bims-amsmem Biomed News
on AMPK signaling mechanism in energy metabolism
Issue of 2022–05–15
sixty-four papers selected by
Dipsikha Biswas, Københavns Universitet
  1. Structure-function analysis of the AMPK activator SC4 and identification of a potent pan AMPK activator.
  2. Profilin upregulation induces autophagy through stabilization of AMP-activated protein kinase.
  3. Electroacupuncture in the treatment of neurogenic urine retention through autophagy mediated by AMPK/mTOR pathway.
  4. Effects of a novel AMP-mimicking prodrug on AMPK and mTORC1 activity in C2C12 cells.
  5. Cardiac AMPK is not required for cardioprotection by metformin in pulmonary hypertension induced right heart failure.
  6. The Effects of Metformin on Skeletal Muscle Differentiation and Satellite Cell Function in C2C12 Myocytes and Juvenile Lean and Obese Mice.
  7. In Vivo Effects of AICAR and Prodrug 39 (P39) on Anabolic Pathways Following Refeeding in Skeletal Muscle.
  8. Intestinal AMPK modulation of gut microbiota mediates cross-talk with brown fat to control metabolism.
  9. AMP-activated protein kinase α1 phosphorylates PHD2 to maintain systemic iron homeostasis.
  10. Combinatorial therapies for rescuing myotonic dystrophy type 1 skeletal muscle defects.
  11. Cilia-localized GID/CTLH ubiquitin ligase complex regulates protein homeostasis of sonic hedgehog signaling components.
  12. Chronic AICAR treatment enhances anabolic signaling in sarcopenic skeletal muscle.
  13. Anti-obesity effect of Premature Citrus Extract in high-fat diet induced obesity mice.
  14. Ethyl Gallate Dual-Targeting PTPN6 and PPARγ Shows Anti-Diabetic and Anti-Obese Effects.
  15. Chronic AICAR treatment enhances anabolic signaling in sarcopenic skeletal muscle.
  16. Defective AMPK regulation of cholesterol metabolism accelerates atherosclerosis by promoting HSPC mobilization and myelopoiesis.
  17. Guanylate-binding protein 2b regulates the AMPK/mTOR/ULK1 signalling pathway to induce autophagy during Mycobacterium bovis infection.
  18. Effect of Long-Term Supplementation with Acetic Acid on the Skeletal Muscle of Aging Sprague Dawley Rats.
  19. Leptin relieves ischemia/reperfusion induced acute kidney injury through inhibiting apoptosis and autophagy.
  20. Bilirubin Improves Gap Junction to Alleviate Doxorubicin-Induced Cardiotoxicity by Regulating AMPK-Axl-SOCS3-Cx43 Axis.
  21. Relationship of PI3K-Akt/mTOR/AMPK signaling pathway genetic mutation with efficacy and prognosis in nasopharyngeal carcinoma.
  22. Salidroside Ameliorates Ischemia-Induced Neuronal Injury through AMPK Dependent and Independent Pathways to Maintain Mitochondrial Quality Control.
  23. Combined effects of exercise training and D-allulose intake on endurance capacity in mice.
  24. Distinct AMPK-Mediated FAS/HSL Pathway Is Implicated in the Alleviating Effect of Nuciferine on Obesity and Hepatic Steatosis in HFD-Fed Mice.
  25. Heat stress-induced changes in the myocardium.
  26. Hempseed (Cannabis sativa) Peptide H3 (IGFLIIWV) Exerts Cholesterol-Lowering Effects in Human Hepatic Cell Line.
  27. Targeting Sirt1, AMPK, Nrf2, CK2, and Soluble Guanylate Cyclase with Nutraceuticals: A Practical Strategy for Preserving Bone Mass.
  28. Changes in AMPKα and Ubiquitin Ligases in Myocyte Reverse Remodeling after Surgical Ventricular Reconstruction in rats with ischemic cardiomyopathy.
  29. Measuring AMPK Activity in Vivo Using a Genetically-Encoded Biosensor.
  30. Dexmedetomidine Can Enhance PINK1/Parkin-Mediated Mitophagy in MPTP-Induced PD Mice Model by Activating AMPK.
  31. Increased Connexin 43 expression in AMPK deficient cardiomyocytes in response to Doxorubicin treatment.
  32. Hernandezine Regulates Proliferation and Autophagy-Induced Apoptosis in Melanoma Cells.
  33. Antitumor activity of pachymic acid in cervical cancer through inducing endoplasmic reticulum stress, mitochondrial dysfunction, and activating the AMPK pathway.
  34. Chitosan Oligosaccharide Ameliorates Kidney Injury by Improving Intestinal Barrier Dysfunction and Lipid Metabolism in Obese-insulin Resistant Rats.
  35. Corrigendum: Metformin Ameliorates Inflammation and Airway Remodeling of Experimental Allergic Asthma in Mice by Restoring AMPKα Activity.
  36. [Dihydromyricetin reduces lipid accumulation in LO2 cells via AMPK/mTOR-mediated lipophagy pathway and inhibits HepG2 cell proliferation in vitro].
  37. Cleaved to keep dying cells alive.
  38. STK11 overexpression prevents glucocorticoid-induced osteoporosis via activating the AMPK/SIRT1/PGC1α axis.
  39. Neuronal growth regulator 1 promotes adipocyte lipid trafficking via interaction with CD36.
  40. Elimination of AKT in Cardiomyocytes Causes Complex Reorganization of Signal Transduction leading to AMPK Inhibition and Fatal Cardiac Atrophy.
  41. Qili Qiangxin, a compound herbal medicine formula, alleviates hypoxia-reoxygenation-induced apoptotic and autophagic cell death via suppression of ROS/AMPK/mTOR pathway in vitro.
  42. Hyperbaric Oxygen Therapy Following Cervical Spinal Cord Injury Increases BDNF/TrkB Signaling in Diaphragm Muscle.
  43. Glucolipotoxicity induces endothelial cell dysfunction by activating autophagy and inhibiting autophagic flow.
  44. Curcumin exerts chondroprotective effects against osteoarthritis by promoting AMPK/PINK1/Parkin-mediated mitophagy.
  45. Nutraceuticals/Drugs Promoting Mitophagy and Mitochondrial Biogenesis May Combat the Mitochondrial Dysfunction Driving Progression of Dry Age-Related Macular Degeneration.
  46. HM-Chromanone Ameliorates Hyperglycemia and Dyslipidemia in Type 2 Diabetic Mice.
  47. Quamoclit angulata extract supplementation attenuates hepatic damage via regulation of AMPK/SIRT associated hepatic lipid metabolism in streptozotocin and high fat diet-induced T2DM mice.
  48. Colon Cancer Cell Death by Synergistic Activity of Phytochemicals is Modulated by Increased ROS, Activation of AMPK, and Inhibition of mTORC1.
  49. Alkaline intracellular pH (pHi) increases mTORC1 and mTORC2 signaling through PI3K to promote protein synthesis and cell survival.
  50. Transcriptome Reveals Granulosa Cells Coping through Redox, Inflammatory and Metabolic Mechanisms under Acute Heat Stress.
  51. Electrical Stimulated Glut4 Trafficking Reduces Critical Illness-Associated Muscle Wasting.
  52. Snf1p/Hxk2p/Mig1p pathway regulates hexose transporters transcript levels, affecting the exponential growth and mitochondrial respiration of Saccharomyces cerevisiae.
  53. A novel phenolic formulation for treating hepatic and peripheral insulin resistance by regulating GLUT4-mediated glucose uptake.
  54. ST segment elevation: the lost measure of early ischemic preconditioning in the landmark study of Murry et al. 1986.
  55. Differences in Cellular Metabolism and Metabolic Regulation between Non-diabetic and Diabetic Human Coronary Artery Endothelial Cells.
  56. Novel Inhibitor for Downstream Targeting of Transforming Growth Factor-β Signaling to Suppress Epithelial to Mesenchymal Transition and Cell Migration.
  57. FADS1 overexpression promotes fatty acid synthesis and triacylglycerol accumulation via inhibiting the AMPK/SREBP1 pathway in goat mammary epithelial cells.
  58. Agomelatine Ameliorates Obesity-Induced Kidney Injury through the Inhibition of Renal Fibrosis and Improvement of Impaired Autophagy.
  59. Characterizing Metabolic Changes in Cancer Cells after Treatment with an Extract Created from Walnuts.
  60. Chronic Intermittent Hypoxia Exposure Alternative to Exercise Alleviates High-Fat-Diet-Induced Obesity and Fatty Liver.
  61. Anthelmintics nitazoxanide protects against experimental hyperlipidemia and hepatic steatosis in hamsters and mice.
  62. Mitochondrial Uncoupling Decreases Sarcopenic Obesity By Activation of Skeletal Muscle Mitochondrial Quality Control and Attenuated ER Stress.
  63. Targeting PFKL with penfluridol inhibits glycolysis and suppresses esophageal cancer tumorigenesis in an AMPK/FOXO3a/BIM-dependent manner.
  64. Tert-Butylhydroquinone alleviates insulin resistance and liver steatosis in diabetes.
  1. Biochem J. 2022 May 13. pii: BCJ20220067. [Epub ahead of print]
    Structure-function analysis of the AMPK activator SC4 and identification of a potent pan AMPK activator.
    Ashley J Ovens, Yi Sing Gee, Naomi X Y Ling, Dingyi Yu, Justin D Hardee, Jin D Chung, Kevin R W Ngoei, Nicholas J Waters, Nolan J Hoffman, John W Scott, Kim Loh, Katrin Spengler, Regine Heller, Michael W Parker, Gordon S Lynch, Fei Huang, Sandra Galic, Bruce Kemp, Jonathan Baell, Jonathan S Oakhill, Christopher G Langendorf.
      The AMP-activated protein kinase (AMPK) αβγ heterotrimer is a primary cellular energy sensor and central regulator of energy homeostasis. Activating skeletal muscle AMPK with small molecule drugs improves glucose uptake and provides opportunity for new strategies to treat type 2 diabetes and insulin resistance, with recent genetic and pharmacological studies indicating the α2β2γ1 isoform combination as the heterotrimer complex primarily responsible. With the goal of developing α2β2-specific activators, here we perform structure/function analysis of the 2-hydroxybiphenyl group of SC4, an activator with tendency for α2-selectivity that is also capable of potently activating β2 complexes. Substitution of the LHS 2-hydroxyphenyl group with polar-substituted cyclohexene-based probes resulted in two AMPK agonists, MSG010 and MSG011, which did not display α2-selectivity when screened against a panel of AMPK complexes. By radiolabel kinase assay, MSG010 and MSG011 activated α2β2γ1 AMPK with one order of magnitude greater potency than the pan AMPK activator MK-8722. A crystal structure of MSG011 complexed to AMPK α2β1γ1 revealed a similar binding mode to SC4 and the potential importance of an interaction between the SC4 2-hydroxyl group and a2-Lys31 for directing α2-selectivity. MSG011 induced robust AMPK signalling in mouse primary hepatocytes and commonly used cell lines, and in most cases this occurred in the absence of changes in phosphorylation of the kinase activation loop residue α-Thr172, a classical marker of AMP-induced AMPK activity. These findings will guide future design of α2β2-selective AMPK activators, that we hypothesise may avoid off-target complications associated with indiscriminate activation of AMPK throughout the body.
    Keywords:  AMPK; drug discovery and design; metabolic disorders; structural biology; type 2 diabetes
    DOI:  https://doi.org/10.1042/BCJ20220067
  2. FEBS Lett. 2022 May 09.
    Profilin upregulation induces autophagy through stabilization of AMP-activated protein kinase.
    Shashank Saurav, Sunil Kumar Manna.
      Profilin regulates actin polymerization, and its balanced expression is required for cellular growth and development. Most tumors have compromised profilin expression, and its overexpression in MDA MB-231 breast cancer cells has been reported to activate AMP-activated protein kinase α (AMPKα), an energy-sensing molecule that affects various cellular processes including autophagy. The present study aims to explore the role of profilin in inducing autophagy. We employed all-trans retinoic acid (ATRA) as an inducer of profilin expression and showed that profilin induces autophagy through mTOR inhibition, autophagy-activating kinase ULK1 upregulation, and AMPK stabilization as well as its activation. Furthermore, evidence from our study indicates physical interaction between profilin and AMPK, which results in AMPK stabilization and induction of prolonged autophagy, thereby leading to apoptosis. This study uncovers a novel mechanism that induces autophagy in triple-negative breast cancer cells.
    Keywords:  AMP-activated protein kinase (AMPK); Autophagy; LAMP 2; LC3; Profilin; Retinoic acid (ATRA)
    DOI:  https://doi.org/10.1002/1873-3468.14372
  3. Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2022 Apr 28. pii: 1672-7347(2022)04-0488-09. [Epub ahead of print]47(4): 488-496
    Electroacupuncture in the treatment of neurogenic urine retention through autophagy mediated by AMPK/mTOR pathway.
    Zhengfei Li, Ren Zhang, Guorui Zhao, Yao Kuang.
       OBJECTIVES: Electroacupuncture can enhance autophagic flow, promote neuronal regeneration, axonal and myelin remodeling to achieve the protection of spinal cord injury, but its role in neurogenic urine retention is not completely clear. This study aims to investigate whether the mechanism of electroacupuncture in the treatment of neurogenic urine retention is through autophagy mediated by adenosine monophosphate activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway.
    METHODS: A rat model of neurogenic urine retention after sacral spinal cord injury was established. The rats with successful model were randomly divided into a model group, an electroacupuncture group (electro-acupuncture for Ciliao, Zhongji, and Sanyinjiao by electronic stimulation, once a day, 20 min each time for 7 days), and an electroacupuncture+AMP-activated protein kinase (AMPK) inhibitor group (on the basis of the treatment of electroacupuncture group, 100 μg of AMPK inhibitor compound C was injected intramuscularly around the L2-3 intervertebral space on the 1st and 4th day). The normal group did not receive any treatment. The maximum bladder volume, bladder basal pressure, leak point pressure, and bladder compliance were recorded by multi-channel physiological recorder; the morphology of bladder tissue was observed by HE staining; autophagy was observed under transmission electron microscope; the expressions of LC3II and Beclin1 protein were observed by immunofluorescence staining; the protein levels of AMPK, phosphorylated-AMPK (p-AMPK), mTOR, phosphorylated-mTOR (p-mTOR), microtubule associated protein 1 light chain 3 (LC3) II and Beclin1 in bladder tissue were detected by Western blotting.
    RESULTS: Compared with the normal group, the maximum bladder capacity, leak point pressure, bladder compliance, p-AMPK, LC3II, Beclin1 protein expressions in the bladder tissue of the model group increased, and the p-mTOR protein expressions were decreased (all P<0.05); compared with the model group, the maximum bladder capacity, bladder compliance, p-mTOR protein expression in the bladder tissue of the electroacupuncture group were decreased, and the p-AMPK, LC3II, and Beclin1 protein expressions were increased (all P<0.05); compared with the electroacupuncture group, the maximum bladder capacity, bladder compliance, p-mTOR protein expression in the bladder tissue of the electroacupuncture+AMPK inhibitor group were increased, the p-AMPK, LC3II, and Beclin1 protein expressions were decreased (all P<0.05). In the model group, the bladder became larger, with unclear and varying degrees of degeneration, severe tissue damage and autophagosome appeared; the bladder of the electroacupuncture group was smaller than that of the model group, and all levels were clearly visible with autophagy bodies; the layers were slightly disordered and damaged in the electroacupuncture + AMPK inhibitor group.
    CONCLUSIONS: Electroacupuncture can activate autophagy through AMPK/mTOR pathway, thereby reducing neurogenic urine retention caused by spinal cord injury.
    Keywords:  adenine monophosphate activated protein kinase; autophagy; electroacupuncture; mammalian target of rapamycin; neurogenic urine retention
    DOI:  https://doi.org/10.11817/j.issn.1672-7347.2022.210260
  4. FASEB J. 2022 May;36 Suppl 1
    Effects of a novel AMP-mimicking prodrug on AMPK and mTORC1 activity in C2C12 cells.
    Jared S Ashby, Tyson W Pace, Avery N Allen, Kody M Wilde, David A Galvan, Alyssa M Beitler, Chad R Hancock, Oliver Saunders, David M Thomson.
      The beneficial effects of exercise on skeletal muscle health and function are well-established. The intracellular signaling that mediates exercise's effects on muscle are undoubtedly complex and are not entirely understood. Nonetheless, increasing adenosine monophosphate (AMP) levels during muscle contraction signals a declining energy charge in the cell and this certainly drives many of the beneficial adaptations of exercise. AMP-activated protein kinase (AMPK) is one of AMP's best-defined targets and has been described as the cell's master energy regulator. Because it is activated by exercise, AMPK has gained recent attention as an attractive pharmacological target in harnessing at least some of the benefits of exercise for those who cannot exercise due to old age or other disorders. The prodrug AICAR effectively activates AMPK as it is taken up into the cell and phosphorylated to form ZMP, an AMP mimetic. However, AICAR's therapeutic potential is severely limited by its poor pharmacokinetics. Here, we describe a novel approach to ZMP delivery in a novel prodrug (Prodrug-39; P39). We found that similar to AICAR, P39 concentrations above ~400 μM effectively achieved significant AMPK activation and glycogen depletion. Surprisingly, given AMPK's well-documented inhibition of the mechanistic target of rapamycin (mTORC1), P39 was also effective at stimulating p70S6k and rpS6 phosphorylation concentrations as low as 10 uM and early in the timecourse after administration of P39 at higher concentrations. Given this evidence, we conclude that Prodrug-39 is indeed an effective AMPK activator and may also have some utility in mTOR activation at lower concentrations.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R5792
  5. FASEB J. 2022 May;36 Suppl 1
    Cardiac AMPK is not required for cardioprotection by metformin in pulmonary hypertension induced right heart failure.
    Ben D McNair, Samantha Shorthill, Sydney Polson, Ross Cook, Danielle Bruns.
