AMP-activated Protein Kinase Expression Research Articles

Oral oligofructose challenge reduces expression of glucose transport-1 and 5′-adenosine monophosphate-activated protein kinase in lamellar wall of Holstein heifer claw

The laminar tissue of bovine laminitis may undergo energy failure. The expression of glucose transport protein-1 (GLUT-1) and 5′-adenosine monophosphate-activated protein kinase (AMPK) affects the energy metabolism of digital laminar tissue. This study aimed to determine the expression of glucose uptake and AMPK in laminar wall corium of Holstein heifer claw by oral administration of oligofructose. A total of twelve clinically healthy Holstein heifers were selected and divided into two groups, including control (CON, n = 6) and experimental (OF, n = 6) groups. The heifers of OF group were given 17 g/kg BW oligofructose dissolved in water (20 mL/kg BW) and the heifers of CON group were given water only (20 mL/kg BW). The laminar tissues were collected after euthanasia. The amount of protein and transcript expression of AMPK and GLUT-1 were determined by western blot and quantitative reverse transcription-polymerase chain reaction (qRT-PCR), respectively. Expressions of phosphoenolpyruvate carboxy-kinase (PEPCK), receptor-c coactivator1-α (PGC-1α) and peroxisome proliferator-activated receptor-γ (PPAR-γ) were determined by qRT-PCR. The heifers of OF group showed no significant change in the expression and concentration of AMPK. The phosphor-(Thr172) AMPK and GLUT-1 were significantly decreased, while the gene contents of PPAR-γ and PGC-1α were significantly increased. The activation of AMPK and GLUT-1 in digital laminar tissues of heifers was inhibited, which may contribute to digital laminar tissue damage.

Irisin protects female mice with LPS-induced endometritis through the AMPK/NF-κB pathway.

Objective(s):This research was designed to determine the role of irisin in lipopolysaccharide (LPS)-induced endometritis in female mice. Materials and Methods:Animals were randomly assigned into sham, sham + irisin, LPS, LPS + irisin (0.1, 1, 10 μg/kg), and LPS + irisin + compound C groups. Histological features and expression of AMPK, NF-κB, inflammatory mediators, and oxidative stress markers were compared among different groups. Results:The results showed that LPS resulted in obvious uterus damage, meanwhile, the inflammatory mediators (COX-2, iNOS, IL-1β, IL-6, and TNF-α), as well as NF-κB in the uterine tissue, were significantly increased and the level of adenosine monophosphate-activated protein kinase (AMPK) was reduced. Nevertheless, pretreatment with irisin reversed the phenomena caused by LPS. Interestingly, compound C (AMPK inhibitor) abolished irisin’s effects on the uterus, which suggested that irisin’s beneficial function was achieved through regulating the AMPK-NF-κB pathway. Moreover, LPS-induced alterations of oxidative factors (MnSOD, GSH, and MDA) were reversed significantly by pretreatment with irisin. This data indicated irisin’s beneficial function was also related to antioxidation besides anti-inflammation. Conclusion:Our study implies that irisin is a potential therapeutic agent for endometritis.

The Role of Oxymatrine in Regulating Hematopoietic Stem Cell (HSC) Mitochondrial Apoptosis and Improving Prognosis of Rats with Liver Cirrhosis Through AMP-Activated Protein Kinase (AMPK) Signal Pathway

Our study was aimed to study the mechanism of oxymatrine in regulating the Hematopoietic Stem Cell (HSC) mitochondrial apoptosis and improving prognosis of rats with liver cirrhosis through AMPK signal pathway. The HSC cells were cultivated and identified by IHC method. The inhibition rates for active HSC cells by oxymatrine at several concentrations were as follows; 0.5, 1, 2, 4 and 8 mg/mL, as detected by MTT method. The optimum interfered concentration was confirmed. The condition for apoptosis in HSC cells interfered with oxymatrine at 0.125, 0.25, and 0.5 mg/mL concentrations. The variation of apoptosis in control group and model group was also observed through TEM. The active presentation of AMPK, PAMPK, Caspsase-3 and BCL-2 was detected with RT-PCR. The index of hepatic function in venous blood of rats’ eyeball was tested, and degree of hepatic fibrosis in liver tissue homogenate was also tested. Expression of AMPK/PAMPK, Caspsase-3 and BCL-2 was detected with Western Blot assay. Results showed that, proliferative rate was elevated along with prolonged cultivation duration. The optimum drug interfered concentration was 0.5 mg/mL. The expression of AMPK, PAMPK, Caspsase-3 and BCL-2 in model group was notably higher than in the control group. There was splitting decomposition in cell nucleus and cohesion in cytoplasm after HSC cells were disposed with oxymatrine. The expression of AMPK, Caspsase-3 and BCL-2 was prompted to induce generation of mitochondrial apoptosis, and level of liver function and indicators of hepatic fibrosis were reduced effectively. The prognosis of rats with liver cirrhosis could then be improved.

