Regulation of headache response and transcriptomic network by the trigeminal ganglion clock.

OBJECTIVE: To characterize the circadian features of the trigeminal ganglion in a mouse model of headache. BACKGROUND: Several headache disorders, such as migraine and cluster headache, are known to exhibit distinct circadian rhythms of attacks. The circadian basis for these rhythmic pain responses, however, remains poorly understood. METHODS: We examined trigeminal ganglion ex vivo and single-cell cultures from Per2::LucSV reporter mice and performed immunohistochemistry. Circadian behavior and transcriptomics were investigated using a novel combination of trigeminovascular and circadian models: a nitroglycerin mouse headache model with mechanical thresholds measured every 6 h, and trigeminal ganglion RNA sequencing measured every 4 h for 24 h. Finally, we performed pharmacogenomic analysis of gene targets for migraine, cluster headache, and trigeminal neuralgia treatments as well as trigeminal ganglion neuropeptides; this information was cross-referenced with our cycling genes from RNA sequencing data to identify potential targets for chronotherapy. RESULTS: The trigeminal ganglion demonstrates strong circadian rhythms in both ex vivo and single-cell cultures, with core circadian proteins found in both neuronal and non-neuronal cells. Using our novel behavioral model, we showed that nitroglycerin-treated mice display circadian rhythms of pain sensitivity which were abolished in arrhythmic Per1/2 double knockout mice. Furthermore, RNA-sequencing analysis of the trigeminal ganglion revealed 466 genes that displayed circadian oscillations in the control group, including core clock genes and clock-regulated pain neurotransmitters. In the nitroglycerin group, we observed a profound circadian reprogramming of gene expression, as 331 of circadian genes in the control group lost rhythm and another 584 genes gained rhythm. Finally, pharmacogenetics analysis identified 10 genes in our trigeminal ganglion circadian transcriptome that encode target proteins of current medications used to treat migraine, cluster headache, or trigeminal neuralgia. CONCLUSION: Our study unveiled robust circadian rhythms in the trigeminal ganglion at the behavioral, transcriptomic, and pharmacogenetic levels. These results support a fundamental role of the clock in pain pathophysiology. PLAIN LANGUAGE SUMMARY: Several headache diseases, such as migraine and cluster headache, have headaches that occur at the same time each day. We learned that the trigeminal ganglion, an important pain structure in several headache diseases, has a 24-hour cycle that might be related to this daily cycle of headaches. Our genetic analysis suggests that some medications may be more effective in treating migraine and cluster headache when taken at specific times of the day.

The Role of Organokines in Obesity and Type 2 Diabetes and Their Functions as Molecular Transducers of Nutrition and Exercise.

Maintaining systemic homeostasis requires the coordination of different organs and tissues in the body. Our bodies rely on complex inter-organ communications to adapt to perturbations or changes in metabolic homeostasis. Consequently, the liver, muscle, and adipose tissues produce and secrete specific organokines such as hepatokines, myokines, and adipokines in response to nutritional and environmental stimuli. Emerging evidence suggests that dysregulation of the interplay of organokines between organs is associated with the pathophysiology of obesity and type 2 diabetes (T2D). Strategies aimed at remodeling organokines may be effective therapeutic interventions. Diet modification and exercise have been established as the first-line therapeutic intervention to prevent or treat metabolic diseases. This review summarizes the current knowledge on organokines secreted by the liver, muscle, and adipose tissues in obesity and T2D. Additionally, we highlighted the effects of diet/nutrition and exercise on the remodeling of organokines in obesity and T2D. Specifically, we investigated the ameliorative effects of caloric restriction, selective nutrients including ω3 PUFAs, selenium, vitamins, and metabolites of vitamins, and acute/chronic exercise on the dysregulation of organokines in obesity and T2D. Finally, this study dissected the underlying molecular mechanisms by which nutrition and exercise regulate the expression and secretion of organokines in specific tissues.

Wheatgrass-and-Aronia-Mixed Extract Suppresses Immunoglobulin E-Mediated Allergic Reactions In Vitro and In Vivo.