      Right ventricle (RV) dysfunction dictates survival in pulmonary hypertension (PH) and other clinical conditions. Metformin, a clinically relevant AMP activated protein kinase (AMPK) activator, has been proposed as a therapy for PH induced right heart failure (PH-RHF) due to its well-reported cardioprotective properties. Indeed, our group has recently shown that metformin is protective in PH-RHF, but the mechanisms of protection in the RV remain unknown. The objective of this study was to use determine whether the protective properties of metformin require cardiac AMPK. We used Cre-Lox recombination to create a cardiomyocyte specific AMPK deleted with temporal regulation of expression. AMPK knockout (KO) and wild type (WT) control mice were placed in a hypobaric hypoxia (HH) chamber (~17,000ft) for 4 weeks to induce PH-RHF, with or without metformin (200mg/kg/day) in drinking water. Cardiomyocyte specific AMPK deletion exacerbated RV and LV remodeling in response to HH compared to WT, with significant sex differences evidenced by males undergoing worse remodeling than females. AMPK deletion exacerbated cardiac output and RV stroke volume compared to WT mice. Metformin rescued cardiac remodeling in male KO mice but did not attenuate remodeling in female mice. We suggest that metformin is a potential therapeutic target to attenuate maladaptive cardiac remodeling induced by PH-RHF, but only in male mice. Metformin does not require cardiac AMPK for cardioprotection. The mechanisms by which metformin affords protection in PH-RHF are still unclear, but may include AMPK activity in non-myocyte cell populations. Ongoing work will elucidate the role of AMPK activation in fibroblasts as well as other systemic and cardiac mechanisms which may explain how metformin protects against PH-RHF.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R5315
  6. FASEB J. 2022 May;36 Suppl 1
    The Effects of Metformin on Skeletal Muscle Differentiation and Satellite Cell Function in C2C12 Myocytes and Juvenile Lean and Obese Mice.
    Stephen Kinsey, Jess Ochab.
      AMP activated protein kinase (AMPK) functions as a cellular energy sensor and it is a target for the treatment of obesity and type II diabetes. Activation of AMPK has well-known benefits such as increased fuel oxidation and enhanced insulin sensitivity, however, it also may lead to compromised satellite cell function. This may be particularly detrimental to muscle development in juveniles, when satellite cell density and activity is high. In this study, the type II diabetic drug metformin was used to pharmacologically activate AMPK in C2C12 mouse skeletal muscle myoblasts and in juvenile lean and obese mice. In C2C12 cells 48 h of metformin treatment led to increased phosphorylation of AMPK and reduced mitochondrial respiration, as expected. Myocytes treated with metformin had a decreased expression of the myogenic markers, MyoD and myogenin, as well as a reduction in the satellite cell marker Pax7, and the myotubes were smaller in metformin treated cells. Further, nuclear recruitment was reduced in the metformin group, consistent with a reduction in satellite cell fusion with myotubes. In gastrocnemius from juvenile lean and obese mice, 6 weeks of metformin treatment had little effect on myogenic markers or on Pax7. However, nuclear recruitment was reduced in gastrocnemius from both lean and obese mice. These results support the hypothesis that AMPK activation with metformin in cultured muscle cells and juvenile lean and obese mice interferes with satellite cell fusion.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R6249
  7. FASEB J. 2022 May;36 Suppl 1
    In Vivo Effects of AICAR and Prodrug 39 (P39) on Anabolic Pathways Following Refeeding in Skeletal Muscle.
    Tyson Pace, Erik Marchant, Cameron Wilkinson, Jared S Ashby, David A Galvan, Joseph Rojas, Derek McClune, Oliver Saunders, Chad R Hancock, David M Thomson.
      Exercise training is known to elicit many beneficial adaptations. However, the mechanisms by which this happens are not entirely understood. During muscle contraction, increased intracellular AMP levels mediate many exercise-induced adaptations by activating AMP-activated protein kinase (AMPK), among other targets. AMP mimicry may, therefore, be an attractive pharmacological approach to stimulate some of exercise's benefits. The prodrug AICAR is a well-established AMPK activator when phosphorylated in the cell to form ZMP, an AMP mimetic. However, it has limited pharmacological potential due to poor pharmacokinetics. Prodrug 39 (P39) uses an alternative approach to deliver ZMP to the cell. The purpose of this study was to compare AMPK and P39 effects on blood glucose levels, AMPK activation, and anabolic signaling in the fasted and refed states. C57BL/6 mice were subjected to a 12 hour fast, followed by 1 hour of refeeding. Following the 12-hour fast, blood glucose was measured, and mice were injected with saline, AICAR or P39 (400 mg/kg BW). Mice were immediately transferred to cages with food, except for a fasted saline group. After 1 hour of refeeding, food was removed from cages. Blood glucose was measured 25 minutes later, and mice were sacrificed. Gastrocnemius-plantaris-soleus complex (GPS) muscles were quick-frozen for protein analysis. Fasting blood glucose levels were 80 ± 4 mg/dL prior to refeeding and were elevated to 144 ± 8 mg/dL (p<0.0001) after food reintroduction. Both AICAR and P39 caused a significant reduction in blood glucose compared to refed controls (p<0.0001, 54 ± 9, and 56 ± 10 mg/dL, respectively). In GPS muscle, AMPK phosphorylation status was unaffected by any treatment. However, ACC phosphorylation, a downstream target of AMPK, was ~3-fold higher in AICAR-treated animals compared to refed controls (p<0.01), with no significant effect of P39. Refeeding induced an increase in anabolic signaling, as indicated by an increase in S6 phosphorylation (~4-fold, p<0.01) and a trend for increased eIF4E-binding protein (4E-BP1) phosphorylation (p = 0.1). This increase was entirely prevented in AICAR and P39-treated animals. Overall, these results suggest that AICAR and P39 are similarly effective in lowering blood glucose and dampening anabolic signaling following refeeding. However, AICAR was more effective in activating AMPK signaling in skeletal muscle at this dose.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L7910
  8. FASEB J. 2022 May;36 Suppl 1
    Intestinal AMPK modulation of gut microbiota mediates cross-talk with brown fat to control metabolism.
    Wendong Huang, Eryun Zhang, Lihua Jin, Yangmeng Wang, Zhipeng Fang, Ismail Al-Abdullah, Sarah Shuck.
      The energy-dissipating capacity of brown adipose tissue through thermogenesis can be targeted to improve energy balance. Mammalian 5'-AMP-activated protein kinase (AMPK), a key nutrient sensor for maintaining cellular energy status, is a known therapeutic target for glucose control in Type II diabetes (T2D). Despite current understandings of its well-established roles in regulating glucose metabolism in various tissues, the functions of AMPK in the intestine, an organ for nutrient processing, remain largely unexplored. In this study, we generated the intestinal AMPKα1 specific knock out mice (IKO) and observed that the AMPKα1 IKO mice displayed impaired thermogenesis function of brown adipose tissue both in chow diet and high fat diet condition. Moreover, HFD-fed AMPKα1 IKO mice showed more severe impaired glucose tolerance and insulin resistance. Mechanistically, we revealed that intestinal AMPK can regulate the anti-microbial peptides (AMPs) Reg3 and result in a shift in the gut microbiota composition, which leading to the elevation of the fermentation products methylglyoxal, and methylglyoxal can significantly suppress the UCP1 expression in the brown adipose tissue. While transplantation of the microbiota from AMPKα1 IKO mice to antibiotic-induced microbiome depletion (AIMD) mice, the mice also showed impaired thermogenesis function of brown adipose tissue and elevated serum methylglyoxal level. In addition, we found that the metabolic effect and the AMP regulation of metformin is dependent on the intestinal AMPK. These results demonstrated that the intestinal nutrient sensor AMPKα1 can regulate the thermogenesis function of brown adipose tissue through modulating AMKP expression and reshaping the gut microbiome. These findings provide a new gut-brown adipose tissue axis mechanism for activating energy expenditure and treating metabolic diseases.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R4074
  9. Clin Transl Med. 2022 May;12(5): e854
    AMP-activated protein kinase α1 phosphorylates PHD2 to maintain systemic iron homeostasis.
    Cheng Wang, Wencheng Zhang, Wenjing Xu, Zhaoyu Liu, Kai Huang.
       BACKGROUND: Iron is essential for all mammalian life, and either a deficiency or excess of iron can cause diseases. AMP-activated protein kinase (AMPK) is a critical regulator of metabolic homeostasis; however, it has not been established whether AMPK regulates iron metabolism.
    METHODS: Iron, hepcidin and ferroportin levels were examined in mice with global and hepatocyte-specific knockout of AMPKα1 and AMPKα2. Primary AMPKα1 or AMPKα2 deleted hepatocytes were isolated and cultured in hypoxia condition to explore PHD2, HIF and hydroxylated HIF1α levels. We performed immunoprecipitation, in vitro AMPK kinase assay and site-direct mutant assay to detect phosphorylation sites of PHD2. We also obtained liver tissues from patients with anaemia of chronic disease undergoing surgery, AMPKα1 and hydroxylated HIF1α levels were measured by immunohistochemical analysis.
    RESULTS: We found that mice with global deficiency of AMPKα1, but not AMPKα2, exhibited hypoferraemia as well as iron sequestration in the spleen and liver. Hepatocyte-specific, but not myeloid-specific, ablation of AMPKα1 also reduced serum iron levels in association with increased hepcidin and decreased ferroportin protein levels. Mechanistically, AMPKα1 directly phosphorylated prolyl hydroxylase domain-containing (PHD)2 at serines 61 and 136, which suppressed PHD2-dependent hydroxylation of hypoxia-inducible factor (HIF)1α and subsequent regulation of hepatic hepcidin-related iron signalling. Inhibition of PHD2 hydroxylation ameliorated abnormal iron metabolism in hepatic AMPKα1-deficient mice. Furthermore, we found hepatic AMPKα/PHD2/HIFα/ hepcidin axes were highly clinically relevant to anaemia of chronic disease.
    CONCLUSION: In conclusion, these observations suggest that hepatic AMPKα1 has an essential role in maintaining iron homeostasis by PHD2-dependent regulation of hepcidin, thus providing a potentially promising approach for the treatment of iron disturbances in chronic diseases.
    Keywords:  AMPK; HIF1α; PHD2; hepcidin; iron
    DOI:  https://doi.org/10.1002/ctm2.854
  10. Trends Mol Med. 2022 May 07. pii: S1471-4914(22)00082-X. [Epub ahead of print]
    Combinatorial therapies for rescuing myotonic dystrophy type 1 skeletal muscle defects.
    Aymeric Ravel-Chapuis, Bernard J Jasmin.
      Myotonic dystrophy type 1 (DM1) is a multisystemic disorder for which there is no cure. In recent years, progress has been made in defining disease mechanisms and in developing novel therapies, especially for skeletal muscle defects. Here, we highlight the potential of activating AMP-activated protein kinase (AMPK) with different approaches in combinatorial therapies.
    Keywords:  AMP-activated protein kinase; combinatorial therapy; exercise; myotonic dystrophy; skeletal muscle
    DOI:  https://doi.org/10.1016/j.molmed.2022.04.004
  11. J Cell Sci. 2022 May 01. pii: jcs259209. [Epub ahead of print]135(9):
    Cilia-localized GID/CTLH ubiquitin ligase complex regulates protein homeostasis of sonic hedgehog signaling components.
    Friederike Hantel, Huaize Liu, Lisa Fechtner, Herbert Neuhaus, Jie Ding, Danilo Arlt, Peter Walentek, Pablo Villavicencio-Lorini, Christoph Gerhardt, Thomas Hollemann, Thorsten Pfirrmann.
      Cilia are evolutionarily conserved organelles that orchestrate a variety of signal transduction pathways, such as sonic hedgehog (SHH) signaling, during embryonic development. Our recent studies have shown that loss of GID ubiquitin ligase function results in aberrant AMP-activated protein kinase (AMPK) activation and elongated primary cilia, which suggests a functional connection to cilia. Here, we reveal that the GID complex is an integral part of the cilium required for primary cilia-dependent signal transduction and the maintenance of ciliary protein homeostasis. We show that GID complex subunits localize to cilia in both Xenopus laevis and NIH3T3 cells. Furthermore, we report SHH signaling pathway defects that are independent of AMPK and mechanistic target of rapamycin (MTOR) activation. Despite correct localization of SHH signaling components at the primary cilium and functional GLI3 processing, we find a prominent reduction of some SHH signaling components in the cilium and a significant decrease in SHH target gene expression. Since our data reveal a critical function of the GID complex at the primary cilium, and because suppression of GID function in X. laevis results in ciliopathy-like phenotypes, we suggest that GID subunits are candidate genes for human ciliopathies that coincide with defects in SHH signal transduction.
    Keywords:  AMPK; Ciliopathies; Cilium; GID complex; Sonic hedgehog; Ubiquitin
    DOI:  https://doi.org/10.1242/jcs.259209
  12. FASEB J. 2022 May;36 Suppl 1
    Chronic AICAR treatment enhances anabolic signaling in sarcopenic skeletal muscle.
    Andrew H Smith, Kole D Brodowski, Shalene H Wilcox, Tyler Darrington, David M Thomson.
      Many fields of study have grappled with the challenge of decelerating the effects of aging. Physical exercise plays a vital role in attenuating the impact of the aging process. While many exercise-induced chemical signaling pathways are not fully understood, AMP-activated protein kinase mediates many beneficial adaptations to exercise training. AMPK is activated by the accumulation of AMP (and ADP) during muscle contraction-induced ATP breakdown. The prodrug AICAR mimics AMP accumulation and leads to AMPK activation by increasing the concentration of the AMP analog, ZMP. The primary purpose of the present study is to determine the effect of chronic (31 days) AICAR treatment on muscle anabolic signaling in old (24 mo.) mice. AICAR improved treadmill running performance and attenuated muscle atrophy in the aged mice. Surprisingly, given it's known acute effects on muscle anabolism, AICAR treatment increased the skeletal muscle concentrations of insulin-like growth factor-1 and apelin, both of which are known to activate anabolic signaling through the mechanistic target of rapamycin complex 1 (mTORC1). Accordingly, ribosomal protein S6 phosphorylation was also elevated in AICAR-treated muscles. Our results suggest that in addition to its other well-established metabolic effects, chronic AICAR treatment may enhance anabolism in sarcopenic muscle and potentially in other muscle-wasting conditions.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L7901
  13. FASEB J. 2022 May;36 Suppl 1
    Anti-obesity effect of Premature Citrus Extract in high-fat diet induced obesity mice.
    Premkumar Natraj, Priyanka Rajan, Sachithra S Ranaweera, Lakshi A Dayarathne, Chang-Hoon Han.
       BACKGROUND: Obesity is a worldwide health threat. Premature citrus extract (PCE) are known to contain high concentrations of flavonoids, polyphenols, and organic acids with health functions. The current study aimed to investigate the anti-obesity effect of PCE on a high-fat diet (HFD)-induced obesity mice.
    METHOD: Mice were fed on chow or HFD with or without Orlistat (ORL), Hesperidin (HES), and PCE treatment for 10 weeks. Triglyceride (TG) content and total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-c), and high-density lipoprotein cholesterol (HDL-c) levels were determined by TG, HDL and LDL Assay Kit (BioAssay System). The phosphorylation levels of AMP-activated protein kinase (AMPK) and acetyl Co-A carboxylase (ACC); the expression of sterol regulatory element binding protein-2 (SREBP-2), 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), Acyl-coenzyme A: cholesterol acyltransferases (ACAT-1) and mitochondrial uncoupling proteins (UCP-3) were observed via western blot analysis.
    RESULTS: PCE and HES administered group significantly reduced the weights of body, liver, and white adipose tissue compared to the HFD group. Supplementation with PCE and HES significantly decreased the levels of TG, TC, and (LDL-c), and increased (HDL-c) levels in serum compared to the HFD group. Furthermore, treatment with PCE and HES significantly decreased the levels of TG and TC in the liver whereas increased the levels of TG and TC in feces compared to the HFD group. Western blot analysis revealed that PCE and HES increased the phosphorylation levels of both AMPK and ACC and significantly decreased the expression levels of SREBP-2, HMGCR, and ACAT-1 in the liver compared to the HFD group. In addition, both PCE and HES increased the expression level of UCP-3 in skeletal muscle tissue.
    CONCLUSION: Overall results suggest that PCE and HES exerts an anti-obesity effect via activation of AMPK and associated signaling cascades related to lipid metabolism in HFD-induced obesity mice.
    Keywords:  AMPK; Anti-obesity; HFD-induced obesity mice; Hesperidin; Premature citrus extract
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.0R745
  14. Int J Mol Sci. 2022 Apr 30. pii: 5020. [Epub ahead of print]23(9):
    Ethyl Gallate Dual-Targeting PTPN6 and PPARγ Shows Anti-Diabetic and Anti-Obese Effects.
    Dohee Ahn, Jinsoo Kim, Gibeom Nam, Xiaodi Zhao, Jihee Kwon, Ji Young Hwang, Jae Kwan Kim, Sun-Young Yoon, Sang J Chung.
      The emergence of the high correlation between type 2 diabetes and obesity with complicated conditions has led to the coinage of the term "diabesity". AMP-activated protein kinase (AMPK) activators and peroxisome proliferator-activated receptor (PPARγ) antagonists have shown therapeutic activity for diabesity, respectively. Hence, the discovery of compounds that activate AMPK as well as antagonize PPARγ may lead to the discovery of novel therapeutic agents for diabesity. In this study, the knockdown of PTPN6 activated AMPK and suppressed adipogenesis in 3T3-L1 cells. By screening a library of 1033 natural products against PTPN6, we found ethyl gallate to be the most selective inhibitor of PTPN6 (Ki = 3.4 μM). Subsequent assay identified ethyl gallate as the best PPARγ antagonist (IC50 = 5.4 μM) among the hit compounds inhibiting PTPN6. Ethyl gallate upregulated glucose uptake and downregulated adipogenesis in 3T3-L1 cells as anticipated. These results strongly suggest that ethyl gallate, which targets both PTPN6 and PPARγ, is a potent therapeutic candidate to combat diabesity.
    Keywords:  3T3-L1 adipocyte; AMPK; PPARγ antagonist; PTPN6; adipogenesis; ethyl gallate; natural product
    DOI:  https://doi.org/10.3390/ijms23095020
  15. FASEB J. 2022 May;36 Suppl 1
    Chronic AICAR treatment enhances anabolic signaling in sarcopenic skeletal muscle.
    Jouber Calixto, Connor Johnson, Andrew Smith, Kole Brodowski, Shalene H Wilcox, Jonathon T Hill, David M Thomson.
