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Colorectal cancer (CRC) is the third most common cancer and the second leading cause of death due to cancer in the world. Therefore, the identification of novel druggable targets is urgently needed. Tribbles proteins belong to a pseudokinase family, previously recognized in CRC as oncogenes and potential therapeutic targets. Here, we analyzed the expression of TRIB1, TRIB2, and TRIB3 simultaneously in 33 data sets from CRC based on available GEO profiles. We show that all three Tribbles genes are overrepresented in CRC cell lines and primary tumors, though depending on specific features of the CRC samples. Higher expression of TRIB2 in the tumor microenvironment and TRIB3 overexpression in an early stage of CRC development, unveil a potential and unexplored role for these proteins in the context of CRC. Differential Tribbles expression was also explored in diverse cellular experimental conditions where either genetic or pharmacological approaches were used, providing novel hints for future research. This comprehensive bioinformatic analysis provides new insights into Tribbles gene expression and transcript regulation in CRC. info:eu-repo/semantics/publishedVersion

Decavanadate is a polyoxometalate (POMs) that has shown extensive biological activities, including antidiabetic and anticancer activity. Importantly, vanadium-based compounds as well as antidiabetic biguanide drugs, such as metformin, have shown to exert therapeutic effects in melanoma. A combination of these agents, the metformin-decavanadate complex, was also recognized for its antidiabetic effects and recently described as a better treatment than the monotherapy with metformin enabling lower dosage in rodent models of diabetes. Herein, we compare the effects of decavanadate and metformin-decavanadate on Ca2+-ATPase activity in sarcoplasmic reticulum vesicles from rabbit skeletal muscles and on cell signaling events and viability in human melanoma cells. We show that unlike the decavanadate-mediated non-competitive mechanism, metformin-decavanadate inhibits Ca2+-ATPase by a mixed-type competitive–non-competitive inhibition with an IC50 value about 6 times higher (87 μM) than the previously described for decavanadate (15 μM). We also found that both decavanadate and metformin-decavanadate exert antiproliferative effects on melanoma cells at 10 times lower concentrations than monomeric vanadate. Western blot analysis revealed that both, decavanadate and metformin-decavanadate increased phosphorylation of extracellular signal-regulated kinase (ERK) and serine/threonine protein kinase AKT signaling proteins upon 24 h drug exposure, suggesting that the anti-proliferative activities of these compounds act independent of growth-factor signaling pathways., This work was funded by the Spanish Ministry of Science, Innovation and Universities through grant RTI2018-094629-B-I00 to WL, and Fundação para a Ciência e a Tecnologia (FCT) (PTDC/MED-ONC/4167/2020 “ENDURING”) to BIF.

Lipid peroxidation (LPO), a process that affects human health, can be induced by exposure to vanadium salts and compounds. LPO is often exacerbated by oxidation stress, with some forms of vanadium providing protective effects. The LPO reaction involves the oxidation of the alkene bonds, primarily in polyunsaturated fatty acids, in a chain reaction to form radical and reactive oxygen species (ROS). LPO reactions typically affect cellular membranes through direct effects on membrane structure and function as well as impacting other cellular functions due to increases in ROS. Although LPO effects on mitochondrial function have been studied in detail, other cellular components and organelles are affected. Because vanadium salts and complexes can induce ROS formation both directly and indirectly, the study of LPO arising from increased ROS should include investigations of both processes. This is made more challenging by the range of vanadium species that exist under physiological conditions and the diverse effects of these species. Thus, complex vanadium chemistry requires speciation studies of vanadium to evaluate the direct and indirect effects of the various species that are present during vanadium exposure. Undoubtedly, speciation is important in assessing how vanadium exerts effects in biological systems and is likely the underlying cause for some of the beneficial effects reported in cancerous, diabetic, neurodegenerative conditions and other diseased tissues impacted by LPO processes. Speciation of vanadium, together with investigations of ROS and LPO, should be considered in future biological studies evaluating vanadium effects on the formation of ROS and on LPO in cells, tissues, and organisms as discussed in this review. info:eu-repo/semantics/publishedVersion

