AP collagen peptides (APCPs) promote hair growth by activating the GSK-3ß/ß-catenin pathway and improve hair condition.

AP collagen peptides (APCPs) are enzymatically decomposed collagen peptides that contain tri-peptides such as glycine-proline-hydroxyproline. We found that APCPs increased the proliferation of both human dermal papilla cells (hDPCs) and human outer root sheath cells (hORSCs). APCPs also stimulated the secretion of several growth factors, including IGFBP-6, PDGF-AB, PIGF and VEGF in hDPCs. Moreover, APCPs enhanced the phosphorylation of Akt(Ser473), GSK-3ß(Ser9) and ß-catenin(Ser675), indicating the activation of the GSK-3ß/ß-catenin signalling pathway. Ex vivo culture of human hair follicles (hHFs) tissue and in vivo patch assay revealed that APCPs promoted the elongation of hHFs and the induction of new hair shafts. In a mouse model, APCPs significantly promoted the transition from telogen to anagen phase and prolonged anagen phase, resulting in increased hair growth. APCPs also improved the thickness, amino acid content (cystine and methionine) and roughness of mouse hair. Taken together, these findings demonstrate that APCPs accelerate hair growth and contribute to overall hair health. Therefore, APCPs have the potential to be utilized as a food supplement and ingredient for preventing hair loss and maintaining hair health.

Exosomes derived from human dermal fibroblasts (HDFn-Ex) alleviate DNCB-induced atopic dermatitis (AD) via PPARα.

Atopic dermatitis (AD) is a chronic inflammatory skin disease. Skin barrier dysfunction is the initial step in the development of AD. Recently, exosomes have been considered as potential cell-free medicine for skin defects such as aging, psoriasis and wounds. The aim of this study was to investigate the effects of human dermal fibroblast-neonatal-derived exosome (HDFn-Ex) on AD. HDFn-Ex increased the expression of peroxisome proliferator activated receptor α (PPARα) and alleviated the 1-chloro-2,4-dinitrobenzene (DNCB)-mediated downregulation of filaggrin, involucrin, loricrin, hyaluronic acid synthase 1 (HAS1) and HAS2 in human keratinocyte HaCaT cells. However, these effects were inhibited by the PPARα antagonist GW6471. In the artificial skin model, HDFn-Ex significantly inhibited DNCB-induced epidermal hyperplasia and the decrease in filaggrin and HAS1 levels via a PPARα. In the DNCB-induced AD-like mouse model, HDFn-Ex administration reduced epidermis thickening and mast cell infiltration into the dermis compared to DNCB treatment. Moreover, the decreases in PPARα, filaggrin and HAS1 expression, as well as the increases in IgE and IL4 levels induced by DNCB treatment were reversed by HDFn-Ex. These effects were blocked by pre-treatment with GW6471. Furthermore, HDFn-Ex exhibited an anti-inflammatory effect by inhibiting the DNCB-induced increases in IκBα phosphorylation and TNF-α expression. Collectively, HDFn-Ex exhibited a protective effect on AD. Notably, these effects were regulated by PPARα. Based on our results, we suggest that HDFn-Ex is a potential candidate for treating AD by recovering skin barrier dysfunction and exhibiting anti-inflammatory activity.

A comparison of dyeing efficacy between hair-oxidation-based and hair-coating-based shampoos for the treatment of gray hair.

BACKGROUND: The process of hair dyeing causes hair damage, and periodic re-dyeing is required for newly grown hair. To avoid these hassles, hair color shampoos have been developed and are widely used. In this study, we compared the effects of two hair color shampoos with different dyeing principles to analyze the function of hair color shampoos. We analyzed hair tresses treated by hair-oxidation- and hair-coating-based shampoos. MATERIALS AND METHODS: We measured the color, tensile properties, softness, elasticity, gloss, moisture content, and protein content of the hair tresses dyed with color shampoos. The hair structures were analyzed by scanning and transmission electron microscopies (SEM and TEM) and a hydroxy radical-based method. RESULTS: The shampoo based on hair coating enhanced the hair dyeing effect and roughness, whereas that based on hair oxidation improved the color retention and moisture content in the hair tresses. Frictional resistance, gloss, and elasticity of the hair tresses were similar for the two products. However, according to the results of the protein loss test, TEM, and hydroxyl radical staining, the shampoo based on hair oxidation showed a longer dyeing retention compared to that based on hair coating but caused cuticle damage. CONCLUSION: These results show that the two shampoos with different dyeing principles exhibit different hair dyeing abilities and hair health indices. Therefore, we recommend that hair color shampoos should be used according to the requirements of an individual.