      Many fields of study have grappled with the challenge of decelerating the effects of aging. Physical exercise plays a vital role in attenuating the impact of the aging process. While many exercise-induced chemical signaling pathways are not fully understood, AMP-activated protein kinase mediates many beneficial adaptations to exercise training. AMPK is activated by the accumulation of AMP (and ADP) during muscle contraction-induced ATP breakdown. The prodrug AICAR mimics AMP accumulation and leads to AMPK activation by increasing the concentration of the AMP analog, ZMP. The primary purpose of the present study is to determine the effect of chronic (31 days) AICAR treatment on muscle anabolic signaling in old (24 mo.) mice. AICAR improved treadmill running performance and attenuated muscle atrophy in the aged mice. Surprisingly, given AICAR's known acute effects on anabolic signaling, AICAR treatment increased the skeletal muscle concentrations of insulin-like growth factor-1 and apelin, both of which are known to activate anabolic signaling through the mechanistic target of rapamycin complex 1 (mTORC1). Accordingly, ribosomal protein S6 phosphorylation was also elevated in AICAR-treated muscles. Our results suggest that in addition to its other well-established metabolic effects, chronic AICAR treatment may enhance anabolism in sarcopenic muscle and potentially in other muscle-wasting conditions.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R5697
  16. Mol Metab. 2022 May 10. pii: S2212-8778(22)00083-7. [Epub ahead of print] 101514
    Defective AMPK regulation of cholesterol metabolism accelerates atherosclerosis by promoting HSPC mobilization and myelopoiesis.
    Man K S Lee, Olivia D Cooney, Xuzhu Lin, Shaktypreya Nadarajah, Dragana Dragoljevic, Kevin Huynh, Danise-Ann Onda, Sandra Galic, Peter J Meikle, Thomas Edlund, Morgan D Fullerton, Bruce E Kemp, Andrew J Murphy, Kim Loh.
       OBJECTIVES: Dysregulation of cholesterol metabolism in the liver and hematopoietic stem and progenitor cells (HSPCs) promotes atherosclerosis development. Previously, it has been shown that HMG-CoA-Reductase (HMGCR), the rate-limiting enzyme in the mevalonate pathway, can be phosphorylated and inactivated by the metabolic stress sensor AMP-activated protein kinase (AMPK). However, the physiological significance of AMPK regulation of HMGCR to atherogenesis has yet to be elucidated. The aim of this study was to determine the role of AMPK/HMGCR axis in the development of atherosclerosis.
    METHODS: We have generated a novel atherosclerotic-prone mouse model with defects in the AMPK regulation of HMGCR (Apoe-/-/Hmgcr KI mice). Atherosclerotic lesion size, plaque composition, immune cell and lipid profiles were assessed in Apoe-/- and Apoe-/-/Hmgcr KI mice.
    RESULTS: In this study, we showed that both male and female atherosclerotic-prone mice with a disruption of HMGCR regulation by AMPK (Apoe-/-/Hmgcr KI mice) display increased aortic lesion size concomitant with an increase in plaque-associated macrophages and lipid accumulation. Consistent with this, Apoe-/-/Hmgcr KI mice exhibited an increase in total circulating cholesterol and atherogenic monocytes, Ly6-Chi subset. Mechanistically, increased circulating atherogenic monocytes in Apoe-/-/Hmgcr KI mice was associated with enhanced egress of bone marrow HSPCs and extramedullary myelopoiesis, driven by a combination of elevated circulating 27-hydroxycholesterol and intracellular cholesterol in HSPCs.
    CONCLUSIONS: Our results uncovered a novel signalling pathway involving AMPK-HMGCR axis in the regulation of cholesterol homeostasis in HSPCs, and that inhibition of this regulatory mechanism accelerates the development and progression of atherosclerosis. These findings provide a molecular basis to support the use of AMPK activators that currently undergoing Phase II clinical trial such as O-3O4 and PXL 770 for reducing atherosclerotic cardiovascular disease risks.
    Keywords:  AMPK; HMG-CoA Reductase; HSPCs; atherosclerosis; cholesterol
    DOI:  https://doi.org/10.1016/j.molmet.2022.101514
  17. Virulence. 2022 May 09.
    Guanylate-binding protein 2b regulates the AMPK/mTOR/ULK1 signalling pathway to induce autophagy during Mycobacterium bovis infection.
    Youli Yu, Jialiang Pan, Mengting Liu, Haiqin Jiang, Jingshu Xiong, Lei Tao, Feng Xue, Fang Tang, Hongsheng Wang, Jianjun Dai.
      Mycobacterium bovis is a major cause of animal tuberculosis that is also highly dangerous to human health. Autophagic isolation and degradation of intracellular pathogens are employed by host cells as primary innate immune defence mechanisms to control intercellular M. bovis infection. In this study, RNA-Seq technology was used to obtain the total mRNA from bone marrow-derived macrophages (BMDMs) infected with M. bovis at 6 and 24 h after infection. One of the differential genes, GBP2b, was also investigated. Analysis of the significant pathway involved in GBP2b-coexpressed mRNA demonstrated that GBP2b was associated with autophagy and autophagy-related mammalian target of rapamycin (mTOR) signalling and AMP-activated protein kinase (AMPK) signalling. The results of in vivo and in vitro experiments showed significant up-regulation of GBP2b during M. bovis infection. For in vitro validation, small interfering RNA-GBP2b plasmids were transfected into BMDMs and RAW264.7 cells lines to down-regulate the expression of GBP2b. The results showed that the down-regulation of GBP2b impaired autophagy via the AMPK/mTOR/ULK1 pathway. Further studies revealed that the activation of AMPK signalling was essential for the regulation of autophagy during M. bovis infection, and the down-regulation of GBP2b promoted the intracellular survival of M. bovis. These findings were validated on two types of macrophages, which extended the knowledge about the involvement of GBP2b in the autophagic process. Based on these observations, GBP2b should be developed as a promising molecular target for intervening on host-pathogen interactions to develop novel therapeutic strategies to control M. bovis infections in humans and animals.
    Keywords:  AMPK; GBP2b; Mycobacterium bovis; RNA-Seq; ULK1; autophagy; mTOR
    DOI:  https://doi.org/10.1080/21505594.2022.2073024
  18. Int J Mol Sci. 2022 Apr 23. pii: 4691. [Epub ahead of print]23(9):
    Effect of Long-Term Supplementation with Acetic Acid on the Skeletal Muscle of Aging Sprague Dawley Rats.
    Hitomi Maruta, Reina Abe, Hiromi Yamashita.
      Mitochondrial function in skeletal muscle, which plays an essential role in oxidative capacity and physical activity, declines with aging. Acetic acid activates AMP-activated protein kinase (AMPK), which plays a key role in the regulation of whole-body energy by phosphorylating key metabolic enzymes in both biosynthetic and oxidative pathways and stimulates gene expression associated with slow-twitch fibers and mitochondria in skeletal muscle cells. In this study, we investigate whether long-term supplementation with acetic acid improves age-related changes in the skeletal muscle of aging rats in association with the activation of AMPK. Male Sprague Dawley (SD) rats were administered acetic acid orally from 37 to 56 weeks of age. Long-term supplementation with acetic acid decreased the expression of atrophy-related genes, such as atrogin-1, muscle RING-finger protein-1 (MuRF1), and transforming growth factor beta (TGF-β), activated AMPK, and affected the proliferation of mitochondria and type I fiber-related molecules in muscles. The findings suggest that acetic acid exhibits an anti-aging function in the skeletal muscles of aging rats.
    Keywords:  AMPK; acetic acid; aging; mitochondria; skeletal muscle; type I fiber
    DOI:  https://doi.org/10.3390/ijms23094691
  19. Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2022 Jan 28. pii: 1672-7347(2022)01-0008-10. [Epub ahead of print]47(1): 8-17
    Leptin relieves ischemia/reperfusion induced acute kidney injury through inhibiting apoptosis and autophagy.
    Siyao Li, Kaiting Zhuang, Yi He, Yunzhen Deng, Jing Xi, Junxiang Chen.
       OBJECTIVES: Acute kidney injury (AKI) can be caused by ischemia/reperfusion (I/R), nephrotoxin, and sepsis, with poor prognosis and high mortality. Leptin is a protein molecule that regulates the body's energy metabolism and reproductive activities via binding to its specific receptor. Leptin can inhibit cardiomyocyte apoptosis caused by I/R, but its effect on I/R kidney injury and the underlying mechanisms are still unclear. This study aims to investigate the effect and mechanisms of leptin on renal function, renal histopathology, apoptosis, and autophagy during acute I/R kidney injury.
    METHODS: Healthy adult male mice were randomly divided into 4 groups: a sham+wild-type mice (ob/+) group, a sham+leptin gene-deficient mice (ob/ob) group, an I/R+ob/+ group, and an I/R+ob/ob group (n=8 per group). For sham operation, a longitudinal incision was made on the back of the mice to expose and separate the bilateral kidneys and renal arteries, and no subsequent treatment was performed. I/R treatment was ischemia for 30 min and reperfusion for 48 h. The levels of BUN and SCr were detected to evaluate renal function; HE staining was used to observe the pathological changes of renal tissue; TUNEL staining was used to observe cell apoptosis, and apoptosis-positive cells were counted; Western blotting was used to detect levels of apoptosis-related proteins (caspase 3, caspase 9), autophagy-related proteins [mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), LC3 I, LC3 II], mTOR-dependent signaling pathway proteins [phosphate and tension homology (PTEN), adenosine monophosphate-activated protein kinase (AMPK), protein kinase B (AKT), extracellular regulated protein kinase (ERK), phosphorylated PTEN (p-PTEN), phosphorylated AMPK (p-AMPK), phosphorylated AKT (p-AKT), phosphorylated ERK (p-ERK)].
    RESULTS: There was no significant difference in the levels of BUN and SCr between the sham+ob/+ group and the sham+ob/ob group (both P>0.05). The levels of BUN and SCr in the I/R+ob/+ group were significantly higher than those in the sham+ob/+ group (both P<0.05). Compared with the mice in the sham+ob/ob group or the I/R+ob/+ group, the levels of BUN and SCr in the I/R+ob/ob group were significantly increased (all P<0.05). There was no obvious damage to the renal tubules in the sham+ob/+ group and the sham+ob/ob group. Compared with sham+ob/+ group and sham+ob/ob group, both the I/R+ob/+ group and the I/R+ob/ob group had cell damage such as brush border shedding, vacuolar degeneration, and cast formation. Compared with the I/R+ob/+ group, the renal tubules of the mice in the I/R+ob/ob group were more severely damaged. The pathological score of renal tubular injury showed that the renal tubular injury was the most serious in the I/R+ob/ob group (P<0.05). Compared with the sham+ob/+ group, the protein levels of caspase 3, caspase 9, PTEN, and LC3 II were significantly up-regulated, the ratio of LC3 II to LC3 I was significantly increased, and the protein levels of p-mTOR, p-PTEN, p-AMPK, p-AKT, and p-ERK were significantly down-regulated in the I/R+ob/+ group (all P<0.05). Compared with the sham+ob/ob group, the protein levels of caspase 3, caspase 9, PTEN, and LC3 II were significantly up-regulated, and the ratio of LC3 II to LC3 I was significantly increased, while the protein levels of p-mTOR, p-PTEN, p-AMPK, p-AKT, and p-ERK were significantly down-regulated in the I/R+ob/ob group (all P<0.05). Compared with the I/R+ob/+ group, the levels of p-mTOR, p-PTEN, p-AMPK, p-AKT were more significantly down-regulated, while the levels of caspase 3, caspase 9, PTEN, and LC3 II were more significantly up-regulated, and the ratio of LC3 II to LC3 I was more significantly increase in the I/R+ob/ob group (all P<0.05).
    CONCLUSIONS: Renal function and tubular damage, and elevated levels of apoptosis and autophagy are observed in mice kidneys after acute I/R. Leptin might relieve I/R induced AKI by inhibiting apoptosis and autophagy that through a complex network of interactions between mTOR-dependent signaling pathways.
    Keywords:  acute kidney injury; apoptosis; autophagy; ischemia/reperfusion; leptin
    DOI:  https://doi.org/10.11817/j.issn.1672-7347.2022.210244
  20. Front Pharmacol. 2022 ;13 828890
    Bilirubin Improves Gap Junction to Alleviate Doxorubicin-Induced Cardiotoxicity by Regulating AMPK-Axl-SOCS3-Cx43 Axis.
    Siqi Zhang, Yixin Fan, Binbin Zheng, Yu Wang, Chen Miao, Yue Su, Kun Li, Yan E, Xueli Wang, Xueming He, Xuefeng Wu, Chenjie Xu, Yulin Tang, Wen-Tao Liu, Xiangqing Kong, Liang Hu.
      Doxorubicin induces severe cardiotoxicity, accompanied by the high level of bilirubin in the blood. The conventional wisdom is that bilirubin is considered as a marker of liver damage. By contrast, here we aim to explore the potential protective effect of bilirubin on doxorubicin-induced cardiotoxicity, and investigate the mechanism for drug development. Doxorubicin was used to establish cardiotoxicity model in vitro and in vivo. The electrocardiogram (ECG), echocardiography and molecular biological methods were used to detect the effects of bilirubin on doxorubicin-induced cardiotoxicity. Consecutive intraperitoneal injection of bilirubin for 7 days significantly attenuated doxorubicin-induced arrhythmia, prolonged survival time and reduced the levels of aspartate aminotransferase (AST), lactate dehydrogenase (LDH), creatine kinase MB (CK-MB) and α-hydroxybutyrate dehydrogenase (α-HBDH) in mice. Bilirubin also markedly inhibited doxorubicin-induced phosphorylation of c-Jun N-terminal kinase (JNK) and connexin 43 (Cx43), and improved gap junction function in vitro and in vivo. In addition, bilirubin activated adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) and induced suppressor of cytokine signaling 3 (SOCS3) expression, which was abolished by Axl inhibition. Moreover, pretreatment with AMPK agonist or AMPK inhibitor could mimic or abolish the cardioprotective effect of bilirubin on H9C2 cells in vitro, respectively. Altogether, bilirubin upregulates gap junctions' function to protect against doxorubicin-induced cardiotoxicity by activating AMPK-Axl-SOCS3 signaling axis. We enrich the physiological function of bilirubin, and provide theoretical support for drug development.
    Keywords:  AMPK; Cx43 gap junction; SOCS3; bilirubin; cardiotoxicity; doxorubicin
    DOI:  https://doi.org/10.3389/fphar.2022.828890
  21. Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2022 Feb 28. pii: 1672-7347(2022)02-0165-09. [Epub ahead of print]47(2): 165-173
    Relationship of PI3K-Akt/mTOR/AMPK signaling pathway genetic mutation with efficacy and prognosis in nasopharyngeal carcinoma.
    Yanzhu Chen, Qian He, Hongzhi Ma, Lin Zhang, Feng Liu, Yaqian Han.
       OBJECTIVES: Genetic mutation is one of the important causes for tumor genesis and development, but genetic mutation in nasopharyngeal carcinoma (NPC) has rarely been reported. This study explored the role of phosphatidylinositol 3 kinase-protein kinase B (PI3K-Akt), mammalian target of rapamycin (mTOR), and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) signaling pathway in the efficacy and prognosis in patients with NPC.
    METHODS: A total of 31 patients with advanced NPC, who came from the Affiliated Cancer Hospital of Xiangya School of Medicine of Central South University/Hunan Provincial Cancer Hospital, were enrolled. All of the exons of 288 genes, introns of 38 genes and promoters or fusion breakpoint regions from the nasopharyngeal biopsy tissues before treatment were detected by the gene sequencing platform Illumina NextSeq CN500. The coding regions of 728 genes were carried out a high-depth sequencing of target region capture, and the 4 variant types of tumor genes (including point mutations, insertion deletions of small fragments, copy number variations, and currently known fusion genes) were detected. All of 31 patients received platinum-based induction chemotherapy combined with concurrent chemoradiotherapy and were followed up for a long time.
    RESULTS: The 3-year regional failure-free survival (RFFS) and disease-free survival (DFS) in patients with PI3K-Akt pathway mutation were significantly lower than those in unmutated patients (χ2=6.647, P<0.05). The 3-year RFFS and DFS in patients with mTOR pathway mutations were significantly lower than those in unmutated patients, and there was significant difference (χ2=5.570, P<0.05). The rate of complete response (CR) in patients with unmutated AMPK pathway was significantly higher than that in patients with mutation at 3 months after treatment (P<0.05), and the 3-year RFFS and DFS in patients with AMPK pathway mutation were significantly lower than those in unmutated patients (χ2=4.553, P<0.05). PI3K-Akt/mTOR/AMPK signaling pathway mutations and pre-treatment EB virus DNA copy numbers were independent prognostic factors for 3-year RFFS and DFS in patients with NPC (both P<0.05).
    CONCLUSIONS: The NPC patients with PI3K-Akt/mTOR/AMPK signaling pathway mutation have poor prognosis, and the detection of PI3K-Akt, mTOR, AMPK driver genes and signaling pathways by next-generation sequencing is expected to provide new idea for basic research and targeted therapy of NPC.
    Keywords:  PI3K-Akt/mTOR/AMPK signaling pathway; efficacy; nasopharyngeal carcinoma; next-generation sequencing; prognosis
    DOI:  https://doi.org/10.11817/j.issn.1672-7347.2022.200821
  22. Am J Chin Med. 2022 May 05. 1-21
    Salidroside Ameliorates Ischemia-Induced Neuronal Injury through AMPK Dependent and Independent Pathways to Maintain Mitochondrial Quality Control.
    Bin Wen, Keru Zhou, Caiyin Hu, Jiehui Chen, Kai Xu, Tao Liang, Benhong He, Ling Chen, Juan Chen.
      Salidroside, an active ingredient in Rhodiola rosea, has potent protective activity against cerebral ischemia. However, the mechanisms underlying its pharmacological actions are poorly understood. In this study, we employed a mouse middle cerebral artery occlusion (MCAO) and cellular oxygen and glucose deprivation (OGD) models to test the hypothesis that salidroside may restore mitochondrial quality control in neurons by modulating the relevant signaling. The results indicated that salidroside mitigated almost 40% the ischemia-induced brain infarct volumes in mice and the OGD-decreased viability of neurons to ameliorate the mitochondrial functions. Furthermore, salidroside treatment alleviated the OGD- or ischemia-induced imbalance of mitochondrial fission and fusion, mitophagy and promoted mitochondrial biogenesis in neurons by attenuating the AMPK activity. Moreover, salidroside alleviated 50% the OGD-promoted mitochondrial calcium fluorescence intensity and 5% mitochondria-associated membrane (MAM) area by down-regulating GRP75 expression independent of the AMPK signaling. Finally, similar findings were achieved in primary mouse neurons. Collectively, these data indicate that salidroside effectively restores the mitochondria dynamics, facilitates mitochondrial biogenesis by attenuating the AMPK signaling, and maintains calcium homeostasis in neurons independent of the AMPK activity.