Background TRIB1, TRIB2 and TRIB3 belong to the mammalian Tribbles family of pseudokinases proteins. Several studies reported Tribbles oncogenic role in different types of cancer, including colorectal cancer (CRC). Though current CRC treatment can be curative, patients are in risk of disease recurrence, meaning novel pharmacological targets and strategies are required. Our goal was to analyze Tribbles gene expression in CRC in response to different drugs. Methods Tribbles transcript levels were obtained from GEO profiles database (NCBI). Gene data sets (GDS) were selected based on experimental drug treatment description. Statistical analysis was performed at GraphPadPrism. Results Compared to non-treated control, TRIB2 expression was ∼2-fold increased in colorectal adenocarcinoma samples from patients treated with cyclooxygenase-2 inhibitor celecoxib (GDS3384), though not statistically significant (P < 0.1). TRIB1 was unaltered and data for TRIB3 was not available. By contrast, all Tribbles showed differential expression after treatment of SW620 colon cancer cells with supercritical rosemary extract in progressive increasing doses (0, 30, 60, 100 μg/mL) (P < 0.01;GDS5416). While both TRIB1 and TRIB3 were moderately increased in a dose-dependent manner (∼18% and 13%, respectively), TRIB2 was maximally down-regulated by ∼15% after 60 μg/mL. Conclusions Although celecoxib exhibits antiproliferative effects in different cancer cell types, TRIB2 gene expression showed a trend to be induced after treatment, in contrast to several genes involved in fatty acid oxidation that were down-regulated, which could result from a compensatory mechanism based on a metabolic shift. Since TRIB1/TRIB3 and TRIB2 were oppositely modulated in response to rosemary extract, additional studies are needed to validate its specific pharmacological potential interest for CRC treatment.

Background A recent repurposing pharmacological screening revealed that vanadium-containing drugs anti-proliferative action in ovarian cancer cells was SLC26A2-dependent. SLC26A2/DTDST is a sulfate transporter, related to chondrodysplasia syndromes. Despite some reports on colon cancer, there are no studies on SLC26A2 performed in melanoma in the literature. Methods To better understand its potential use as biomarker for therapeutic decisions in melanoma, we performed gene expression analyses of the data available at GEO profiles (NCBI). Gene data sets that allowed analysis of SLC26A2 expression (1) in melanoma; (2) in response to drugs; (3) regulated by other proteins, were selected. Results Our results showed that, compared to normal skin or benign nevi, SLC26A2 expression was 2.5-fold higher in malignant melanoma (P = 0.019). Compared to the primary tumor, SLC26A2 expression tripled in melanoma (P = 0.022). We found a 6% decrease of SLC26A2 expression in A375 melanoma cells treated with BRAF inhibitor Vemurafenib (P < 0.001). After treatment of A375 cells with MLN4924, a selective inhibitor of the activating enzyme of Nedd8, SLC26A2 decreased in a time-dependent manner ( > 80% at 24 h; P < 0.001). In Sk-Mel-2 cells overexpressing E2F-1, a transcription factor that induces apoptosis in cancer cells, SLC26A2 levels were reduced by 76.4% (P = 0.067). In A375P cells depleted of PGC1α, an important metabolic co-activator in mitochondrial biogenesis and function, SLC26A2 levels increased 16% (P = 0.013). Conclusions From this work, we unveiled, for the first time, potential clues to better understand the regulation and role of SLC26A2 in melanoma. Though, it is still to be determined whether SLC26A2 is a driver or a passenger in the disease.