Mast cells are an important component of immune responses. Immunoglobulin (Ig) E-sensitized mast cells release substances within minutes of allergen exposure, triggering allergic responses. Until now, numerous pharmacological effects of wheatgrass and aronia have been verified, but the effects of wheatgrass and aronia (TAAR)-mixed extract on allergic reactions have not been identified. Therefore, the aim of this study was to demonstrate the anti-allergic effect of TAAR extract on mast cell activation and cutaneous anaphylaxis. In this study, we investigated the anti-allergic effects and related mechanisms of TAAR extract in IgE-activated mast cells in vitro. We also assessed the ameliorating effect of TAAR extract on IgE-mediated passive cutaneous anaphylaxis mice in vivo. The TAAR extract significantly reduced the expression of ß-hexosaminidase, histamine, and pro-inflammatory cytokines, which are mediators related to mast cell degranulation, via the regulation of various signaling pathways. The TAAR extract also regulated oxidative-stress-related factors through the Nrf2 signaling pathway. Additionally, treatment of TAAR extract to the passive cutaneous anaphylaxis mouse model improved ear thickness and local ear pigmentation. Taken together, our results suggest that TAAR extract is a potential candidate natural product to treat overall IgE-mediated allergic inflammation and oxidative-stress-related diseases by suppressing mast cell activity.

ß-Cryptoxanthin Attenuates Cigarette-Smoke-Induced Lung Lesions in the Absence of Carotenoid Cleavage Enzymes (BCO1/BCO2) in Mice.

High dietary intake of ß-cryptoxanthin (BCX, an oxygenated provitamin A carotenoid) is associated with a lower risk of lung disease in smokers. BCX can be cleaved by ß-carotene-15,15'-oxygenase (BCO1) and ß-carotene-9',10'-oxygenase (BCO2) to produce retinol and apo-10'-carotenoids. We investigated whether BCX has protective effects against cigarette smoke (CS)-induced lung injury, dependent or independent of BCO1/BCO2 and their metabolites. Both BCO1-/-/BCO2-/- double knockout mice (DKO) and wild type (WT) littermates were supplemented with BCX 14 days and then exposed to CS for an additional 14 days. CS exposure significantly induced macrophage and neutrophil infiltration in the lung tissues of mice, regardless of genotypes, compared to the non-exposed littermates. BCX treatment significantly inhibited CS-induced inflammatory cell infiltration, hyperplasia in the bronchial epithelium, and enlarged alveolar airspaces in both WT and DKO mice, regardless of sex. The protective effects of BCX were associated with lower expression of IL-6, TNF-α, and matrix metalloproteinases-2 and -9. BCX treatment led to a significant increase in hepatic BCX levels in DKO mice, but not in WT mice, which had significant increase in hepatic retinol concentration. No apo-10'-carotenoids were detected in any of the groups. In vitro BCX, at comparable doses of 3-OH-ß-apo-10'-carotenal, was effective at inhibiting the lipopolysaccharide-induced inflammatory response in a human bronchial epithelial cell line. These data indicate that BCX can serve as an effective protective agent against CS-induced lung lesions in the absence of carotenoid cleavage enzymes.

The circadian E3 ligase FBXL21 regulates myoblast differentiation and sarcomere architecture via MYOZ1 ubiquitination and NFAT signaling.

Numerous molecular and physiological processes in the skeletal muscle undergo circadian time-dependent oscillations in accordance with daily activity/rest cycles. The circadian regulatory mechanisms underlying these cyclic processes, especially at the post-transcriptional level, are not well defined. Previously, we reported that the circadian E3 ligase FBXL21 mediates rhythmic degradation of the sarcomere protein TCAP in conjunction with GSK-3ß, and Psttm mice harboring an Fbxl21 hypomorph allele show reduced muscle fiber diameter and impaired muscle function. To further elucidate the regulatory function of FBXL21 in skeletal muscle, we investigated another sarcomere protein, Myozenin1 (MYOZ1), that we identified as an FBXL21-binding protein from yeast 2-hybrid screening. We show that FBXL21 binding to MYOZ1 led to ubiquitination-mediated proteasomal degradation. GSK-3ß co-expression and inhibition were found to accelerate and decelerate FBXL21-mediated MYOZ1 degradation, respectively. Previously, MYOZ1 has been shown to inhibit calcineurin/NFAT signaling important for muscle differentiation. In accordance, Fbxl21 KO and MyoZ1 KO in C2C12 cells impaired and enhanced myogenic differentiation respectively compared with control C2C12 cells, concomitant with distinct effects on NFAT nuclear localization and NFAT target gene expression. Importantly, in Psttm mice, both the levels and diurnal rhythm of NFAT2 nuclear localization were significantly diminished relative to wild-type mice, and circadian expression of NFAT target genes associated with muscle differentiation was also markedly dampened. Furthermore, Psttm mice exhibited significant disruption of sarcomere structure with a considerable excess of MYOZ1 accumulation in the Z-line. Taken together, our study illustrates a pivotal role of FBXL21 in sarcomere structure and muscle differentiation by regulating MYOZ1 degradation and NFAT2 signaling.

