Determination of α-Dicarbonyl compounds in traditional Chinese herbal medicines.

α-DCs (α-dicarbonyls) have been proven to be closely related to aging and the onset and development of many chronic diseases. The wide presence of this kind of components in various foods and beverages has been unambiguously determined, but their occurrence in various phytomedicines remains in obscurity. In this study, we established and evaluated an HPLC-UV method and used it to measure the contents of four α-DCs including 3-deoxyglucosone (3-DG), glyoxal (GO), methylglyoxal (MGO), and diacetyl (DA) in 35 Chinese herbs after they have been derivatized with 4-nitro-1,2-phenylenediamine. The results uncover that 3-DG is the major component among the α-DCs, being detectable in all the selected herbs in concentrations ranging from 22.80 µg/g in the seeds of Alpinia katsumadai to 7032.75 µg/g in the fruit of Siraitia grosuenorii. The contents of the other three compounds are much lower than those of 3-DG, with GO being up to 22.65 µg/g, MGO being up to 55.50 µg/g, and DA to 18.75 µg/g, respectively. The data show as well the contents of the total four α-DCs in the herbs are generally in a comparable level to those in various foods, implying that herb medicines may have potential risks on human heath in view of the α-DCs.

Lesion of NPY Receptor-expressing Neurons in Perifornical Lateral Hypothalamus Attenuates Glucoprivic Feeding.

Glucoprivic feeding is one of several counterregulatory responses (CRRs) that facilitates restoration of euglycemia following acute glucose deficit (glucoprivation). Our previous work established that glucoprivic feeding requires ventrolateral medullary (VLM) catecholamine (CA) neurons that coexpress neuropeptide Y (NPY). However, the connections by which VLM CA/NPY neurons trigger increased feeding are uncertain. We have previously shown that glucoprivation, induced by an anti-glycolygic agent 2-deoxy-D-glucose (2DG), activates perifornical lateral hypothalamus (PeFLH) neurons and that expression of NPY in the VLM CA/NPY neurons is required for glucoprivic feeding. We therefore hypothesized that glucoprivic feeding and possibly other CRRs require NPY-sensitive PeFLH neurons. To test this, we used the ribosomal toxin conjugate NPY-saporin (NPY-SAP) to selectively lesion NPY receptor-expressing neurons in the PeFLH of male rats. We found that NPY-SAP destroyed a significant number of PeFLH neurons, including those expressing orexin, but not those expressing melanin-concentrating hormone. The PeFLH NPY-SAP lesions attenuated 2DG-induced feeding but did not affect 2DG-induced increase in locomotor activity, sympathoadrenal hyperglycemia, or corticosterone release. The 2DG-induced feeding response was also significantly attenuated in NPY-SAP-treated female rats. Interestingly, PeFLH NPY-SAP lesioned male rats had reduced body weights and decreased dark cycle feeding, but this effect was not seen in female rats. We conclude that a NPY projection to the PeFLH is necessary for glucoprivic feeding, but not locomotor activity, hyperglycemia, or corticosterone release, in both male and female rats.

Phenylboronic Acid-Modified Near-Infrared Region II Excitation Donor-Acceptor-Donor Molecule for 2-Deoxy-d-Glucose Improved Starvation/Chemo/Photothermal Combination Therapy.

Synergistic chemotherapy and photothermal therapy (PTT) have emerged as a promising anticancer paradigm to achieve expected therapeutic effects while mitigating side effects. However, the chemo/PTT combination therapy suffers from limited penetration depth, thermoresistance performance of tumor cells, and low drug bioavailability. Herein, multifunctional nanoparticles (BTP/DOX/2DG NPs) coloaded with near-infrared region II (NIR-II) light excitation donor-acceptor-donor (D-A-D) small molecules, doxorubicin (DOX), and 2-deoxy-d-glucose (2-DG) are developed for reinforced starvation/chemo/NIR-II PTT combination therapy. The synthesized phenylboronic acid (PBA)-modified water-soluble D-A-D molecule (BBT-TF-PBA) not only exhibits high binding ability to DOX and 2-DG through donor-acceptor coordination interactions PBA-diol bonds but also serves as a photoactive agent for NIR-II fluorescence imaging, NIR-II photoacoustic imaging, and NIR-II PTT. Under the acidic and oxidizing conditions in the tumor microenvironment, donor-acceptor coordination interactions and PBA-diol bond are decomposed, simultaneously releasing DOX and 2-DG from BTP/DOX/2DG NPs to achieve effective chemotherapy and starvation therapy. 2-DG also effectively inhibits the expression of heat shock protein and further enhances NIR-II PTT and chemotherapy efficiency. In vitro and in vivo experiments demonstrate the combination effect of BTP/DOX/2DG NPs for chemotherapy, NIR-II PTT, and starvation therapy.