The application of metals in biological systems has been a rapidly growing branch of science. Vanadium has been investigated and reported as an anticancer agent. Melanoma is the most aggressive type of skin cancer, the incidence of which has been increasing annually worldwide. It is of paramount importance to identify novel pharmacological agents for melanoma treatment. Herein, a systematic review of publications including “Melanoma and Vanadium” was performed. Nine vanadium articles in several melanoma cells lines such as human A375, human CN-mel and murine B16F10, as well as in vivo studies, are described. Vanadium-based compounds with anticancer activity against melanoma include: (1) oxidovanadium(IV); (2) XMenes; (3) vanadium pentoxide, (4) oxidovanadium(IV) pyridinonate compounds; (5) vanadate; (6) polysaccharides vanadium(IV/V) complexes; (7) mixed-metal binuclear ruthenium(II)–vanadium(IV) complexes; (8) pyridoxal-based oxidovanadium(IV) complexes and (9) functionalized nanoparticles of yttrium vanadate doped with europium. Vanadium compounds and/or vanadium materials show potential anticancer activities that may be used as a useful approach to treat melanoma.

© 2021 by the authors., The Tribbles family of pseudokinases controls a wide number of processes during cancer on-set and progression. However, the exact contribution of each of the three family members is still to be defined. Their function appears to be context-dependent as they can act as oncogenes or tumor suppressor genes. They act as scaffolds modulating the activity of several signaling pathways involved in different cellular processes. In this review, we discuss the state-of-knowledge for TRIB1, TRIB2 and TRIB3 in the development and progression of colorectal cancer. We take a perspective look at the role of Tribbles proteins as potential biomarkers and therapeutic targets. Specifically, we chronologically systematized all available articles since 2003 until 2020, for which Tribbles were associated with colorectal cancer human samples or cell lines. Herein, we discuss: (1) Tribbles amplification and overexpression; (2) the clinical significance of Tribbles overexpression; (3) upstream Tribbles gene and protein expression regulation; (4) Tribbles pharmacological modulation; (5) genetic modulation of Tribbles; and (6) downstream mechanisms regulated by Tribbles; establishing a comprehensive timeline, essential to better consolidate the current knowledge of Tribbles’ role in colorectal cancer., This work was supported by the Spanish Ministry of Science, Innovation and Universities through Grant RTI2018-094629-B-I00 to W.L. This work was supported by the European Commission project TRIBBLES–748585 and Fundação para a Ciência e a Tecnologia (FCT) (grant PTDC/MEDONC/4167/2020 “ENDURING”) to B.I.F.

Background Polyoxometalates are a diverse family of metal-oxo anions of early transitional metal ions in high oxidation states, including vanadium. Such compounds are gaining increasing interest in biomedicine due to their anti-cancer activity. Regarding melanoma, available therapies’ effectiveness is still currently limited, commonly leading to relapse. For this reason, it is vital to identify novel anti-cancer agents. Methods To better understand the mechanism of action involved in the anti-cancer vanadium-based compounds in melanoma, we conducted a systematic review. Search was performed in Clinical Trials, Cochrane Library, Pubmed, Scielo, and Web of Science (20/4-8/5/2020). Review articles were excluded. Inclusion criteria considered articles between 2000 and 2020, written in English, in which vanadium compounds were evaluated alone (not combined with other drugs), allowing to recognise the effects exclusively derived from vanadium. Results Based on the inclusion/exclusion criteria, a total of 5 articles, published between 2013 and 2020, were analyzed. All tested compounds, including Oxidovanadium(IV) complexes [VIVO(mpp)2] and [VIVO(ppp)2] (VS3, VS4), reduced cellular viability. Moreover, Vanadium pentoxide (V2O5) and Xyloglucan from Copaifera Langsdorffii complexed with Oxovanadium(IV/V) (XGC:VO), in murine B16F10 cells, and Inorganic anion vanadate (VN), [VIVO (dhp)2] (VS2), and N,N’-ethylenebis (pyridoxylideneiminato) vanadium(IV) complex [Pyr2enV(IV)], in A375 cells, induced apoptosis. VN, VS2 and Pyr2enV(IV) increased reactive oxygen species (ROS) levels in A375 cells. In vivo experiments demonstrated that V2O5 prolonged the life of mice implanted with B16F10 cells, with no systemic toxicity. Conclusions Several different vanadium-based compounds have anti-cancer potential in melanoma, mainly increasing apoptosis and ROS production. Specific metal-based complexes are potential melanoma treatments, which might have been previously neglected.

