificial sweeteners do not affect gut hormone levels and are metabolically inert in mice on a high-fat diet. In contrast, prebiotic oligosaccharides (OFS) prevent body weight gain but not glucose intolerance. Alterations in sweet and short-chain fatty acid receptors (FFAR) (studied in WT and α-gust-/- mice) that regulate gut hormone levels are not mandatory for the positive effects of OFS. Enhanced uptake of SCFAs may favor interaction with FFAR2/3 on adipose tissue to induce weight loss.

The Sweetener-Sensing Mechanisms of the Ghrelin Cell.

Carbohydrate administration decreases plasma levels of the 'hunger hormone' ghrelin. The ghrelin cell is co-localized with the sweet taste receptor subunit, TAS1R3, and the gustatory G-protein, gustducin, both involved in the sensing of sweeteners by entero-endocrine cells. This study investigated the role of gustducin-mediated sweet taste receptor signaling on ghrelin secretion in a gastric ghrelinoma cell line, tissue segments and mice. The monosaccharide d-glucose and low-intensity sweetener oligofructose (OFS) decreased (p < 0.001) ghrelin secretion while the high-intensity sweetener sucralose increased (p < 0.001) ghrelin secretion in vitro. These effects were not mediated via the sweet taste receptor or glucose transporters (the sodium-dependent glucose cotransporter SGLT-1 and GLUT2). The effect of these compounds was mimicked ex vivo in gastric and jejunal segments from both wild type (WT) and α-gustducin knockout (α-gust-/-) mice. In vivo, the sensing of d-glucose was polarized since intragastric but not intravenous administration of d-glucose decreased (p < 0.05) ghrelin levels in an α-gustducin independent manner which involved inhibition of duodenal ghrelin release. In contrast, neither OFS nor sucralose affected ghrelin secretion in vivo. In conclusion, α-gustducin-mediated sweet taste receptor signaling does not play a functional role in the sensing of carbohydrates, or low- or high-intensity sweeteners by the ghrelin cell.

The Gustatory Signaling Pathway and Bitter Taste Receptors Affect the Development of Obesity and Adipocyte Metabolism in Mice.

Intestinal chemosensory signaling pathways involving the gustatory G-protein, gustducin, and bitter taste receptors (TAS2R) have been implicated in gut hormone release. Alterations in gut hormone profiles may contribute to the success of bariatric surgery. This study investigated the involvement of the gustatory signaling pathway in the development of diet-induced obesity and the therapeutic potential of targeting TAS2Rs to induce body weight loss. α-gustducin-deficient (α-gust-/-) mice became less obese than wild type (WT) mice when fed a high-fat diet (HFD). White adipose tissue (WAT) mass was lower in α-gust-/- mice due to increased heat production as a result of increases in brown adipose tissue (BAT) thermogenic activity, involving increased protein expression of uncoupling protein 1. Intra-gastric treatment of obese WT and α-gust-/- mice with the bitter agonists denatonium benzoate (DB) or quinine (Q) during 4 weeks resulted in an α-gustducin-dependent decrease in body weight gain associated with a decrease in food intake (DB), but not involving major changes in gut peptide release. Both WAT and 3T3-F442A pre-adipocytes express TAS2Rs. Treatment of pre-adipocytes with DB or Q decreased differentiation into mature adipocytes. In conclusion, interfering with the gustatory signaling pathway protects against the development of HFD-induced obesity presumably through promoting BAT activity. Intra-gastric bitter treatment inhibits weight gain, possibly by directly affecting adipocyte metabolism.

Shifting the circadian rhythm of feeding in mice induces gastrointestinal, metabolic and immune alterations which are influenced by ghrelin and the core clock gene Bmal1.

BACKGROUND: In our 24-hour society, an increasing number of people are required to be awake and active at night. As a result, the circadian rhythm of feeding is seriously compromised. To mimic this, we subjected mice to restricted feeding (RF), a paradigm in which food availability is limited to short and unusual times of day. RF induces a food-anticipatory increase in the levels of the hunger hormone ghrelin. We aimed to investigate whether ghrelin triggers the changes in body weight and gastric emptying that occur during RF. Moreover, the effect of genetic deletion of the core clock gene Bmal1 on these physiological adaptations was studied. METHODS: Wild-type, ghrelin receptor knockout and Bmal1 knockout mice were fed ad libitum or put on RF with a normal or high-fat diet (HFD). Plasma ghrelin levels were measured by radioimmunoassay. Gastric contractility was studied in vitro in muscle strips and in vivo (13C breath test). Cytokine mRNA expression was quantified and infiltration of immune cells was assessed histologically. RESULTS: The food-anticipatory increase in plasma ghrelin levels induced by RF with normal chow was abolished in HFD-fed mice. During RF, body weight restoration was facilitated by ghrelin and Bmal1. RF altered cytokine mRNA expression levels and triggered contractility changes resulting in an accelerated gastric emptying, independent from ghrelin signaling. During RF with a HFD, Bmal1 enhanced neutrophil recruitment to the stomach, increased gastric IL-1α expression and promoted gastric contractility changes. CONCLUSIONS: This is the first study demonstrating that ghrelin and Bmal1 regulate the extent of body weight restoration during RF, whereas Bmal1 controls the type of inflammatory infiltrate and contractility changes in the stomach. Disrupting the circadian rhythm of feeding induces a variety of diet-dependent metabolic, immune and gastrointestinal alterations, which may explain the higher prevalence of obesity and immune-related gastrointestinal disorders among shift workers.