Investigations into the Signaling Pathways Involving Glucose-Stimulated Zinc Secretion (GSZS) from Prostate Epithelial Cells In Vitro and In Vivo.

PURPOSE: Recently, we reported that exposure of prostate cells in vitro or the in vivo prostate to high glucose results in release of Zn2+ ions, a process now referred to as glucose-stimulated zinc secretion (GSZS). To our knowledge, the metabolic event(s) that trigger GSZS remain largely unknown. Here, we explore several signaling pathways both in vitro using a prostate epithelial cell line and in vivo from the rat prostate. METHODS: PNT1A cells grown to confluence were washed and tagged with ZIMIR to monitor zinc secretion by optical methods. The expression levels of GLUT1, GLUT4, and Akt in cells cultured in either zinc-rich or zinc-poor media and after exposure to high versus low glucose were determined. Zinc secretion from the rat prostate in vivo as detected by MRI was compared in control animals after injection of glucose, deoxyglucose, or pyruvate to initiate zinc secretion and in animals pre-treated with WZB-117 (a GLUT1 inhibitor) or S961 (a peripheral insulin receptor inhibitor). RESULTS: PNT1A cells exposed to high levels of glucose secrete zinc whereas cells exposed to an equivalent amount of deoxyglucose or pyruvate do not. Expression of Akt was dramatically altered by zinc supplementation of the culture media but not after exposure to glucose while GLUT1 and GLUT4 levels were less affected. Rats pre-treated with WZB-117 prior to imaging showed a reduction in GSZS from the prostate compared to controls whereas rats pre-treated with S961 showed no difference. Interestingly, in comparison to PNT1A cells, pyruvate and deoxyglucose also stimulate zinc secretion in vivo likely through indirect mechanisms. CONCLUSIONS: GSZS requires metabolism of glucose both in vitro (PNT1A cells) and in vivo (rat prostate). Pyruvate also stimulates zinc secretion in vivo but likely via an indirect pathway involving rapid production of glucose via gluconeogenesis. These combined results support the conclusion that glycolytic flux is required to trigger GSZS in vivo.

Role of Glycolysis and Fatty Acid Synthesis in the Activation and T Cell-Modulating Potential of Dendritic Cells Stimulated with a TLR5-Ligand Allergen Fusion Protein.

Trained immune responses, based on metabolic and epigenetic changes in innate immune cells, are de facto innate immune memory and, therefore, are of great interest in vaccine development. In previous studies, the recombinant fusion protein rFlaA:Betv1, combining the adjuvant and toll-like receptor (TLR)5-ligand flagellin (FlaA) and the major birch pollen allergen Bet v 1 into a single molecule, significantly suppressed allergic sensitization in vivo while also changing the metabolism of myeloid dendritic cells (mDCs). Within this study, the immune-metabolic effects of rFlaA:Betv1 during mDC activation were elucidated. In line with results for other well-characterized TLR-ligands, rFlaA:Betv1 increased glycolysis while suppressing oxidative phosphorylation to different extents, making rFlaA:Betv1 a suitable model to study the immune-metabolic effects of TLR-adjuvanted vaccines. In vitro pretreatment of mDCs with cerulenin (inhibitor of fatty acid biosynthesis) led to a decrease in both rFlaA:Betv1-induced anti-inflammatory cytokine Interleukin (IL) 10 and T helper cell type (TH) 1-related cytokine IL-12p70, while the pro-inflammatory cytokine IL 1ß was unaffected. Interestingly, pretreatment with the glutaminase inhibitor BPTES resulted in an increase in IL-1ß, but decreased IL-12p70 secretion while leaving IL-10 unchanged. Inhibition of the glycolytic enzyme hexokinase-2 by 2-deoxyglucose led to a decrease in all investigated cytokines (IL-10, IL-12p70, and IL-1ß). Inhibitors of mitochondrial respiration had no effect on rFlaA:Betv1-induced IL-10 level, but either enhanced the secretion of IL-1ß (oligomycin) or decreased IL-12p70 (antimycin A). In extracellular flux measurements, mDCs showed a strongly enhanced glycolysis after rFlaA:Betv1 stimulation, which was slightly increased after respiratory shutdown using antimycin A. rFlaA:Betv1-stimulated mDCs secreted directly antimicrobial substances in a mTOR- and fatty acid metabolism-dependent manner. In co-cultures of rFlaA:Betv1-stimulated mDCs with CD4+ T cells, the suppression of Bet v 1-specific TH2 responses was shown to depend on fatty acid synthesis. The effector function of rFlaA:Betv1-activated mDCs mainly relies on glycolysis, with fatty acid synthesis also significantly contributing to rFlaA:Betv1-mediated cytokine secretion, the production of antimicrobial molecules, and the modulation of T cell responses.

