Monophosphate-activated Protein Kinase-α Research Articles

Hypothalamic regulation of energy homoeostasis when consuming diets of different energy concentrations: comparison between Tibetan and Small-tailed Han sheep.

Seasonal energy intake of Tibetan sheep on the harsh Qinghai-Tibetan Plateau (QTP) fluctuates greatly and is often well below maintenance requirements. The aim of this study was to gain insight into how the hypothalamus regulates energy homoeostasis in Tibetan sheep. We compared Tibetan and Small-tailed Han sheep (n 24 of each breed), which were each allocated randomly into four groups and offered one of four diets that differed in digestible energy densities: 8·21, 9·33, 10·45 and 11·57 MJ/kg DM. Sheep were weighed every 2 weeks, and it was assumed that the change in body weight (BW) reflected the change in energy balance. The arcuate nucleus of the hypothalamus in Tibetan sheep had greater protein expressions of neuropeptide Y (NPY) and agouti-related peptide (AgRP) when in negative energy balance, but lesser protein expressions of proopiomelanocortin (POMC) and cocaine and amphetamine-regulated transcript (CART) when in positive energy balance than Small-tailed Han sheep. As a result, Tibetan sheep had a lesser BW loss when in negative energy balance and stored more energy and gained more BW when in positive energy balance than Small-tailed Han sheep with the same dietary intake. Moreover, in the hypothalamic adenosine monophosphate-activated protein kinase (AMPK) regulation pathway, Tibetan sheep had greater adenosine monophosphate-activated protein kinase-α 2 protein expression than Small-tailed Han sheep, which supported the premise of a better ability to regulate energy homoeostasis and better growth performance. These differences in the hypothalamic NPY/AgRP, POMC/CART and AMPK pathways between breeds conferred an advantage to the Tibetan over Small-tailed Han sheep to cope with low energy intake on the harsh QTP.

Empagliflozin and Liraglutide Differentially Modulate Cardiac Metabolism in Diabetic Cardiomyopathy in Rats.

Glucagon-like peptide 1 receptor agonists (GLP-1RAs) and sodium-glucose cotransporter-2 inhibitors (SGLT2is) are antihyperglycemic agents with cardioprotective properties against diabetic cardiomyopathy (DCM). However, the distinctive mechanisms underlying GLP-1RAs and SGLT2is in DCM are not fully elucidated. The purpose of this study was to investigate the impacts of GLP1RAs and/or SGLT2is on myocardial energy metabolism, cardiac function, and apoptosis signaling in DCM. Biochemistry and echocardiograms were studied before and after treatment with empagliflozin (10 mg/kg/day, oral gavage), and/or liraglutide (200 μg/kg every 12 h, subcutaneously) for 4 weeks in male Wistar rats with streptozotocin (65 mg/kg intraperitoneally)-induced diabetes. Cardiac fibrosis, apoptosis, and protein expression of metabolic and inflammatory signaling molecules were evaluated by histopathology and Western blotting in ventricular cardiomyocytes of different groups. Empagliflozin and liraglutide normalized myocardial dysfunction in diabetic rats. Upregulation of phosphorylated-acetyl coenzyme A carboxylase, carnitine palmitoyltransferase 1β, cluster of differentiation 36, and peroxisome proliferator-activated receptor-gamma coactivator, and downregulation of glucose transporter 4, the ratio of phosphorylated adenosine monophosphate-activated protein kinase α2 to adenosine monophosphate-activated protein kinase α2, and the ratio of phosphorylated protein kinase B to protein kinase B in diabetic cardiomyocytes were restored by treatment with empagliflozin or liraglutide. Nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3, interleukin-1β, tumor necrosis factor-α, and cleaved caspase-1 were significantly downregulated in empagliflozin-treated and liraglutide-treated diabetic rats. Both empagliflozin-treated and liraglutide-treated diabetic rats exhibited attenuated myocardial fibrosis and apoptosis. Empagliflozin modulated fatty acid and glucose metabolism, while liraglutide regulated inflammation and apoptosis in DCM. The better effects of combined treatment with GLP-1RAs and SGLT2is may lead to a potential strategy targeting DCM.