Melatonin and doxorubicin synergistically enhance apoptosis via autophagy-dependent reduction of AMPK\u03b11 transcription in human breast cancer cells

Doxorubicin is one of the most effective agents used to treat various cancers, including breast cancer, but its usage is limited by the risk of adverse effects, including cardiotoxicity. Melatonin, a natural hormone that functions as a major regulator of circadian rhythms, has been considered a supplemental component for doxorubicin due to its potential to improve its effectiveness. However, the mechanisms and biological targets of the combination of melatonin and doxorubicin with respect to cancer cell death are not well understood. In the present study, we found that melatonin synergized with doxorubicin to induce apoptosis of breast cancer cells by decreasing the expression of AMP-activated protein kinase α1 (AMPK α1), which acts as a critical survival factor for cancer cells. This cotreatment-induced reduction in AMPKα1 expression occurred at the transcriptional level via an autophagy-dependent mechanism. The synergistic effects of the combined treatment were evident in many other cancer cell lines, and melatonin was also highly effective in inducing cancer death when combined with other cancer drugs, including cisplatin, 5-fluorouracil, irinotecan, and sorafenib. AMPKα1 expression was decreased in all of these cases, suggesting that reducing AMPKα1 can be considered an effective method to increase the sensitivity of cancer cells to doxorubicin treatment.

Lycopsamine Attenuates Diabetes Mellitus via The Nuclear Factor Kappa B-Induced 5′ Adenosine Monophosphate-Activated Protein Kinase Signal Pathway

Diabetes mellitus is a metabolic disorder characterized by inflammation, abnormal glycolipid metabolism, insulin resistance, and mitochondrial dysfunction leading to hyperglycemia. The aim of the present investigation was to determine the efficacy of lycopsamine in a rat model of diabetes mellitus to understand its mechanism. Lycopsamine treatment markedly lowered the level of total cholesterol, triglyceride, nonesterified fatty acids, and low-density lipoprotein in diabetic rats. There was also a reduction in interleukin-6, interleukin-10, C-reactive protein, and tumor necrosis factor-α levels. Lycopsamine treatment normalized the metabolism of lipid and glucose, insulin resistance, and body weight of diabetic rats. Findings of immunohistochemical analyses exhibited rise in precipitation of immunocytes in renal cells. Results potentially demonstrated that lycopsamine treatment remarkably reduced the nuclear factor-kappa B level and enhanced the 5′ adenosine monophosphate-activated protein kinase expression. Altogether, administration of lycopsamine suppressed the expression of inflammatory cytokines and attenuated the metabolic symptoms in diabetes mellitus experimental rats.

Roles of AMPK and Its Downstream Signals in Pain Regulation.

Pain is an unpleasant sensory and emotional state that decreases quality of life. A metabolic sensor, adenosine monophosphate-activated protein kinase (AMPK), which is ubiquitously expressed in mammalian cells, has recently attracted interest as a new target of pain research. Abnormal AMPK expression and function in the peripheral and central nervous systems are associated with various types of pain. AMPK and its downstream kinases participate in the regulation of neuron excitability, neuroinflammation and axonal and myelin regeneration. Numerous AMPK activators have reduced pain behavior in animal models. The current understanding of pain has been deepened by AMPK research, but certain issues, such as the interactions of AMPK at each step of pain regulation, await further investigation. This review examines the roles of AMPK and its downstream kinases in neurons and non-neuronal cells, as well as their contribution to pain regulation.

Piperazine ferulate prevents high-glucose-induced filtration barrier injury of glomerular endothelial cells.