Mast cell modulates tumorigenesis caused by repeated bowel inflammation condition in azoxymethane/dextran sodium sulfate-induced colon cancer mouse model.

Mast cells infiltrate the inflammatory microenvironment and regulate the production of many pro-inflammatory cytokines and mediators of inflammatory cell production to promote tumor development and growth in intestinal lesions. Currently, there are insufficient studies of the mediators and signaling pathways regulated by mast cells that influence the pathogenesis of colon cancer in inflamed colon tissue. This study aimed to confirm the role of mast cells in the incidence and growth of colitis-associated colon cancer (CAC) and to identify inflammation-mediated factors and signaling pathways related to tumor development. CAC was induced by the administration of azoxymethane (AOM) and dextran sodium sulfate (DSS) in mast cell-deficient (WBB6F1/J-W/WV) and mast cell-sufficient control (WBB6F1_+/+) mice. The results confirmed that mast cell-deficient mice exhibited less tumor formation than normal mice under the same conditions, and down-regulated expression of pro-inflammatory cytokines and mediators. Mast cells play an important role in tumor formation by regulating pro-inflammatory cytokines and inflammatory mediators in CAC, indicating that they can act as new targets for the prevention and treatment of CAC.

Extract of Wheatgrass and Aronia Mixture Ameliorates Atopic Dermatitis-Related Symptoms by Suppressing Inflammatory Response and Oxidative Stress In Vitro and In Vivo.

Atopic dermatitis is regulated by the production of pro-inflammatory cytokines and chemokines via the nuclear factor kappa B or mitogen-activated protein kinase signaling pathways, as well as, the release of oxidative stress-related factors via the NF-E2 p45-related factor 2 signaling pathway. Both wheatgrass (Triticum aestivum L., TA) and aronia (Aronia melanocarpa, AR) are known for their anti-inflammatory and antioxidant properties, however, the anti-inflammatory and antioxidant effects of TA and AR (TAAR) mixture extract have not been elucidated in an atopic dermatitis model. In this study, we assessed the inhibitory effects and underlying molecular mechanism of TAAR extract against lipopolysaccharide-induced inflammation and tumor necrosis factor-α/interferon-γ-induced inflammation and oxidative stress in vitro. We also investigated the alleviating effect of TAAR extract on DNCB-induced atopic dermatitis-like skin lesions in mice in vivo. We found that TAAR extract treatment inhibited inflammatory mediators in both RAW 264.7 cells and HaCaT cells, and increased the expression of oxidative stress defense enzymes in HaCaT cells. Furthermore, treatment of the DNCB-induced mouse model with TAAR extract ameliorated the overall symptoms of atopic dermatitis. Therefore, TAAR extract as a novel natural therapeutic agent may be used for the treatment of atopic dermatitis.

Extract of Triticum aestivum Sprouts Suppresses Acetaminophen-Induced Hepatotoxicity in Mice by Inhibiting Oxidative Stress.

Wheat (Triticum aestivum L.) is the oldest known food crop, and many studies have reported that wheat shoots (i.e., wheatgrass) possess anti-cancer, anti-inflammatory, and antioxidant activities. However, the potentially ameliorative effect of wheat shoots on hepatotoxicity caused by high doses of N-acetyl-para-aminophenol (acetaminophen, APAP) has yet to be reported. C57BL/6 mice received daily oral TAE (100 or 200 mg/kg), positive control (silymarin 100 mg/kg), or negative control (saline vehicle) treatments for 7 days prior to intraperitoneal APAP injection. Histological, serum (ELISA), Western blotting, and quantitative PCR analyses of excised liver tissues were then performed. Pre-treatment with TAE (100 or 200 mg/kg) ameliorated APAP-induced pathological damage (i.e., hepatotoxic lesions), reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, and also ameliorated APAP-induced increases in oxidative stress, thereby inhibiting oxidative liver damage and reducing the expression of inflammatory cytokines. In addition, TAE pre-treatment inhibited the expression of Cytochrome P4502E1 (CYP2E1), which is a key enzyme in the onset of APAP-induced hepatotoxicity, suppressed the expression of the target proteins regulated by the antioxidant enzyme Nrf2, and suppressed hepatocyte apoptosis. These findings suggest that TAE is an attractive therapeutic candidate that exhibits potential hepatoprotective activity by inhibiting oxidative stress, inflammation, apoptosis, and liver damage.