    Keywords:  AMPK Signaling; Cerebral Ischemia; Mice; Mitochondrial Quality Control; Salidroside
    DOI:  https://doi.org/10.1142/S0192415X2250046X
  23. Physiol Rep. 2022 May;10(9): e15297
    Combined effects of exercise training and D-allulose intake on endurance capacity in mice.
    Takamasa Tsuzuki, Ryo Suzuki, Risa Kajun, Takako Yamada, Tetsuo Iida, Bingyang Liu, Teruhiko Koike, Yukiyasu Toyoda, Takayuki Negishi, Kazunori Yukawa.
      This study investigated the combined effects of exercise training and D-allulose intake on endurance capacity in mice. Male C57BL/6J mice were fed either a control diet (Con) or a 3% D-allulose diet (Allu) and further divided into the sedentary (Sed) or exercise training (Ex) groups (Con-Sed, Con-Ex, Allu-Sed, Allu-Ex, respectively; n = 6-7/group). The mice in the Ex groups were trained on a motor-driven treadmill 5 days/week for 4 weeks (15-18 m/min, 60 min). After the exercise training period, all mice underwent an exhaustive running test to assess their endurance capacity. At 48 h after the running test, the mice in the Ex groups were subjected to run at 18 m/min for 60 min again. Then the gastrocnemius muscle and liver were sampled immediately after the exercise bout. The running time until exhaustion tended to be higher in the Allu-Ex than in the Con-Ex group (p = 0.08). The muscle glycogen content was significantly lower in the Con-Ex than in the Con-Sed group and was significantly higher in the Allu-Ex than in the Con-Ex group (p < 0.05). Moreover, exercise training increased the phosphorylation levels of adenosine monophosphate-activated protein kinase (AMPK) in the muscle and liver. The phosphorylation levels of acetyl coenzyme A carboxylase (ACC), a downstream of AMPK, in the muscle and liver were significantly higher in the Allu-Ex than in the Con-Sed group (p < 0.05), suggesting that the combination of exercise training and D-allulose might have activated the AMPK-ACC signaling pathway, which is associated with fatty acid oxidation in the muscle and liver. Taken together, our data suggested the combination of exercise training and D-allulose intake as an effective strategy to upregulate endurance capacity in mice. This may be associated with sparing glycogen content and enhancing activation of AMPK-ACC signaling in the skeletal muscle.
    Keywords:  endurance capacity; exercise training; glycogen; rare sugar
    DOI:  https://doi.org/10.14814/phy2.15297
  24. Nutrients. 2022 Apr 30. pii: 1898. [Epub ahead of print]14(9):
    Distinct AMPK-Mediated FAS/HSL Pathway Is Implicated in the Alleviating Effect of Nuciferine on Obesity and Hepatic Steatosis in HFD-Fed Mice.
    Hanyuan Xu, Xiaorui Lyu, Xiaonan Guo, Hongbo Yang, Lian Duan, Huijuan Zhu, Hui Pan, Fengying Gong, Linjie Wang.
      Nuciferine (Nuci), the main aporphine alkaloid component in lotus leaf, was reported to reduce lipid accumulation in vitro. Herein we investigated whether Nuci prevents obesity in high fat diet (HFD)-fed mice and the underlying mechanism in liver/HepG2 hepatocytes and epididymal white adipose tissue (eWAT) /adipocytes. Male C57BL/6J mice were fed with HFD supplemented with Nuci (0.10%) for 12 weeks. We found that Nuci significantly reduced body weight and fat mass, improved glycolipid profiles, and enhanced energy expenditure in HFD-fed mice. Nuci also ameliorated hepatic steatosis and decreased the size of adipocytes. Furthermore, Nuci remarkably promoted the phosphorylation of AMPK, suppressed lipogenesis (SREBP1, FAS, ACC), promoted lipolysis (HSL, ATGL), and increased the expressions of adipokines (FGF21, ZAG) in liver and eWAT. Besides, fatty acid oxidation in liver and thermogenesis in eWAT were also activated by Nuci. Similar results were further observed at cellular level, and these beneficial effects of Nuci in cells were abolished by an effective AMPK inhibitor compound C. In conclusion, Nuci supplementation prevented HFD-induced obesity, attenuated hepatic steatosis, and reduced lipid accumulation in liver/hepatocytes and eWAT/adipocytes through regulating AMPK-mediated FAS/HSL pathway. Our findings provide novel insight into the clinical application of Nuci in treating obesity and related complications.
    Keywords:  AMP-activated protein kinase (AMPK); Nuciferine (Nuci); fatty acid synthase (FAS); hepatic steatosis; hormone sensitive lipase (HSL); obesity
    DOI:  https://doi.org/10.3390/nu14091898
  25. FASEB J. 2022 May;36 Suppl 1
    Heat stress-induced changes in the myocardium.
    Melissa Roths, Tori Rudolph, Alyssa Freestone, Lance Baumgard, Joshua Selsby.
      Prolonged exposure to heat can lead to environment-induced heat stress (EIHS), which may be a threat to human health. How EIHS affects cardiac morphology and the myocardium are unknown. We hypothesized that EIHS would alter cardiac morphology and cause cellular dysfunction. To test this hypothesis, crossbred female pigs were exposed to thermoneutral (TN; 20.6 ± 0.2 ºC; n=8) or EIHS (37.4 ± 0.2 ºC; n=8) conditions for 24 h. Rectal temperature (RT), skin temperature (ST), and respiratory rate (RR) were recorded every 4 h during the environmental challenge. Pigs were euthanized following the environmental challenge and hearts were collected. Hearts were weighed and heart length (apex to base), width (left/right dimension), and left ventricle (LV) and right ventricle (RV) wall thickness were measured with calipers. Portions of LV and RV were lyophilized to measure tissue water content or homogenized for protein extraction and western blotting. Environment-induced heat stress increased RT by 1.3 ºC (p<0.01), ST by 11 °C (p<0.01) and RR by 72 breaths per minute (p<0.01) compared to TN. Heart weight tended to be decreased (7.6%; p=0.07) and heart length was decreased (8.5%; p=0.01) by EIHS, but heart width was similar between groups. Left ventricle wall thickness was increased (22%; p=0.02) and RV thickness was decreased (26%; p=0.04) in EIHS compared to TN. Water content in the RV was similar between groups, however, in LV it was increased (8.6%; p<0.01) in EIHS compared to TN, suggesting edema contributed to increased LV thickness. Lastly, using a western blot approach we discovered that pathways regulating energy homeostasis were impacted by EIHS and sometimes differed between RV and LV. In RV and LV, phosphorylation (p) of AMP-activated protein kinase (AMPK) was decreased by EIHS (RV - 73%, p<0.02; LV - 54%, p=0.04), and in the RV was accompanied by increased protein phosphate type 2A (PP2A; 15.5%, p<0.01), which regulates pAMPK, and decreased p-Acetyl-CoA carboxylase (ACC; 40%, p<0.01), a client protein of pAMPK, whereas these were similar in LV. In RV, mTOR signaling appeared to be blunted, mitochondrial content increased, and markers of mitophagy increased by EIHS compared to TN, however, LV was resistant to these changes. In total, these data demonstrate that a single bout of EIHS caused cardiac morphological changes and biochemical changes in the myocardium and that EIHS affects the LV and RV differently.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R4526
  26. Nutrients. 2022 Apr 26. pii: 1804. [Epub ahead of print]14(9):
    Hempseed (Cannabis sativa) Peptide H3 (IGFLIIWV) Exerts Cholesterol-Lowering Effects in Human Hepatic Cell Line.
    Jianqiang Li, Carlotta Bollati, Martina Bartolomei, Angelica Mazzolari, Anna Arnoldi, Giulio Vistoli, Carmen Lammi.
      Hempseed (Cannabis sativa) protein is an important source of bioactive peptides. H3 (IGFLIIWV), a transepithelial transported intestinal peptide obtained from the hydrolysis of hempseed protein with pepsin, carries out antioxidant and anti-inflammatory activities in HepG2 cells. In this study, the main aim was to assess its hypocholesterolemic effects at a cellular level and the mechanisms behind this health-promoting activity. The results showed that peptide H3 inhibited the 3-hydroxy-3-methylglutaryl co-enzyme A reductase (HMGCoAR) activity in vitro in a dose-dependent manner with an IC50 value of 59 μM. Furthermore, the activation of the sterol regulatory element binding proteins (SREBP)-2 transcription factor, followed by the increase of low-density lipoprotein (LDL) receptor (LDLR) protein levels, was observed in human hepatic HepG2 cells treated with peptide H3 at 25 µM. Meanwhile, peptide H3 regulated the intracellular HMGCoAR activity through the increase of its phosphorylation by the activation of AMP-activated protein kinase (AMPK)-pathways. Consequently, the augmentation of the LDLR localized on the cellular membranes led to the improved ability of HepG2 cells to uptake extracellular LDL with a positive effect on cholesterol levels. Unlike the complete hempseed hydrolysate (HP), peptide H3 can reduce the proprotein convertase subtilisin/kexin 9 (PCSK9) protein levels and its secretion in the extracellular environment via the decrease of hepatic nuclear factor 1-α (HNF1-α). Considering all these evidences, H3 may represent a new bioactive peptide to be used for the development of dietary supplements and/or peptidomimetics for cardiovascular disease (CVD) prevention.
    Keywords:  LDLR; PCSK9; SREBP-2; cholesterol metabolism; hempseed protein
    DOI:  https://doi.org/10.3390/nu14091804
  27. Int J Mol Sci. 2022 Apr 26. pii: 4776. [Epub ahead of print]23(9):
    Targeting Sirt1, AMPK, Nrf2, CK2, and Soluble Guanylate Cyclase with Nutraceuticals: A Practical Strategy for Preserving Bone Mass.
    Mark F McCarty, Lidianys Lewis Lujan, Simon Iloki Assanga.
      There is a vast pre-clinical literature suggesting that certain nutraceuticals have the potential to aid the preservation of bone mass in the context of estrogen withdrawal, glucocorticoid treatment, chronic inflammation, or aging. In an effort to bring some logical clarity to these findings, the signaling pathways regulating osteoblast, osteocyte, and osteoclast induction, activity, and survival are briefly reviewed in the present study. The focus is placed on the following factors: the mechanisms that induce and activate the RUNX2 transcription factor, a key driver of osteoblast differentiation and function; the promotion of autophagy and prevention of apoptosis in osteoblasts/osteoclasts; and the induction and activation of NFATc1, which promotes the expression of many proteins required for osteoclast-mediated osteolysis. This analysis suggests that the activation of sirtuin 1 (Sirt1), AMP-activated protein kinase (AMPK), the Nrf2 transcription factor, and soluble guanylate cyclase (sGC) can be expected to aid the maintenance of bone mass, whereas the inhibition of the serine kinase CK2 should also be protective in this regard. Fortuitously, nutraceuticals are available to address each of these targets. Sirt1 activation can be promoted with ferulic acid, N1-methylnicotinamide, melatonin, nicotinamide riboside, glucosamine, and thymoquinone. Berberine, such as the drug metformin, is a clinically useful activator of AMPK. Many agents, including lipoic acid, melatonin, thymoquinone, astaxanthin, and crucifera-derived sulforaphane, can promote Nrf2 activity. Pharmacological doses of biotin can directly stimulate sGC. Additionally, certain flavonols, notably quercetin, can inhibit CK2 in high nanomolar concentrations that may be clinically relevant. Many, though not all, of these agents have shown favorable effects on bone density and structure in rodent models of bone loss. Complex nutraceutical regimens providing a selection of these nutraceuticals in clinically meaningful doses may have an important potential for preserving bone health. Concurrent supplementation with taurine, N-acetylcysteine, vitamins D and K2, and minerals, including magnesium, zinc, and manganese, plus a diet naturally high in potassium, may also be helpful in this regard.
    Keywords:  AMPK; NFATc1; Nrf2; RUNX2; Sirt1; nutraceuticals; osteoblasts; osteoclasts; osteoporosis; soluble guanylate cyclase
    DOI:  https://doi.org/10.3390/ijms23094776
  28. Mol Biol Rep. 2022 May 08.
    Changes in AMPKα and Ubiquitin Ligases in Myocyte Reverse Remodeling after Surgical Ventricular Reconstruction in rats with ischemic cardiomyopathy.
    Yasushige Shingu, Tetsuya Hieda, Satoshi Sugimoto, Hidetsugu Asai, Tomoji Yamakawa, Satoru Wakasa.
       BACKGROUND: The change in myocardial protein degradation systems after ventricular unloading has been unknown. We aimed to evaluate the anti-hypertrophic protein adenosine monophosphate-activated protein kinase (AMPK) and two major protein degradation systems (ubiquitin proteasome system and autophagy) in a model of surgical ventricular reconstruction (SVR) in rats with ischemic cardiomyopathy.
    METHODS AND RESULTS: Rats were randomized into the following groups: sham/sham (control group), myocardial infarction (MI)/sham (sham group) and MI/SVR (SVR group), with an interval of 4 weeks. Two (early, n = 5 for each) and 28 days (late, n = 5 for each) after SVR, ventricular size, and wall stress were assessed. Myocyte area, protein expression of AMPKα and autophagy markers, and gene expression of ubiquitin ligases (Atrogin-1 and Murf-1) were evaluated in the late phase. In the early phase, left ventricular dimensions and wall stress were smaller in the SVR group than in the sham group, whereas they were comparable in the late period. Myocyte area in the SVR group was reduced to the value in the control group, while it was larger in the sham group than in the control group. Total-AMPKα, p-AMPKα, and AMPKα phosphorylation rates were higher, and Atrogin-1 and Murf-1 were lower in the SVR group than in the sham group, while the autophagy markers were not different between the groups. p-AMPKα had strong negative correlations with myocyte area, Atrogin-1, and Murf-1.
    CONCLUSIONS: In myocyte reverse remodeling after SVR, AMPKα phosphorylation increased in association with reduced gene expression of ubiquitin ligases.
    Keywords:  AMPKα; Atrogin-1; Murf-1; Myocyte reverse remodeling; Surgical ventricular reconstruction
    DOI:  https://doi.org/10.1007/s11033-022-07347-8
  29. FASEB J. 2022 May;36 Suppl 1
    Measuring AMPK Activity in Vivo Using a Genetically-Encoded Biosensor.
    Lynly Carman, Matthew D Nelson.
      Adenosine monophosphate-activated kinase (AMPK) is a highly conserved heterotrimeric protein complex that functions in a broad spectrum of cellular stress response pathways. Most work investigating the cellular dynamics of AMPK activity has been limited to either whole-organ/tissue or, in the case of smaller genetic models like Caenorhabditis elegans, whole-organism analyses. Some limitations of these approaches include: 1) An inability to observe cellular heterogeneity of AMPK activation; 2) a lack of the temporal dynamics; 3) an inability to correlate AMPK dynamics with physiological states of a living organism. Thus, we sought to adapt a genetically-coded AMPK biosensor, called AMPKAR-EV (Kongaya et al., 2017), for use in C. elegans, which is transparent and genetically tractable. The AMPKAR-EV biosensor uses fluorescence resonance energy transfer (FRET) to report AMPK phosphorylation of substrates. We expressed codon-adapted AMPKAR-EV in C. elegans and made three separate transgenic lines expressing it in intestines, body wall muscles, and neurons. As expected, we find that the biosensor responds to conditions that promote AMPK activation. We crossed the lines expressing AMPKAR-EV in the intestines into a number of different genetic mutants, which we either generated using CRISPR or obtained from previous studies. For context, work in mammals suggests that protein kinase A (PKA) phosphorylates AMPK at serine 173 (S173 - inhibitory site) and prevents catalytic phosphorylation at threonine 172 (T172 - activation site). These sites are conserved to C. elegans but located at serine 244 (S244) and threonine 243 (T243) respectively; however, the significance of these regulatory sites have not been specifically studied. To test these mechanisms more precisely, we made two C. elegans strains that also express AMPKAR-EV, one that alters the activation site (aak-2(T243A)) and another that alters the inhibitory site (aak-2(S244G)). The third strain we constructed expresses AMPKAR-EV in the kin-2(ce179) mutant background, which exhibits constitutively active PKA. Using these strains we are optimizing AMPKAR-EV in vivo and testing the hypothesis that PKA inhibits AMPK at the S244 inhibitory site, which prevents its phosphorylation at the activation site. We envision that AMPKAR-EV will be of broad use in C. elegans as an organism well-known for its study of behavior, aging, and longevity.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L8058
  30. Oxid Med Cell Longev. 2022 ;2022 7511393
    Dexmedetomidine Can Enhance PINK1/Parkin-Mediated Mitophagy in MPTP-Induced PD Mice Model by Activating AMPK.
    Cheng Chen, Yaohua Chen, Tingting Liu, Dan Song, Di Ma, Oumei Cheng.
      Parkinson's disease (PD) is a common neurodegenerative disease characterized by the degeneration of dopaminergic (DA) neurons in the substantia nigra (SN). Our previous study has shown that dexmedetomidine (Dex) can protect mitochondrial function and reduce apoptosis in MPP+-induced SH-SY5Y cells. Evidences have shown that mitophagy is related to the development of PD. In this study, we investigated whether Dex can enhance mitophagy in MPTP-induced mice to play a neuroprotective effect. In our experiment, mice were injected with MPTP 30 mg/kg intraperitoneally for 5 consecutive days to establish a PD subacute model. Dex (30, 50, and 100 μg/kg) was injected intraperitoneally 30 minutes before each injection of MPTP, respectively. Our results showed that Dex (50 μg/kg) most significantly attenuated MPTP-induced motor dysfunction and restored TH-positive neurons in the SN, increased the expression of the antiapoptotic protein Bcl-2, and decreased the expression of apoptotic proteins cleaved casepase3, cleaved casepase9, and Bax. Moreover, Dex increased the activity of mitochondrial Complexes I-IV and decreased the level of oxidative stress, manifesting as decreased MDA levels and increased SOD and GSH-PX levels. Besides, under transmission electron microscopy, Dex increased the mitophagosome which is an autophagosome with a mitochondrion-like structure inside under the electron microscope. In addition, Dex could also increase the expression of mitophagy-related proteins p-AMPK, LC3II/I, PINK1, and Parkin and decrease P62. However, after using Compound C (CC, 10 mg/kg, AMPK inhibitor), the effects of Dex on increasing PINK1/Parkin-induced mitophagy and neuroprotection were attenuated. In conclusion, Dex may improve mitochondrial function by activating AMPK to enhance PINK1/Parkin-induced mitophagy, thereby protecting dopaminergic neurons.