Metabolic modulation of synaptic failure and thalamocortical hypersynchronization with preserved consciousness in Glut1 deficiency.

Individuals with glucose transporter type I deficiency (G1D) habitually experience nutrient-responsive epilepsy associated with decreased brain glucose. However, the mechanistic association between blood glucose concentration and brain excitability in the context of G1D remains to be elucidated. Electroencephalography (EEG) in G1D individuals revealed nutrition time-dependent seizure oscillations often associated with preserved volition despite electrographic generalization and uniform average oscillation duration and periodicity, suggesting increased facilitation of an underlying neural loop circuit. Nonlinear EEG ictal source localization analysis and simultaneous EEG/functional magnetic resonance imaging converged on the thalamus-sensorimotor cortex as one potential circuit, and 18F-deoxyglucose positron emission tomography (18F-DG-PET) illustrated decreased glucose accumulation in this circuit. This pattern, reflected in a decreased thalamic to striatal 18F signal ratio, can aid with the PET imaging diagnosis of the disorder, whereas the absence of noticeable ictal behavioral changes challenges the postulated requirement for normal thalamocortical activity during consciousness. In G1D mice, 18F-DG-PET and mass spectrometry also revealed decreased brain glucose and glycogen, but preserved tricarboxylic acid cycle intermediates, indicating no overall energy metabolism failure. In brain slices from these animals, synaptic inhibition of cortical pyramidal neurons and thalamic relay neurons was decreased, and neuronal disinhibition was mitigated by metabolic sources of carbon; tonic-clonic seizures were also suppressed by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor inhibition. These results pose G1D as a thalamocortical synaptic disinhibition disease associated with increased glucose-dependent neuronal excitability, possibly in relation to reduced glycogen. Together with findings in other metabolic defects, inhibitory neuron dysfunction is emerging as a modulable mechanism of hyperexcitability.

Anti-epileptic effect of 2-deoxy-D-glucose by activation of miR-194/KATP signaling pathway. / 2-脱氧-D-葡萄糖激活miR-194/KATP信号通路发挥抗癫痫作用.