Gastrodia elata Blume and Zanthoxylum schinifolium Siebold & Zucc Mixed Extract Suppress Platelet Aggregation and Thrombosis.

Background and objectives: Blood vessel thrombosis causes blood circulation disorders, leading to various diseases. Currently, various antiplatelet and anticoagulant drugs, such as aspirin, warfarin, heparin, and non-vitamin K antagonist oral anticoagulants (NOACs), are used as the major drugs for the treatment of a wide range of thrombosis. However, these drugs have a side effect of possibly causing internal bleeding due to poor hemostasis when taken for a long period of time. Materials and Methods: Gastrodia elata Blume (GE) and Zanthoxylum schinifolium Siebold & Zucc (ZS) are known to exhibit hemostatic and antiplatelet effects as traditional medicines that have been used for a long time. In this study, we investigated the effect of a mixed extract of GE and ZS (MJGE09) on platelet aggregation and plasma coagulation. Results: We found that MJGE09 inhibited collagen-and ADP-induced platelet aggregation in vitro. In addition, collagen- and ADP-induced platelet aggregation were also inhibited in a dose-dependent manner on the platelets of mice that were orally administered MJGE09 ex vivo. However, compared with aspirin, MJGE09 did not prolong the rat tail vein bleeding time in vivo and did not show a significant effect on the increase in the prothrombin time (PT) and activated partial thromboplastin time (aPTT). Conclusions: These results suggest that MJGE09 can be used as a potential anticoagulant with improved antithrombotic efficacy.

Effects of the Clock Modulator Nobiletin on Circadian Rhythms and Pathophysiology in Female Mice of an Alzheimer's Disease Model.

Alzheimer's disease (AD) is an age-related neurodegenerative disorder and the most common cause of dementia. Various pathogenic mechanisms have been proposed to contribute to disease progression, and recent research provided evidence linking dysregulated circadian rhythms/sleep and energy metabolism with AD. Previously, we found that the natural compound Nobiletin (NOB) can directly activate circadian cellular oscillators to promote metabolic health in disease models and healthy aging in naturally aged mice. In the current study, using the amyloid-ß AD model APP/PS1, we investigated circadian, metabolic and amyloid characteristics of female mice and the effects of NOB. Female APP/PS1 mice showed reduced sleep bout duration, and NOB treatment exhibited a trend to improve it. While glucose tolerance was unchanged, female APP/PS1 mice displayed exaggerated oxygen consumption and CO2 production, which was mitigated by NOB. Likewise, cold tolerance in APP/PS1 was impaired relative to WT, and interestingly was markedly enhanced in NOB-treated APP/PS1 mice. Although circadian behavioral rhythms were largely unchanged, real-time qPCR analysis revealed altered expression of several core clock genes by NOB in the cerebral cortex, notably Bmal1, Npas2, and Rora. Moreover, NOB was also able to activate various clock-controlled metabolic genes involved in insulin signaling and mitochondrial function, including Igf1, Glut1, Insr, Irs1, Ucp2, and Ucp4. Finally, we observed that NOB attenuated the expression of several AD related genes including App, Bace1, and ApoE, reduced APP protein levels, and strongly ameliorated Aß pathology in the cortex. Collectively, these results reveal novel genotype differences and importantly beneficial effects of a natural clock-enhancing compound in biological rhythms and related pathophysiology, suggesting the circadian clock as a modifiable target for AD.

Inhibitory effects of nodakenin on inflammation and cell death in lipopolysaccharide-induced liver injury mice.