Effect of 4-week electroacupuncture intervention on "Browning" of white fat in rats and its mechanism

Objective: To investigate the effects of 4-week electroacupuncture intervention on "Browning" of white fat in rats, and to explore its molecular mechanisms. Methods: Twenty-four 8-week-old male SD rats were randomly divided into sedentary group (Sed), aerobic exercise group (Exe) and electroacupuncture group (ElA), 8 rats in each group. Exe group used 65% Max oxygen uptake intensity treadmill exercise, 1 h/d,6 d/w, while the ElA group used electric acupuncture to stimulate "zusanli" and "tianshu" points, 20 min/d,6 d/w, and the weight of rats was recorded every week. After 4 weeks of intervention, blood samples were collected from the apex and abdominal aorta. The wet weight of scapular fat and perirenal fat of rats was detected and the body fat and the serum levels of Irisin were determined. What's more, the expressions of adenosine 5'-monophosphate activated protein kinase-α (AMPKα), phosphorylation of adenosine 5'-monophosphate activated protein kinase-α (p-AMPKα), peroxisome proliferator activated receptor γ coactivator-1α (PGC-1α), fibronectin type III domain containing 5 (FNDC5) and uncoupling protein (UCP1) and brown adipose tissue lipid droplets in rats were detected. Results: Compared with the Sed group, the weight gain of rats in the Exe and ElA groups was decreased significantly from the second week. After 4 weeks, the body weight and body fat of the Exe and ElA groups were decreased significantly (P＜0.01), while there was no significant difference between the Exe and ElA groups (P＞0.05). ②After 4 weeks of intervention, the white fat wet weight of ElA group and Exe group was significantly was reduced (P＜0.05 or P＜0.01). ③The serum levels of Irisin in Exe and ElA groups were significantly increased (P＜0.05), and the expressions of p-AMPKα/AMPKα, PGC-1α, FNDC5 in the gastrocnemius and the expressions of UCP1 in white fat and brown adipose tissue of Exe and ElA groups were increased (P＜0.05). ④The area of lipid droplets in white and brown adipose tissue of Exe and ElA groups was significantly reduced (P＜ 0.01). Conclusion: 4 weeks of electroacupuncture intervention can effectively control the weight of rats and induce "browning" of white fat, and its effect was similar to aerobic exercise, which may be release Irisin through the AMPKα-PGC-1α-FNDC5-Irisin signaling pathway, and then "crosstalk" with adipose tissue, up-regulate the expression of UCP1, and induce "browning" of white adipose tissue.

Vanillic Acid Alleviates Acute Myocardial Hypoxia/Reoxygenation Injury by Inhibiting Oxidative Stress.

Oxidative stress is an important factor of myocardial hypoxia/reoxygenation (H/R) injury. Our research focuses on how to reduce the cardiac toxicity caused by oxidative stress through natural plant extracts. Vanillic acid (VA) is a phenolic compound found in edible plants and rich in the roots of Angelica sinensis. Experimental studies have provided evidence for this compound's effectiveness in cardiovascular diseases; however, its mechanism is still unclear. In this study, molecular mechanisms related to the protective effects of VA were investigated in H9c2 cells in the context of H/R injury. The results showed that pretreatment with VA significantly increased cell viability and decreased the percentage of apoptotic cells, as well as lactate dehydrogenase and creatine phosphokinase activity, in the supernatant, accompanied by reduced levels of reactive oxygen species and reduced caspase-3 activity. VA pretreatment also restored mitochondrial membrane potentials. Moreover, preincubation with VA significantly attenuated mitochondrial permeability transition pore activity. VA administration upregulated adenosine monophosphate-activated protein kinase α2 (AMPKα2) protein expression, and interestingly, pretreatment with AMPKα2-siRNA lentivirus effectively attenuated the cardioprotective effects of VA in response to H/R injury.

Gaudichaudione H Inhibits Inflammatory Responses in Macrophages and Dextran Sodium Sulfate-Induced Colitis in Mice.

Macrophages-involved inflammation is considered to induce the damage in various diseases. Herein, novel therapeutics inhibiting over-activation of macrophages could prove an effective strategy to prevent inflammation-related diseases. Gaudichaudione H (GH), which is a natural small molecular compound isolated from Garcinia oligantha Merr. (Clusiaceae) has previously been demonstrated its anti-cancer effects on several cancer cell lines. However, no report has been published about the anti-inflammatory effect of GH to date. This study aims to examine the anti-inflammatory effects and potential molecular mechanism of GH, and provide new insights toward the treatment of inflammation. GH inhibited nitric oxide (NO) production, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression, cytokine interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) production, and messenger RNA (mRNA) expression to attenuate inflammatory responses in lipopolysaccharide (LPS)-induced RAW 264.7 cells or stimulated bone marrow-derived macrophages (BMDMs). GH inhibited nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathways, the nuclear translocation of transcription factors NF-κB and activator protein 1 (AP-1), as well as upstream signaling of the toll-like receptor 4 (TLR4)-myeloid differentiation primary response 88 (MyD88) pathway in stimulated macrophages. Furthermore, the result of the intracellular signaling array showed that the phosphorylation of adenosine 5'-monophosphate-activated protein kinase-α (AMPKα), proline-rich Akt substrate of 40 kDa (PRAS40), and p38 could be down regulated by GH in BMDMs, indicating that the mechanism by which GH inhibited inflammation may be also associated with the energy metabolism pathway, PRAS40-mediated NF-κB pathway, cell proliferation, apoptosis, and autophagy, etc. In addition, GH alleviated dextran sodium sulfate (DSS)-induced colitis in mice by ameliorating weight loss, stool consistency change, blood in the stool, and colon shortening. GH decreased the protein and mRNA levels of IL-6 and TNF-α, iNOS and COX-2 mRNA expression, the activation of NF-κB and MAPK pathways, the phosphorylation of AMPKα and PRAS40, histological damage, and infiltration of macrophages in the colons of mice with DSS-induced colitis. Taken together, our results support that GH exerts the anti-inflammatory effects in macrophages in vitro through regulation of NF-κB and MAPK pathways, and DSS-induced colitis mouse model in vivo. These findings suggest that GH may be a promising candidate in treating macrophage-related inflammatory disease.