Filtration barrier injury induced by high glucose (HG) levels leads to the development of diabetic nephropathy. The endothelial glycocalyx plays a critical role in glomerular barrier function. In the present study, the effects of piperazine ferulate (PF) on HG-induced filtration barrier injury of glomerular endothelial cells (GEnCs) were investigated and the underlying mechanism was assessed. Immunofluorescence was used to observe the distribution of the glycocalyx as well as the expression levels of syndecan-1 and Zonula occludens-1 (ZO-1). Endothelial permeability assays were performed to assess the effects of PF on the integrity of the filtration barrier. Protein and mRNA expression levels were measured by western blotting and reverse transcription-quantitative PCR analyses, respectively. In vitro experiments revealed that adenosine monophosphate-activated protein kinase (AMPK) mediated HG-induced glycocalyx degradation and endothelial barrier injury. PF inhibited the HG-induced endothelial barrier injury and restored the expression levels of heparanase-1 (Hpa-1), ZO-1 and occludin-1 by AMPK. In vivo assays demonstrated that PF reduced the expression levels of Hpa-1, increased the expression levels of ZO-1 and attenuated glycocalyx degradation in the glomerulus. These data suggested that PF attenuated HG-induced filtration barrier injury of GEnC by regulating AMPK expression.

MicroRNA-mediated inhibition of AMPK coordinates tissue-specific downregulation of skeletal muscle metabolism in hypoxic naked mole-rats.

Naked mole-rats reduce their metabolic requirements to tolerate severe hypoxia. However, the regulatory mechanisms that underpin this metabolic suppression have yet to be elucidated. 5'-AMP-activated protein kinase (AMPK) is the cellular 'master' energy effector and we hypothesized that alterations in the AMPK pathway contribute to metabolic reorganization in hypoxic naked mole-rat skeletal muscle. To test this hypothesis, we exposed naked mole-rats to 4 h of normoxia (21% O2) or severe hypoxia (3% O2), while indirectly measuring whole-animal metabolic rate and fuel preference. We then isolated skeletal muscle and assessed protein expression and post-translational modification of AMPK, and downstream changes in key glucose and fatty acid metabolic proteins mediated by AMPK, including acetyl-CoA carboxylase (ACC1), glycogen synthase (GS) and glucose transporters (GLUTs) 1 and 4. We found that in hypoxic naked mole-rats (1) metabolic rate decreased ∼80% and fuel use switched to carbohydrates, and that (2) levels of activated phosphorylated AMPK and GS, and GLUT4 expression were downregulated in skeletal muscle, while ACC1 was unchanged. To explore the regulatory mechanism underlying this hypometabolic state, we used RT-qPCR to examine 55 AMPK-associated microRNAs (miRNAs), which are short non-coding RNA post-transcriptional silencers. We identified changes in 10 miRNAs (three upregulated and seven downregulated) implicated in AMPK downregulation. Our results suggest that miRNAs and post-translational mechanisms coordinately reduce AMPK activity and downregulate metabolism in naked mole-rat skeletal muscle during severe hypoxia. This novel mechanism may support tissue-specific prioritization of energy for more essential organs in hypoxia.

Effect of Deer Antler Extract on Muscle Differentiation and 5-Aminoimidazole-4-Carboxamide Ribonucleoside (AICAR)-Induced Muscle Atrophy in C2C12 Cells.

The effect of deer antler extract on muscle differentiation and muscle atrophy were evaluated to minimize muscle loss following aging. Various deer antler extracts (HWE, hot water extract of deer antler; FE, HWE of fermented deer antler; ET, enzyme-assisted extract of deer antler; UE, extract prepared by ultrasonication of deer antler) were evaluated for their effect on muscle differentiation and inhibition of 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR)-induced muscle atrophy in C2C12 cells. Morphological changes according to the effect of antler extracts on muscle differentiation were confirmed by Jenner-Giemsa staining. In addition, the expression levels of genes related to muscle differentiation and atrophy were confirmed through qRT-PCR. In the presence of antler extracts, the length and thickness of myotubes and myogenin differentiation 1 (MyoD1) and myogenic factor 5 (Myf5) gene expression were increased compared to those in the control group (CON). Gene expression of AMP-activated protein kinase (AMPK), MyoD1, and myogenin, along with the muscle atrophy factors muscle RING finger-1 (MuRF-1) and forkhead box O3a (FoxO3a) upon addition of deer antler extracts to muscle-atrophied C2C12 cells was determined by qRT-PCR after treatment with AICAR. The expression of MuRF-1 and FoxO3a decreased in the groups treated with antler extracts compared to that in the group treated with AICAR alone. In addition, gene expression of MyoD1 and myogenin in the muscle atrophy cell model was significantly increased compared that into the CON. Therefore, our findings indicate that antler extract can increase the expression of MyoD1, Myf5 and myogenin, inhibit muscle atrophy, and promote muscle differentiation.