BACKGROUND: Nodakenin, a coumarin glucoside isolated from the roots of Angelica biserrata, has been reported to have anti-inflammatory, antibacterial, anticancer effects. However, despite these studies, the potential liver protective effects of nodakenin in inflammatory liver injury models have not been reported. METHODS: A mouse model of inflammatory liver injury was induced by injection of lipopolysaccharide (LPS) (15 mg/kg, intraperitoneally (i.p)). Liver tissue AST, ALT, ROS, T-GSH and T-SOD were analyzed by ELISA. The concentrations of TNF-α, IL-6, and IL-1ß in serum of LPS-induced inflammatory liver injury mice were analyzed. The mRNA expression levels of GPx1, catalase, SOD1, SOD2, TNF-α, IL-6, IL-1ß, iNOS and COX-2 were analyzed using real-time PCR. The expressions of MAPK, IRF3, NF-κB, Nrf2, HO-1, caspase-3 and caspase-7 were analyzed using western blotting. Liver tissue was stained with IHC to confirm NF-κB, Nrf-2, HO-1, caspase-3, Bax, and Bcl2. Tunnel analysis was performed to confirm the fragmented nuclear DNA characteristics of apoptosis. RESULTS: The administration of nodakenin (10 and 30 mg/kg) reduced serum aminotransferase levels compared to LPS-induced liver damage and significantly improved the oxidative state of liver tissue and pathological damage. Moreover, inhibited the phosphorylation of transforming growth factor beta (TGF-ß)-activated kinase (TAK)-1 in LPS-induced inflammatory liver injury model, and significantly inhibited the transcriptional of nuclear factor-kappa B (NF-kB) and the secretion of pro-inflammatory mediators. In addition nodakenin pre-treatment also attenuated hepatocyte death by regulating apoptosis-related mitochondrial proteins, such as cysteinyl aspartate specific proteinase 3 (caspase 3), poly (ADP-ribose) polymerase (PARP), B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X (Bax). CONCLUSION: Our findings suggest that nodakenin has anti-inflammatory, anti-oxidant and anti-apoptotic activity and may be an adjunctive prevention agent for liver injury.

Extract of Boehmeria nivea Suppresses Mast Cell-Mediated Allergic Inflammation by Inhibiting Mitogen-Activated Protein Kinase and Nuclear Factor-κB.

Mast cells are effector cells that initiate allergic inflammatory immune responses by inducing inflammatory mediators. Boehmeria nivea (Linn.) Gaudich is a natural herb in the nettle family Urticaceae that possesses numerous pharmacological properties. Despite the various pharmacological benefits of Boehmeria nivea, its effects on allergic inflammation have not yet been determined. Here, we investigated the effect of the ethanol extract of Boehmeria nivea (BNE) on degranulation rat basophilic leukemia (RBL)-2H3 mast cells stimulated with anti-dinitrophenyl (anti-DNP) and bovine serum albumin (BSA) during immunoglobulin E (IgE)-mediated allergic immune response. The results showed inhibition of the release of ß-hexosaminidase and histamine from the cells. BNE suppressed pro-inflammatory cytokines (Tumor necrosis factor (TNF)-α, Interleukin (IL)-1ß, and IL-6) and reduced T helper (Th)2 cytokine IL-4 expression and/or secretion correlated with the downregulation of p38, extracellular signal-regulated kinases (ERK) mitogen-activated protein kinase (MAPK), and nuclear factor-κB (NF-κB) signaling pathways in treated RBL-2H3 mast cells. In passive cutaneous anaphylaxis, treatment with BNE during IgE-mediated local allergic reaction triggered a reduction in mouse ear pigmentation and thickness. Taken together, these results indicated that BNE suppressed mast cell-mediated inflammation, suggesting that BNE might be a candidate for the treatment of various allergic disorders.

Chijabyukpi-Tang Inhibits Pro-Inflammatory Cytokines and Chemokines via the Nrf2/HO-1 Signaling Pathway in TNF-α/IFN-γ-Stimulated HaCaT Cells and Ameliorates 2,4-Dinitrochlorobenzene-Induced Atopic Dermatitis-Like Skin Lesions in Mice.

Chijabyukpi-tang (CBT) is an oriental herbal formula consisting of three herbs (Gardeniae Fructus (Gardenia jasminoides J.Ellis.), Phellodendri Cortex (Phellodendron amurense Rupr.), Glycyrrhizae Radix (Glycyrrhiza uralensis Fisch. ex DC.) at the ratio of 2: 2: 1. CBT has traditionally been used to treat eczema with inflammation in Northeast Asia. The components of CBT have been shown to have anti-inflammatory and anti-oxidant properties, but the exact role and mechanism of CBT on atopic dermatitis (AD) remain unclear. In this study, we investigated the anti-inflammatory effect and mechanism of CBT in the HaCaT human keratinocyte cell line and investigated the anti-atopic effect in mice models of atopic dermatitis-like skin lesions. In the tumor necrosis factor alpha (TNF)-α/interferon (IFN)-γ-stimulated HaCaT cells, CBT inhibited the production of pro-inflammatory cytokines and chemokines and elevated the nuclear translocation of NF-E2 p45 related factors 2 (Nrf2) and subsequent production of heme oxygenase-1 (HO-1). CBT improved the symptoms of atopic dermatitis-like lesions in 2,4-dinitrochlorobenzene (DNCB)-treated mice by suppressing the levels of serum immunoglobulin E (IgE), and various pro-inflammatory cytokines and chemokines. The improvement effect of CBT on atopic dermatitis-like lesions can be predicted to be due to increased Nrf2 and HO-1 gene expression. These results suggest that CBT is an herbal medicine with the potential for use as a therapeutic agent for inflammatory skin diseases such as atopic dermatitis.