Testosterone Increases the Expression and Phosphorylation of AMP Kinase α in Men With Hypogonadism and Type 2 Diabetes.

Adenosine 5'-monophosphate-activated protein kinase-α (AMPKα) is a mediator of exercise-induced glucose uptake in skeletal muscle. We evaluated whether AMPKα expression and phosphorylation are reduced in skeletal muscle and adipose tissue of patients with hypogonadotropic hypogonadism (HH), and whether testosterone replacement therapy results in restoration of the expression and phosphorylation of AMPKα. This is a secondary analysis of a previously completed trial that showed an insulin-sensitizing effect of testosterone therapy in men with type 2 diabetes and HH. Clinical research center at university. Thirty-two men with HH and 32 eugonadal men were compared at baseline. Men with HH were treated with intramuscular injections of testosterone or placebo every 2 weeks for 22 weeks. Quadriceps muscle biopsies and subcutaneous abdominal fat biopsies were obtained before and after 4-hour euglycemic hyperinsulinemic clamp, prior to and after testosterone or placebo therapy. mRNA expression of AMPKα in hypogonadal men was lower by 37% in adipose tissue and 29% in skeletal muscle, respectively, compared with levels in eugonadal men, while phosphorylated AMPKα was lower by 22% and 28%, respectively. Following testosterone replacement, the expression of AMPKα did not alter in the fasting state but increased markedly by 41% and 46% in adipose tissue and muscle, respectively, after the clamp. In contrast, phosphorylated AMPKα increased by 69% in muscle after testosterone therapy but did not change following the clamp. Testosterone modulates the expression of AMPKα and phosphorylated AMPKα. These effects may contribute to the improved insulin sensitivity following testosterone therapy.

Liraglutide improves insulin sensitivity in high fat diet induced diabetic mice through multiple pathways

Glucagon like peptide-1 (GLP-1) promotes postprandial insulin secretion. Liraglutide, a full agonist of the GLP-1 receptor, reduces body weight, improve insulin sensitivity, and alleviate Non Alcoholic Fatty Liver Disease (NAFLD). However, the underlying mechanisms remain unclear. This study aims to explore the underlying mechanisms and cell signaling pathways involved in the anti-obesity and anti-inflammatory effects of liraglutide.Mice were fed a high fat high sucrose diet to induce diabetes, diabetic mice were divided into two groups and injected with liraglutide or vehicle for 14 days. Liraglutide treatment improved insulin sensitivity, accompanied with reduced expression of the phosphorylated Acetyl-CoA carboxylase-2 (ACC2) and upregulation of long chain acyl CoA dehydrogenase (LCAD) in insulin sensitive tissues. Furthermore, liraglutide induced adenosine monophosphate-activated protein kinase-α (AMPK-α) and Sirtuin-1(Sirt-1) protein expression in liver and perigonadal fat. Liraglutide induced elevation of fatty acid oxidation in these tissues may be mediated through the AMPK-Sirt-1 cell signaling pathway. In addition, liraglutide induced brown adipocyte differentiation in skeletal muscle, including induction of uncoupling protein-1 (UCP-1) and PR-domain-containing-16 (PRDM-16) protein in association with induction of SIRT-1. Importantly, liraglutide displayed anti-inflammation effect. Specifically, liraglutide led to a significant reduction in circulating interleukin-1 β (IL-1 β) and interleukin-6 (IL-6) as well as hepatic IL-1 β and IL-6 content. The expression of inducible nitric oxide synthase (iNOS-1) and cyclooxygenase-2 (COX-2) in insulin sensitive tissues was also reduced following liraglutide treatment. In conclusion, liraglutide improves insulin sensitivity through multiple pathways resulting in reduction of inflammation, elevation of fatty acid oxidation, and induction of adaptive thermogenesis.

Protective Effects of Combination of Radix Astragali and Radix Salviae Miltiorrhizae on Kidney of Spontaneously Hypertensive Rats and Renal Intrinsic Cells.