Biglycan reduces body weight by regulating food intake in mice and improves glucose metabolism through AMPK/AKT dual pathways in skeletal muscle.

While biglycan (BGN) is suggested to direct diverse signaling cascades, the effects of soluble BGN as a ligand on metabolic traits have not been studied. Herein, we tested the effects of BGN on obesity in high-fat diet (HFD)-induced obese animals and glucose metabolism, with the underlying mechanism responsible for observed effects in vitro. Our results showed that BGN administration (1 mg/kg body weight, intraperitoneally) significantly prevented HFD-induced obesity, and this was mainly attributed to reduced food intake. Also, intracerebroventricular injection of BGN reduced food intake and body weight. The underlying mechanism includes modulation of neuropeptides gene expression involved in appetite in the hypothalamus in vitro and in vivo. In addition, BGN regulates glucose metabolism as shown by improved glucose tolerance in mice as well as AMPK/AKT dual pathway-driven enhanced glucose uptake and GLUT4 translocation in L6 myoblast cells. In conclusion, our results suggest BGN as a potential therapeutic target to treat risk factors for metabolic diseases.

Low-intensity cold atmospheric plasma reduces wrinkles on photoaged skin through hormetic induction of extracellular matrix protein expression in dermal fibroblasts.

BACKGROUND: Recent evidence indicates that cold atmospheric plasma (CAP) can upregulate the production of extracellular matrix (ECM) proteins in dermal fibroblasts and enhance transdermal drug delivery when applied at a low intensity. OBJECTIVES: The aim of this study was to evaluate the effect of low-intensity CAP (LICAP) on photoaging-induced wrinkles in an animal model and the expression profiles of ECM proteins in human dermal fibroblasts. METHODS: Each group was subjected to photoaging induction and allocated to therapy (LICAP, topical polylactic acid (PLA), or both). The wrinkles were evaluated via visual inspection, quantitative analysis, and histology. The expression of collagen I/III and fibronectin was assessed using reverse transcription-quantitative polymerase chain reaction, western blot analysis, and immunofluorescence. The amount of aqueous reactive species produced by LICAP using helium and argon gas was also measured. RESULTS: Wrinkles significantly decreased in all treatment groups compared to those in the untreated control. The differences remained significant for at least 4 weeks. Dermal collagen density increased following LICAP and PLA application. LICAP demonstrated a hormetic effect on ECM protein expression in human dermal fibroblasts. The production of reactive species increased, showing a biphasic pattern, with an initial linear phase and a slow saturation phase. The initial linearity was sustained for a longer time in the helium plasma (~60 s) than in the argon plasma (~15 s). CONCLUSION: LICAP appears to be a novel treatment option for wrinkles on the photodamaged skin. This treatment effect seems to be related to its hormetic effect on dermal ECM production.

Dehydrozingerone inhibits renal lipotoxicity in high-fat diet-induced obese mice.

Ectopic fat accumulation in the kidneys causes oxidative stress, inflammation and cell death. Dehydrozingerone (DHZ) is a curcumin analog that exhibits antitumour, antioxidant and antidiabetic effects. However, the efficacy of DHZ in diabetic nephropathy (DN) is unknown. Here, we verified the efficacy of DHZ on DN. We divided the experimental animals into three groups: regular diet, 60% high-fat diet (HFD) and HFD with DHZ for 12 weeks. We analysed levels of renal triglycerides and urinary albumin and albumin-creatinine ratio, renal morphological changes and molecular changes via real-time polymerase chain reaction and immunoblotting. Furthermore, high glucose (HG)- or palmitate (PA)-stimulated mouse mesangial cells or mouse podocytes were treated with DHZ for 24 h. As a result, DHZ markedly reduced renal glycerol accumulation and albuminuria excretion through improvement of thickened glomerular basement membrane, podocyte loss and slit diaphragm reduction. In the renal cortex in the HFD group, phospho-AMPK and nephrin expression reduced, whereas arginase 2 and CD68 expression increased; however, these changes were recovered after DHZ administration. Increased reactive oxygen species (ROS) stimulated by HG or PA in podocytes was inhibited by DHZ treatment. Collectively, these findings indicate that DHZ ameliorates DN via inhibits of lipotoxicity-induced inflammation and ROS formation.

