The role of Clock in the plasticity of circadian entrainment

The mammalian circadian clock lying in suprachiasmatic nucleus (SCN) is synchronized to about 24 h by the environmental light-dark cycle (LD). The circadian clock exhibits limits of entrainment above and below 24 h, beyond which it will not entrain. Little is known about the mechanisms regulating the limits of entrainment. In this study, we show that wild-type mice entrain to only an LD 24 h cycle, whereas Clock mutant mice can entrain to an LD 24, 28, and 32 h except for LD 20 h and LD 36 h cycle. Under an LD 28 h cycle, Clock mutant mice showed a clear rhythm in Per2 mRNA expression in the SCN and behavior. Light response was also increased. This is the first report to show that the Clock mutation makes it possible to adapt the circadian oscillator to a long period cycle and indicates that the clock gene may have an important role for the limits of entrainment of the SCN to LD cycle.     

Involvement of circadian clock gene Clock in diabetes-induced circadian augmentation of plasminogen activator inhibitor-1 (PAI-1) expression in the mouse heart

Diabetes is associated with an excess risk of cardiac events, and one of the risk factors for infarction is the elevated-levels of plasminogen activator inhibitor-1 (PAI-1). To evaluate how the molecular clock mechanism is involved in the diabetes-induced circadian augmentation of PAI-1 gene expression, we examined the expression profiles of PAI-1 mRNA in the hearts of Clock mutant mice with streptozotocin-induced diabetes. Circadian expression of PAI-1 mRNA was blunted to low levels under both normal and diabetic conditions in Clock mutant mice, although the expression rhythm was augmented in diabetic wild-type (WT) mice. Furthermore, plasma PAI-1 levels became significantly higher in WT mice than in Clock mutant mice after STZ administration. Our results suggested that the circadian clock component, CLOCK, is involved in the diabetes-induced circadian augmentation of PAI-1 expression in the mouse heart.     

Combination of meal and exercise timing with a high-fat diet influences energy expenditure and obesity in mice

In mice, obesity has been observed not only in those freely fed a high-fat diet (HFD) but also in those fed while physically inactive. In contrast, a HFD during physically active periods protects against obesity and the impairments in the circadian rhythm induced by free feeding of a HFD. Although exercise is known to be effective for obesity prevention and management, the optimal timing of exercise has not yet been determined. In the present experiments, we aimed to determine the best combination of daily timing of HFD consumption and exercise for the prevention of HFD-induced weight gain in mice. In this experiment, “morning” refers to the beginning of the active phase (the “morning” for nocturnal animals). Increases in body weight related to free feeding of a HFD was significantly reduced with 4?h of exercise during the late (evening) or middle (noon) active period compared to 4?h of exercise during the early (morning) active period or free access to exercise, which resulted in hours of exercise similar to that of morning exercise. These results suggested that eating in the morning or at noon followed by exercise in the evening could prevent weight gain more effectively than exercise in the morning followed by eating at noon or in the evening. The group fed a HFD for 4?h in the morning had lower body weight than the group fed a HFD for 4?h in the evening without exercise. The last group of experiments tested the hypothesis that there would be an interaction between mealtime and exercise time (i.e. time of day) versus order (i.e. which comes first) effects. We compared groups that exercised for 4?h at noon and were fed either in the morning or evening and groups that were fed for 4?h at noon and either exercised in the morning or evening. We found that the groups that were fed before exercise gained less body and fat weight and more skeletal muscle weight compared to the groups that exercised before eating. Corresponding to the body and fat weight changes, the respiratory exchange ratio (RER) was lower and energy expenditure was higher in the groups fed before exercise than in the groups fed after exercise, and these effects on energy metabolism were also observed in the early stage of HFD feeding before obesity. When obese mice fed a HFD for 12 weeks were exposed to a combination of feeding and exercise timing in an effort to reduce body weight, eating followed by exercise resulted in greater weight loss, similar to the experiments conducted to prevent weight gain. These results demonstrate that a combination of daily timing of eating and exercise may influence weight gain and that eating followed by exercise may be effective for minimizing increases in body and fat weight as well as maximizing increases in skeletal muscle weight.     