Mechanistic understanding of ß-cryptoxanthin and lycopene in cancer prevention in animal models.

To better understand the potential function of carotenoids in the chemoprevention of cancers, mechanistic understanding of carotenoid action on genetic and epigenetic signaling pathways is critically needed for human studies. The use of appropriate animal models is the most justifiable approach to resolve mechanistic issues regarding protective effects of carotenoids at specific organs and tissue sites. While the initial impetus for studying the benefits of carotenoids in cancer prevention was their antioxidant capacity and pro-vitamin A activity, significant advances have been made in the understanding of the action of carotenoids with regards to other mechanisms. This review will focus on two common carotenoids, provitamin A carotenoid ß-cryptoxanthin and non-provitamin A carotenoid lycopene, as promising chemopreventive agents or chemotherapeutic compounds against cancer development and progression. We reviewed animal studies demonstrating that ß-cryptoxanthin and lycopene effectively prevent the development or progression of various cancers and the potential mechanisms involved. We highlight recent research that the biological functions of ß-cryptoxanthin and lycopene are mediated, partially via their oxidative metabolites, through their effects on key molecular targeting events, such as NF-κB signaling pathway, RAR/PPARs signaling, SIRT1 signaling pathway, and p53 tumor suppressor pathways. The molecular targets by ß-cryptoxanthin and lycopene, offer new opportunities to further our understanding of common and distinct mechanisms that involve carotenoids in cancer prevention. This article is part of a Special Issue entitled Carotenoids recent advances in cell and molecular biology edited by Johannes von Lintig and Loredana Quadro.

Xanthophyll ß-Cryptoxanthin Inhibits Highly Refined Carbohydrate Diet-Promoted Hepatocellular Carcinoma Progression in Mice.

SCOPE: ß-Cryptoxanthin (BCX) can be cleaved by both ß-carotene 15,15'-oxygenase (BCO1) and ß-carotene 9',10'-oxygenase (BCO2), generating biological active vitamin A and apocarotenoids. We examined whether BCX feeding could inhibit diethylnitrosamine (DEN)-initiated, highly refined carbohydrate diet (HRCD)-promoted hepatocellular carcinoma (HCC) development, dependent or independent of BCO1/BCO2 activity. METHODS AND RESULTS: Two-week-old male wild-type (WT) and BCO1-/- /BCO2-/- double knockout (DKO) mice are given a single intraperitoneal injection of DEN (25 mg kg-1 body weight) to initiate hepatic carcinogenesis. At 6 weeks of age, all animals are fed HRCD (66.5% of energy from carbohydrate) with or without BCX for 24 weeks. BCX feeding increases hepatic vitamin A levels in WT mice, but not in DKO mice that shows a significant accumulation of hepatic BCX. Compared to their respective HRCD littermates, both WT and DKO fed BCX have significantly lower HCC multiplicity, average tumor size, and total tumor volume, and the steatosis scores. The chemopreventive effects of BCX are associated with increased p53 protein acetylation and decreased protein levels of lactate dehydrogenase and hypoxia-inducible factor-1α in tumors. CONCLUSION: This study suggests that BCX feeding may alleviate HRCD-promoted HCC progression by modulating the acetylation of p53, hypoxic tumor microenvironment, and glucose metabolism, independent of BCO1/BCO2.

Umbelliferone reduces the expression of inflammatory chemokines in HaCaT cells and DNCB/DFE-induced atopic dermatitis symptoms in mice.