OBJECTIVES: Epilepsy is a syndrome of central nervous system dysfunction caused by many reasons, which is mainly characterized by abnormal discharge of neurons in the brain. Therefore, finding new targets for epilepsy therapy has always been the focus and hotspot in neurological research field. Studies have found that 2-deoxy-D-glucose (2-DG) exerts anti-epileptic effect by up-regulation of KATP channel subunit Kir6.1, Kir6.2 mRNA and protein. By using the database of TargetScan and miRBase to perform complementary pairing analysis on the sequences of miRNA and related target genes, it predicted that miR-194 might be the upstream signaling molecule of KATP channel. This study aims to explore the mechanism by which 2-DG exerts its anti-epileptic effect by regulating KATP channel subunits Kir6.1 and Kir6.2 via miR-194. METHODS: A magnesium-free epilepsy model was established and randomly divided into a control group, an epilepsy group (EP group), an EP+2-DG group, and miR-194 groups (including EP+miR-194 mimic, EP+miR-194 mimic+2-DG, EP+miR-194 mimic control, EP+miR-194 inhibitor, EP+miR-194 inhibitor+2-DG, and EP+miR-194 inhibitor control groups). The 2-DG was used to intervene miR-194 mimics, patch-clamp method was used to detect the spontaneous recurrent epileptiform discharges, real-time PCR was used to detect neuronal miR-194, Kir6.1, and Kir6.2 expressions, and the protein levels of Kir6.1 and Kir6.2were detected by Western blotting. RESULTS: Compared with the control group, there was no significant difference in the amplitude of spontaneous discharge potential in the EP group (P>0.05), but the frequency of spontaneous discharge was increased (P<0.05). Compared with the EP group, the frequency of spontaneous discharge was decreased (P<0.05). Compared with the EP+miR-194 mimic control group, the mRNA and protein expressions of Kir6.1 and Kir6.2 in the EP+miR-194 mimic group were down-regulated (all P<0.05). Compared with the EP+miR-194 inhibitor control group, the mRNA and protein expressions of Kir6.1 and Kir6.2 in the EP+miR-194 inhibitor group were up-regulated (all P<0.05). After pretreatment with miR-194 mimics, the mRNA and protein expression levels of KATP channel subunits Kir6.1 and Kir6.2 were decreased (all P<0.05). Compared with the EP+2-DG group, the mRNA and protein expression levels of Kir6.1 and Kir6.2 in the EP+miR-194 mimic+2-DG group were down-regulated (all P<0.05) and the mRNA and protein expression levels of Kir6.1 and Kir6.2 in the EP+miR-194 inhibitor+2-DG group were up-regulated (all P<0.05). CONCLUSIONS: The 2-DG might play an anti-epilepsy effect by up-regulating KATP channel subunits Kir6.1 and Kir6.2via miR-194.

Blood-based untargeted metabolomics in relapsing-remitting multiple sclerosis revealed the testable therapeutic target.

Metabolic aberrations impact the pathogenesis of multiple sclerosis (MS) and possibly can provide clues for new treatment strategies. Using untargeted metabolomics, we measured serum metabolites from 35 patients with relapsing-remitting multiple sclerosis (RRMS) and 14 healthy age-matched controls. Of 632 known metabolites detected, 60 were significantly altered in RRMS. Bioinformatics analysis identified an altered metabotype in patients with RRMS, represented by four changed metabolic pathways of glycerophospholipid, citrate cycle, sphingolipid, and pyruvate metabolism. Interestingly, the common upstream metabolic pathway feeding these four pathways is the glycolysis pathway. Real-time bioenergetic analysis of the patient-derived peripheral blood mononuclear cells showed enhanced glycolysis, supporting the altered metabolic state of immune cells. Experimental autoimmune encephalomyelitis mice treated with the glycolytic inhibitor 2-deoxy-D-glucose ameliorated the disease progression and inhibited the disease pathology significantly by promoting the antiinflammatory phenotype of monocytes/macrophage in the central nervous system. Our study provided a proof of principle for how a blood-based metabolomic approach using patient samples could lead to the identification of a therapeutic target for developing potential therapy.

In Vitro Evaluation of Antibacterial, Antioxidant, and Antidiabetic Activities and Glucose Uptake through 2-NBDG by Hep-2 Liver Cancer Cells Treated with Green Synthesized Silver Nanoparticles.