Controlling access time to a high-fat diet during the inactive period protects against obesity in mice

Free feeding (FF) with a high fat diet (HFD) causes excessive body weight gain, whereas restricted feeding (RF) with a HFD attenuates body weight gain. The effects of timing of feeding with a HFD (day vs. night) and feeding duration on energy homeostasis have not yet been investigated. In this study, we fed mice a HFD or a normal diet (ND) twice a day, during their active and inactive periods, on a schedule. The amount of food was regulated by feeding duration (2, 4 or 8?h). First, we investigated the effects of 4-h RF during active–inactive periods (ND–ND, HFD–HFD, ND–HFD or HFD–ND). Among all the 4-h RF groups, mice consumed almost the same amount of calories as those in the FF[ND] group, even those fed a HFD. Body weight and visceral fat in these three groups were lower than that in the FF[HFD] group. Second, we investigated the effects of RF duration. Body weight and visceral fat were higher in the 8-h groups than in the 4-h groups. Body weight and visceral fat were higher in the 2-h groups than in the 4-h groups even though the 2-h groups had less food. Third, we investigated the effects of eating a HFD during the inactive period, when RF duration was extended (2, 6 or 12?h). Mice were fed with a HFD during the inactive period for 2?h and fed with a ND during the active period for 2, 6 or 12?h. Body weight and visceral fat in these mice were comparable to those in the FF[ND] mice. The results of our first set of experiments suggest that 4-h RF was an adequate feeding duration to control the effect of a HFD on obesity. The results of our second set of experiments suggest 2-h RF (such as speed-eating) and 8-h RF, representative of eating disorders, are unhealthy feeding patterns related to obesity. The results of our third set of experiments suggest that eating a HFD for a short period during the night does not affect body weight and visceral fat. Taken together, these results indicate that consideration to feeding with a HFD during the inactive period and restricting eating habits relieve the risks of body weight gain and visceral fat accumulation.     

Dietary modulation of clostridial cluster XIVa gut bacteria (Roseburia spp.) by chitin-glucan fiber improves host metabolic alterations induced by high-fat diet in mice

Recent studies have provided new evidence that alterations in the composition of the gut microbiota — known as dysbiosis — participate in the development of obesity. The aim of the present study was to investigate the ability of chitin-glucan (CG) from a fungal source to modulate both the gut microbiota and glucose and lipid metabolism in high-fat (HF) diet-induced obese mice. Supplementation of the HF diet with fungal CG (10% w/w) induced caecal enlargement with prominent changes in gut microbiota: it restored the number of bacteria from clostridial cluster XIVa including Roseburia spp., which were decreased due to HF feeding. Furthermore, CG treatment significantly decreased HF-induced body weight gain, fat mass development, fasting hyperglycemia, glucose intolerance, hepatic triglyceride accumulation and hypercholesterolemia, independently of the caloric intake. All those parameters were negatively correlated with specific bacteria of clostridial cluster XIVa, i.e., Roseburia spp. (Pearson's correlations analysis). In contrast to prebiotics that more specifically target the bifidobacteria species, CG effects on obesity appear to be independent of the incretin glucagon-like peptide 1 (GLP-1) production, since portal GLP-1 and proglucagon (its precursor) expression were not modified by the dietary intervention. In conclusion, our findings support the view that chronic consumption of CG has potential beneficial effects with respect to the development of obesity and associated metabolic diabetes and hepatic steatosis, through a mechanism related to the restoration of the composition and/or the activity of gut bacteria, namely, bacteria from clostridial cluster XIVa.     

Ilex paraguariensis extract ameliorates obesity induced by high-fat diet: potential role of AMPK in the visceral adipose tissue

The aim of present study was to investigate the anti-obesity effect of Ilex paraguariensis extract and its molecular mechanism in rats rendered obese by a high-fat diet (HFD). I. paraguariensis extract supplementation significantly lowered body weight, visceral fat-pad weights, blood and hepatic lipid, glucose, insulin, and leptin levels of rats administered HFD. Feeding I. paraguariensis extract reversed the HFD-induced downregulation of the epididymal adipose tissue genes implicated in adipogenesis or thermogenesis, such as peroxisome proliferators’ activated receptor γ2, adipocyte fatty acid binding protein, sterol-regulatory-element-binding protein-1c, fatty acid synthase, HMG-CoA reductase, uncoupling protein 2, and uncoupling protein 3. Dietary supplementation with I. paraguariensis extract protected rats from the HFD-induced decreases in the phospho-AMP-activated protein kinase (AMPK)/AMPK and phospho-acetyl-CoA carboxylase (ACC)/ACC protein ratio related to fatty acid oxidation in the edipidymal adipose tissue. The present study reports that the I. paraguariensis extract can have a protective effect against a HFD-induced obesity in rats through an enhanced expression of uncoupling proteins and elevated AMPK phosphorylation in the visceral adipose tissue.     