Human placenta-derived mesenchymal stem cells induce trophoblast invasion via dynamic effects on mitochondrial function.

The trophoblast is a critical cell for placental development and embryo implantation in the placenta. We previously reported that placenta-derived mesenchymal stem cells (PD-MSCs) increase trophoblast invasion through several signaling pathways. However, the paracrine effects of PD-MSCs on mitochondrial function in trophoblasts are still unclear. Therefore, the objective of the study was to analyze the mitochondrial function of trophoblasts in response to cocultivation with PD-MSCs. The results showed that PD-MSCs regulate the balance between cell survival and death and protect damaged mitochondria in trophoblasts from oxidative stress. Moreover, PD-MSCs upregulate factors involved in mitochondrial autophagy in trophoblast cells. Finally, PD-MSCs improve trophoblast invasion. Taken together, the data indicate that PD-MSCs can regulate trophoblast invasion through dynamic effects on mitochondrial energy metabolism. These results support the fundamental role of mitochondrial energy mechanism in trophoblast invasion and suggest a new therapeutic strategy for infertility.

Effect of chitinase-3-like protein 1 on glucose metabolism: In vitro skeletal muscle and human genetic association study.

We investigated the effect of chitinase-3-like protein 1 (CHI3L1) on glucose metabolism and its underlying mechanisms in skeletal muscle cells, and evaluated whether the observed effects are relevant in humans. CHI3L1 was associated with increased glucose uptake in skeletal muscles in an AMP-activated protein kinase (AMPK)-dependent manner, and with increased intracellular calcium levels via PAR2. The improvement in glucose metabolism observed in an intraperitoneal glucose tolerance test on male C57BL/6J mice supported this association. Inhibition of the CaMKK was associated with suppression of CHI3L1-mediated glucose uptake. Additionally, CHI3L1 was found to influence glucose uptake through the PI3K/AKT pathway. Results suggested that CHI3L1 stimulated the phosphorylation of AS160 and p38 MAPK downstream of AMPK and AKT, and the resultant GLUT4 translocation. In primary myoblast cells, stimulation of AMPK and AKT was observed in response to CHI3L1, underscoring the biological relevance of CHI3L1. CHI3L1 levels were elevated in cells under conditions that mimic exercise in vitro and in exercised mice in vivo, indicating that CHI3L1 is secreted during muscle contraction. Finally, similar associations between CHI3L1 and metabolic parameters were observed in humans alongside genotype associations between CHI3L1 and diabetes at the population level. CHI3L1 may be a potential therapeutic target for the treatment of diabetes.

ATP synthase inhibitory factor 1 (IF1), a novel myokine, regulates glucose metabolism by AMPK and Akt dual pathways.

ATPase inhibitory factor 1 (IF1) is an ATP synthase-interacting protein that suppresses the hydrolysis activity of ATP synthase. In this study, we observed that the expression of IF1 was up-regulated in response to electrical pulse stimulation of skeletal muscle cells and in exercized mice and healthy men. IF1 stimulates glucose uptake via AMPK in skeletal muscle cells and primary cultured myoblasts. Reactive oxygen species and Rac family small GTPase 1 (Rac1) function in the upstream and downstream of AMPK, respectively, in IF1-mediated glucose uptake. In diabetic animal models, the administration of recombinant IF1 improved glucose tolerance and down-regulated blood glucose level. In addition, IF1 inhibits ATP hydrolysis by ß-F1-ATPase in plasma membrane, thereby increasing extracellular ATP and activating the protein kinase B (Akt) pathway, ultimately leading to glucose uptake. Thus, we suggest that IF1 is a novel myokine and propose a mechanism by which AMPK and Akt contribute independently to IF1-mediated improvement of glucose tolerance impairment. These results demonstrate the importance of IF1 as a potential antidiabetic agent.-Lee, H. J., Moon, J., Chung, I., Chung, J. H., Park, C., Lee, J. O., Han, J. A., Kang, M. J., Yoo, E. H., Kwak, S.-Y., Jo, G., Park, W., Park, J., Kim, K. M., Lim, S., Ngoei, K. R. W., Ling, N. X. Y., Oakhill, J. S., Galic, S., Murray-Segal, L., Kemp, B. E., Mantzoros, C. S., Krauss, R. M., Shin, M.-J., Kim, H. S. ATP synthase inhibitory factor 1 (IF1), a novel myokine, regulates glucose metabolism by AMPK and Akt dual pathways.