The alarming rise in diabetes owing to drug resistance necessitates the implementation of prompt countermeasures in the treatment module of diabetes. Due to their unique physicochemical features, silver nanoparticles may have potential applications in the medical and pharmaceutical industries. Silver nanoparticles (AgNPs) were synthesized from the culture filtrate of Salmonella enterica (ATCC-14028). UV-Vis spectrophotometry, FTIR, SEM, and energy dispersive X-rays were used in the characterization of the nanoparticles. Transmission electron microscopy (TEM) revealed that AgNPs are spherical and highly scattered and vary in size from 7.18 nm to 13.24 nm. AgNP stability and protein loss were confirmed by thermogravimetric analysis (TGA) at different temperatures. The AgNPs had excellent antibacterial activity and a strong synergistic effect against methicillin-resistant bacteria Staphylococcus aureus (MRSA) ATCC-4330 and Streptococcus epidermis (MRSE) ATCC-51625. The DPPH experiment revealed that the AgNPs had high antioxidant activity. The antidiabetic assay revealed that these AgNPs had an IC50 for alpha-amylase of 428.60 µg/ml and an IC50 for alpha-glucosidase of 562.02 µg/ml. Flow cytometry analysis of Hep-2 cells treated with AgNPs (40 µg/ml) revealed higher expression of 2-NBDG glucose absorption (uptake) compared to control metformin. These AgNPs have promising antidiabetic properties and could be used in pharmaceuticals and biomedical industries.

Reduction of 3-Deoxyglucosone by Epigallocatechin Gallate Results Partially from an Addition Reaction: The Possible Mechanism of Decreased 5-Hydroxymethylfurfural in Epigallocatechin Gallate-Treated Black Garlic.

5-Hydroxymethylfurfural (5-HMF) is a harmful substance generated during the processing of black garlic. Our previous research demonstrated that impregnation of black garlic with epigallocatechin gallate (EGCG) could reduce the formation of 5-HMF. However, there is still a lack of relevant research on the mechanism and structural identification of EGCG inhibiting the production of 5-HMF. In this study, an intermediate product of 5-HMF, 3-deoxyglucosone (3-DG), was found to be decreased in black garlic during the aging process, and impregnation with EGCG for 24 h further reduced the formation of 3-DG by approximately 60% in black garlic compared with that in the untreated control. The aging-mimicking reaction system of 3-DG + EGCG was employed to determine whether the reduction of 3-DG was the underlying mechanism of decreased 5-HMF formation in EGCG-treated black garlic. The results showed that EGCG accelerated the decrease of 3-DG and further attenuated 5-HMF formation, which may be caused by an additional reaction with 3-DG, as evidenced by LC-MS/MS analysis. In conclusion, this study provides new insights regarding the role of EGCG in blocking 5-HMF formation.

Effect of biochanin A on the rumen microbial community of Holstein steers consuming a high fiber diet and subjected to a subacute acidosis challenge.

Subacute rumen acidosis (SARA) occurs when highly fermentable carbohydrates are introduced into the diet, decreasing pH and disturbing the microbial ecology of the rumen. Rumen amylolytic bacteria rapidly catabolize starch, fermentation acids accumulate in the rumen and reduce environmental pH. Historically, antibiotics (e.g., monensin, MON) have been used in the prevention and treatment of SARA. Biochanin A (BCA), an isoflavone produced by red clover (Trifolium pratense), mitigates changes associated with starch fermentation ex vivo. The objective of the study was to determine the effect of BCA on amylolytic bacteria and rumen pH during a SARA challenge. Twelve rumen fistulated steers were assigned to 1 of 4 treatments: HF CON (high fiber control), SARA CON, MON (200 mg d-1), or BCA (6 g d-1). The basal diet consisted of corn silage and dried distiller's grains ad libitum. The study consisted of a 2-wk adaptation, a 1-wk HF period, and an 8-d SARA challenge (d 1-4: 40% corn; d 5-8: 70% cracked corn). Samples for pH and enumeration were taken on the last day of each period (4 h). Amylolytic, cellulolytic, and amino acid/peptide-fermenting bacteria (APB) were enumerated. Enumeration data were normalized by log transformation and data were analyzed by repeated measures ANOVA using the MIXED procedure of SAS. The SARA challenge increased total amylolytics and APB, but decreased pH, cellulolytics, and in situ DMD of hay (P < 0.05). BCA treatment counteracted the pH, microbiological, and fermentative changes associated with SARA challenge (P < 0.05). Similar results were also observed with MON (P < 0.05). These results indicate that BCA may be an effective alternative to antibiotics for mitigating SARA in cattle production systems.

Cytotoxic activity of high dose ascorbic acid is enhanced by 2-deoxy-d-glucose in glycolytic melanoma cells.