Circadian Clock genes Per2 and clock regulate steroid production, cell proliferation, and luteinizing hormone receptor transcription in ovarian granulosa cells

Circadian Clock genes are associated with the estrous cycle in female animals. Treatment with Per2 and Clock siRNAs decreased the number of granulosa cells and LHr expression in follicle-stimulating hormone FSH-treated granulosa cells. Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom, whereas Clock siRNA treatment inhibited the production of estradiol and expression of P450arom mRNA. Per2 and Clock siRNA treatment increased and unchanged, respectively, progesterone production in FSH-treated granulosa cells. Similarly, expression of StAR mRNA was increased by Per2 siRNA and unchanged by Clock siRNA. Our data provide a new insight that Per2 and Clock have different action on ovarian granulosa cell functions.     

A water-soluble extract from Cucurbita moschata shows anti-obesity effects by controlling lipid metabolism in a high fat diet-induced obesity mouse model

During the screening of a variety of plant sources for their anti-obesity activity, it was found that a water-soluble extract, named PG105, prepared from stem parts of Cucurbita moschata, contains potent anti-obesity activities in a high fat diet-induced obesity mouse model. In this animal model, increases in body weight and fat storage were suppressed by 8-week oral administration of PG105 at 500 mg/kg, while the overall amount of food intake was not affected. Furthermore, PG105 protected the development of fatty liver and increased the hepatic beta-oxidation activity. Results from blood analysis showed that the levels of triglyceride and cholesterol were significantly lowered by PG105 administration, and also that the level of leptin was reduced, while that of adiponectin was increased. To understand the underlying mechanism at the molecular level, the effects of PG105 were examined on the expression of the genes involved in lipid metabolism by Northern blot analysis. In the liver of PG105-treated mice, the mRNA level of lipogenic genes such as SREBP-1c and SCD-1 was decreased, while that of lipolytic genes such as PPARalpha, ACO-1, CPT-1, and UCP-2 was modestly increased. Our data suggest that PG105 may have great potential as a novel anti-obesity agent in that both inhibition of lipid synthesis and acceleration of fatty acid breakdown are induced by this reagent.     

Chlorella modulates insulin signaling pathway and prevents high-fat diet-induced insulin resistance in mice

Aims

The search for natural agents that minimize obesity-associated disorders is receiving special attention. In this regard, the present study aimed to evaluate the prophylactic effect of Chlorella vulgaris (CV) on body weight, lipid profile, blood glucose and insulin signaling in liver, skeletal muscle and adipose tissue of diet-induced obese mice.

Main methods

Balb/C mice were fed either with standard rodent chow diet or high-fat diet (HFD) and received concomitant treatment with CV for 12 consecutive weeks. Triglyceride, free fatty acid, total cholesterol and fractions of cholesterol were measured using commercial assay. Insulin and leptin levels were determined by enzyme-linked immunosorbent assay (ELISA). Insulin and glucose tolerance tests were performed. The expression and phosphorylation of IRβ, IRS-1 and Akt were determined by Western blot analyses.

Key findings

Herein we demonstrate for the first time in the literature that prevention by CV of high-fat diet-induced insulin resistance in obese mice, as shown by increased glucose and insulin tolerance, is in part due to the improvement in the insulin signaling pathway at its main target tissues, by increasing the phosphorylation levels of proteins such as IR, IRS-1 and Akt. In parallel, the lower phosphorylation levels of IRS-1^ser307 were observed in obese mice. We also found that CV administration prevents high-fat diet-induced dyslipidemia by reducing triglyceride, cholesterol and free fatty acid levels.

Significance

We propose that the modulatory effect of CV treatment preventing the deleterious effects induced by high-fat diet is a good indicator for its use as a prophylactic–therapeutic agent against obesity-related complications.     