    DOI:  https://doi.org/10.1155/2022/7511393
  31. FASEB J. 2022 May;36 Suppl 1
    Increased Connexin 43 expression in AMPK deficient cardiomyocytes in response to Doxorubicin treatment.
    Mandeep Singh, Naunihal Singh, Tamayo Kobayashi, Saturo Kobayashi, Qiangrong Liang.
       CONTEXT: Doxorubicin (DOX) is an anthracycline anticancer drug that can cause dose-dependent cardiomyopathy and chronic heart failure. DOX cardiotoxicity has been attributed to excess production of ROS and genotoxicity that leads to cell death. AMPK is a kinase composed of α, β and γ subunits. AMPK has been proposed to modulate DOX-induced cardiotoxicity, but the downstream mediators or effectors remain unclear. Connexin 43 (Cx43) is a gap junction protein abundantly expressed in the cardiomyocyte and is essential for heart function. Studies have suggested a role for Cx43 in DOX cardiotoxicity, but it remains unknown whether AMPK interacts with Cx43 to regulate cardiac response to DOX chemotherapy.
    OBJECTIVE: To determine the role of AMPK in the regulation of the Cx43 in response to DOX-induced cardiotoxicity.
    METHODS: H9c2 rat cardiomyocytes were seeded on 6-well plates and cultured in DMEM with 6.5% fetal bovine serum. To investigate the role of AMPK, the cells were transfected with siRNAs targeting different AMPK isoforms using Lipofectamine RNAiMAX and Opti-MEM I reduced serum medium. After 48 hours of transfection, half of the wells received 0.5 µM DOX treatment, and the other half received equal amounts of saline. Protein samples were collected 16 hours later using the Cell lysis buffer. Western blot analysis was performed to confirm the knockdown of AMPK isoforms and to determine the protein expression levels of Cx43.
    RESULTS: Our findings showed that siRNA transfections effectively reduced the protein levels of AMPK isoform a1 and a2, respectively, as indicated by the Western blot analysis. DOX treatment increased the expression levels of the Cx43 in cardiomyocytes. Also, knockdown (KD) of AMPKα1 isoform resulted in a similar increase in Cx43 expression levels as did DOX, but AMPKα2 KD did not significantly alter Cx43 levels, suggesting an AMPKa1 specific effect on Cx43 expression. Interestingly, AMPKα1 KD, together with DOX treatment, did not produce a synergistic effect to increase Cx43 expression levels further, suggesting the possibility that DOX might have elevated the Cx43 expression through its effect on AMPK.
    CONCLUSION: Our results confirmed the previous studies showing that treatment of the cardiomyocytes with DOX increased the expression of Cx43. However, our investigations produced a novel finding that knocking down AMPKα1 also increased the Cx43 but did not enhance the effect of DOX on Cx43 expression, suggesting a role for AMPK in the pathway that regulates the Cx43 to mediate the DOX-induced cardiotoxicity. Our investigations also showed an isoform-specific effect of AMPK on Cx43 expression since only α1 but not α2 KD increased Cx43 levels. Whether the increase in Cx43 is due to increased gene expression or inhibited protein degradation remains to be determined. Further investigation is also warranted to elucidate the roles of other AMPK subunits or isoforms in regulating the Connexin 43 expression in the context of doxorubicin cardiotoxicity.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R4237
  32. J Nat Prod. 2022 May 11.
    Hernandezine Regulates Proliferation and Autophagy-Induced Apoptosis in Melanoma Cells.
    Xiaoting Wang, Xiaojie Li, Yuan Xia, Danyang Wang, Xiaohan Li, Ling Liu, Qiusheng Zheng, Defang Li, Qingling Jiang.
      Hernandezine is isolated from an herbal medicine that selectively inhibits multidrug resistance and improves the efficacy of drugs for cancer treatment. To date, no studies on hernandezine in melanoma have been conducted. In this study, hernandezine was found to inhibit proliferation and induce apoptosis in melanoma A375 cells and B16 cells. In hernandezine-treated melanoma cells, G0/G1 cycle arrest occurred accompanied by significantly downregulated levels of phosphorylated JAK2 and STAT3. In addition, the cycle arrest could be enhanced by AG490 (JAK2 inhibitor), suggesting that the JAK2/STAT3 pathway is involved in cell cycle regulation in hernandezine-treated melanoma cells. Hernandezine-treated melanoma cells exhibited autophagy-specific structures, autophagy markers (LC3II/LC3-I), and autophagic flow over time. Moreover, 3-MA (autophagy inhibitor) significantly inhibited apoptosis, indicating that hernandezine promotes apoptosis by inducing autophagy. Combined with differential expression of P-AMPK, P-ACC (downstream targets of adenine monophosphate activated protein kinase, AMPK), and P-p70S6K (downstream targets of mammalian target of rapamycin, mTOR) and significant inhibition of apoptosis by AMPK inhibitor complex C (CC) in hernandezine-treated melanoma cells suggested that hernandezine could induce autophagy via the AMPK-mTOR pathway, thereby inducing apoptosis. This study first analyzed the effect of melanoma cells by hernandezine and provided a theory for hernandezine in the treatment of melanoma.
    DOI:  https://doi.org/10.1021/acs.jnatprod.2c00098
  33. Environ Toxicol. 2022 May 07.
    Antitumor activity of pachymic acid in cervical cancer through inducing endoplasmic reticulum stress, mitochondrial dysfunction, and activating the AMPK pathway.
    Ting Yang, Sijuan Tian, Yaohui Wang, Jing Ji, Juan Zhao.
      Pachymic acid has various pharmacological effects, including anti-inflammatory, antioxidant, immunomodulatory, and antitumor. However, the role of pachymic acid in cervical cancer remains unclear. So, we investigated the effects of pachymic acid in cervical cancer and elucidated the underlying mechanisms. We treated HeLa cells and normal cervical epithelial cells (HUCECs) with pachymic acid (0, 10, 20, 40, 80, or 160 μM) for 72 h, and found the cell activity was decreased in cells treated with 160 μM pachymic acid for 48 h or 80 μM pachymic acid for 72 h, while HUCECs viability without effect. Next, we observed that endoplasmic reticulum (ER) related gene expression, mitochondrial membrane potential (MMP) changes, ATP depletion, reactive oxygen species (ROS) generation and apoptosis were increased. Moreover, we observed that cytochrome C (Cytc) expression was increased and apoptosis-inducing factor (AIF) was decreased in the cytoplasm of pachymic acid-treated HeLa cells. Tauroursodeoxycholic acid (TUDCA) of ER stress inhibitor reversed the effects of pachymic acid on HeLa cells. Phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) of the AMPK pathway key protein was upregulated in pachymic acid-induced HeLa cells. Finally, we subcutaneously implanted HeLa cells into female nude mice and treated them with pachymic acid (50 mg/kg) for 3 weeks (5 days/week), and observed in pachymic acid induced xenograft mice, tumor growth was suppressed, cell apoptosis, ER-related gene expression, and ROS levels in tumor tissues were increased. Therefore, these findings demonstrated that pachymic acid plays an anti-tumor activity in cervical cancer through inducing ER stress, mitochondrial dysfunction, and activating the AMPK pathway.
    Keywords:  AMPK pathway; ER stress; cervical cancer; mitochondrial dysfunction; pachymic acid
    DOI:  https://doi.org/10.1002/tox.23555
  34. FASEB J. 2022 May;36 Suppl 1
    Chitosan Oligosaccharide Ameliorates Kidney Injury by Improving Intestinal Barrier Dysfunction and Lipid Metabolism in Obese-insulin Resistant Rats.
    Prempree Sutthasupha, Sasivimon Promsan, Nichakorn Phengpol, Rath Pichyangkura, Chatchai Muanprasat, Anusorn Lungkaphin.
       INTRODUCTION: Obesity remains a global health problem. High-fat diet (HFD) consumption increases the risk of type 2 diabetes and initiates kidney injury. Chitosan Oligosaccharide (COS) provides anti-obesity effects through several mechanisms. However, the effects of COS on preventing obesity-related kidney injury in obese-insulin resistance have not been studied. A novel action of COS in activating intestinal tight junction assembly was reported in in vitro via calcium sensing receptor-mediated AMP-activated protein kinase (CaSR/AMPK) activation. HFD-induced gut dysbiosis is related with intestinal barrier dysfunction and systemic low-grade inflammation. Therefore, AMPK activation might be a therapeutic target for preventing obesity-related complications. Here, we explored the protective effects of COS in obesity-induced kidney dysfunction and the involved molecular mechanisms. We hypothesized that COS might exert renoprotection against obesity-induced kidney injury through increasing intestinal barrier integrity and lipid excretion.
    METHODS: Male Wistar rats were fed with HFD for 16 weeks to induce obesity. Next, HFD rats were randomly divided into 4 groups: HFD, HFD treated with COS 5 or 10 mg/kg/day (COS5 or COS10) and HFD treated with metformin 30 mg/kg/day (MET). The interventions were orally treated for 8 weeks. At the end of experiment, blood, feces, intestinal and renal tissue samples were collected for further investigations.
    RESULTS: After 16 weeks of high-fat diet feeding, obesity and glucose intolerance were presented in HFD compared to normal diet (ND) rats, as indicated by the significant increases in body weight (BW), plasma cholesterol and total area under the curve for glucose (TAUCg). Kidney injury were observed at week 24, as shown by elevated kidney injury score and serum creatinine. Treatment with COS5 or COS10 significantly decreased BW, hypercholesterolemia, glucose intolerance and kidney dysfunction. Interestingly, COS10 markedly increased fecal lipid excretion compared with HFD and MET. In addition, intestinal CaSR, p-AMPK/AMPK and tight junction protein claudin-1 expressions were downregulated in HFD which were upregulated by COS10 and MET. Renal TNFαR activation and increased proinflammatory cytokines were detected in HFD. Moreover, renal apoptosis markers, Bcl-2-associated X protein (Bax) and cleaved caspase-3, were increased, along with the suppressing of anti-apoptotic B-cell lymphoma-2 (Bcl-2) in HFD. Treatment with COS10 or MET markedly alleviated renal inflammation and apoptosis.
    CONCLUSION: COS exerted renoprotection through attenuating renal inflammation and apoptosis as a result of improving intestinal barrier via CaSR/AMPK activation and lipid absorption in obese rats. Therefore, COS might be an effective new dietary supplement for treating obesity and its related complications in clinical use.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.0R803
  35. Front Pharmacol. 2022 ;13 900127
    Corrigendum: Metformin Ameliorates Inflammation and Airway Remodeling of Experimental Allergic Asthma in Mice by Restoring AMPKα Activity.
    Wenxian Ma, Qiaoyan Jin, Haiqin Guo, Xinpeng Han, Lingbin Xu, Shemin Lu, Changgui Wu.
      [This corrects the article DOI: 10.3389/fphar.2022.780148.].
    Keywords:  AMPK; airway inflammation; airway remodeling; asthma; metformin
    DOI:  https://doi.org/10.3389/fphar.2022.900127
  36. Nan Fang Yi Ke Da Xue Xue Bao. 2022 Apr 20. 42(4): 518-527
    [Dihydromyricetin reduces lipid accumulation in LO2 cells via AMPK/mTOR-mediated lipophagy pathway and inhibits HepG2 cell proliferation in vitro].
    X Liao, Y Hao, M Wu, H Liu, L Jiang, Z Ye, W Liao, H Deng.
       OBJECTIVE: To explore the mechanism underlying the hepatoprotective effect of dihydromyricetin (DMY) against lipid accumulation in light of the lipophagy pathway and the inhibitory effect of DMY on HepG2 cell proliferation.
    METHODS: LO2 cells were cultured in the presence of 10% FBS for 24 h and treated with 100 μg/mL DMY, or exposed to 50% FBS for 24 h followed by treatment with 50, 100, or 200 μg/mL DMY; the cells in recovery group were cultured in 50% FBS for 24 h and then in 10% FBS for another 24 h. Oil red O staining was used to observe the accumulation of lipid droplets in the cells, and the levels of TC, TG, and LDL and activities of AST, ALT and LDH were measured. The expression of LC3 protein was detected using Western blotting. AO staining and transmission electron microscopy were used to determine the numbers of autophagolysosomes and autophagosomes, respectively. The formation of autophagosomes was observed with MDC staining, and the mRNA expression levels of LC3, ATG7, AMPK, mTOR, p62 and Beclin1 were determined with q-PCR. Flow cytometry was performed to analyze the effect of 50, 100, and 200 μg/mL DMY on cell cycle and apoptosis of HepG2 cells; DNA integrity in the treated cells was examined with cell DNA fragmentation test.
    RESULTS: DMY treatment and pretreatment obviously inhibited lipid accumulation and reduced the levels of TC, TG, LDL and enzyme activities of AST, ALT and LDH in LO2 cells (P < 0.05). In routinely cultured LO2 cells, DMY significantly promoted the formation of autophagosomes and autophagolysosomes and upregulated the expression of LC3 protein. DMY obviously attenuated high FBS-induced inhibition of autophagosome formation in LO2 cells, up- regulated the mRNA levels of LC3, ATG7, Beclin1 and AMPK, and downregulated p62 and mTOR mRNA levels (P < 0.05 or 0.01). In HepG2 cells, DMY caused obvious cell cycle arrest, inhibited cell proliferation, and induced late apoptosis and DNA fragmentation.
    CONCLUSION: DMY reduces lipid accumulation in LO2 cells by regulating the AMPK/ mTOR-mediated lipophagy pathway and inhibits the proliferation of HepG2 by causing cell cycle arrest and promoting apoptosis.
    Keywords:  AMPK/mTOR; dihydromyricetin; lipophagy; nonalcoholic fatty liver disease; proliferation inhibition
    DOI:  https://doi.org/10.12122/j.issn.1673-4254.2022.04.07
  37. Sci Signal. 2022 May 10. 15(733): eabq8516
    Cleaved to keep dying cells alive.
    Leslie K Ferrarelli.
      Upon DNA damage, caspase-3 sends the kinase AMPK to the nucleus to prevent apoptotic progression.
    DOI:  https://doi.org/10.1126/scisignal.abq8516
  38. Hum Cell. 2022 May 11.
    STK11 overexpression prevents glucocorticoid-induced osteoporosis via activating the AMPK/SIRT1/PGC1α axis.
    Jiao Xiao, Wenjin Li, Guojuan Li, Jiankai Tan, Na Dong.
      Osteoporosis (OP) is a frequent orthopedic disease characterized by pain, fractures and deformities. Glucocorticoids are the most common cause of secondary osteoporosis. Here, we aim to explore the function and mechanism of STK11 in glucocorticoid (GC)-induced OP. Human mesenchymal stromal cells (hMSCs) were differentiated under osteogenic or adipogenic culture medium. An in-vitro OP model was induced by dexamethasone (DEX). The viability, differentiation, apoptosis, and ROS level were evaluated for investigating the functions of SKT11 on hMSCs. The SIRT1 inhibitor EX-527, PGC1α inhibitor SR-18292, and AMPK activator metformin were administered into hMSCs for confirming the mechanism of SKT11. Our results showed that STK11 was down-regulated in OP tissues, as well as DEX-treated hMSCs. Overexpressing STK11 attenuated DEX-mediated inhibition of osteogenic differentiation and heightened the activation of the AMPK/SIRT1/PGC1α pathway, whereas STK11 knockdown exerted opposite effects. Inhibiting SIRT1 or PGC1α repressed the promotive effect of STK11 on osteogenic differentiation of hMSCs, while activation of AMPK abated the inhibitory effect of STK11 knockdown on osteogenic differentiation of hMSCs. In conclusion, this study revealed that overexpressing STK11 dampened GC-induced OP by activating the AMPK/SIRT1/PGC1α axis.
    Keywords:  AMPK; Glucocorticoids; Osteoporosis; PGC1α; SIRT1; STK11
    DOI:  https://doi.org/10.1007/s13577-022-00704-6
  39. J Lipid Res. 2022 May 05. pii: S0022-2275(22)00054-2. [Epub ahead of print] 100221
    Neuronal growth regulator 1 promotes adipocyte lipid trafficking via interaction with CD36.
    Ara Yoo, Yeonhee Joo, Yeongmi Cheon, Sung Joong Lee, Soojin Lee.
      Neuronal growth regulator 1 (NEGR1) is a glycosylphosphatidylinositol (GPI)-anchored membrane protein associated with several human pathologies, including obesity, depression, and autism. Recently, significantly enlarged white adipose tissue (WAT), hepatic lipid accumulation, and decreased muscle capacity were reported in Negr1-deficient mice. However, the mechanism behind these phenotypes was not clear. In the present study, we found NEGR1 to interact with cluster of differentiation 36 (CD36), the major fatty acid translocase in the plasma membrane. Binding assays with a soluble form of NEGR1 and in-situ proximal ligation assays indicated that NEGR1-CD36 interaction occurs at the outer leaflet of the cell membrane. Furthermore, we show that NEGR1 overexpression induced CD36 protein destabilization in vitro. Both mRNA and protein levels of CD36 were significantly elevated in the WAT and liver tissues of Negr1-/- mice. Accordingly, fatty acid uptake rate increased in NEGR1-deficient primary adipocytes. Finally, we demonstrated that Negr1-/- mouse embryonic fibroblasts (MEFs) showed elevated reactive oxygen species levels and decreased adenosine monophosphate-activated protein kinase activation compared with control MEFs. Based on these results, we propose that NEGR1 regulates cellular fat content by controlling the expression of CD36.
    Keywords:  AMPK activation; CD36; ROS; adipose tissue; diabetes; fatty acid/transport; lipid rafts; obesity; protein-protein interaction; proximal ligation assay
    DOI:  https://doi.org/10.1016/j.jlr.2022.100221
  40. FASEB J. 2022 May;36 Suppl 1
    Elimination of AKT in Cardiomyocytes Causes Complex Reorganization of Signal Transduction leading to AMPK Inhibition and Fatal Cardiac Atrophy.