Although, numerous in vitro studies showed that cancer cells are killed after exposure to pharmacological doses of ascorbic acid (AA), significant clinical data proving the efficacy of AA is still absent. A hallmark of most tumor cells is an altered glucose metabolism characterized by an upregulation of glycolysis despite normoxic conditions (Warburg effect). Since pyruvate is capable of detoxifying hydrogen peroxide (H2O2), the alleged mediator of AA-induced toxicity, it seems likely that enhanced glycolysis and subsequent higher pyruvate formation might be an explanation for the attenuated effect of pharmacological AA in vivo. Therefore, inhibition of glycolysis might be a promising approach to enhance the anticancer effect of AA by diminishing the generation of pyruvate. Considering the altered metabolism of cancer cells, we examined the cytotoxic potential of 2-DG and/or AA using SRB assay in two different cell lines: a glycolytic human melanoma (451Lu) and a non-glycolytic breast cancer (MCF-7) cell line. Inhibition of glycolysis increased AA cytotoxicity in 451Lu cells, whereas same treatment had a marginal effect on MCF-7 cells. We also investigated the influence of glycolysis inhibition on H2O2 generation. H2O2 concentrations were higher in presence of 451Lu cells when pretreated with 2-DG, but not in MCF-7 cells. Treatment with 10 mM 2-DG decreased pyruvate and lactate concentrations in both cell lines in a concentration-dependent manner. In summary, 2-DG enhances the cytotoxic effect of AA in glycolytic 451Lu cells by increasing AA-induced H2O2 concentration. This result indicates that lower pyruvate levels, as a result of glycolysis inhibition, may be responsible for the enhanced effect of 2-DG on AA toxicity. Further experiments are needed to clarify the underlying mechanism and the potential use in cancer therapy.

HM-chromanone, a component of Portulaca oleracea L., stimulates glucose uptake and glycogen synthesis in skeletal muscle cell.

BACKGROUND: Diabetes mellitus is a chronic metabolic disease characterized by increased blood glucose levels. In order to lower blood glucose, it is important to stimulate glucose uptake and glycogen synthesis in the muscle. (E)-5-hydroxy-7-methoxy-3-(2'-hydroxybenzyl)-4-chromanone (HM-chromanone), a constituent isolated from Portulaca oleracea L., exhibits anti-diabetic effects; however, its mechanisms are not yet clearly understood on glucose uptake and glycogen synthesis in muscle cells. PURPOSE: In the present study, we examined the effects of HM-chromanone on glucose uptake into L6 skeletal muscle cells and elucidated the underlying mechanisms. METHODS: The effects of HM-chromanone on glucose uptake into L6 skeletal muscle cells were assessed by 2-Deoxyglucose uptake assay. Western blot analysis was carried out to elucidate the underlying molecular mechanisms. RESULTS: We found that HM-chromanone promoted glucose uptake into L6 skeletal muscle cells in a dose-dependent manner. Moreover, HM-chromanone induced the phosphorylation of IRS-1Tyr612 and AKTSer473, and the activation of PI3K. HM-chromanone also stimulated the phosphorylation of AMPKThr172, AS160Thr642, TBC1D1Ser237, and ACC via the CaMKKß pathway. Furthermore, HM-chromanone increased glycogen synthesis through the inactivation of glycogen synthase kinase 3 α/ß. CONCLUSION: The results of this study indicate that HM-chromanone stimulates glucose uptake through the activation of the PI3K/AKT and CaMKKß-AMPK pathways and glycogen synthesis via the GSK3 α/ß pathway in L6 skeletal muscle cells.

Rosemary extract activates AMPK, inhibits mTOR and attenuates the high glucose and high insulin-induced muscle cell insulin resistance.