Mitochondrial Dynamics in Placenta-Derived Mesenchymal Stem Cells Regulate the Invasion Activity of Trophoblast.

Mitochondrial dynamics are involved in many cellular events, including the proliferation, differentiation, and invasion/migration of normal as well as cancerous cells. Human placenta-derived mesenchymal stem cells (PD-MSCs) were known to regulate the invasion activity of trophoblasts. However, the effects of PD-MSCs on mitochondrial function in trophoblasts are still insufficiently understood. Therefore, the objectives of this study are to analyze the factors related to mitochondrial function and investigate the correlation between trophoblast invasion and mitophagy via PD-MSC cocultivation. We assess invasion ability and mitochondrial function in invasive trophoblasts according to PD-MSC cocultivation by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and extracellular flux (XF) assay. Under PD-MSCs co-cultivation, invasion activity of a trophoblast is increased via activation of the Rho signaling pathway as well as Matrix metalloproteinases (MMPs). Additionally, the expression of mitochondrial function (e.g., reactive oxygen species (ROS), calcium, and adenosine triphosphate (ATP) synthesis) in trophoblasts are increased via PD-MSCs co-cultivation. Finally, PD-MSCs regulate mitochondrial autophagy factors in invasive trophoblasts via regulating the balance between PTEN-induced putative kinase 1 (PINK1) and parkin RBR E3 ubiquitin protein ligase (PARKIN) expression. Taken together, these results demonstrate that PD-MSCs enhance the invasion ability of trophoblasts via altering mitochondrial dynamics. These results support the fundamental mechanism of trophoblast invasion via mitochondrial function and provide a new stem cell therapy for infertility.

Metformin overcomes resistance to cisplatin in triple-negative breast cancer (TNBC) cells by targeting RAD51.

BACKGROUND: Chemotherapy is a standard therapeutic regimen to treat triple-negative breast cancer (TNBC); however, chemotherapy alone does not result in significant improvement and often leads to drug resistance in patients. In contrast, combination therapy has proven to be an effective strategy for TNBC treatment. Whether metformin enhances the anticancer effects of cisplatin and prevents cisplatin resistance in TNBC cells has not been reported. METHODS: Cell viability, wounding healing, and invasion assays were performed on Hs 578T and MDA-MB-231 human TNBC cell lines to demonstrate the anticancer effects of combined cisplatin and metformin treatment compared to treatment with cisplatin alone. Western blotting and immunofluorescence were used to determine the expression of RAD51 and gamma-H2AX. In an in vivo 4T1 murine breast cancer model, a synergistic anticancer effect of metformin and cisplatin was observed. RESULTS: Cisplatin combined with metformin decreased cell viability and metastatic effect more than cisplatin alone. Metformin suppressed cisplatin-mediated RAD51 upregulation by decreasing RAD51 protein stability and increasing its ubiquitination. In contrast, cisplatin increased RAD51 expression in an ERK-dependent manner. In addition, metformin also increased cisplatin-induced phosphorylation of γ-H2AX. Overexpression of RAD51 blocked the metformin-induced inhibition of cell migration and invasion, while RAD51 knockdown enhanced cisplatin activity. Moreover, the combination of metformin and cisplatin exhibited a synergistic anticancer effect in an orthotopic murine model of 4T1 breast cancer in vivo. CONCLUSIONS: Metformin enhances anticancer effect of cisplatin by downregulating RAD51 expression, which represents a novel therapeutic target in TNBC management.

Intermittent restraint-induced sympathetic activation attenuates hepatic steatosis and inflammation in a high-fat diet-fed mouse model.