    Stefanie Gödecke, Lukas Breuling, Phil-Torben Müller, Uli Flögel, Andre Heinen, Axel Gödecke.
      Akt1 and Akt2 are the main protein kinase B (Akt) isoforms expressed in the mammalian heart. Tamoxifen(OHTx) -inducible, cardiomyocyte-specific Akt1/ Akt2 double knockout mice (iCM-Akt12) show progressive cardiac atrophy and loss of contractile function leading to terminal heart failure 23.9 ± 2 days after first OHTx injection. TUNEL staining of iCM-Akt12 hearts revealed that cardiomyocyte apoptosis did not contribute substantially to cardiac atrophy. Rather, cellular atrophy caused the loss of cardiac mass. The cellular area (a), width (w), and length (l) declined between day 9 and day 14 [-18 (a)/-18 (w)/-9% (l)] as compared to WT controls. This size reduction was retarded between d14 and d21 (-20 (a)/-24 (w)/-8% (l). In vivo 31 P-imaging identified a progressive energetic deficiency of iCM-Akt12 hearts on d15 and d20 after OHTx injection. Further analyses showed that in the early phase up to day 14 increased autophagic activity and reduced de novo protein synthesis seemed to cause cellular atrophy. As expected, deletion of Akt led to impaired mTORC1 signaling indicated by reduced phosphorylation of S6K, RPS6, 4E-BP1, all involved in translation. Concomitantly, we measured reduced protein synthesis. Autophagy was increased in iCM-Akt12 hearts, as shown by increased LC3-II/I ratio and higher expression of autophagic genes. From day 14 after KO induction (late phase), protein synthesis appeared to increase and mTORC1 substrates involved in translation (S6K, RPS6 and 4E-BP1) were higher phosphorylated. Of note, also a higher phosphorylation of the inhibitory mTORC1 target site in the central autophagy kinase Ulk1 (S757) was found. Moreover, autophagy appeared to be disturbed, as p62 accumulated. Despite increasing energetic depletion, the cellular energy sensor AMPK was not activated in iCM-Akt12 hearts. Rather, AMPK was inhibited by phosphorylation of the α-subunit on serine 485/491. The S485/491 phosphorylation has been attributed to several kinases (Akt, PKD1, S6K, PKC, PKA). Akt deletion led to a pronounced hyperactivation of Pdk1 signaling upstream of Akt, causing an activation of protein kinases regulated by Pdk1 (S6K, PKA, PKC, PKD). This was demonstrated by increased phosphorylation of specific kinase substrates and consensus motifs in late iCM-Akt12 hearts and might promote the AMPK inactivation. To test to what extent a non-inhibitable AMPK might attenuate the fatal phenotype of cardiac Akt loss, AAV-mediated infection of iCM-Akt12 mice with a phospho-defective AMPKα2 S491A mutant prior to KO induction was performed. This led to an increased survival and improved heart function, demonstrating that the AMPK inhibition contributes to the lethal phenotype. In conclusion, loss of Akt signaling leads to a widespread dysregulation of intracellular signal transduction. Whereas enhanced autophagy and inhibition of mTORC1 are direct consequences of Akt deletion, the later reactivation of mTORC1 and inhibition of AMPK causes attenuation of autophagy, increased translation, deceleration of cellular atrophy, worsenes energetic shortage, and aggravates cardiac dysfunction.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R5496
  41. J Integr Med. 2022 Apr 27. pii: S2095-4964(22)00051-6. [Epub ahead of print]
    Qili Qiangxin, a compound herbal medicine formula, alleviates hypoxia-reoxygenation-induced apoptotic and autophagic cell death via suppression of ROS/AMPK/mTOR pathway in vitro.
    Cai-Lian Fan, Wan-Jun Cai, Meng-Nan Ye, Miao Chen, Yi Dai.
       OBJECTIVE: Qili Qiangxin (QLQX), a compound herbal medicine formula, is used effectively to treat congestive heart failure in China. However, the molecular mechanisms of the cardioprotective effect are still unclear. This study explores the cardioprotective effect and mechanism of QLQX using the hypoxia-reoxygenation (H/R)-induced myocardial injury model.
    METHODS: The main chemical constituents of QLQX were analyzed using high-performance liquid chromatography-evaporative light-scattering detection. The model of H/R-induced myocardial injury in H9c2 cells was developed to simulate myocardial ischemia-reperfusion injury. Apoptosis, autophagy, and generation of reactive oxygen species (ROS) were measured to assess the protective effect of QLQX. Proteins related to autophagy, apoptosis and signalling pathways were detected using Western blotting.
    RESULTS: Apoptosis, autophagy and the excessive production of ROS induced by H/R were significantly reduced after treating the H9c2 cells with QLQX. QLQX treatment at concentrations of 50 and 250 μg/mL caused significant reduction in the levels of LC3II and p62 degradation (P < 0.05), and also suppressed the AMPK/mTOR signalling pathway. Furthermore, the AMPK inhibitor Compound C (at 0.5 μmol/L), and QLQX (250 μg/mL) significantly inhibited H/R-induced autophagy and apoptosis (P < 0.01), while AICAR (an AMPK activator, at 0.5 mmol/L) increased cardiomyocyte apoptosis and autophagy and abolished the anti-apoptotic effect of QLQX. Similar phenomena were also observed on the expressions of apoptotic and autophagic proteins, demonstrating that QLQX reduced the apoptosis and autophagy in the H/R-induced injury model via inhibiting the AMPK/mTOR pathway. Moreover, ROS scavenger, N-Acetyl-L-cysteine (NAC, at 2.5 mmol/L), significantly reduced H/R-triggered cell apoptosis and autophagy (P < 0.01). Meanwhile, NAC treatment down-regulated the ratio of phosphorylation of AMPK/AMPK (P < 0.01), which showed a similar effect to QLQX.
    CONCLUSION: QLQX plays a cardioprotective role by alleviating apoptotic and autophagic cell death through inhibition of the ROS/AMPK/mTOR signalling pathway.
    Keywords:  AMPK/mTOR pathway; Apoptosis; Autophagy; Herbal medicine; Hypoxia–reoxygenation; Qili Qiangxin formula; Reactive oxygen species
    DOI:  https://doi.org/10.1016/j.joim.2022.04.005
  42. FASEB J. 2022 May;36 Suppl 1
    Hyperbaric Oxygen Therapy Following Cervical Spinal Cord Injury Increases BDNF/TrkB Signaling in Diaphragm Muscle.
    Franccesco P Boeno, Michael D Sunshine, Victoria E Bindi, David D Fuller, Ashley J Smuder.
      Respiratory insufficiency is the leading cause of morbidity and mortality following cervical spinal cord injury (SCI). Diaphragm atrophy following SCI occurs as a result of damage to respiratory neural circuitry, which leads to impaired muscle contractility. Disruption of redox balance occurs after SCI and increased mitochondrial reactive oxygen species production is an upstream trigger for diaphragm muscle atrophy and weakness. Recently, we demonstrated that increased antioxidant capacity in response to hyperbaric oxygen (HBO) therapy may play a role in preserving mitochondrial redox homeostasis following SCI. However, a precise understanding of the mechanisms by which HBO therapy can prevent diaphragm dysfunction is lacking. In this regard, evidence indicates that the secretion of neurotrophic factors, such as brain derived neurotrophic factor (BDNF), may also contribute to enhanced functional recovery when HBO therapy is initiated acutely following cervical SCI. Through activation of the 5' adenosine monophosphate-activated protein kinase (AMPK) and serine-threonine protein kinase (AKT) signaling via the phosphorylation of its receptor tropomyosin receptor kinase B (TrKb), BDNF expression has been shown to modulate mitochondrial biogenesis and skeletal muscle proteolysis. Therefore, we tested the hypothesis that 10 days of HBO therapy initiated in the acute phase following cervical SCI can increase diaphragm mitochondrial biogenesis and prevent diaphragm proteolytic signaling by activating BDNF/TrkB signaling. To test this, adult male Sprague-Dawley rats were separated into three groups (n=8/group): (1) non-injured, room air exposure (CON); (2) lateral-cervical spinal cord contusion, room air exposure (SCI); and (3) lateral-cervical spinal cord contusion, HBO therapy (SCI+HBO). Animals in the SCI and SCI+HBO groups were anesthetized, and following laminectomy at the C3-C4 level, a lateral-cervical spinal contusion was made using a mechanical impactor. HBO therapy consisted of 10 days of exposure starting on the day of SCI, using a 40 L chamber flushed with 100% O2 and pressurized to 3 ATA. Our results indicate that HBO therapy delivered in the acute phase of SCI positively affected BDNF/TrkB signaling. Specifically, diaphragm protein expression of BDNF, p-AKT/AKT and p-AMPK/AMPK were increased after 10 days of HBO therapy compared to CON animals (p<0.05). Additionally, HBO therapy prevented the SCI-induced reduction in diaphragm peroxisome proliferator-activated receptor-gamma coactivator (PGC1-α) protein expression (p<0.05), as well as the caspase-3-specific cleavage of α-II-spectrin observed in the SCI group (p<0.05). In summary, these findings demonstrate that HBO therapy can induce BDNF signaling and highlight an additional mechanism for the protective effects of HBO therapy to preserve diaphragm muscle function following SCI.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R2130
  43. Diab Vasc Dis Res. 2022 May-Jun;19(3):19(3): 14791641221102513
    Glucolipotoxicity induces endothelial cell dysfunction by activating autophagy and inhibiting autophagic flow.
    Yulan Liu, Hong Xiang, Wenfang Xiong, Jie Ouyang, Hengdao Liu, Shaoli Zhao, Jie Xiao, Jialing Li, Zhihao Shu, Xuewen Wang, Huiqin Liu, Jing Zhang, Jianing Fan, Ying Li, Shuhua Chen, Hongwei Lu.
       OBJECTIVES: This study aims to determine the role and mechanism of autophagy in endothelial cell dysfunction by glucolipotoxicity.
    METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with high glucose and high palmitic acid. The number of autophagosomes was evaluated by monodansylcadaverine (MDC) staining and transmission electron microscopy (TEM). The expression of autophagy-related proteins (LC3 and P62) was assessed by Western blotting. Capillary tube-like formation was evaluated on Matrigel. Reactive oxygen species (ROS) production was detected by DCFH-DA. Cell apoptosis was measured by Hoechst 33258 staining and flow cytometry. Phosphorylation of AMPK, mTOR, and ULK1 was also analyzed by Western blotting.
    RESULTS: We found that glucolipotoxicity induced autophagy initiation and hindered autophagosomes degradation. Moreover, glucolipotoxicity increased the production of intracellular ROS, decreased the ability of tubular formation, and increased cell apoptosis. However, endothelial cell dysfunction was alleviated by 3-methyladenine, an early-stage autophagy inhibitor. Additionally, glucolipotoxicity promoted the phosphorylation of AMPK and ULK1 and inhibited the phosphorylation of mTOR.
    CONCLUSIONS: Glucolipotoxicity initiates autophagy through the AMPK/mTOR/ULK1 signaling pathway and inhibits autophagic flow, leading to the accumulation of autophagosomes, thereby inducing apoptosis and impairing endothelial cell function.
    Keywords:  AMPK/mTOR; Glucolipotoxicity; autophagy; diabetes; endothelial cell dysfunction
    DOI:  https://doi.org/10.1177/14791641221102513
  44. Biomed Pharmacother. 2022 May 09. pii: S0753-3322(22)00481-4. [Epub ahead of print]151 113092
    Curcumin exerts chondroprotective effects against osteoarthritis by promoting AMPK/PINK1/Parkin-mediated mitophagy.
    Zhuangzhuang Jin, Bohan Chang, Yingliang Wei, Yue Yang, He Zhang, Jiabao Liu, Longhuan Piao, Lunhao Bai.
      Osteoarthritis (OA), a chronic degenerative disease with heterogeneous properties, is difficult to cure due to its complex pathogenesis. Curcumin possesses excellent anti-inflammatory and antioxidant properties and may have potential therapeutic value in OA. In this study, we investigated the action targets of curcumin and identified potential anti-OA targets for curcumin. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway analyses were performed to evaluate these targets. Furthermore, we established a sodium monoiodoacetate-induced rat knee OA model and IL-1β induced OA chondrocyte model to verify the effect and mechanism of curcumin against OA. The GO and KEGG analyses screened seven hub genes involved in metabolic processes and the AMPK signaling pathway. Curcumin can significantly attenuate OA characteristics according to Osteoarthritis Research Society International (OARSI) and Mankin scores in OA rats. Additionally, curcumin is notably employed as an activator of mitophagy in maintaining mitochondrial homeostasis (ROS, Ca2+, ATP production, and mitochondrial membrane potential). The expression levels of mitophagy-related proteins were increased not only in articular cartilage but also in chondrocytes with curcumin intervention. Combining validation experiments and network pharmacology, we identified the importance of mitophagy in the curcumin treatment of OA. The chondroprotective effects of curcumin against OA are mediated by the AMPK/PINK1/Parkin pathway, and curcumin may serve as a potential novel drug for OA management.
    Keywords:  AMPK; Curcumin; Mitochondria; Mitophagy; Osteoarthritis; PINK1/Parkin
    DOI:  https://doi.org/10.1016/j.biopha.2022.113092
  45. Nutrients. 2022 May 09. pii: 1985. [Epub ahead of print]14(9):
    Nutraceuticals/Drugs Promoting Mitophagy and Mitochondrial Biogenesis May Combat the Mitochondrial Dysfunction Driving Progression of Dry Age-Related Macular Degeneration.
    Lidianys María Lewis Luján, Mark F McCarty, James J Di Nicolantonio, Juan Carlos Gálvez Ruiz, Ema Carina Rosas-Burgos, Maribel Plascencia-Jatomea, Simon Bernard Iloki Assanga.
      In patients with age-related macular degeneration (AMD), the crucial retinal pigment epithelial (RPE) cells are characterized by mitochondria that are structurally and functionally defective. Moreover, deficient expression of the mRNA-editing enzyme Dicer is noted specifically in these cells. This Dicer deficit up-regulates expression of Alu RNA, which in turn damages mitochondria-inducing the loss of membrane potential, boosting oxidant generation, and causing mitochondrial DNA to translocate to the cytoplasmic region. The cytoplasmic mtDNA, in conjunction with induced oxidative stress, triggers a non-canonical pathway of NLRP3 inflammasome activation, leading to the production of interleukin-18 that acts in an autocrine manner to induce apoptotic death of RPE cells, thereby driving progression of dry AMD. It is proposed that measures which jointly up-regulate mitophagy and mitochondrial biogenesis (MB), by replacing damaged mitochondria with "healthy" new ones, may lessen the adverse impact of Alu RNA on RPE cells, enabling the prevention or control of dry AMD. An analysis of the molecular biology underlying mitophagy/MB and inflammasome activation suggests that nutraceuticals or drugs that can activate Sirt1, AMPK, Nrf2, and PPARα may be useful in this regard. These include ferulic acid, melatonin urolithin A and glucosamine (Sirt1), metformin and berberine (AMPK), lipoic acid and broccoli sprout extract (Nrf2), and fibrate drugs and astaxanthin (PPARα). Hence, nutraceutical regimens providing physiologically meaningful doses of several or all of the: ferulic acid, melatonin, glucosamine, berberine, lipoic acid, and astaxanthin, may have potential for control of dry AMD.
    Keywords:  AMPK; Nrf2; Sirt1; age-related macular degeneration; astaxanthin; berberine; ferulic acid; melatonin; mitochondrial biogenesis; nutraceuticals
    DOI:  https://doi.org/10.3390/nu14091985
  46. Nutrients. 2022 May 06. pii: 1951. [Epub ahead of print]14(9):
    HM-Chromanone Ameliorates Hyperglycemia and Dyslipidemia in Type 2 Diabetic Mice.
    Jae Eun Park, Jaemin Son, Youngwan Seo, Ji Sook Han.
      The effects of (E)-5-hydroxy-7-methoxy-3-(2-hydroxybenzyl)-4-chromanone (HMC) on hyperglycemia and dyslipidemia were investigated in diabetic mice. Mice were separated into three groups: db/db, rosiglitazone and HMC. Blood glucose or glycosylated hemoglobin values in HMC-treated mice were significantly lower compared to db/db mice. Total cholesterol, LDL-cholesterol, and triglyceride values were lower, and HDL-C levels were higher, in the HMC group compared to the diabetic and rosiglitazone groups. HMC markedly increased IRS-1Tyr612, AktSer473 and PI3K levels and plasma membrane GLUT4 levels in skeletal muscle, suggesting improved insulin resistance. HMC also significantly stimulated AMPKThr172 and PPARα in the liver, and ameliorated dyslipidemia by inhibiting SREBP-1c and FAS. Consequently, HMC reduced hyperglycemia by improving the expression of insulin-resistance-related genes and improved dyslipidemia by regulating fatty acid synthase and oxidation-related genes in db/db mice. Therefore, HMC could ameliorate hyperglycemia and dyslipidemia in type 2 diabetic mice.
    Keywords:  AMPK; HM-chromanone; db/db mice; high blood glucose; type 2 diabetes
    DOI:  https://doi.org/10.3390/nu14091951
  47. Nutr Res. 2022 Mar 27. pii: S0271-5317(22)00028-8. [Epub ahead of print]104 10-19
    Quamoclit angulata extract supplementation attenuates hepatic damage via regulation of AMPK/SIRT associated hepatic lipid metabolism in streptozotocin and high fat diet-induced T2DM mice.
    Heaji Lee, Younmi Kim, Sun Yeou Kim, Yunsook Lim.
