NAMPT and BMAL1 Are Independently Involved in the Palmitate-Mediated Induction of Neuroinflammation in Hypothalamic Neurons.

Obesity is a prominent metabolic disease that predisposes individuals to multiple comorbidities, including type 2 diabetes mellitus, cardiovascular diseases, and cancer. Elevated circulating levels of fatty acids contribute to the development of obesity, in part, by targeting the hypothalamus. Palmitate, the most abundant circulating saturated fatty acid, has been demonstrated to dysregulate NAMPT and circadian clock proteins, as well as induce neuroinflammation. These effects ultimately result in hypothalamic dysregulation of feeding behavior and energy homeostasis. NAMPT is the rate-limiting enzyme of the NAD+ salvage pathway and its expression is under the control of the circadian clock. NAD+ produced from NAMPT can modulate the circadian clock, demonstrating bidirectional interactions between circadian and metabolic pathways. Using NPY/AgRP-expressing mHypoE-46 neurons as well as the novel mHypoA-BMAL1-WT/F and mHypoA-BMAL1-KO/F cell lines, we studied whether there were any interactions between NAMPT and the core circadian clock protein BMAL1 in the palmitate-mediated induction of neuroinflammation. We report that palmitate altered Nampt, Bmal1, Per2 and the inflammatory genes Nf-κb, IκBα, Il-6, and Tlr4. Contrary to studies performed with peripheral tissues, the palmitate-mediated induction in Nampt was independent of BMAL1, and basal Nampt levels did not appear to exhibit rhythmic expression. Palmitate-induced downregulation of Bmal1 and Per2 was independent of NAMPT. However, NAMPT and BMAL1 were both involved in the regulation of Nf-κb, IκBα, Il-6, and Tlr4, as NAMPT inhibition resulted in the repression of basal Nf-κb and IκBα and normalized palmitate-mediated increases in Il-6, and Tlr4. On the other hand, BMAL1 deletion repressed basal Nf-κb, but increased basal Il-6. We conclude that NAMPT and BMAL1 do not interact at the transcriptional level in hypothalamic neurons, but are independently involved in the expression of inflammatory genes.

Analysis of Western diet, palmitate and BMAL1 regulation of neuropeptide Y expression in the murine hypothalamus and BMAL1 knockout cell models.

Western diets that are high in saturated fat and sugar disrupt circadian rhythms, induce weight gain, and lead to metabolic diseases including obesity. However, the mechanistic link between altered circadian rhythms and energy homeostasis remains poorly understood. In C57BL/6J mice, consuming a Western diet for 16 weeks significantly reduced food intake (at zeitgeber 12-16), in association with decreases in hypothalamic expression of the orexigenic neuropeptides, neuropeptide Y (Npy) and agouti-related peptide (AgRP). To examine the acute effects of the most prevalent saturated fatty acid in a Western diet, palmitate, and the role of the core clock gene, Bmal1, in the regulation of hypothalamic feeding neuropeptides, we used heterogeneous and clonal BMAL1 knockout (KO) immortalized hypothalamic cell lines, expressing specific neuropeptides, derived from male (M) and female (F) mice. Both mHypoA-BMAL1-KO/F and mHypoA-BMAL1-KO/M cells demonstrated a loss of circadian rhythmicity in expression of the clock gene, Per2, as compared to wild-type (control) cultures. Loss of BMAL1 also altered the time-dependent expression of Npy and proopiomelanocortin, and disrupted AgRP rhythmicity. Furthermore, palmitate increased BMAL1 binding to the Npy promotor region, and palmitate treatment (50 µM for 24 h) stimulated Npy expression in a BMAL1-dependent manner in both heterogeneous and clonal NPY-expressing female-derived cell models. The results of this study demonstrate that circadian expression of Bmal1 serves as a mechanistic link between Western diet- and palmitate-induced disruptions of the normal rhythmic patterns in hypothalamic feeding-related neuropeptides.

[Evaluation of Naturally Occurring Compounds Regulating Brown/Beige Adipocyte Differentiation].