The influence of thyroid state on hypothalamic AMP-activated protein kinase pathways in broilers

To investigate whether there are important interactions in play in broilers between thyroid hormones and the central regulation of energy homeostasis through AMP-activated protein kinase (AMPK), we induced a functional hyperthyroid and hypothyroid state in broiler chicks, and quantified systemic and hypothalamic AMPK related gene expression and related protein. Thyroid state was manipulated through dietary supplementation of triiodothyronine (T3) or methimazole (MMI) for 7 days. A hypothalamic AMPK suppressor, 0.1% α-lipoic acid (α-LA) was used to assess the effects of the T3 and MMI feed formulations on the AMPK pathways. Feed intake and body weight were reduced in both hypothyroid and hyperthyroid conditions. In hyperthyroid conditions (T3 supplementation) expression of the AMPKα1 subunit increased, while in hypothyroid conditions (MMI supplementation) active phosphorylated AMPK levels in the hypothalamus dropped, but gene expression of the AMPKα1 and α2 subunit increased. For FAS and ACC (involved in fatty acid metabolism), and CRH, TRH and CNR1 (anorexigenic neuropeptides stimulating energy expenditure) there were indications that their regulation in response to thyroid state might be modulated through AMPK pathways. Our results indicate that the expression of hypothalamic AMPK as well as that of several other genes from AMPK pathways are involved in thyroid-hormone-induced changes in appetite, albeit differently according to thyroid state.

Phosphatase and Tensin Homolog Deleted on Chromosome Ten Knockdown Attenuates Cognitive Deficits by Inhibiting Neuroinflammation in a Mouse Model of Perioperative Neurocognitive Disorder

Phosphatase and tensin homolog deleted on chromosome ten (PTEN) is a crucial regulator of neuronal development, neuronal survival, axonal regeneration, and synaptic plasticity. In this study we examined the potential role of PTEN in cognitive function in a mouse model of perioperative neurocognitive disorder (PND). Adult male C57BL/6J mice received intracerebroventricular injections of small interfering RNA (siRNA) against PTEN or control siRNA 3 days prior to exploratory laparotomy (n = 8 per group). A group of healthy mice not undergoing surgery included as additional control. Barnes maze and fear conditioning tests were conducted 7 days after surgery. Mice were then sacrificed to examine the expression of PTEN, AMP-activated protein kinase (AMPK), ionized calcium binding adaptor molecule (Iba)-1, B-cell lymphoma (Bcl)-2, Bcl2-associated X protein (Bax), interleukin (IL)-1β, and tumor necrosis factor (TNF)-α in the hippocampus. The microglial activation was examined by immunohistochemistry using Iba-1 as a microglia maker. Nissl and terminal transferase deoxyuridine triphosphate nick-end labeling (TUNEL) staining were used to measure cell death and apoptosis. In comparison to the healthy controls, surgically treated mice had longer latency to identify the target box in both training and testing sessions in the Barnes maze test and shorter freezing time in the fear conditioning test. Surgically treated mice had increased expression of PTEN, AMPK, Bax, IL-1β, and TNF-α, as well as increasing number of activated microglia and apoptosis neurons in the hippocampus. PTEN knockdown significantly attenuated the behavioral deficits in Barnes maze and fear conditioning tests, as well as over-expression of PTEN, AMPK, Bax, IL-1β, and TNF-α induced by surgery. PTEN knockdown could attenuate cognitive deficits induced by trauma, likely through inhibiting the activation of microglia.