Simple Summary Poor survival and treatment failure of patients with cancer are mainly due to resistance to therapy. Tribbles homologue 2 (TRIB2) has recently been identified as a protein that promotes resistance to several anti-cancer drugs. In this study, RNA sequencing and bioinformatics analysis were used with the aim of characterizing the impact of TRIB2 on the expression of genes and developing pharmacological strategies to revert these TRIB2-mediated changes, thereby overcoming therapy resistance. We show that two naturally occurring alkaloids, harmine and piperlongumine, inverse the gene expression profile produced by TRIB2 and sensitize cancer cells to anti-cancer drugs. Our data suggest that harmine and piperlongumine or similar compounds might have the potential to overcome TRIB2-mediated therapy resistance in cancer patients. Abstract Therapy resistance is responsible for most relapses in patients with cancer and is the major challenge to improving the clinical outcome. The pseudokinase Tribbles homologue 2 (TRIB2) has been characterized as an important driver of resistance to several anti-cancer drugs, including the dual ATP-competitive PI3K and mTOR inhibitor dactolisib (BEZ235). TRIB2 promotes AKT activity, leading to the inactivation of FOXO transcription factors, which are known to mediate the cell response to antitumor drugs. To characterize the downstream events of TRIB2 activity, we analyzed the gene expression profiles of isogenic cell lines with different TRIB2 statuses by RNA sequencing. Using a connectivity map-based computational approach, we identified drug-induced gene-expression profiles that invert the TRIB2-associated expression profile. In particular, the natural alkaloids harmine and piperlongumine not only produced inverse gene expression profiles but also synergistically increased BEZ235-induced cell toxicity. Importantly, both agents promote FOXO nuclear translocation without interfering with the nuclear export machinery and induce the transcription of FOXO target genes. Our results highlight the great potential of this approach for drug repurposing and suggest that harmine and piperlongumine or similar compounds might be useful in the clinic to overcome TRIB2-mediated therapy resistance in cancer patients.

Flow-limiting atherosclerotic lesions of arteries supplying the limbs are a cause of symptoms in patients with peripheral artery disease (PAD). Musculoskeletal metabolic factors also contribute to the pathophysiology of claudication, which is manifest as leg discomfort that impairs walking capacity. Accordingly, we conducted a case-control study to determine whether skeletal muscle metabolic gene expression is altered in PAD. Calf skeletal muscle gene expression of patients with PAD and healthy subjects was analyzed using microarrays. The top-ranking gene differentially expressed between PAD and controls (FDR < 0.001) wasPLA2G16, which encodes adipose-specific phospholipase A2 (AdPLA) and is implicated in the maintenance of insulin sensitivity and regulation of lipid metabolism. Differential expression was confirmed by qRT-PCR;PLA2G16was downregulated by 68% in patients with PAD (p< 0.001). Expression ofPla2g16was then measured in control (db/+) and diabetic (db/db) mice that underwent unilateral femoral artery ligation. There was significantly reduced expression ofPla2g16in the ischemic leg of both control and diabetic mice (by 51%), with significantly greater magnitude of reduction in the diabetic mice (by 79%). We conclude that AdPLA is downregulated in humans with PAD and in mice with hindlimb ischemia. Reduced AdPLA may contribute to impaired walking capacity in patients with PAD via its effects on skeletal muscle metabolism. Further studies are needed to fully characterize the role of AdPLA in PAD and to investigate its potential as a therapeutic target for alleviating symptoms of claudication. American Diabetes AssociationAmerican Diabetes Association [K12HL083786] National Heart, Lung, and Blood InstituteUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Heart Lung & Blood Institute (NHLBI) [K12HL083786] American Heart AssociationAmerican Heart Association [10SDG4200060, 18SFRN33900085] Joslin Diabetes Research Center [P30 DK036836] info:eu-repo/semantics/publishedVersion