Impaired action of insulin in skeletal muscle, termed insulin resistance, leads to increased blood glucose levels resulting in compensatory increase in insulin levels. The elevated blood glucose and insulin levels exacerbate insulin resistance and contribute to the pathogenesis of type 2 diabetes mellitus. In previous studies we found attenuation of free fatty acid-induced muscle cell insulin resistance by rosemary extract (RE). In the present study we investigated the effects of RE on high glucose (HG) and high insulin (HI)-induced muscle cell insulin resistance. Exposure of L6 myotubes to 25 mmol/L glucose and 100 nmol/L insulin for 24 h, to mimic hyperglycemia and hyperinsulinemia, abolished the acute insulin-stimulated glucose uptake, increased the serine phosphorylation of IRS-1 and the phosphorylation/activation of mTOR and p70S6K. Treatment with RE significantly improved the insulin-stimulated glucose uptake and increased the acute insulin-stimulated tyrosine phosphorylation while reducing the HG+HI-induced serine phosphorylation of IRS-1 and phosphorylation of mTOR and p70S6K. Additionally, treatment with RE significantly increased the phosphorylation of AMPK, its downstream effector ACC and the plasma membrane GLUT4 levels. Our data indicate a potential of RE to counteract muscle cell insulin resistance and more studies are required to investigate its effectiveness in vivo. Novelty: RE phosphorylated muscle cell AMPK and ACC under both normal and HG+HI conditions. The HG+HI-induced serine phosphorylation of IRS-1 and activation of mTOR and p70S6K were attenuated by RE. RE restored the insulin-stimulated glucose uptake by enhancing GLUT4 glucose transporter translocation to plasma membrane.

KD025 Shifts Pulmonary Endothelial Cell Bioenergetics and Decreases Baseline Lung Permeability.

KD025 is a ROCK2 inhibitor currently being tested in clinical trials for the treatment of fibrotic lung diseases. The therapeutic effects of KD025 are partly due to its inhibition of profibrotic pathways and fat metabolism. However, whether KD025 affects pulmonary microvascular endothelial cell (PMVEC) function is unknown, despite evidence that alveolar-capillary membrane disruption constitutes major causes of death in fibrotic lung diseases. We hypothesized that KD025 regulates PMVEC metabolism, pH, migration, and survival, a series of interrelated functional characteristics that determine pulmonary barrier integrity. We used PMVECs isolated from Sprague Dawley rats. KD025 dose-dependently decreased lactate production and glucose consumption. The inhibitory effect of KD025 was more potent compared with other metabolic modifiers, including 2-deoxy-glucose, extracellular acidosis, dichloroacetate, and remogliflozin. Interestingly, KD025 increased oxidative phosphorylation, whereas 2-deoxy-glucose did not. KD025 also decreased intracellular pH and induced a compensatory increase in anion exchanger 2. KD025 inhibited PMVEC migration, but fasudil (nonspecific ROCK inhibitor) did not. We tested endothelial permeability in vivo using Evans Blue dye in the bleomycin pulmonary fibrosis model. Baseline permeability was decreased in KD025-treated animals independent of bleomycin treatment. Under hypoxia, KD025 increased PMVEC necrosis as indicated by increased lactate dehydrogenase release and propidium iodide uptake and decreased ATP; it did not affect Annexin V binding. ROCK2 knockdown had no effect on PMVEC metabolism, pH, and migration, but it increased nonapoptotic caspase-3 activity. Together, we report that KD025 promotes oxidative phosphorylation; decreases glycolysis, intracellular pH, and migration; and strengthens pulmonary barrier integrity in a ROCK2-independent manner.

Cytoprotective effects of a proprietary red maple leaf extract and its major polyphenol, ginnalin A, against hydrogen peroxide and methylglyoxal induced oxidative stress in human keratinocytes.

Phytochemicals from functional foods are common ingredients in dietary supplements and cosmetic products for anti-skin aging effects due to their antioxidant activities. A proprietary red maple (Acer rubrum) leaf extract (Maplifa™) and its major phenolic compound, ginnalin A (GA), have been reported to show antioxidant, anti-melanogenesis, and anti-glycation effects but their protective effects against oxidative stress in human skin cells remain unknown. Herein, we investigated the cytoprotective effects of Maplifa™ and GA against hydrogen peroxide (H2O2) and methylglyoxal (MGO)-induced oxidative stress in human keratinocytes (HaCaT cells). H2O2 and MGO (both at 400 µM) induced toxicity in HaCaT cells and reduced their viability to 59.2 and 61.6%, respectively. Treatment of Maplifa™ (50 µg mL-1) and GA (50 µM) increased the viability of H2O2- and MGO-treated cells by 22.0 and 15.5%, respectively. Maplifa™ and GA also showed cytoprotective effects by reducing H2O2-induced apoptosis in HaCaT cells by 8.0 and 7.2%, respectively. The anti-apoptotic effect of Maplifa™ was further supported by the decreased levels of apoptosis associated enzymes including caspases-3/7 and -8 in HaCaT cells by 49.5 and 19.0%, respectively. In addition, Maplifa™ (50 µg mL-1) and GA (50 µM) reduced H2O2- and MGO-induced reactive oxygen species (ROS) by 84.1 and 56.8%, respectively. Furthermore, flow cytometry analysis showed that Maplifa™ and GA reduced MGO-induced total cellular ROS production while increasing mitochondria-derived ROS production in HaCaT cells. The cytoprotective effects of Maplifa™ and GA in human keratinocytes support their potential utilization for cosmetic and/or dermatological applications.