Improving the post-thaw quality of rooster semen using the extender supplemented with resveratrol

Avian spermatozoa are highly susceptible to reactive oxygen species (ROS) produced during the cryopreservation. The aim of the current study was to investigate the antioxidant effects of resveratrol (RSV) during rooster semen cryopreservation. Changes in expression of AMP-activated protein kinase as a possible mechanism behind the beneficial effects of resveratrol were also evaluated. Semen samples were collected from ten Ross broiler breeders (52-wk) using abdominal massage, then divided into 4 equal aliquots and cryopreserved in Beltsville extender that contained different concentrations (0 µM, 0.01µM, 0.1µM, and 1µM) of RSV. higher percentage (P < 0.05) of total motility and membrane integrity was observed in RSV-0.1 compared to the other frozen groups. Moreover, higher percentage of sperm mitochondrial activity was observed in the RSV-0.01 and RSV-0.1 compared to the frozen control (P < 0.05). The lowest percentage of apoptotic like changes was found in the RSV-0.1 in comparison to the other groups (P < 0.05). RSV-0.01 and RSV-1 groups produced the lowest levels of H2O2 and O2− compared to the other frozen groups, respectively. Malondialdehyde (MDA) concentration, velocity average path (VAP), and linearity (LIN) were not affected by different concentrations of RSV (P > 0.05). We observed a dose-dependent increase in AMP-activated protein kinase expression in groups exposed to RSV. Thus, RSV-1 increased AMP-activated protein kinase phosphorylation but had no positive effects on post thaw sperm parameters. Our findings suggest that RSV-0.1 improve thawed sperm functions, and these effects might be mediated through activation of AMP-activated protein kinase.

Synergetic Action of Forskolin and Mevastatin Induce Normalization of Lipids Profile in Dyslipidemic Rats through Adenosine Monophosphate Kinase Upregulation.

In the present study, we examined the synergetic effect of forskolin and mevastatin administration on lipid profile and lipid metabolism in omental adipose tissue in dyslipidemic rats. The study was conducted on forty male albino rats. The rats were randomly classified into four main groups of ten animals in each group as follows: group A, served as control nontreated; group B, rats that received Triton WR 1339 (500 mg/kg); group C, rats that received Triton WR 1339 with forskolin (100% FSK extract 0.5 mg/kg/day) for four weeks; and group D, dyslipidemic rats received both mevastatin and forskolin. At the end of the experimental period, blood and omental adipose tissue samples were collected, preserved, and used for biochemical determination of lipid profile and mRNA expression profile of adenylate cyclase (AC), hormone-sensitive lipase, respectively (HSL), and adenosine monophosphate-activated protein kinase (AMPK). The results showed a significant decline in the serum concentration of total cholesterol, LDL-cholesterol, and triglycerides, although there was a significant increase in serum levels of HDL-cholesterol and glycerol in rats received forskolin alone or with mevastatin when compared with control and dyslipidemic groups. The mRNA expression levels of AC, HSL, and AMPK were significantly increased in omental adipose tissue of rats received forskolin when compared with other groups. In conclusion, forskolin acts synergistically with mevastatin to lower lipid profile and improve lipid metabolism in dyslipidemic rats through upregulation of AMPK expression.

The Effect of Endurance Training with Crocin Consumption on the Levels of AMPK and NRF1 Gene Expression in the Soleus Muscles of Diabetic Rats

Background: Exercise training and crocin, as a herbal antioxidant, help improve metabolism and insulin sensitivity in type 2 diabetes mellitus (T2D). AMP-activated protein kinase (AMPK) and nuclear respiratory factor 1 (NRF1) involve in the mitochondrial biogenesis pathway and diabetes control. Objective: This study aimed to investigate the effects of high-intensity interval training (HIIT), and low intensity continued training (LICT) in combination with crocin consumption on AMPK and NRF1 gene expression in the muscle tissue of diabetic rats. Methods: In this experimental study, 56 adult rats were divided into eight groups (n = 7), including diabetic (crocin utilization, HIIT, LICT, HIIT with crocin, LICT with crocin, control, sham groups) and healthy control group. The HIIT and LICT groups performed five sessions of training per week for eight weeks, and the crocin consumption groups received 25 mg/kg of crocin daily for eight weeks. Moreover, AMPK and NRF1 genes expression levels were evaluated in different groups using real-time polymerase chain reaction (RT-PCR). Results: According to our results, while induction of diabetes significantly decreased the levels of NRF1 and AMPK gene expression (P ≤ 0.05), training and crocin consumption significantly increased the levels of AMPK and NRF1 gene expression (P ≤ 0.05). Conclusions: It appears that crocin consumption along with HIIT improves diabetes symptoms through AMPK and NRF1 mechanism in diabetic rats.