Nonalcoholic fatty liver disease (NAFLD) is very prevalent worldwide and is associated with insulin resistance and metabolic syndrome. Stress is a physiological and biological response to maintain homeostasis of the body against stressors while severe stress response is an important contributor to various illnesses, including metabolic syndrome and brain disorders. We have evaluated the effects of intermittent restraint stress on NAFLD in a high-fat diet (HFD)-fed mouse model. C57/BL6 mice had free access to a 60% HFD for 8 wk, with or without intermittent restraint stress (3 h) conducted three times a week. HFD administration increased fat accumulation in liver tissues. Unlike the stressed standard diet group, the levels of hepatic total cholesterol and triglycerides were significantly ameliorated in the HFD with stress group compared with the HFD alone group. These beneficial results were in accordance with serum levels of liver enzymes (aspartate transaminase, alanine transaminase) and hepatic levels of TNF-α and oxidative stress parameters (reactive oxygen species, nitric oxide, and malondialdehyde). The intermittent restraint stress significantly attenuated the HFD-derived alterations in serum insulin levels, hepatic protein kinase B activity, and gene expression, especially related to lipogenesis. This intermittent restraint stress also elevated the serum epinephrine concentration and activated the adrenergic receptor ß2 or ß3 in livers or white adipose tissue (WAT). Activation of energy expenditure markers (uncoupling protein 1, peroxisome proliferator-activated receptor-γ coactivator-1α) in brown adipose tissue and the browning of WAT were also observed in the HFD with stress group. Taken together, our findings showed the beneficial effects of sympathetic activation by intermittent restraint stress on HFD-induced hepatic steatosis and partial inflammation.NEW & NOTEWORTHY In modern society, stress is a part of daily life, and a certain level of stress is inevitable to most of the general population. Uncontrolled severe stress is obviously harmful; however, certain kind/level of stress could be beneficial on lipid metabolism via sympathetic activation. Our data suggest that a sympathetic activation by intermittent restraint stress could play a positive role in maintaining the balance of hepatic lipid metabolism, especially under high-fat diet conditions.

LIF, a Novel Myokine, Protects Against Amyloid-Beta-Induced Neurotoxicity via Akt-Mediated Autophagy Signaling in Hippocampal Cells.

BACKGROUND: Leukemia inhibitory factor, a novel myokine, is known to be associated with neural function, but the underlying molecular mechanism remains unclear. METHODS: HT-22 mouse hippocampal cells, primary hippocampal cells, and Drosophila Alzheimer's disease model were used to determine the effect of leukemia inhibitory factor on neurons. Immunoblot analysis and immunofluorescence method were used to analyze biological mechanism. RESULTS: Leukemia inhibitory factor increased Akt phosphorylation in a phosphoinositide-3-kinase-dependent manner in hippocampal cells. Leukemia inhibitory factor also increased the phosphorylation of the mammalian target of rapamycin and the downstream S6K. Leukemia inhibitory factor stimulated the phosphorylation of signal transducer and activator of transcription via extracellular signal-regulated kinases. Leukemia inhibitory factor increased c-fos expression through both Akt and extracellular signal-regulated kinases. Leukemia inhibitory factor blocked amyloid ß-induced neural viability suppression and inhibited amyloid ß-induced glucose uptake impairment through the block of amyloid ß-mediated insulin receptor downregulation. Leukemia inhibitory factor blocked amyloid ß-mediated induction of the autophagy marker, microtubule-associated protein 1A/1B-light chain 3. Additionally, in primary prepared hippocampal cells, leukemia inhibitory factor stimulated Akt and extracellular signal-regulated kinase, demonstrating that leukemia inhibitory factor has physiological relevance in vivo. Suppression of the autophagy marker, light chain 3II, by leukemia inhibitory factor was observed in a Drosophila model of Alzheimer's disease. CONCLUSIONS: These results demonstrate that leukemia inhibitory factor protects against amyloid ß-induced neurotoxicity via Akt/extracellular signal-regulated kinase-mediated c-fos induction, and thus suggest that leukemia inhibitory factor is a potential drug for Alzheimer's disease.

Endogenous Ligand for GPR120, Docosahexaenoic Acid, Exerts Benign Metabolic Effects on the Skeletal Muscles via AMP-activated Protein Kinase Pathway.