Acacetin enhances glucose uptake through insulin-independent GLUT4 translocation in L6 myotubes.

BACKGROUND: Lowering blood glucose levels by increasing glucose uptake in insulin target tissues, such as skeletal muscle and adipose tissue, is one strategy to discover and develop antidiabetic drugs from natural products used as traditional medicines. PURPOSE: Our goal was to reveal the mechanism and activity of acacetin (5,7-dihydroxy-4'-methoxyflavone), one of the major compounds in Agastache rugose, in stimulating glucose uptake in muscle cells. METHODS: To determine whether acacetin promotes GLUT4-dependent glucose uptake in cultured L6 skeletal muscle cells, we performed a [14C] 2-deoxy-D-glucose (2-DG) uptake assay after treating differentiated L6-GLUT4myc cells with acacetin. RESULTS: Acacetin dose-dependently increased 2-DG uptake by enhancing GLUT4 translocation to the plasma membrane. Our results revealed that acacetin activated the CaMKII-AMPK pathway by increasing intracellular calcium concentrations. We also found that aPKCλ/ζ phosphorylation and intracellular reactive oxygen species (ROS) production were involved in acacetin-induced GLUT4 translocation. Moreover, acacetin-activated AMPK inhibited intracellular lipid accumulation and increased 2-DG uptake in HepG2 cells. CONCLUSION: Taken together, these results suggest that acacetin might be useful as an antidiabetic functional ingredient. Subsequent experiments using disease model animals are needed to verify our results.

5-Hydroxymethylfurfural accumulation plays a critical role on acrylamide formation in coffee during roasting as confirmed by multiresponse kinetic modelling.

This study aims to investigate in depth the mechanism of acrylamide formation in coffee during roasting. For this purpose, a comprehensive kinetic model including the elementary steps for acrylamide formation was proposed. The changes in sucrose, reducing sugars, free amino acids, asparagine, acrylamide, 3-deoxyglucosone, methylglyoxal, glyoxal, and 5-hydroxymethylfurfural were monitored in coffee during roasting at 200, 220 and 240 °C. Dominant pathways of complex reactions leading to acrylamide were unravelled by means of multiresponse kinetic modelling approach. The results of the model indicated that sucrose degrades into glucose and a reactive fructofuranosyl cation. Interestingly, glucose takes part mostly in the formation of intermediates, glyoxal and especially 3-deoxyglucosone rather than acrylamide formation. On the other hand, fructofuranosyl cation contributed mostly to the formation of 5-hydroxymethylfurfural which was found to be the most important intermediate precursor of acrylamide formed in coffee during roasting.

Glycation and Antioxidants: Hand in the Glove of Antiglycation and Natural Antioxidants.

The non-enzymatic interaction of sugar and protein resulting in the formation of advanced glycation end products responsible for cell signaling alterations ultimately leads to the human chronic disorders such as diabetes mellitus, cardiovascular diseases, cancer, etc. Studies suggest that AGEs upon interaction with receptors for advanced glycation end products (RAGE) result in the production of pro-inflammatory molecules and free radicals that exert altered gene expression effect. To date, many studies unveiled the potent role of synthetic and natural agents in inhibiting the glycation reaction at a lesser or greater extent. This review focuses on the hazards of glycation reaction and its inhibition by natural antioxidants, including polyphenols.