Brown adipose tissue is a critical regulator of metabolic health, and contributes to thermogenesis by uncoupling oxidative phosphorylation through the action of mitochondrial uncoupling protein 1 (Ucp1). Recent studies have shown that cold exposure and the stimulation of ß3-adrenergic receptors induce the development of brown cell-like "beige" adipocytes in white adipose tissue. Brown and/or beige adipocyte-mediated thermogenesis suppresses high-fat diet-associated obesity. Therefore, the development of brown/beige adipocytes may prevent obesity and metabolic diseases. In the present study, we elucidated whether naturally occurring compounds contribute to regulating the cellular differentiation of brown/beige adipocytes. We screened for the up-regulated expression of Ucp1 during beige adipogenesis using extracts of crude herbal drugs frequently used in Kampo prescriptions (therapeutic drugs in Japanese traditional medicine). This screening revealed that the extract prepared from Citri Unshiu Pericarpium [the peel of Citrus unshiu (Swingle) Marcov.] increased the expression of Ucp1 in beige adipocytes. We also focused on the function of clock genes in regulating brown/beige adipogenesis. Therefore, another aim of the present study was to evaluate naturally occurring compounds that regulate brain and muscle Arnt-like 1 (Bmal1) gene expression. In this review, we focus on naturally occurring compounds that affect regulatory processes in brown/beige adipogenesis, and discuss better preventive strategies for the management of obesity and other metabolic disorders.

The epigenetic language of circadian clocks.

Epigenetic control, which includes DNA methylation and histone modifications, leads to chromatin remodeling and regulated gene expression. Remodeling of chromatin constitutes a critical interface of transducing signals, such as light or nutrient availability, and how these are interpreted by the cell to generate permissive or silenced states for transcription. CLOCK-BMAL1-mediated activation of clock-controlled genes (CCGs) is coupled to circadian changes in histone modification at their promoters. Several chromatin modifiers, such as the deacetylases SIRT1 and HDAC3 or methyltransferase MLL1, have been shown to be recruited to the promoters of the CCGs in a circadian manner. Interestingly, the central element of the core clock machinery, the transcription factor CLOCK, also possesses histone acetyltransferase activity. Rhythmic expression of the CCGs is abolished in the absence of these chromatin modifiers. Here we will discuss the evidence demonstrating that chromatin remodeling is at the crossroads of circadian rhythms and regulation of metabolism and cellular proliferation.

[Chronobiological aspects of obesity and metabolic syndrome]. / Aspectos cronobiológicos de la obesidad y el síndrome metabólico.

Circadian rhythms (approximately 24h) are widely characterized at molecular level and their generation is acknowledged to originate from oscillations in expression of several clock genes and from regulation of their protein products. While general entrainment of organisms to environmental light-dark cycles is mainly achieved through the master clock of the suprachiasmatic nucleus in mammals, this molecular clockwork is functional in several organs and tissues. Some studies have suggested that disruption of the circadian system (chronodisruption (CD)) may be causal for manifestations of the metabolic syndrome. This review summarizes (1) how molecular clocks coordinate metabolism and their specific role in the adipocyte; (2) the genetic aspects of and scientific evidence for obesity as a chronobiological illness; and (3) CD and its causes and pathological consequences. Finally, ideas about use of chronobiology for the treatment of obesity are discussed.

Antioxidant N-acetyl-L-cysteine ameliorates symptoms of premature aging associated with the deficiency of the circadian protein BMAL1.

Deficiency of the circadian clock protein BMAL1 leads to premature aging and increased levels of reactivate oxygen species in several tissues of mice. In order to investigate the role of oxidative stress in accelerated aging and development of age-related pathologies, we continuously administered the antioxidant N-acetyl-L-cysteine toBmal1-deficient mice through their entire lifespan by supplementing drinking water. We found that the life long treatment with antioxidant significantly increased average and maximal lifespan and reduced the rate of age-dependent weight loss and development of cataracts. At the same time, it had no effect on time of onset and severity of other age-related pathologies characteristic of Bmal1-/- mice, such as joint ossification, reduced hair regrowth and sarcopenia. We conclude that chronic oxidative stress affects longevity and contributes to the development of at least some age-associated pathology, although ROS-independent mechanisms may also play a role. Our bioinformatics analysis identified the presence of a conservative E box element in the promoter regions of several genes encoding major antioxidant enzymes. We speculate that BMAL1 controls antioxidant defense by regulating the expression of major antioxidant enzymes.