To evaluate the effects of combination of Radix Astragali (RA) and Radix Salviae Miltiorrhizae (RS) on kidney of spontaneously hypertensive rats (SHRs) and renal intrinsic cells. SHRs were intragastrically administrated with RA (5.09 g/kg) and RS (2.55 g/kg) either alone or with combination for 4 weeks; valsartan (13.35 mg/kg) was used as a positive control. Blood pressure and renal ultrasonography were monitored periodically. The biomarkers [microalbumin (mALB), cystatin ^C, angiotensin II (Ang II), interleukin-1 beta (IL-1β), and β2-microglobulin (β2-Mg), etc.] in serum and urine were measured by enzyme-linked immunosorbent assay (ELISA). The protein expressions [phosphorylated adenosine 5'-monophosphate-activated protein kinase-α1 (p-AMPKα1), sestrin-β, calcium/calmodulin-dependent protein kinase kinase-β (CaMKK-β), phosphoinositide 3-kinases (PI3K), serine-threonine protein kinase 1 (AKT1), and vascular endothelial growth factor receptor 2 (VEGFR2)] in renal cortex were determined by Western blot. In vitro, the hypertensive cellular model was established by applying 2×10-6 mol/L Ang ^II. The primary human podocytes, human glomerular endothelial cells (HRGECs), and human proximal tubular epithelial cells (HK-2s) were pre-incubated with sulfotanshinone sodium (Tan, 10 μg/mL) and/or calycosin-7-O-β-D-glucoside (Cal, 5 μg/mL). The cellular viability and apoptosis were assayed by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) and Annexin V/PI staining, respectively. The level of endothelial nitric oxide synthase (eNOS) in culture supernatant was determined by ELISA. RA+RS signifificantly decreased the diastolic blood pressure, renal vascular resistance index, and parenchymal thickness, increased 24 h urinary volume as well as lowered the levels of urine mALB and serum cystatin ^C, IL-1β and β2-Mg of SHRs (P <0.05 vs. SHRs). The decreased protein levels of p-AMPKα1, sestrinβ and CaMKK-β and the increased protein levels of PI3K, AKT1 and VEGFR2 in renal cortex of SHRs were normalized after RA+RS treatment (P <0.05). In vitro, Tan and Cal attenuated the Ang II-induced abnormal proliferation and increased the apoptosis of HRGECs and HK-2s and improved the level of eNOS in culture supernatant. Whereas, neither of them showed powerful effect on podocyte. The combination of RA and RS had potential effects on alleviating the renal damages of SHRs and the renoprotection was independent of blood pressure level.

Chromium acetate stimulates adipogenesis through regulation of gene expression and phosphorylation of adenosine monophosphate-activated protein kinase in bovine intramuscular or subcutaneous adipocytes

ObjectiveWe hypothesized that Cr source can alter adipogenic-related transcriptional regulations and cell signaling. Therefore, the objective of the study was to evaluate the biological effects of chromium acetate (CrAc) on bovine intramuscular (IM) and subcutaneous (SC) adipose cells.MethodsBovine preadipocytes isolated from two different adipose tissue depots; IM and SC were used to evaluate the effect of CrAc treatment during differentiation on adipogenic gene expression. Adipocytes were incubated with various doses of CrAc: 0 (differentiation media only, control), 0.1, 1, and 10 μM. Cells were harvested and then analyzed by real-time quantitative polymerase chain reaction in order to measure the quantity of adenosine monophosphate-activated protein kinase-α (AMPK-α), CCAAT enhancer binding protein-β (C/EBPβ), G protein-coupled receptor 41 (GPR41), GPR43, peroxisome proliferator-activated receptor-γ (PPARγ), and stearoyl CoA desaturase (SCD) mRNA relative to ribosomal protein subunit 9 (RPS9). The ratio of phosphorylated-AMPK (pAMPK) to AMPK was determined using a western blot technique in order to determine changing concentration.ResultsThe high dose (10 μM) of CrAc increased C/EBPβ, in both IM (p = 0.02) and SC (p = 0.02). Expression of PPARγ was upregulated by 10 μM of CrAc in IM but not in SC. Expression of SCD was also increased in both IM and SC with 10 μM of CrAc treatment. Addition of CrAc did not alter gene expression of glucose transporter 4, GPR41, or GPR43 in both IM and SC adipocytes. Addition of CrAc, resulted in a decreased pAMPKα to AMPKα ration (p<0.01) in IM.ConclusionThese data may indicate that Cr source may influence lipid filling in IM adipocytes via inhibitory action of AMPK phosphorylation and upregulating expression of adipogenic genes.

Effect of Marine Microalga Chlorella pyrenoidosa Ethanol Extract on Lipid Metabolism and Gut Microbiota Composition in High-Fat Diet-Fed Rats.