AdipoRon, an Orally Active, Synthetic Agonist of AdipoR1 and AdipoR2 Receptors Has Gastroprotective Effect in Experimentally Induced Gastric Ulcers in Mice

Introduction: Adiponectin is a hormone secreted by adipocytes, which exhibits insulin-sensitizing and anti-inflammatory properties and acts through adiponectin receptors: AdipoR1 and AdipoR2. The aim of the study was to evaluate whether activation of adiponectin receptors AdipoR1 and AdipoR2 with an orally active agonist AdipoRon has gastroprotective effect and to investigate the possible underlying mechanism. Methods: We used two well-established mouse models of gastric ulcer (GU) induced by oral administration of EtOH (80% solution in water) or diclofenac (30 mg/kg, p.o.). Gastroprotective effect of AdipoRon (dose 5 and 50 mg/kg p.o.) was compared to omeprazole (20 mg/kg p.o.) or 5% DMSO solution (control). Clinical parameters of gastroprotection were assessed using macroscopic (gastric lesion area) and microscopic (evaluation of the gastric mucosa damage) scoring. To establish the molecular mechanism, we measured: myeloperoxidase (MPO), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX) activities; glutathione (GSH) level; and IL-1β, adenosine monophosphate-activated protein kinase (AMPK), and phosphorylated AMPK expression in gastric tissue. Results: AdipoRon produced a gastroprotective effect in both GU mouse models as evidenced by significantly lower macroscopic and microscopic damage scores. AdipoRon exhibited anti-inflammatory effect by reduction in MPO activity and IL-1β expression in the gastric tissue. Moreover, AdipoRon induced antioxidative action, as demonstrated with higher GSH levels, and increased SOD and GPX activity. Conclusions: Activation of AdipoR1 and AdipoR2 using AdipoRon reduced gastric lesions and enhanced cell response to oxidative stress. Our data suggest that AdipoR1 and AdipoR2 activation may be an attractive therapeutic strategy to inhibit development of gastric ulcers.

Effect of electroacupuncture on the expression of autophagy related protein in lung tissue of rats with chronic obstructive pulmonary disease

To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) and "Feishu" (BL13) on the expression of autophagy related proteins in the lung tissue of rats with chronic obstructive pulmonary disease (COPD), so as to explore the mechanism of EA underlying improvement of COPD. Thirty male SD rats were randomly divided into normal, model and EA groups (n=10 in each group). The COPD model was established by intratracheal infusion of Lipopolysaccharide (LPS, 1 mg/kg) and exposure in cigarette smoke. EA was applied to bilateral ST36 and BL13 for 30 min, once every other day for 2 weeks. The pulmonary function (forced vital capacity ［FVC］, forced expiratory volume in 0.1 s and 0.3 s ［FEV0.1, FEV0.3］, FEV0.1/FVC and FEV0.3/FVC) was detected by animal pulmonary function analysis system. Histopathological changes of the airway and lung were displayed by H.E. staining. Autophagosomes in the airway and lung tissues were observed by electron microscope. The expression of AMP activated protein kinase (AMPK), mammalian target of rapamycin (mTOR), Unc-51 like autophagy activating kinase 1(ULK1), autophagy related protein ATG6(Beclin1)mRNAs in lung tissue were examined by quantitative real-time PCR. The expression of AMPK, mTOR, ULK1, Beclin1 and microtubule-associated protein 1 light chain 3 (LC3)proteins in lung tissue were examined by Western blot. The contents of tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) in the broncho alveolar lavage fluid (BALF) were assayed by ELISA. Following modeling, the FVC, FEV0.1, FEV0.3, FEV0.1/FVC and FEV0.3/FVC levels were significantly decreased (P<0.01), the infiltration of inflammatory cells and the increase of autophagosomes were obvious in airway and lung tissue, the mRNA and protein expression of AMPK, ULK1, Beclin1 and the ratio of LC3Ⅱ/LC3Ⅰ were increased (P<0.01), while the mRNA and protein expression of mTOR were decreased (P<0.01), the contents of TNF-α and IL-6 in the BALF were increased in the model group compared with the normal group (P<0.01). After EA intervention, all the indexes mentioned above were completely reversed in the EA group relevant to the model group (P<0.01, P<0.05). EA at ST36 and BL13 can improve the lung function of COPD rats, which may be related to its effects in inhibiting the autophagy level and reducing the inflammation response in the lung.