Poor paternal diet has emerged as a risk factor for metabolic disease in offspring, and alterations in sperm may be a major mechanism mediating these detrimental effects of diet. Although exercise in the general population is known to improve health, the effects of paternal exercise on sperm and offspring metabolic health are largely unknown. Here, we studied 7-week-old C57BL/6 male mice fed a chow or high-fat diet and housed either in static cages (sedentary) or cages with attached running wheels (exercise trained). After 3 weeks, one cohort of males was sacrificed and cauda sperm obtained, while the other cohort was bred with chow-fed sedentary C57BL/6 females. Offspring were chow fed, sedentary, and studied during the first year of life. We found that high-fat feeding of sires impairs glucose tolerance and increases the percentage of fat mass in both male and female offspring at 52 weeks of age. Strikingly, paternal exercise suppresses the effects of paternal high-fat diet on offspring, reversing the observed impairment in glucose tolerance, percentage of fat mass, and glucose uptake in skeletal muscles of the offspring. These changes in offspring phenotype are accompanied by changes in sperm physiology, as, for example, high-fat feeding results in decreased sperm motility, an effect normalized in males subject to exercise training. Deep sequencing of sperm reveals pronounced effects of exercise training on multiple classes of small RNAs, as multiple changes to the sperm RNA payload observed in animals consuming a high-fat diet are suppressed by exercise training. Thus, voluntary exercise training of male mice results in pronounced improvements in the metabolic health of adult male and female offspring. We provide the first in-depth analysis of small RNAs in sperm from exercise-trained males, revealing a marked change in the levels of multiple small RNAs with the potential to alter phenotypes in the next generation.

Objective Plasma levels of branched-chain amino acids (BCAA) are consistently elevated in obesity and type 2 diabetes (T2D) and can also prospectively predict T2D. However, the role of BCAA in the pathogenesis of insulin resistance and T2D remains unclear. Methods To identify pathways related to insulin resistance, we performed comprehensive gene expression and metabolomics analyses in skeletal muscle from 41 humans with normal glucose tolerance and 11 with T2D across a range of insulin sensitivity (SI, 0.49 to 14.28). We studied both cultured cells and mice heterozygous for the BCAA enzyme methylmalonyl-CoA mutase (Mut) and assessed the effects of altered BCAA flux on lipid and glucose homeostasis. Results Our data demonstrate perturbed BCAA metabolism and fatty acid oxidation in muscle from insulin resistant humans. Experimental alterations in BCAA flux in cultured cells similarly modulate fatty acid oxidation. Mut heterozygosity in mice alters muscle lipid metabolism in vivo, resulting in increased muscle triglyceride accumulation, increased plasma glucose, hyperinsulinemia, and increased body weight after high-fat feeding. Conclusions Our data indicate that impaired muscle BCAA catabolism may contribute to the development of insulin resistance by perturbing both amino acid and fatty acid metabolism and suggest that targeting BCAA metabolism may hold promise for prevention or treatment of T2D., Highlights • Human insulin resistance is associated with perturbed muscle BCAA metabolism. • Experimental modulation of BCAA metabolic flux alters fatty acid oxidation in vitro. • Mut heterozygosis leads to increased body weigh and muscle TAG accumulation in mice.