      Quamoclit angulata (QA) is a species of the Convolvulaceae family and has a regulatory effect on glucose homeostasis. However, the effects of QA on hyperglycemia-induced hepatic damage has not been elucidated. We hypothesized that QA extract (QAE) would alleviate hepatic damage through regulation of hepatic lipid accumulation in type 2 diabetes mellitus (T2DM). T2DM was induced by streptozotocin-high-fat diet in C57BL6 male mice for 8 weeks. The diabetic mice were supplemented with QAE at low dose (5 mg/kg) or high dose (HQ, 10 mg/kg) by oral gavage every day for 12 weeks. Histopathological changes in hepatic tissue were examined using hematoxylin and eosin staining, and the protein levels of biomarkers related to AMP-activation kinase (AMPK)/sirtuin-1 (SIRT1)-associated lipid metabolism were measured using Western blotting. QAE supplementation ameliorated plasma insulin and glycated hemoglobin in diabetic mice. Furthermore, QAE decreased hepatic lipid accumulation demonstrated by hepatic triglyceride and cholesterol levels. QAE supplementation induced hepatic AMPK, which activates SIRT1 accompanied by reduced lipogenesis in the HQ group. These changes were partially explained by the amelioration of advanced glycation end product, hepatic oxidative stress, inflammation, and fibrosis in diabetic mice. Altogether, QAE would be a potential nutraceutical to prevent hepatic damage by regulation of AMPK/SIRT1-associated lipid metabolism through oxidative stress, inflammation, and fibrosis in T2DM.
    Keywords:  Energy metabolism; Fatty liver; Lipid metabolism; Quamoclit angulata; Type 2 diabetes mellitus
    DOI:  https://doi.org/10.1016/j.nutres.2022.03.012
  48. FASEB J. 2022 May;36 Suppl 1
    Colon Cancer Cell Death by Synergistic Activity of Phytochemicals is Modulated by Increased ROS, Activation of AMPK, and Inhibition of mTORC1.
    Vaishnavi S Srirama, Rubina H Thomas, Uthayashanker Ezekiel.
      Colorectal cancer (CRC) is the second leading cause of cancer-related death in the United States, and has high prevalence in both men and women. Consumption of foods high in fruits, vegetables, and spices is strongly associated with a reduced risk of developing cancer, and may be attributable to the synergy of phytochemicals present in the diet. Previously, we have shown that combinatorial treatment with curcumin and silibinin B (CS) led synergistically to higher rates of colon cancer cell death in comparis on to single compound treated cells. Our goal is to elucidate the molecular and signaling mechanisms of the synergistically enhanced anticancer activity mediated by CS in CRC. Cancer cells maintain high energy levels by higher rates of glycolysis and lactic acid fermentation, occurring in the abundance of oxygen, and have elevated levels of reactive oxygen species (ROS) compared to normal cells. Cancer cells adapt to increased levels of ROS by developing ROS scavenging mechanisms. Despite this, the ROS levels are higher than in normal cells. Phytochemicals have been shown to increase ROS by interfering with enzymes involved in reducing ROS. Our hypothesis is that phytochemical-based disruption of ROS scavenging mechanisms increases ROS to higher levels, which leads to the activation of AMPK. Activated AMPK, in turn, inhibits the mTOR pathway thereby increasing cell death by apoptosis and autophagy. Our initial results showed increased ROS and activated AMPK in combinatorial treated cells compared to single compound and control treated cells.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.0R812
  49. FASEB J. 2022 May;36 Suppl 1
    Alkaline intracellular pH (pHi) increases mTORC1 and mTORC2 signaling through PI3K to promote protein synthesis and cell survival.
    Dubek Kazyken, Stephen Lentz, Maxwell Wadley, Diane C Fingar.
      The conserved kinase mTOR (mechanistic target of rapamycin) regulates cell metabolism and promotes cell growth, proliferation, and survival in response to diverse environmental cues (e.g., nutrients; growth factors; hormones). mTOR forms the catalytic core of two multiprotein complexes, mTORC1 and mTORC2, which possess unique downstream targets and cellular functions. While mTORC1 and mTORC2 often respond to distinct upstream cues, they share a requirement for PI3K in their activation by growth factors. While many studies agree that amino acids activate mTORC1 but not mTORC2, several studies reported paradoxical activation of mTORC2 by amino acids. We noted that stimulating amino acid starved cells with a commercial mixture of amino acids increased mTORC2-dependent Akt S473 phosphorylation rapidly while re-feeding cells with complete DMEM containing amino acids failed to do so. Interestingly, we found the pH of the commercial amino acid mixture to be ~ pH 10. Upon controlling for pH, stimulating starved cells with amino acids at pH 10 but not 7.4 increased mTORC2 signaling. Moreover, DMEM at alkaline pH was sufficient to increase mTORC2 catalytic activity and signaling. Using a fluorescent pH-sensitive dye (cSNARF-1-AM) coupled to ratio-metric live cell imaging, we confirmed that alkaline extracellular pH (pHe) translated into a rapid increase in intracellular pH (pHi). Moreover, blunting this increase with a pharmacological inhibitor of an H+ transporter attenuated the increase in mTORC2 signaling by pHe. Alkaline pHi also activated AMPK, a canonical sensor of energetic stress that promotes mTORC2 signaling, as reported previously by us. Functionally, we found that alkaline pHi attenuated apoptosis caused by growth factor withdrawal through activation of AMPK-mTORC2 signaling. These results indicate that alkaline pHi augments mTORC2 signaling to promote cell survival, in part through AMPK. In the course of this work, we noted that pHi increased phosphorylation of several downstream targets of PI3K (e.g., Akt P-T308 and P-S473; S6K1 P-T389 and P-T229; PRAS40 P-T246; Tsc2 P-S939), suggesting that PI3K itself responds to changes in pHi. Indeed, alkaline pHi increased PI-3',4',5'-P3 levels in a manner sensitive to the PI3K inhibitor BYL-719. Thus, alkaline pHi elevates PI3K activity, which increases both mTORC1 and mTORC2 signaling. Mechanistically, we found that activation of PI3K by alkaline pHi induced dissociation of Tsc2 from lysosomal membranes, thereby relieving TSC-mediated suppression of Rheb, a mTORC1-activating GTPase. Functionally, we found that activation of PI3K by alkaline pHi increased mTORC1-mediated 4EBP1 phosphorylation, which initiates cap-dependent translation by eIF4E. Alkaline pHi also increased mTORC1-driven protein synthesis. Taken together, these findings reveal alkaline pHi as a previously unrecognized activator of PI3K-mTORC1/2 signaling that promotes protein synthesis and cell survival. As elevated pHi represents an under-appreciated hallmark of cancer cells, these findings suggest that by alkaline pHi sensing by the PI3K-mTOR axis and AMPK-mTORC2 axes may contribute to tumorigenesis.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L7803
  50. Cells. 2022 Apr 25. pii: 1443. [Epub ahead of print]11(9):
    Transcriptome Reveals Granulosa Cells Coping through Redox, Inflammatory and Metabolic Mechanisms under Acute Heat Stress.
    Abdul Sammad, Hanpeng Luo, Lirong Hu, Huabin Zhu, Yachun Wang.
      Heat stress affects granulosa cells (GCs) and the ovarian follicular microenvironment, causing poor oocyte developmental competence and fertility. This study aimed to investigate the physical responses and global transcriptomic changes in bovine GCs to acute heat stress (43 °C for 2 h) in vitro. Heat-stressed GCs exhibited transient proliferation senescence and resumed proliferation at 48 h post-stress, while post-stress immediate culture-media change had a relatively positive effect on proliferation resumption. Increased accumulation of reactive oxygen species and apoptosis was observed in the heat-stress group. In spite of the upregulation of inflammatory (CYCS, TLR2, TLR4, IL6, etc.), pro-apoptotic (BAD, BAX, TNFSF9, MAP3K7, TNFRSF6B, FADD, TRADD, RIPK3, etc.) and caspase executioner genes (CASP3, CASP8, CASP9), antioxidants and anti-apoptotic genes (HMOX1, NOS2, CAT, SOD, BCL2L1, GPX4, etc.) were also upregulated in heat-stressed GCs. Progesterone and estrogen hormones, along with steroidogenic gene expression, declined significantly, in spite of the upregulation of genes involved in cholesterol synthesis. Out of 12,385 differentially expressed genes (DEGs), 330 significant DEGs (75 upregulated, 225 downregulated) were subjected to KEGG functional pathway annotation, gene ontology enrichment, STRING network analyses and manual querying of DEGs for meaningful molecular mechanisms. High inflammatory response was found to be responsible for oxidative-stress-mediated apoptosis of GCs and nodes towards the involvement of the NF-κB pathway and repression of the Nrf2 pathway. Downregulation of MDM4, TP53, PIDD1, PARP3, MAPK14 and MYC, and upregulation of STK26, STK33, TGFB2, CDKN1A and CDKN2A, at the interface of the MAPK and p53 signaling pathway, can be attributed to transient cellular senescence and apoptosis in GCs. The background working of the AMPK pathway through upregulation of AKT1, AMPK,&amp;nbsp;SIRT1, PYGM, SLC2A4 and SERBP1 genes, and downregulation of PPARGCIA, IGF2,&amp;nbsp;PPARA, SLC27A3, SLC16A3, TSC1/2, KCNJ2, KCNJ16, etc., evidence the repression of cellular transcriptional activity and energetic homeostasis modifications in response to heat stress. This study presents detailed responses of acute-heat-stressed GCs at physical, transcriptional and pathway levels and presents interesting insights into future studies regarding GC adaptation and their interaction with oocytes and the reproductive system at the ovarian level.
    Keywords:  AMPK pathway; NF-κB pathway; apoptosis; granulosa cells; heat stress; oxidative stress; p53 pathway; steroidogenesis
    DOI:  https://doi.org/10.3390/cells11091443
  51. FASEB J. 2022 May;36 Suppl 1
    Electrical Stimulated Glut4 Trafficking Reduces Critical Illness-Associated Muscle Wasting.
    Alex B Addinsall, Nicola Cacciani, Anders Backéus, Yvette Hedstöm, Lars Larsson.
      Critical illness myopathy (CIM) is a debilitating condition characterized by the preferential loss of the motor protein myosin. CIM is a consequence of critical care, impairs recovery and provides long-term complications, and mortality. CIM pathophysiology is complex and remains incompletely understood, however loss of mechanical stimuli appears central to critical illness associated muscle atrophy and weakness. Passive mechanical loading (ML) and electrical stimulation (ES) augment muscle mass and function, however the mechanisms underpinning these therapies are less known. Here we aimed to assess the hypothesis that chronic supramaximal ES would ameliorate CIM in a unique experimental rat model of critical care. Rats were subjected to deep sedation, controlled mechanical ventilation, and immobilization with and without direct soleus ES for 8 days. Critical care reduced soleus muscle mass, cross-sectional area (CSA), Myosin: Actin ratio and single muscle fibre specific force. ES reduced the loss of soleus muscle fibre CSA and Myosin: Actin ratio yet failed to effect specific force. Insulin signalling gene pathway was downregulated following critical care and GLUT4 trafficking was reduced leading to muscle glycogen depletion. ES promoted phosphofructokinase and insulin signalling pathways and maintained GLUT4 translocation and glycogen levels. ES evoked AMPK, but not AKT, signalling pathway, where the downstream target TBC1D4 and AMPK-specific P-TBC1D4 levels tended to be increased. Reduced muscle protein degradation promoted soleus CSA, as ES reduced E3 ligases Atrogin-1 and MuRF1 downstream of AMPK-FoxO3. These results demonstrate chronic supramaximal ES reduces critical care associated muscle wasting, preserved glucose signalling and reduced muscle protein degradation in CIM.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R3369
  52. Fungal Genet Biol. 2022 May 05. pii: S1087-1845(22)00045-7. [Epub ahead of print]161 103701
    Snf1p/Hxk2p/Mig1p pathway regulates hexose transporters transcript levels, affecting the exponential growth and mitochondrial respiration of Saccharomyces cerevisiae.
    Andres Carrillo-Garmendia, Cecilia Martinez-Ortiz, Jairo Getzemani Martinez-Garfias, Susana Estefania Suarez-Sandoval, Juan Carlos González-Hernández, Gerardo M Nava, Miguel David Dufoo-Hurtado, Luis Alberto Madrigal-Perez.
      The Crabtree effect molecular regulation comprehension could help to improve ethanol production with biotechnological purposes and a better understanding of cancer etiology due to its similarity with the Warburg effect. Snf1p/Hxk2p/Mig1p pathway has been linked with the transcriptional regulation of the hexose transporters and phenotypes associated with the Crabtree effect. Nevertheless, direct evidence linking the genetic control of the hexose transporters with modulation of the Crabtree effect phenotypes by the Snf1p/Hxk2p/Mig1p pathway is still lacking. In this sense, we provide evidence that SNF1 and HXK2 genes deletion affects exponential growth, mitochondrial respiration, and transcript levels of hexose transporters in a glucose-dependent manner. The Vmax of the hexose transporters with the high transcript levels was correlated positively with the exponential growth and negatively with the mitochondrial respiration. HXT2 gene transcript levels were the most affected by the deletion of the SNF1/HXK2/MIG1 pathway. Deleting the orthologous genes SNF1 and HXK2 in Kluyveromyces marxianus (Crabtree negative yeast) has an opposite effect compared to Saccharomyces cerevisiae in growth and mitochondrial respiration. Overall, these results indicate that the SNF1/HXK2/MIG1 pathway regulates transcript levels of the hexose transporters, which shows an association with the exponential growth and mitochondrial respiration in a glucose-dependent manner.
    Keywords:  AMP-activated kinase (AMPK); Cell growth; Cell signaling; Mitochondrial metabolism; Mitochondrial respiration; Saccharomyces cerevisiae
    DOI:  https://doi.org/10.1016/j.fgb.2022.103701
  53. J Tradit Complement Med. 2022 Mar;12(2): 195-205
    A novel phenolic formulation for treating hepatic and peripheral insulin resistance by regulating GLUT4-mediated glucose uptake.
    Bean Bu Kang, Been Huang Chiang.
      Chronic insulin resistance suppresses muscle and liver response to insulin, which is partially due to impaired vesicle trafficking. We report here that a formula consisting of resveratrol, ferulic acid and epigallocatechin-3-O-gallate is more effective in ameliorating muscle and hepatic insulin resistance than the anti-diabetic drugs, metformin and AICAR. The formula enhanced glucose transporter-4 (GLUT4) translocation to the plasma membrane in the insulin-resistant muscle cells by regulating both insulin-independent (calcium and AMPK) and insulin-dependent (PI3K) signaling molecules. Particularly, it regulated the subcellular location of GLUT4 through endosomes to increase glucose uptake under insulin-resistant condition. Meanwhile, this phytochemicals combination increased glycogen synthesis and decreased glucose production in the insulin-resistant liver cells. On the other hand, this formula also showed anti-diabetic potential by the reduction of lipid content in the myotubes, hepatocytes, and adipocytes. This study demonstrated that the three phenolic compounds in the formula could work in distinct mechanisms and enhance both insulin-dependent and independent vesicles trafficking and glucose transport mechanisms to improve carbohydrate and lipid metabolism.
    Keywords:  ACC, acetyl-CoA carboxylase; Abbreviations: 2DG, 2-deoxyglucose; BSA, bovine serum albumin; Bt2-cAMP, dibutyryl cAMP; CB, cytochalasin B; DAB, diaminobenzidine; DMEM, Dulbecco's modified Eagle's medium; DMSO, dimethyl sulfoxide; ECL, enhanced chemiluminescence; EGCG, epigallocatechin-3-O-gallate; Endosome; FBS, fetal bovine serum; FER, ferulic acid; GLUT4; GLUT4, glucose transporter-4; Glucose uptake; Hepatic glucose production; IDV, indinavir; IMCL, intramyocellular lipid; ISP, insulin signaling pathway; Insulin resistance; KHB, Krebs-Henseleit buffer; Metabolic syndrome; ORO, Oil Red O; PBS, phosphate-buffered saline; PVDF, polyvinylidene difluoride; Phenolic phytochemicals; RSV, resveratrol; SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis; T2DM, type 2 diabetes mellitus; Tf-HRP, Transferrin conjugated to horseradish peroxidase; Type 2 diabetes; α-MEM, α-minimal essential medium
    DOI:  https://doi.org/10.1016/j.jtcme.2021.08.004
  54. FASEB J. 2022 May;36 Suppl 1
    ST segment elevation: the lost measure of early ischemic preconditioning in the landmark study of Murry et al. 1986.
    Michael Sturek, James Byrd, Mouhamad Alloosh.
       BACKGROUND: In their landmark study Murry et al. (Circulation 74:1124, 1986) showed dramatic ST segment elevation of the ECG during the first occlusion of a dog coronary artery. They made the insightful observation that "… in subsequent occlusions, these changes were slower to develop and frequently did not reach the magnitude seen during the first occlusion." This electrophysiological preconditioning has been almost completely dismissed as irrelevant to ischemic preconditioning in animal models. We found in the Ossabaw miniature swine model of metabolic syndrome that brief coronary occlusion often led to ST elevation that quickly progressed to ventricular tachycardia and fibrillation.
    HYPOTHESIS: An AMPKγ3 mutation in Ossabaw swine will explain the extreme ST elevation and provide evidence for the relevance of ST segment ischemic preconditioning.
    METHODS AND RESULTS: Lean mutants had impaired glucose tolerance compared to wild type Ossabaw swine. Myocardial ischemia induced by balloon occlusion of the circumflex artery elicited profound electrocardiographic ST elevation in lean mutant compared to wild type swine. Infusion of an AMPK agonist, AICAR (0.5 mM), prior to occlusion significantly reduced ST elevation in lean mutants, while completely preventing ST segment elevation in wild type. Ischemic preconditioning of the ST segment changes occurred in wild type pigs during the course of 5 occlusions, but only occurred in mutant pigs with AICAR infusion. Infusion of compound C (30 μM), an AMPK antagonist, prior to occlusion blocked the AICAR-induced attenuation of ST elevation in lean mutants. The KATP channel agonist pinacidil increased ST elevation and the antagonist glibenclamide attenuated ST elevation during occlusion. While the degree of diet-induced MetS (obesity, further impaired glucose tolerance, insulin resistance, and hypertension) was not different between mutant and wild type swine, except for greater plasma lipids in mutants, there was no difference in ST elevation during myocardial occlusion.
    CONCLUSIONS: Lean Ossabaw swine with the V199l AMPK mutation have impaired glucose tolerance and exacerbated KATP channel-mediated ST elevation during myocardial ischemia. Excess calorie diet induces MetS and abolishes cardioprotection in wild type AMPK swine. ST segment changes are an excellent measure of ischemic preconditioning in this model, thus showing the relevance of Murry's landmark discovery.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R6373
  55. FASEB J. 2022 May;36 Suppl 1
    Differences in Cellular Metabolism and Metabolic Regulation between Non-diabetic and Diabetic Human Coronary Artery Endothelial Cells.