Docosahexaenoic acid (DHA) is an endogenous ligand of G protein-coupled receptor 120 (GPR120). However, the mechanisms underlying DHA action are poorly understood. In this study, DHA stimulated glucose uptake in the skeletal muscles in an AMP-activated protein kinase (AMPK)-dependent manner. GPR120-mediated increase in intracellular Ca(2+) was critical for DHA-mediated AMPK phosphorylation and glucose uptake. In addition, DHA stimulated GLUT4 translocation AMPK-dependently. Inhibition of AMPK and Ca(2+)/calmodulin-dependent protein kinase kinase blocked DHA-induced glucose uptake. DHA and GW9508, a GPR120 agonist, increased GPR120 expression. DHA-mediated glucose uptake was not observed in GPR120 knockdown conditions. DHA increased AMPK phosphorylation, glucose uptake, and intracellular Ca(2+) concentration in primary cultured myoblasts. Taken together, these results indicated that the beneficial metabolic role of DHA was attributed to its ability to regulate glucose via the GPR120-mediated AMPK pathway in the skeletal muscles.

Dehydrozingerone exerts beneficial metabolic effects in high-fat diet-induced obese mice via AMPK activation in skeletal muscle.

Dehydrozingerone (DHZ) exerts beneficial effects on human health; however, its mechanism of action remains unclear. Here, we found that DHZ suppressed high-fat diet-induced weight gain, lipid accumulation and hyperglycaemia in C57BL/6 mice and increased AMP-activated protein kinase (AMPK) phosphorylation and stimulated glucose uptake in C2C12 skeletal muscle cells. DHZ activated p38 mitogen-activated protein kinase (MAPK) signalling in an AMPK-dependent manner. Inhibiting AMPK or p38 MAPK blocked DHZ-induced glucose uptake. DHZ increased GLUT4 (major transporter for glucose uptake) expression in skeletal muscle. Glucose clearance and insulin-induced glucose uptake increased in DHZ-fed animals, suggesting that DHZ increases systemic insulin sensitivity in vivo. Thus, the beneficial health effects of DHZ could possibly be explained by its ability to activate the AMPK pathway in skeletal muscle.

[6]-Gingerol Affects Glucose Metabolism by Dual Regulation via the AMPKα2-Mediated AS160-Rab5 Pathway and AMPK-Mediated Insulin Sensitizing Effects.

[6]-Gingerol has been used to control diabetes and dyslipidemia; however, its metabolic role is poorly understood. In this study, [6]-gingerol increased adenosine monophosphate (AMP)-activated protein kinase (AMPK) phosphorylation in mouse skeletal muscle C2C12 cells. Stimulation of glucose uptake by [6]-gingerol was dependent on AMPKα2. Moreover, both Inhibition and knockdown of AMPKα2 blocked [6]-gingerol-induced glucose uptake. [6]-Gingerol significantly decreased the activity of protein phosphatase 2A (PP2A). Inhibition of PP2A activity with okadaic acid enhanced the phosphorylation of AMPKα2. Moreover, the interaction between AMPKα2 and PP2A was increased by [6]-gingerol, suggesting that PP2A mediates the effect of [6]-gingerol on AMPK phosphorylation. In addition, [6]-gingerol increased the phosphorylation of Akt-substrate 160 (AS160), which is a Rab GTPase-activating protein. Inhibition of AMPKα2 blocked [6]-gingerol-induced AS160 phosphorylation. [6]-gingerol increased the Rab5, and AMPKα2 knockdown blocked [6]-gingerol-induced expression of Rab5, indicating AMPK play as an upstream of Rab5. It also increased glucose transporter 4 (GLUT4) mRNA and protein expression and stimulated GLUT4 translocation. Furthermore, insulin-mediated glucose uptake and Akt phosphorylation were further potentiated by [6]-gingerol treatment. This potentiation was not observed in the presence of AMPK inhibitor compound C. In summary, our results suggest that [6]-gingerol plays an important role in glucose metabolism via the AMPKα2-mediated AS160-Rab5 pathway and through potentiation of insulin-mediated glucose regulation.

E3 ubiquitin ligase, WWP1, interacts with AMPKα2 and down-regulates its expression in skeletal muscle C2C12 cells.