Context: Increased circulating free fatty acids (FFAs) have been proposed to contribute to insulin resistance in obesity. Short-term studies have investigated the effects of acipimox, an inhibitor of hormone-sensitive lipase, on glucose homeostasis, but longer-term studies have not been performed. Objective: To test the hypothesis that long-term treatment with acipimox would reduce FFA and improve insulin sensitivity among nondiabetic, insulin-resistant, obese subjects. Design, Setting, Patients, and Intervention: At an academic medical center, 39 obese men and women were randomized to acipimox 250 mg thrice-daily vs identical placebo for 6 months. Main Outcome Measures: Plasma lipids, insulin sensitivity, adiponectin, and mitochondrial function via assessment of the rate of post-exercise phosphocreatine recovery on 31P-magnetic resonance spectroscopy as well as muscle mitochondrial density and relevant muscle gene expression. Results: Fasting glucose decreased significantly in acipimox-treated individuals (effect size, −6 mg/dL; P = .02), in parallel with trends for reduced fasting insulin (effect size, −6.8 μU/mL; P = .07) and HOMA-IR (effect size, −1.96; P = .06), and significantly increased adiponectin (effect size, +668 ng/mL; P = .02). Acipimox did not affect insulin-stimulated glucose uptake, as assessed by euglycemic, hyperinsulinemic clamp. Effects on muscle mitochondrial function and density and on relevant gene expression were not seen. Conclusion: These data shed light on the long-term effects of FFA reduction on insulin sensitivity, other metabolic parameters, and muscle mitochondrial function in obesity. Reduced FFA achieved by acipimox improved fasting measures of glucose homeostasis, lipids, and adiponectin but had no effect on mitochondrial function, mitochondrial density, or muscle insulin sensitivity.

Rationale, aims, and objectives Health literacy (HL) has been widely referenced as a determinant of health outcomes, making the assessment of low HL a fundamental step to plan educational interventions. This study aimed to translate and adapt the Short Assessment of Health Literacy-Spanish and English (SAHL-S&E) questionnaire into European Portuguese. Methods The SAHL-S&E questionnaire was translated using the recommendations of the International Society for Pharmacoeconomics and Outcomes Research. One hundred fifty-three Portuguese native speakers aged over 18 years old were included in this study, enrolled among users of community pharmacies in the Algarve region (Portugal). Results The translation of the questionnaire used showed a good internal consistency (Cronbach alpha: .812), and a statistically significant (F= 5.05P< .001) interrater reliability. Over a third of subjects (37.9%) achieved a score less than or equal to 14, which is indicative of low HL. Conclusion This tool, intended to be used in the European Portuguese population, can be used for low HL screening. info:eu-repo/semantics/publishedVersion

Nutrient deprivation or starvation frequently correlates with amino acid limitation. Amino acid starvation initiates a signal transduction cascade starting with the activation of the kinase GCN2 (general control non-derepressible 2) phosphorylation of eIF2 (eukaryotic initiation factor 2), global protein synthesis reduction and increased ATF4 (activating transcription factor 4). ATF4 modulates a wide spectrum of genes involved in the adaptation to dietary stress. The hormone FGF21 (fibroblast growth factor 21) is induced during fasting in liver and its expression induces a metabolic state that mimics long-term fasting. Thus FGF21 is critical for the induction of hepatic fat oxidation, ketogenesis and gluconeogenesis, metabolic processes which are essential for the adaptive metabolic response to starvation. In the present study, we have shown that FGF21 is induced by amino acid deprivation in both mouse liver and cultured HepG2 cells. We have identified the human FGF21 gene as a target gene for ATF4 and we have localized two conserved ATF4-binding sequences in the 5′ regulatory region of the human FGF21 gene, which are responsible for the ATF4-dependent transcriptional activation of this gene. These results add FGF21 gene induction to the transcriptional programme initiated by increased levels of ATF4 and offer a new mechanism for the induction of the FGF21 gene expression under nutrient deprivation.