Effects of marine microalga Chlorella pyrenoidosa 55% ethanol extract (CPE55) on lipid metabolism, gut microbiota and regulation mechanism in high fat diet-fed induced hyperlipidaemia rats were investigated. Structure characterizations of major compounds in CPE55 were determined by ultra-performance liquid chromatography-quadrupole/time of flight mass spectrometry (UPLC-Q-TOF-MS/MS). The compositions of gut microbiota in rats were analyzed by high-throughput next-generation 16S rRNA gene sequencing. Oral administration with CPE55 markedly alleviated dyslipidemia through improving adverse blood lipid profile and inhibiting hepatic lipid accumulation and steatosis. CPE55 has downregulated the gene expression levels of acetyl CoA carboxylase, sterol regulatory element-binding transcription factor-1c, and 3-hydroxy-3-methyl glutaryl coenzyme A reductase and upregulated adenosine 5′-monophosphate-activated protein kinase-α. It has also improved the abundance of bacteria Alistipes, Prevotella, Alloprevotella, and Ruminococcus1 and decreased the abundances of Turicibacter and Lachnospira. Turicibacter and Lachnospira were both positive correlations of metabolic phenotypes. The findings above illustrated that CPE55 might be developed as food ingredients to ameliorate lipid metabolic disorders and hyperlipidaemia.

Regulatory Efficacy of the Polyunsaturated Fatty Acids from Microalgae Spirulina platensis on Lipid Metabolism and Gut Microbiota in High-Fat Diet Rats.

Ultra-high performance liquid chromatography coupled with photo-diode array detector and electrospray ionization-mass spectrometry was employed to analyze the major fatty acids in Spirulina platensis 95% ethanol extract (SPL95). The effects of SPL95 on hepatoprotection were evaluated, including liver tissue histopathology, liver, and serum biochemical analysis. The active principle of SPL95 revealed a hypolipidemic effect, as indicated by down-regulating the mRNA and protein levels of sterol regulatory element-binding transcription factor-1c, 3-hydroxy-3-methyl glutaryl coenzyme A reductase, acetyl CoA carboxylase pathway, and upregulating adenosine 5′-monophosphate-activated protein kinase-α in liver. SPL95 enriched the beneficial bacteria, including Prevotella, Alloprevotella, Porphyromonadaceae, Barnesiella, and Paraprevotella. Treatment with SPL95 led to a decrease in microbes, such as Turicibacter, Romboutsia, Phascolarctobacterium, Olsenella, and Clostridium XVIII, which were positively correlated with serum triglyceride, total cholesterol, and low-density-lipoprotein cholesterol levels, but negatively correlated with the serum high-density-lipoprotein cholesterol levels. These results provide evidence that the fatty acid from SPL95 may be used as a novel adjuvant therapy and functional food to regulate gut microbiota in obese and diabetic individuals.

Are the Effects of the Cholera Toxin and Isoproterenol on Human Keratinocytes' Proliferative Potential Dependent on Whether They Are Co-Cultured with Human or Murine Fibroblast Feeder Layers?

Human keratinocyte culture has provided the means to treat burns, wounds and skin pathologies. To date, to efficiently culture keratinocytes, cells are cultured on an irradiated feeder layer (iFL), either comprising human (iHFL) or murine (i3T3FL) fibroblasts, and the culture medium is supplemented with a cyclic adenosine monophosphate (cAMP) accumulation inducing agent such as isoproterenol (ISO) or cholera toxin (CT). Previous studies have characterized how the feeder layer type and the cAMP inducer type influence epithelial cells’ phenotype independently from one another, but it is still unknown if an optimal combination of feeder layer and cAMP inducer types exists. We used sophisticated statistical models to search for a synergetic effect of feeder layer and cAMP inducer types on human keratinocytes’ proliferative potential. Our data suggests that, when culturing human keratinocytes, using iHFL over i3T3FL increases population doublings and colony-forming efficiency through signaling pathways involving Ak mouse strain thymoma (Akt, also known as protein kinase B) isoforms 1 to 3, signal transducer and activator of transcription 5 (STAT5), p53, and adenosine monophosphate activated protein kinase α1 (AMPKα1). Both tested cAMP inducers ISO and CT yielded comparable outcomes. However, no significant synergy between feeder layer and cAMP inducer types was detected. We conclude that, to promote human keratinocyte growth in the early passages of culture, co-culturing them with a human feeder layer is preferable to a murine feeder layer.

Chronic Exercise Training Improved Aortic Endothelial and Mitochondrial Function via an AMPKα2-Dependent Manner.