It is known that the activity of AMP-activated protein kinase (AMPKα2) was depressed under high glucose conditions. However, whether protein expression of AMPKα2 is also down-regulated or not remains unclear. In this study, we showed that the expression of AMPKα2 was down-regulated in cells cultured under high glucose conditions. Treatment of proteasome inhibitor, MG132, blocked high glucose-induced AMPKα2 down-regulation. Endogenous AMPKα2 ubiquitination was detected by immunoprecipitation of AMPKα2 followed by immunoblotting detection of ubiquitin. The yeast-two hybrid (YTH) approach identified WWP1, an E3 ubiquitin ligase, as the AMPKα2-interacting protein in skeletal muscle cells. Interaction between AMPKα2 and WWP1 was validated by co-immunoprecipitation. Knockdown of WWP1 blocked high glucose-induced AMPKα2 down-regulation. The overexpression of WWP1 down-regulated AMPKα2. In addition, the expression of WWP1 is increased under high glucose culture conditions in both mRNA and protein levels. The level of AMPKα2 was down-regulated in the quadriceps muscle of diabetic animal model db/db mice. Expression of WWP1 blocked metformin-induced glucose uptake. Taken together, our results demonstrated that WWP1 down-regulated AMPKα2 under high glucose culture conditions via the ubiquitin-proteasome pathway.

Alcohol induces cell proliferation via hypermethylation of ADHFE1 in colorectal cancer cells.

BACKGROUND: The hypermethylation of Alcohol dehydrogenase iron containing 1 (ADHFE1) was recently reported to be associated with colorectal cancer (CRC) differentiation. However, the effect of alcohol on ADHFE1 hypermethylation in CRC is still unclear. METHODS: The methylation status and expression levels of ADHFE1 were investigated in primary tumor tissues and adjacent normal tissues of 73 patients with CRC, one normal colon cell line, and 4 CRC cell lines (HT-29, SW480, DLD-1, and LoVo) by quantitative methylation-specific polymerase chain reaction (QMSP) and real-time reverse transcription polymerase chain reaction (real time PCR), respectively. The effect of alcohol on the methylation status of ADHFE1 was analyzed in HT-29, SW480, DLD-1, and CCD18Co cells using QMSP, real-time PCR, immunoblot, and cell proliferation assay. RESULTS: ADHFE1 was hypermethylated in 69 of 73 CRC tissues (95%) compared to adjacent normal tissues (p<0.05). The mRNA expression of ADHFE1 was significantly reduced in CRC compared to adjacent normal tissues (p<0.05) and its expression was decreased in the alcohol consumption group (p<0.05). ADHFE1 was hypermethylated and its expression was decreased in 4 CRC cell lines compared with normal colon cell line. Alcohol induced hypermethylation of ADHFE1, decreased its expression, and stimulated cell proliferation of HT-29, SW480, and DLD-1cells. CONCLUSION: These results demonstrate that the promoter hypermethylation of ADHFE1 is frequently present in CRC and alcohol induces methylation-mediated down expression of ADHFE1 and proliferation of CRC cells.

Rhus verniciflua extract modulates survival of MCF-7 breast cancer cells through the modulation of AMPK-pathway.

Rhus verniciflua STOKES (RVS) is used as an anti-cancer agent in traditional herbal medicine. However, the underlying molecular mechanism of its action is poorly understood. Here, we elucidated the mechanism of the anti-cancer mechanism of RVS in MCF-7 human breast cancer cells. We found that RVS increased the phosphorylation of AMP-activated protein kinase (AMPK) and downstream acetyl-CoA carboxylase (ACC) and suppressed cell viability in an AMPK-dependent fashion. RVS also induced an increase in reactive oxygen species (ROS) levels. RVS-induced AMPK phosphorylation was not observed in the presence of N-acetyl-cysteine (NAC), which indicated that ROS is associated with RVS-induced AMPK phosphorylation. In addition, fluorescent staining (Annexin V/propidium iodide) revealed that RVS increased the expression of Annexin V, which indicates that RVS leads to cancer-induced apoptosis. Moreover, RVS increased the phosphorylation of p53 and the expression of Bax. The inhibition of AMPK blocked RVS-induced p53 phosphorylation and Bax expression, which suggests that AMPK is involved in RVS-induced cancer apoptosis. Taken together, these results demonstrate that RVS has anti-tumor effects on MCF-7 cells through an AMPK-signaling pathway.