Perfluorooctanesulfonic Acid (PFOS) Thwarts the Beneficial Effects of Calorie Restriction and Metformin.

A combination of calorie restriction (CR), dietary modification, and exercise is the recommended therapy to reverse obesity and nonalcoholic fatty liver disease. In the liver, CR shifts hepatic metabolism from lipid storage to lipid utilization pathways, such as AMP-activated protein kinase (AMPK). Perfluorooctanesulfonic acid (PFOS), a fluorosurfactant previously used in stain repellents and anti-stick materials, can increase hepatic lipids in mice following relatively low-dose exposures. To test the hypothesis that PFOS administration interferes with CR, adult male C57BL/6N mice were fed ad libitum or a 25% reduced calorie diet concomitant with either vehicle (water) or 100 μg PFOS/kg/day via oral gavage for 6 weeks. CR alone improved hepatic lipids and glucose tolerance. PFOS did not significantly alter CR-induced weight loss, white adipose tissue mass, or liver weight over 6 weeks. However, PFOS increased hepatic triglyceride accumulation, in both mice fed ad libitum and subjected to CR. This was associated with decreased phosphorylated AMPK expression in liver. Glucagon (100 nM) treatment induced glucose production in hepatocytes, which was further upregulated with PFOS (2.5 μM) co-treatment. Next, to explore whether the observed changes were related to AMPK signaling, HepG2 cells were treated with metformin or AICAR alone or in combination with PFOS (25 μM). PFOS interfered with glucose-lowering effects of metformin, and AICAR treatment partially impaired PFOS-induced increase in glucose production. In 3T3-L1 adipocytes, metformin was less effective with PFOS co-treatment. Overall, PFOS administration disrupted hepatic lipid and glucose homeostasis and interfered with beneficial glucose-lowering effects of CR and metformin.

Ventromedial hypothalamic nucleus glycogen regulation of metabolic-sensory neuron AMPK and neurotransmitter expression: role of lactate.

Astrocyte glycogen is dynamically remodeled during metabolic stability and provides oxidizable l-lactate equivalents during neuroglucopenia. Current research investigated the hypothesis that ventromedial hypothalamic nucleus (VMN) glycogen metabolism controls glucostimulatory nitric oxide (NO) and/or glucoinhibitory gamma-aminobutyric acid (GABA) neuron 5'-AMP-activated protein kinase (AMPK) and transmitter marker, e.g., neuronal nitric oxide synthase (nNOS), and glutamate decarboxylase65/67 (GAD) protein expression. Adult ovariectomized estradiol-implanted female rats were injected into the VMN with the glycogen phosphorylase inhibitor 1,4-dideoxy-1,4-imino-d-arabinitol (DAB) before vehicle or l-lactate infusion. Western blot analysis of laser-catapult-microdissected nitrergic and GABAergic neurons showed that DAB caused lactate-reversible upregulation of nNOS and GAD proteins. DAB suppressed or increased total AMPK content of NO and GABA neurons, respectively, by lactate-independent mechanisms, but lactate prevented drug enhancement of pAMPK expression in nitrergic neurons. Inhibition of VMN glycogen disassembly caused divergent changes in counter-regulatory hormone, e.g. corticosterone (increased) and glucagon (decreased) secretion. Outcomes show that VMN glycogen metabolism controls local glucoregulatory transmission by means of lactate signal volume. Results implicate glycogen-derived lactate deficiency as a physiological stimulus of corticosterone release. Concurrent normalization of nitrergic neuron nNOS and pAMPK protein and corticosterone secretory response to DAB by lactate infers that the hypothalamic-pituitary-adrenal axis may be activated by VMN NO-mediated signals of cellular energy imbalance.