HMGCS2 (hydroxymethylglutaryl CoA synthase 2), the gene that regulates ketone body production, is barely expressed in cultured cell lines. In this study, we restored HMGCS2 expression and activity in HepG2 cells, thus showing that the wild type enzyme can induce fatty acid β-oxidation (FAO) and ketogenesis, whereas a catalytically inactive mutant C166A did not generate either process. Peroxisome proliferator-activated receptor (PPAR) α expression also induces fatty acid β-oxidation and endogenous HMGCS2 expression. Interestingly, PPARα-mediated induction was abolished when HMGCS2 expression was down-regulated by RNAi. These results indicate that HMGCS2 expression is both sufficient and necessary to the control of fatty acid oxidation in these cells. Next, we examined the expression pattern of several PPARα target genes in this now “ketogenic” HepG2 cell line. FGF21 (fibroblast growth factor 21) expression was specifically induced by HMGCS2 activity or by the inclusion of the oxidized form of ketone bodies (acetoacetate) in the culture medium. This effect was blunted by SirT1 (sirtuin 1) RNAi, so we propose a SirT1-dependent mechanism for FGF21 induction by acetoacetate. These data suggest a novel feed-forward mechanism by which HMGCS2 could regulate adaptive metabolic responses during fasting. This mechanism could be physiologically relevant, because fasting-mediated induction of liver FGF21 was dependent on SirT1 activity in vivo.

Acetoacetyl-CoA synthetase (AACS) is a cytoplasmic enzyme that activates the ketone body acetoacetate for lipogenesis. Aacs is expressed in liver, thus this tissue produces and consumes ketone bodies. To understand this apparent paradox we studied the expression of Aacs in liver during the transition from fasting to feeding, and characterized the kinetic parameters of the human enzyme. We show that Aacs liver expression is down-regulated by fasting and only up-regulated after 5 h of refeeding, while the levels of circulating ketone bodies returned to basal levels within 20 min of food availability. Since the human enzyme has a high affinity for the ketone body acetoacetate (KM = 37.6 μM), these results indicates that AACS utilizes ketones during periods of feeding. Human AACS was also characterized as an acetylated enzyme in vivo, where lysine 633 (from a P4XGK domain) plays a critical role in the catalytic activity but not in the acetylation state of the protein.

FSP27 [cell death-inducing DFFA-like effector c (CIDEC) in humans] is a protein associated with lipid droplets that downregulates the fatty acid oxidation (FAO) rate when it is overexpressed. However, little is known about its physiological role in liver. Here, we show that fasting regulates liver expression of Fsp27 in a time-dependent manner. Thus, during the initial stages of fasting, a maximal induction of 800-fold was achieved, whereas during the later phase of fasting, Fsp27 expression decreased. The early response to fasting can be explained by a canonical PKA-CREB-CRTC2 signaling pathway because: i) CIDEC expression was induced by forskolin, ii) Fsp27 promoter activity was increased by CREB, and iii) Fsp27 expression was upregulated in the liver of Sirt1 knockout animals. Interestingly, pharmacological (etomoxir) or genetic (Hmgcs2 interference) inhibition of the FAO rate increases the in vivo expression of Fsp27 during fasting. Similarly, CIDEC expression was upregulated in HepG2 cells by either etomoxir or HMGCS2 interference. Our data indicate that there is a kinetic mechanism of autoregulation between short- and long-term fasting, by which free FAs delivered to the liver during early fasting are accumulated/exported by FSP27/CIDEC, whereas over longer periods of fasting, they are degraded in the mitochondria through the carnitine palmitoyl transferase system.

Lipogenic gene expression in liver is repressed in mice upon leucine deprivation. The hormone fi broblast growth factor 21 (FGF21), which is critical to the adaptive metabolic response to starvation, is also induced under amino acid deprivation. Upon leucine deprivation, we found that FGF21 is needed to repress expression of lipogenic genes in liver and white adipose tissue, and stimulate phosphorylation of hormone-sensitive lipase in white adipose tissue. The increased expression of Ucp1 in brown adipose tissue under these circumstances is also impaired in FGF21- defi cient mice. Our results demonstrate the important role of FGF21 in the regulation of lipid metabolism during amino acid starvation. ¿De Sousa-Coelho, A. L., J. Relat, E. Hondares, A. Pérez-Martí, F. Ribas, F. Villarroya, P. F. Marrero, and D. Haro. FGF21 mediates the lipid metabolism response to amino acid starvation.