Umbelliferone (UMB) is a coumarin derivative present in roots and barks of plants, such as Angelica decursiva, Artemisia capillaris, and orange. UMB has been previously reported to exhibit anti-inflammatory, anti-diabetic, and anti-cancer effects. However, the effect of UMB on atopic dermatitis (AD) remains unknown. The purpose of this study was to investigate the anti-atopic effects of UMB on 2,4-dinitrochlorobenzene (DNCB)- and house dust mite extract (Dermatophagoides farinae extract, DFE)-treated mice with AD-like skin lesions and on tumor necrosis factor (TNF)-α/interferon (IFN)-γ-treated HaCaT cells. In DNCB/DFE-treated mice, oral administration of UMB (20 and 40 mg/kg) for 28 days led to a significant decrease in ear thickness, spleen size and weight, serum levels of immunoglobulin E (IgE), IgG1, IgG2a, TNF-α, and interleukin 4 (IL-4), and mast cell infiltration; it also led to the suppression of pro-inflammatory cytokines and chemokines. In addition, UMB reduced the secretion of pro-inflammatory cytokines and chemokines in TNF-α/IFN-γ-treated HaCaT cells via regulation of MAPK, IkB-α/NF-κB, and STAT1 signaling pathways. Taken together, these results indicate that UMB ameliorates AD-associated symptoms and inflammation via regulation of various signaling pathways, suggesting that UMB might be a potential therapeutic agent of AD.

Dietary ß-Cryptoxanthin Inhibits High-Refined Carbohydrate Diet-Induced Fatty Liver via Differential Protective Mechanisms Depending on Carotenoid Cleavage Enzymes in Male Mice.

BACKGROUND: ß-Cryptoxanthin (BCX), a provitamin A carotenoid shown to protect against nonalcoholic fatty liver disease (NAFLD), can be cleaved by ß-carotene-15,15'-oxygenase (BCO1) to generate vitamin A, and by ß-carotene-9',10'-oxygenase (BCO2) to produce bioactive apo-carotenoids. BCO1/BCO2 polymorphisms have been associated with variations in plasma carotenoid amounts in both humans and animals. OBJECTIVES: We investigated whether BCX feeding inhibits high refined-carbohydrate diet (HRCD)-induced NAFLD, dependent or independent of BCO1/BCO2. METHODS: Six-week-old male wild-type (WT) and BCO1-/-/BCO2-/- double knockout (DKO) mice were randomly fed HRCD (66.5% of energy from carbohydrate) with or without BCX (10 mg/kg diet) for 24 wk. Pathological and biochemical variables were analyzed in the liver and mesenteric adipose tissues (MATs). Data were analyzed by 2-factor ANOVA. RESULTS: Compared to their respective HRCD controls, BCX reduced hepatic steatosis severity by 33‒43% and hepatic total cholesterol by 43‒70% in both WT and DKO mice (P < 0.01). Hepatic concentrations of BCX, but not retinol and retinyl palmitate, were 33-fold higher in DKO mice than in WT mice (P < 0.001). BCX feeding increased the hepatic fatty acid oxidation protein peroxisome proliferator-activated receptor-α, and the cholesterol efflux gene ATP-binding cassette transporter5, and suppressed the lipogenesis gene acetyl-CoA carboxylase 1 (Acc1) in the MAT of WT mice but not DKO mice (P < 0.05). BCX feeding decreased the hepatic lipogenesis proteins ACC and stearoyl-CoA desaturase-1 (3-fold and 5-fold) and the cholesterol synthesis genes 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase and HMG-CoA synthase 1 (2.7-fold and 1.8-fold) and increased the cholesterol catabolism gene cholesterol 7α-hydroxylase (1.9-fold) in the DKO but not WT mice (P < 0.05). BCX feeding increased hepatic protein sirtuin1 (2.5-fold) and AMP-activated protein kinase (9-fold) and decreased hepatic farnesoid X receptor protein (80%) and the inflammatory cytokine gene Il6 (6-fold) in the MAT of DKO mice but not WT mice (P < 0.05). CONCLUSION: BCX feeding mitigates HRCD-induced NAFLD in both WT and DKO mice through different mechanisms in the liver-MAT axis, depending on the presence or absence of BCO1/BCO2.

Ablation of carotenoid cleavage enzymes (BCO1 and BCO2) induced hepatic steatosis by altering the farnesoid X receptor/miR-34a/sirtuin 1 pathway.