Chronic exercise training is known to protect the vasculature; however, the underlying mechanisms remain obscure. The present study hypothesized that exercise may improve aortic endothelial and mitochondrial function through an adenosine monophosphate-activated protein kinase α2 (AMPKα2)-dependent manner. Ten-week-old AMPKα2 knockout (AMPKα2−/−) mice and age-matched wild-type (WT) mice were subjected to daily treadmill running for 6 weeks, and the thoracic aorta from these mice were used for further examination. Our results showed that exercise significantly promoted vasodilatation and increased expression and phosphorylation of endothelial nitric oxide synthase (eNOS), concomitant with increased AMPKα2 expression in WT mice. These effects were not observed in AMPKα2−/− mice. Furthermore, exercise training increased thoracic aortic mitochondrial content as indicated by increased Complex I and mitochondrial DNA (mtDNA) in WT mice but not in AMPKα2−/− mice. This may be caused by decreased mitochondrial autophagy since the expression of BH3 domain-containing BCL2 family members BNIP3-like (BNIP3L) and LC3B were decreased in WT mice with exercise. And these changes were absent with AMPKα2 deletion in mice. Importantly, exercise increased the expression of manganous superoxide dismutase (MnSOD) and catalase, suggesting that mitochondrial antioxidative capacity was increased. Notably, the improved antioxidative capacity was lost in AMPKα2−/− mice with exercise. In conclusion, this study illustrated that AMPKα2 plays a critical role in exercise-related vascular protection via increasing endothelial and mitochondrial function in the artery.

Postexercise High-Fat Feeding Suppresses p70S6K1 Activity in Human Skeletal Muscle.

This study aimed to examine the effects of reduced CHO but high postexercise fat availability on cell signaling and expression of genes with putative roles in regulation of mitochondrial biogenesis, lipid metabolism, and muscle protein synthesis. Ten males completed a twice per day exercise model (3.5 h between sessions) comprising morning high-intensity interval training (8 × 5 min at 85% V˙O2peak) and afternoon steady-state (SS) running (60 min at 70% V˙O2peak). In a repeated-measures design, runners exercised under different isoenergetic dietary conditions consisting of high-CHO (HCHO: 10 g·kg CHO, 2.5 g·kg protein, and 0.8 g·kg fat for the entire trial period) or reduced-CHO but high-fat availability in the postexercise recovery periods (HFAT: 2.5 g·kg CHO, 2.5 g·kg protein, and 3.5 g·kg fat for the entire trial period). Muscle glycogen was lower (P < 0.05) at 3 h (251 vs 301 mmol·kg dry weight) and 15 h (182 vs 312 mmol·kg dry weight) post-SS exercise in HFAT compared with HCHO. Adenosine monophosphate-activated protein kinase α2 activity was not increased post-SS in either condition (P = 0.41), although comparable increases (all P < 0.05) in PGC-1α, p53, citrate synthase, Tfam, peroxisome proliferator-activated receptor, and estrogen-related receptor α mRNA were observed in HCHO and HFAT. By contrast, PDK4 (P = 0.003), CD36 (P = 0.05), and carnitine palmitoyltransferase 1 (P = 0.03) mRNA were greater in HFAT in the recovery period from SS exercise compared with HCHO. Ribosomal protein S6 kinase activity was higher (P = 0.08) at 3 h post-SS exercise in HCHO versus HFAT (72.7 ± 51.9 vs 44.7 ± 27 fmol·min·mg). Postexercise high-fat feeding does not augment the mRNA expression of genes associated with regulatory roles in mitochondrial biogenesis, although it does increase lipid gene expression. However, postexercise ribosomal protein S6 kinase 1 activity is reduced under conditions of high-fat feeding, thus potentially impairing skeletal muscle remodeling processes.

Hypoxia, AMPK activation and uterine artery vasoreactivity.

Genes near adenosine monophosphate-activated protein kinase-α1 (PRKAA1) have been implicated in the greater uterine artery (UtA) blood flow and relative protection from fetal growth restriction seen in altitude-adapted Andean populations. Adenosine monophosphate-activated protein kinase (AMPK) activation vasodilates multiple vessels but whether AMPK is present in UtA or placental tissue and influences UtA vasoreactivity during normal or hypoxic pregnancy remains unknown. We studied isolated UtA and placenta from near-term C57BL/6J mice housed in normoxia (n = 8) or hypoxia (10% oxygen, n = 7-9) from day 14 to day 19, and placentas from non-labouring sea level (n = 3) or 3100 m (n = 3) women. Hypoxia increased AMPK immunostaining in near-term murine UtA and placental tissue. RT-PCR products for AMPK-α1 and -α2 isoforms and liver kinase B1 (LKB1; the upstream kinase activating AMPK) were present in murine and human placenta, and hypoxia increased LKB1 and AMPK-α1 and -α2 expression in the high- compared with low-altitude human placentas. Pharmacological AMPK activation by A769662 caused phenylephrine pre-constricted UtA from normoxic or hypoxic pregnant mice to dilate and this dilatation was partially reversed by the NOS inhibitor l-NAME. Hypoxic pregnancy sufficient to restrict fetal growth markedly augmented the UtA vasodilator effect of AMPK activation in opposition to PE constriction as the result of both NO-dependent and NO-independent mechanisms. We conclude that AMPK is activated during hypoxic pregnancy and that AMPK activation vasodilates the UtA, especially in hypoxic pregnancy. AMPK activation may be playing an adaptive role by limiting cellular energy depletion and helping to maintain utero-placental blood flow in hypoxic pregnancy.