POS-382 A New AMPK Activator PA-S14 Attenuates Renal Injury through Promoting Autophagy

Autophagy deficiency plays a critical role in the pathogenesis of chronic kidney disease (CKD). AMP-activated protein kinase (AMPK), plays a central role in modulating metabolic stability. Several studies have shown that AMPK activator could ameliorate renal fibrosis, however, the underlying mechanisms have not been elucidated. In this study, we constructed a new AMPK activator, PA-S14, which is an allosteric compound from piericidin A (PA). The therapeutic effects of PA-S14 were tested in several CKD models. Furthermore, the autophagy status was also assessed. Bal/bc mice were performed unilateral ureteral obstruction (UUO), unilateral ischemia-reperfusion injury (UIRI) surgery, or injected with adriamycin (ADR). Some mice were intraperitoneally injected with PA-S14 (1mg/kg/d). In vitro, human renal tubular epithelial cells (HKC-8) were treated with transforming-growth factor (TGF)-β1, Bafilomycin A1(an autophagy inhibitor), and PA-S14. Kidney fibrosis, mitochondrial function, autophagy and AMPK/mTOR signaling pathway were assessed. Autophagosomes was visualized by the confocal microscopy after mRFP-EGFP-LC3 lentivirus infection and transmission electron microscopy. The expression of AMPK was decreased in various CKD models (UUO, 5/6 nephrectomy and adriamycin nephropathy). In vivo, administration of PA-S14 alleviated proteinuria and serum creatinine, and attenuated kidney fibrosis in unilateral ishemia-reperfusion (UIRI) and ADR mice. Moreover, PA-S14 upregulated p-AMPK expression and downregulated p-mTOR expression. PA-S14 also triggered the reversal of peroxisome proliferator-activated receptor γ co-activator 1 (PGC-1α), mitochondrial transcription factor A (TFAM), the important mitochondrial biogenesis factors. PA-S14 treatment also induced autophagosome formation, increased LC3-II lipidation, and decreased p62. In HKC-8 cells, PA-S14 inhibited TGF-β1-mediated fibrosis, alleviated the mitochondrial ROS production, and improved mitochondrial membrane potential. In addition, PA-S14 also activated AMPK and inhibited mTOR pathway, further leading to increased autophagic flux. In addition, Bafilomycin A1-suppressed autophagic flux could be reversed by PA-S14. PA-S14 also preserved the expression of phosphorylated AMPK in PP2C, an AMPK inhibitor, -treated cells. In summary, our results demonstrate that PA-S14 acts as a new AMPK activator to protect against renal fibrosis. The underlying mechanisms are related to PA-S14-induced AMPK activation, which further promotes autophagy through regulating mTOR pathway.

Dapagliflozin Regulates Cardiac Metabolic Remodeling Partially via Uncoupling Protein 3 in a Nondiabetic Thoracic Aortic Constriction-Induced Mouse Model

Objectives: We aimed to examine the protective effects of dapagliflozin (dapa) on thoracic aortic constriction (TAC)-induced heart failure in a nondiabetic mouse model. More specifically, we determined the effects of dapa on uncoupling protein 3 (UCP3) activation and subsequent metabolic remodeling. Methods: Sixty C57BL/6J mice were divided into six groups for TAC surgery and received different doses of dapa via gavage for 4 weeks. Echocardiography was performed to evaluate cardiac structure and function. Histological and molecular markers of cardiac remodeling and metabolic changes were assessed through staining assays, RT-PCR, western blot, and ELISA. HL-1 cells were used to explore the role of UCP3 in metabolic remodeling through transfection with UCP3 siRNA. Results: Mice that received TAC exhibited elevated heart weight/body weight ratios (HW/BW), left ventricular (LV) hypertrophy, impaired LV ejection fraction, and increased rates of fibrosis and apoptosis, unlike mice that received sham operation. Treatment with dapa after TAC restored HW/BW, improved LV parameters, and reduced fibrosis and apoptosis. dapa changed the expression levels of enzymes involved in glucose and fatty acid (FA) metabolism, such as pyruvate dehydrogenase lipoamide kinase isozyme 4, glucose transporter 4, carnitine palmitoyltransferase-1α, carnitine O-acetyltransferase, carnitine O-octanoyltransferase, acyl-CoA thioesterase 1, acyl-CoA thioesterase 2, peroxisome proliferator-activated receptors α and β, proliferator-activated receptor-gamma coactivator-1α, and UCP3, relative to the levels in mice in the TAC group. UCP3 siRNA reduced the expression of AMP-activated protein kinase and of factors involved in FA oxidation in vitro. Conclusions: Dapa exhibits cardioprotective effects in the model and augments expression of UCP3, which may be involved in metabolic remodeling in the failing heart.