ß-Carotene-15, 15'-oxygenase (BCO1) and ß-carotene-9', 10'-oxygenase (BCO2) are essential enzymes in carotenoid metabolism. While BCO1/BCO2 polymorphisms have been associated with alterations to human and animal carotenoid levels, experimental studies have suggested that BCO1 and BCO2 may have specific physiological functions beyond the cleavage of carotenoids. In the present study, we investigated the effect of ablation of both BCO1/BCO2 in the development of non-alcoholic fatty liver disease (NAFLD) and its underlying molecular mechanism(s). BCO1/BCO2 double knock out (DKO) mice developed hepatic steatosis (8/8) and had significantly higher levels of hepatic and plasma triglyceride and total cholesterol compared to WT (0/8). Hepatic changes in the BCO1/BCO2 DKO mice were associated with significant: 1) increases in lipogenesis markers, and decreases in fatty acid ß-oxidation markers; 2) upregulation of cholesterol metabolism markers; 3) alterations to microRNAs related to TG accumulation and cholesterol metabolism; 4) increases in an hepatic oxidative stress marker (HO-1) but decreases in anti-oxidant enzymes; and 5) decreases in farnesoid X receptor (FXR), small heterodimer partner (SHP), and sirtuin 1 (SIRT1). The present study provided novel experimental evidence that BCO1 and BCO2 could play a significant role in maintaining normal hepatic lipid and cholesterol homeostasis, potentially through the regulation of the FXR/miR-34a/SIRT1 pathway.

Xylobiose, an Alternative Sweetener, Ameliorates Diabetes-Related Metabolic Changes by Regulating Hepatic Lipogenesis and miR-122a/33a in db/db Mice.

Type 2 diabetes is a major public health concern worldwide. Xylobiose (XB) consists of two molecules of d-xylose and is a major disaccharide in xylooligosaccharides that are used as prebiotics. We hypothesized that XB could regulate diabetes-related metabolic and genetic changes via microRNA expression in db/db mice. For six weeks, C57BL/KsJ-db/db mice received 5% XB as part of the total sucrose content of their diet. XB supplementation improved glucose tolerance with reduced levels of OGTT AUC, fasting blood glucose, HbA1c, insulin, and HOMA-IR. Furthermore, XB supplementation decreased the levels of total triglycerides, total cholesterol, and LDL-C. The expression levels of miR-122a and miR-33a were higher and lower in the XB group, respectively. In the liver, expressions of the lipogenic genes, including, fatty acid synthase (FAS), peroxisome proliferator activated receptor γ (PPARγ), sterol regulatory element-binding protein-1C (SREBP-1C), sterol regulatory element-binding protein-2 (SREBP-2), acetyl-CoA carboxylase (ACC), HMG-CoA reductase (HMGCR), ATP-binding cassette transporter G5/G8 (ABCG5/8), cholesterol 7 alpha-hydroxylase (CYP7A1), and sterol 12-alpha-hydroxylase (CYP8B1), as well as oxidative stress markers, including superoxide dismutase 1 (SOD1), superoxide dismutase 2 (SOD2), glutathione peroxidase (GPX), and catalase, were also regulated by XB supplementation. XB supplementation inhibited the mRNA expressions levels of the pro-inflammatory cytokines, tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, interleukin (IL)-6, and monocyte chemoattractant protein (MCP)-1, as well as phosphorylation of c-Jun N-terminal kinase/stress activated protein kinase (JNK/SAPK), p38 mitogen-activated protein kinases (MAPK), and extracellular signal-regulated kinases 1/2 (ERK1/2). These data demonstrate that XB exhibits anti-diabetic, hypolipogenic, and anti-inflammatory effects via regulation of the miR-122a/33a axis in db/db mice.

Enzymatic Process for High-Yield Turanose Production and Its Potential Property as an Adipogenesis Regulator.

Turanose is a sucrose isomer naturally existing in honey and a promising functional sweetener due to its low glycemic response. In this study, the extrinsic fructose effect on turanose productivity was examined in Neisseria amylosucrase reaction. Turanose was produced, by increasing the amount of extrinsic fructose as a reaction modulator, with high concentration of sucrose substrate, which resulted in 73.7% of production yield. In physiological functionality test, lipid accumulation in 3T3-L1 preadipocytes in the presence of high amounts of pure glucose was attenuated by turanose substitution in a dose-dependent manner. Turanose treatments at concentrations representing 50%, 75%, and 100% of total glucose concentration in cell media significantly reduced lipid accumulation by 18%, 35%, and 72%, respectively, as compared to controls. This result suggested that turanose had a positive role in controlling adipogenesis, and enzymatic process of turanose production has a potential to develop a functional food ingredient for controlling obesity and related chronic diseases.