Effects of paeonol on anti-neuroinflammatory responses in microglial cells.

Increasing studies suggest that inflammatory processes in the central nervous system mediated by microglial activation plays an important role in numerous neurodegenerative diseases. Development of planning for microglial suppression is considered a key strategy in the search for neuroprotection. Paeonol is a major phenolic component of Moutan Cortex, widely used as a nutrient supplement in Chinese medicine. In this study, we investigated the effects of paeonol on microglial cells stimulated by inflammagens. Paeonol significantly inhibited the release of nitric oxide (NO) and the expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Treatment with paeonol also reduced reactive oxygen species (ROS) production and inhibited an ATP-induced increased cell migratory activity. Furthermore, the inhibitory effects of neuroinflammation by paeonol were found to be regulated by phosphorylated adenosine monophosphate-activated protein kinase-α (AMPK-α) and glycogen synthase kinase 3 α/β (GSK 3α/β). Treatment with AMPK or GSK3 inhibitors reverse the inhibitory effect of neuroinflammation by paeonol in microglial cells. Furthermore, paeonol treatment also showed significant improvement in the rotarod performance and microglial activation in the mouse model as well. The present study is the first to report a novel inhibitory role of paeonol on neuroinflammation, and presents a new candidate agent for the development of therapies for inflammation-related neurodegenerative diseases.

Energy-regulated molecules maintain young status in the trophocytes and fat cells of old queen honeybees.

Queen honeybees (Apis mellifera) have much longer lifespans than worker bees. Energy-regulated molecules in the trophocytes and fat cells of workers during aging have been determined, but are unknown in queen bees. In the present study, energy-regulated molecules were evaluated in the trophocytes and fat cells of young and old queen bees. Adenosine monophosphate-activated protein kinase α2 (AMPK-α2), phosphorylated AMPK-α2 (pAMPK-α2), and cAMP-specific phosphodiesterases activity increased with aging. The pAMPK-α2/AMPK-α2 ratio and AMPK activity; adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) concentrations; the ADP/ATP ratio and the AMP/ATP ratio; the cyclic adenosine monophosphate concentration; forkhead box protein O expression; Silent information regulator T1 (SirT1) expression and activity; and peroxisome proliferator-activated receptor-α (PPAR-α)expression were not significantly different between young and old queen bees. These results show that energy-regulated molecules maintain a youthful status in the trophocytes and fat cells of queen bees during aging. These cells seem to have longevity-promoting mechanisms and may clarify the secret of longevity in queen bees.

Hepatic oxidative stress in ovariectomized transgenic mice expressing the hepatitis C virus polyprotein is augmented through suppression of adenosine monophosphate-activated protein kinase/proliferator-activated receptor gamma co-activator 1 alpha signali

Oxidative stress plays an important role in hepatocarcinogenesis of hepatitis C virus (HCV)-related chronic liver diseases. Despite the evidence of an increased proportion of females among elderly patients with HCV-related hepatocellular carcinoma (HCC), it remains unknown whether HCV augments hepatic oxidative stress in postmenopausal women. The aim of this study was to determine whether oxidative stress was augmented in ovariectomized (OVX) transgenic mice expressing the HCV polyprotein and to investigate its underlying mechanisms. OVX and sham-operated female transgenic mice expressing the HCV polyprotein and non-transgenic littermates were assessed for the production of reactive oxygen species (ROS), expression of inflammatory cytokines and antioxidant potential in the liver. Compared with OVX non-transgenic mice, OVX transgenic mice showed marked hepatic steatosis and ROS production without increased induction of inflammatory cytokines, but there was no increase in ROS-detoxifying enzymes such as superoxide dismutase 2 and glutathione peroxidase 1. In accordance with these results, OVX transgenic mice showed less activation of peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α), which is required for the induction of ROS-detoxifying enzymes, and no activation of adenosine monophosphate-activated protein kinase-α (AMPKα), which regulates the activity of PGC-1α. Our study demonstrated that hepatic oxidative stress was augmented in OVX transgenic mice expressing the HCV polyprotein by attenuation of antioxidant potential through inhibition of AMPK/PGC-1α signaling. These results may account in part for the mechanisms by which HCV-infected women are at high risk for HCC development when some period has passed after menopause.