Arthrospira maxima prevents the acute fatty liver induced by the administration of simvastatin,ethanol and a hypercholesterolemic diet to mice

An evident fatty liver, corroborated morphologically and chemically, was produced in CD-1 mice after five daily doses of simvastatin 75 mg/Kg body weight, a hypercholesterolemic diet and 20 percent ethanol in the drinking water. After treating the animals, they presented serum triacylglycerols levels five times higher than the control mice, total lipids, cholesterol and triacylglycerols in the liver were 2, 2 and 1.5 times higher, respectively, than in control animals. When Arthrospira maxima was given with diet two weeks prior the onset of fatty liver induction, there was a decrement of liver total lipids (40%), liver triacylglycerols (50%) and serum triacylglycerols (50%) compared to the animals with the same treatment but without Arthrospira maxima. In addition to the mentioned protective effect, the administration of this algae, produced a significant increase (45%) in serum high density lipoproteins. The mechanism for this protective effect was not established in these experiments.     

Sodium Pentaborate Pentahydrate ameliorates lipid accumulation and pathological damage caused by high fat diet induced obesity in BALB/c mice

BackgroundObesity is one of the most popular topic in the field of research. In order to defeat this highly widespread disease, the mechanism of fat accumulation at the molecular level and its elimination are crucial. The use of boron has been showing promising results during the recent years.MethodsIn this study, anti-obesity potential of Sodium Pentaborate Pentahydrate (SPP) used as a dietary supplement on BALB/c mice fed with a high-fat diet was evaluated. Mice were divided into four groups with different diets, consisting of a normal diet, a high-fat diet (HFD) (containing 60 % fat), a HFD-supplemented with 0.5 mg/g body weight (BW) of SPP and a HFD-supplemented with 1.5 mg/g body weight (BW) of SPP. The animals were then observed for 10 weeks and physically monitored, and were sacrificed at the end of the experiment for physical and physicochemical evaluation.ResultsAccording to the physical parameters measured -body weight, food and water intake ratios-, the results indicate that SPP decreased weight gain in a dose dependent manner. Measurement of the hormone levels in the blood and fat accumulation in organs of mice also supported the anti-obesity effects of SPP. Expressions of adipogenesis related genes were also negatively regulated by SPP administration in white adipose tissue (WAT) tissue.ConclusionThese findings promise a treatment approach and drug development that can be used against obesity when SPP is used in the right doses. As a future aspect, clinical studies with SPP will reveal the effect of boron derivatives on obesity.     

Yarrowia lipolytica: A model and a tool to understand the mechanisms implicated in lipid accumulation

The oleaginous yeast Yarrowia lipolytica is known to inhabit various lipid-containing environments. One of the most striking features in this yeast is the presence of several multigene families involved in the metabolic pathways of hydrophobic substrate utilization. The complexity and the multiplicity of these genes give Y. lipolytica a wide capability range towards hydrophobic substrate (HS) utilization and storage. The combination of the increasing knowledge of this yeast's metabolism and the development of more efficient genetic tools is offering new perspectives in using Y. lipolytica as a model organism to study the mechanisms involved in lipid metabolism associated to fat uptake, storage, deposition, mobilization and regulation. Nutrient status and culture conditions seem to play a major role in obesity.     

Circadian regulation of egg-laying behavior in fruit flies Drosophila melanogaster

Significant progress has been made in our understanding of the neurogenetics of circadian clocks in fruit flies Drosophila melanogaster. Several pacemaker neurons and clock genes have now been identified and their roles in the cellular and molecular clockwork established. Some recent findings suggest that the basic architecture of the clock is multi-oscillatory; the clock mechanisms in the ventral lateral neurons (LN(v)s) of the fly brain govern locomotor activity and adult emergence rhythms, while the peripheral oscillators located in antennal cells regulate olfactory rhythm. Among circadian phenomena exhibited by Drosophila, the egg-laying rhythm is unique in many ways: (i) this rhythm persists under constant light (LL), while locomotor activity and adult emergence become arrhythmic, (ii) its circadian periodicity is much longer than 24h, and (iii) while egg-laying is rhythmic under constant darkness, the expression of two core clock genes period (per) and timeless (tim), is non-oscillatory in the ovaries. In this paper, we review our current knowledge of the circadian regulation of egg-laying behavior in Drosophila, and provide some possible explanations for its self-sustained nature. We conclude by discussing the existing limitations in our understanding of the regulatory mechanisms and propose few approaches to address them.