Light-to-Moderate Ethanol Feeding Augments AMPK-α Phosphorylation and Attenuates SREBP-1 Expression in the Liver of Rats

Fatty liver disease, a hepatic manifestation of metabolic syndrome, is one of the major causes of chronic liver diseases. Epidemiological studies suggest that regular light-to-moderate ethanol consumption lowers the risk of developing metabolic disorders including dislipidemia, insulin resistance, type 2 diabetes and fatty liver disease. However, the mechanism(s) of the protective effect of light-to-moderate ethanol consumption on the liver remains unknown. In the present study, we investigated the effects of light (6%, 0.94 g/kg/day) and moderate (12%, 1.88 g/kg/day) ethanol feeding in rats for 3 weeks on the circulating and hepatic biochemical profiles and on the hepatic protein expression and phosphorylation status of adenosine monophosphate-activated protein kinase-α (AMPK-α) and other down-stream targets of this enzyme including sterol regulatory element-binding protein-1 (SREBP-1), SREBP cleavage-activating protein (SCAP) and 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA reductase). Despite no significant difference in food-intake among the groups, light ethanol treatment significantly increased the body weight compared to control rats. Serum glucose, insulin, total cholesterol, triglycerides, phospholipids and hepatic cholesterol and triglycerides were not significantly different among the groups. However, serum free fatty acids were significantly reduced with light ethanol treatment. Both light and moderate ethanol treatment significantly increased the hepatic levels of phosphorylated AMPK-α protein and this was associated with significant reduction of SREBP-1 protein expression, suggesting an enhanced fatty acid oxidation. In addition, light ethanol treatment significantly decreased the SCAP protein expression in the liver. However, liver HMG-CoA protein expression was not significantly different with ethanol consumption. Chronic light-to-moderate ethanol consumption increased AMPK activation which was associated with decreased expression of SREBP-1 and SCAP in the liver. Thus, our studies provide mechanistic evidence for the earlier epidemiological studies that indicate light-to-moderate ethanol intake lowers the risk of development of fatty liver disease and other metabolic disorders. Our studies demonstrate that the protective effects of light-to-moderate ethanol arise at least in part by increased phosphorylation of AMPK-α and decreased SREBP-1 expression in the liver. Further studies are warranted to determine the effects of light-to-moderate ethanol on intracellular up-stream and down-stream targets of AMPK and also on the implications of light-to-moderate ethanol in protecting non-alcoholic fatty liver disease.

Antineuroinflammatory effects of lycopene via activation of adenosine monophosphate-activated protein kinase-α1/heme oxygenase-1 pathways

Microglia play an important role in the immune defense in the central nervous system. Activation of microglia leads to the production of excessive inflammatory molecules and deleterious consequences, including neuronal death. Lycopene, 1 of the major carotenoids present in tomatoes, has been shown to exert antioxidant properties and to inhibit cancer cell proliferation. However, the effects of lycopene on neuroinflammatory responses in microglia remain unknown. In this study, we investigated the signaling pathways involved in lycopene-inhibited expression of cyclooxygenase (COX)-2 and inflammation mediators in BV-2 microglia, mouse primary cultured microglia, and rat primary cultured microglia. Lycopene inhibited the enhancement of lipopolysaccharide (LPS)-induced nuclear factor-kappaB (NF-κB) and activator protein 1 (AP-1) DNA binding activity. In the present study, we demonstrated that lycopene inhibits LPS-induced COX-2 expression through heme oxygenase-1 (HO-1) activation. Our results also demonstrate that stimulation with lycopene increases the phosphorylation of liver kinase B1 (LKB1), calmodulin-dependent protein kinase II (CaMKII), and adenosine monophosphate-activated protein kinase (AMPK)-α1. Treatment with AMPK inhibitors effectively antagonized lycopene-stimulated HO-1 expression. Interestingly, we also found that lycopene increased phospho-AMPKα1 accumulation in the nucleus in microglia. Preincubation of cells with HO-1 and AMPK selective pharmacological inhibitors dramatically reversed the inhibitory effect of lycopene on LPS-induced COX-2 and prostaglandin E2 production. Transfection of microglia with HO-1 and AMPKα small interfering RNA (siRNA) also effectively reversed the inhibitory effect of lycopene on LPS-induced COX-2 expression. In a mouse model, lycopene showed significant antineuroinflammatory effects on microglial activation and motor behavior deficits. These findings suggest that lycopene-inhibited LPS-induced COX-2 expression is mediated by HO-1 activation through the AMPK pathway. Therefore, lycopene might be useful as a therapeutic agent for the treatment of neuroinflammation-associated disorders.