    Cynthia M Xu, Catherine Karbasiafshar, Karla M S Braga, Rayane B Teixiera, Frank W Sellke, M R Abid.
       OBJECTIVE: To determine the differences in cellular metabolism and metabolic regulation in non-diabetic and diabetic human coronary artery endothelial cells (HCAEC and DM-HCAEC) to further the understanding of the effects of metabolic syndrome on endothelial dysfunction in the setting of cardiovascular disease.
    METHODS: Both HCAEC (n = 5) and DM-HCAEC (n = 5) were cultured to passage 6. The cells were then starved for 18 hours. After treatment, a Reactive Oxygen Species (ROS) Assay was performed, and cell lysates were made for proteomics analysis and western blotting.
    RESULTS: The ROS assay demonstrated that the total ROS was significantly higher in DM-HCAEC (p<0.0001) than in HCAEC. Proteomics analysis and western blotting showed that DM-HCAECs had significantly altered cellular transport, RNA processing, protein biogenesis, and antioxidant pathways. Proteomics analysis demonstrated that there were about 1,500 proteins significantly upregulated or downregulated in DM-HCAEC compared to HCAEC. Analysis using Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed a significant increase in the "vesicle-mediated transport" pathway and significant decrease in the "mRNA metabolic process" pathway in DM-HCAEC with false discovery rates (FDR) of 1.40 x 10-58 and 4.90 x 10-92 , respectively. Western blot analysis showed a significant changes in phosphorylated AMPK/total AMPK, phosphorylated mTOR (serine 2481), total mTOR, and several enzymes responsible for the regulation or elimination of ROS (Figure 1). The increased levels of AMPK activation and dysregulation of mTOR may be due to the excessive ROS in DM-HCAEC, which are consistent with the ROS assay findings and significant changes in expression of enzymes that control ROS levels. Further research is needed to determine the mechanisms of downstream effects.
    CONCLUSIONS: AMPK and mTOR are key regulators of energy homeostasis, cellular growth and metabolism, and are altered in diabetes in the setting of increased oxidative stress. These findings will guide further experiments to assess the effect of metabolic syndrome and diabetes on therapeutics targeted at improving the outcomes of both acute and chronic myocardial ischemia. This also underlines the need for further development of therapies or the use of current therapies to address the metabolic disturbances found in diabetes to improve clinical outcomes in cardiovascular disease.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R3934
  56. Int J Mol Sci. 2022 May 02. pii: 5047. [Epub ahead of print]23(9):
    Novel Inhibitor for Downstream Targeting of Transforming Growth Factor-β Signaling to Suppress Epithelial to Mesenchymal Transition and Cell Migration.
    Tsugumasa Toma, Hiroshi Tateishi, Kensaku Kawakami, Taha F S Ali, Masahiro Kamo, Kazuaki Monde, Yuta Nakashima, Mikako Fujita, Masami Otsuka.
      Cancer metastasis accounts for most of the mortality associated with solid tumors. However, antimetastatic drugs are not available on the market. One of the important biological events leading to metastasis is the epithelial to mesenchymal transition (EMT) induced by cytokines, namely transforming growth-factor-β (TGF-β). Although several classes of inhibitors targeting TGF-β and its receptor have been developed, they have shown profound clinical side effects. We focused on our synthetic compound, HPH-15, which has shown anti-fibrotic activity via the blockade of the TGF-β Smad-dependent signaling. In this study, 10 μM of HPH-15 was found to exhibit anti-cell migration and anti-EMT activities in non-small-cell lung cancer (NSCLC) cells. Although higher concentrations are required, the anti-EMT activity of HPH-15 has also been observed in 3D-cultured NSCLC cells. A mechanistic study showed that HPH-15 inhibits downstream TGF-β signaling. This downstream inhibition blocks the expression of cytokines such as TGF-β, leading to the next cycle of Smad-dependent and -independent signaling. HPH-15 has AMPK-activation activity, but a relationship between AMPK activation and anti-EMT/cell migration was not observed. Taken together, HPH-15 may lead to the development of antimetastatic drugs with a new mechanism of action.
    Keywords:  Smad; anti-cell migration; anti-epithelial to mesenchymal transition (EMT); non-small-cell lung cancer (NSCLC) cells; transforming growth factor-β (TGF-β)
    DOI:  https://doi.org/10.3390/ijms23095047
  57. Food Funct. 2022 May 12.
    FADS1 overexpression promotes fatty acid synthesis and triacylglycerol accumulation via inhibiting the AMPK/SREBP1 pathway in goat mammary epithelial cells.
    Jiangtao Huang, Yuexin Shao, Xueyang Zong, Huawen Zhang, Xian Zhang, Zhifei Zhang, Huaiping Shi.
      Delta-5 desaturase (D5D), encoded by the fatty acid desaturase 1 (FADS1) gene, is a rate-limiting enzyme in polyunsaturated fatty acid (PUFA) synthesis that influences the PUFA levels in milk fat. However, the function and molecular mechanism of FADS1 in milk fat metabolism remain largely unknown. The FADS1 overexpression increased the triglyceride content, lipid droplet size, and expression of genes related to fatty acid de novo synthesis (SREBP1 and ACC), intracellular fatty acid transporters (FABP3 and FABP4) and triacylglycerol synthesis gene (DGAT2). It also significantly promoted the SREBP1 nuclear translocation by inhibiting the AMPK activation. In addition, FADS1 overexpression inhibited cell proliferation and arrested cell cycle at the G1 phase. These findings reveal a novel FADS1-AMPK-SREBP1 pathway regulating milk fat production in the goat mammary gland.
    DOI:  https://doi.org/10.1039/d2fo00246a
  58. FASEB J. 2022 May;36 Suppl 1
    Agomelatine Ameliorates Obesity-Induced Kidney Injury through the Inhibition of Renal Fibrosis and Improvement of Impaired Autophagy.
    Sasivimon Promsan, Nichakorn Phengpol, Prempree Sutthasupha, Anusorn Lungkaphin.
       INTRODUCTION: Chronic kidney disease is the crucial complication that could occur in obesity. Prolong high-fat diet consumption can stimulate reactive oxygen species (ROS) and contribute to kidney injury through various pathways. Autophagy plays an important role for maintaining intracellular homeostasis. ROS could induce impaired autophagy, subsequently led to kidney damage. Agomelatine (AGOM), a melatonin analogue, is indicated for the treatment of major depressive disorders. The antioxidant and anti-inflammation effects have been reported in AGOM treatment in chronic mild stress-induced depression. Previous study found that AGOM inhibited apoptosis and ER stress in the kidney under high-fat diet condition in rats. However, the effect of AGOM supplementation on renal fibrosis and impaired autophagy in obese condition has never been elucidated.
    AIMS: To evaluate the renoprotective effect of AGOM on kidney function in high-fat diet-induced kidney injury, as well as to explore the underlying mechanisms involving autophagy signaling pathway.
    MATERIALS AND METHODS: Male Wistar rats were received normal diet (ND) or high-fat diet (HF) for 16 weeks. Then, the HF rats were separated into 4 subgroups including (1) HF; (2) agomelatine20 (AGOM20); (3) AGOM40, the rats were received AGOM at the dose of 20 and 40 mg/kg/day, respectively; and (4) N-acetylcysteine (NAC), the rats were received NAC at the dose of 100 mg/kg/day by oral gavage for 4 weeks. After 4 weeks of treatment, all rats were sacrificed. Blood, urine and kidney tissues were collected for further investigations.
    RESULTS: The results demonstrated that HF rats developed kidney dysfunction as shown by the raising in serum creatinine, creatinine clearance along with the changes of glomerular structure and the elevating of positive area of α-SMA, which is renal fibrosis indicator. These alterations were reversed by AGOM and NAC treatment. Moreover, HF rats showed renal oxidative stress as indicated by the increasing in PKCα and NOX4 expression and decreasing in antioxidant enzyme, GCLC. AGOM and NAC administrations reduced oxidative stress and restored antioxidant enzyme. Furthermore, high-fat diet consumption markedly reduced AMPK, along with the elevation of mTOR expression. Additionally, we found that HF rats aggravated impaired autophagy which could induce oxidative stress, fibrosis and finally led to kidney cell death. AGOM administration restored autophagy process as shown by the increasing in Beclin-1, LC3B and Atg5 protein. These results demonstrated that AGOM improved kidney injury through the regulation of oxidative stress, fibrosis and autophagy process.
    CONCLUSIONS: These findings indicated that AGOM prevented kidney injury under obese condition via the inhibition of oxidative stress and fibrosis and improved impaired autophagy via regulating AMPK-mTOR-autophagy signaling pathways.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.0R423
  59. FASEB J. 2022 May;36 Suppl 1
    Characterizing Metabolic Changes in Cancer Cells after Treatment with an Extract Created from Walnuts.
    Codey Y Huang, J B White.
      One of the main hallmarks of cancer cells is the reprogramming of energy metabolism to support the high energy and metabolite demands of constant proliferation. Although previously observed as mainly a shift from oxidative phosphorylation to glycolysis, also known as the Warburg Effect, recent research has shown that cancer cells can utilize both pathways to make energy, which has been referred to as metabolic flux. This suggests that cancer cells can adapt to their environment by reactivating or continuing to use the oxidative phosphorylation pathway to make energy. Thus, the energy metabolism of cancer cells is of therapeutic interest. The triple-negative breast cancer cell line MDA-MB-231 can undergo metabolic flux, making it an ideal cell line to study the effects of metabolism-altering drugs. Previous work from our lab showed that treatment with a Walnut Extract (WE) induced cell death in MDA-MB-231 cells, as well as HeLa cells. We also observed that the WE treatment targets the mitochondria by destabilizing the mitochondrial outer membrane potential, suggesting that the extract may be able to impact oxidative phosphorylation. To examine the effects on energy production, we have taken advantage of the Agilent Seahorse XFe96 Cell Metabolic Analyzer to monitor, in real time, metabolic changes in MDA-MB-231 and HeLa cells treated with WE. WE treatment had impacts on both glycolytic and oxidative phosphorylation processes in both cell lines. Both time- and dose-dependent responses were observed, with WE treated MDA-MB-231 cells exhibiting a 70% reduction in basal oxygen consumption rate compared to vehicle treated cells. Additionally, basal glycolytic rate was reduced by up to 41%, and compensatory glycolysis was reduced 70%. In HeLa cells, basal oxygen consumption rate was reduced by 74%, with a 56% reduction in basal glycolytic rate and a 76% reduction in compensatory glycolysis. Quantification of ATP production rate showed an overall reduction of 73%, with a 91% reduction in mitochondrially contributed ATP production, as well as a 47% reduction in glycolytic ATP production in MDA-MB-231 cells. Quantitation of total ATP within the cell using a separate assay shows an 62% decrease in ATP in treated HeLa cells. We have also looked at the status of mTOR and AMPK in the cells after treatment with the WE, because of their role in energy sensing and energy production. Interestingly, our preliminary data suggests that phosphorylated mTOR levels do not change and we also do not detect any phosphorylation of AMPK, suggesting that the WE is not effecting its changes on these proteins despite the reduction in overall ATP in the cell. Taken together, our results suggest that the WE treatment can alter and reduce the rates of glycolysis and oxidative phosphorylation, and by extension, the metabolic flux of these cancer cells.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.R2465
  60. Int J Mol Sci. 2022 May 06. pii: 5209. [Epub ahead of print]23(9):
    Chronic Intermittent Hypoxia Exposure Alternative to Exercise Alleviates High-Fat-Diet-Induced Obesity and Fatty Liver.
    Yunfei Luo, Qiongfeng Chen, Junrong Zou, Jingjing Fan, Yuanjun Li, Zhijun Luo.
      Obesity often concurs with nonalcoholic fatty liver disease (NAFLD), both of which are detrimental to human health. Thus far, exercise appears to be an effective treatment approach. However, its effects cannot last long and, moreover, it is difficult to achieve for many obese people. Thus, it is necessary to look into alternative remedies. The present study explored a noninvasive, easy, tolerable physical alternative. In our experiment, C57BL/6 mice were fed with a high-fat diet (HFD) to induce overweight/obesity and were exposed to 10% oxygen for one hour every day. We found that hypoxia exerted protective effects. First, it offset HFD-induced bodyweight gain and insulin resistance. Secondly, hypoxia reversed the HFD-induced enlargement of white and brown adipocytes and fatty liver, and protected liver function. Thirdly, HFD downregulated the expression of genes required for lipolysis and thermogenesis, such as UCP1, ADR3(beta3-adrenergic receptor), CPT1A, ATGL, PPARα, and PGC1α, M2 macrophage markers arginase and CD206 in the liver, and UCP1 and PPARγ in brown fat, while these molecules were upregulated by hypoxia. Furthermore, hypoxia induced the activation of AMPK, an energy sensing enzyme. Fourthly, our results showed that hypoxia increased serum levels of epinephrine. Indeed, the effects of hypoxia on bodyweight, fatty liver, and associated changes in gene expression ever tested were reproduced by injection of epinephrine and prevented by propranolol at varying degrees. Altogether, our data suggest that hypoxia triggers stress responses where epinephrine plays important roles. Therefore, our study sheds light on the hope to use hypoxia to treat the daunting disorders, obesity and NAFLD.
    Keywords:  AMPK; chronic intermittent hypoxia; epinephrine; fatty liver; inflammation; obesity
    DOI:  https://doi.org/10.3390/ijms23095209
  61. Acta Pharm Sin B. 2022 Mar;12(3): 1322-1338
    Anthelmintics nitazoxanide protects against experimental hyperlipidemia and hepatic steatosis in hamsters and mice.
    Fengfeng Li, Man Jiang, Minghui Ma, Xuyang Chen, Yidan Zhang, Yixin Zhang, Yuanyuan Yu, Yunfeng Cui, Jiahui Chen, Hui Zhao, Zhijie Sun, Deli Dong.
      Lipid metabolism disorders contribute to hyperlipidemia and hepatic steatosis. It is ideal to develop drugs simultaneous improving both hyperlipidemia and hepatic steatosis. Nitazoxanide is an FDA-approved oral antiprotozoal drug with excellent pharmacokinetic and safety profile. We found that nitazoxanide and its metabolite tizoxanide induced mild mitochondrial uncoupling and subsequently activated AMPK in HepG2 cells. Gavage administration of nitazoxanide inhibited high-fat diet (HFD)-induced increases of liver weight, blood and liver lipids, and ameliorated HFD-induced renal lipid accumulation in hamsters. Nitazoxanide significantly improved HFD-induced histopathologic changes of hamster livers. In the hamsters with pre-existing hyperlipidemia and hepatic steatosis, nitazoxanide also showed therapeutic effect. Gavage administration of nitazoxanide improved HFD-induced hepatic steatosis in C57BL/6J mice and western diet (WD)-induced hepatic steatosis in Apoe -/- mice. The present study suggests that repurposing nitazoxanide as a drug for hyperlipidemia and hepatic steatosis treatment is promising.
    Keywords:  AMPK; Autophagy; Hepatic steatosis; Hyperlipidemia; Mitochondrial uncoupling; Nitazoxanide; SQSTM1/P62; Tizoxanide
    DOI:  https://doi.org/10.1016/j.apsb.2021.09.009
  62. FASEB J. 2022 May;36 Suppl 1
    Mitochondrial Uncoupling Decreases Sarcopenic Obesity By Activation of Skeletal Muscle Mitochondrial Quality Control and Attenuated ER Stress.
    Wagner S Dantas, Elizabeth R M Zunica, Elizabeth C Heintz, Charles L Hoppel, Christopher L Axelrod, John P Kirwan.
       BACKGROUND: Sarcopenic obesity is a highly prevalent disease with poor survival and ineffective medical interventions. Mitochondrial dysfunction is purported to be central in the pathogenesis of sarcopenic obesity by impairing both organelle biogenesis and quality control. We have previously identified an orally available mitochondrial-targeted furazano[3,4-b]pyrazine named BAM15 that selectively lowers respiratory coupling efficiency and protects against diet-induced obesity in mice. Here, we tested the hypothesis that mitochondrial uncoupling simultaneously attenuates loss of muscle function and weight gain in a mouse model of sarcopenic obesity.
    METHODS: 80-week-old male C57BL/6J mice with obesity were randomized to 10 weeks of high fat diet (CTRL) or BAM15 (BAM15; 0.1% w/w in high fat diet) treatment. Body composition, muscle function, energy expenditure, and locomotor activity were determined after treatment. Skeletal muscle was harvested and evaluated for histology, gene expression, protein signaling, and mitochondrial structure and function.
    RESULTS: BAM15 decreased body weight (~25% reduction, P<0.001) which was attributable to increased energy expenditure (~20% increment, P<0.001). BAM15 increased muscle mass (~13% increment, P<0.001), strength (~37% increment, P<0.0001), and locomotor activity (~25% increment, P<0.001). Improvements in physical function were mediated in part by reductions in skeletal muscle inflammation (IL-6 and gp130, both P<0.05), enhanced mitochondrial function, and improved endoplasmic reticulum homeostasis and reduced inflammation. Specifically, BAM15 activated mitochondrial quality control through AMPK (PINK1-ubiquitin binding and LC3II, P<0.01), increased electron transport chain activity (citrate synthase and complex II activity, all P<0.05), restricted endoplasmic reticulum (ER) misfolding (decreased oligomer A11 insoluble/soluble ratio, P<0.0001) while limiting ER stress (decreased PERK signaling, P<0.0001), apoptotic signaling (decreased cytochrome C release and Caspase-3/9 activation, all P<0.001), and muscle protein degradation (decreased 14-kDa actin fragment insoluble/soluble ratio, P<0.001).
    CONCLUSIONS: Mitochondrial uncoupling agents such as BAM15 may mitigate age-related decline in muscle mass and function by molecular and cellular bioenergetic adaptations that confer protection against sarcopenic obesity through activation of mitochondrial quality control and attenuation of ER stress.
    DOI:  https://doi.org/10.1096/fasebj.2022.36.S1.L7532
  63. Acta Pharm Sin B. 2022 Mar;12(3): 1271-1287
    Targeting PFKL with penfluridol inhibits glycolysis and suppresses esophageal cancer tumorigenesis in an AMPK/FOXO3a/BIM-dependent manner.
    Cancan Zheng, Xiaomei Yu, Yiyao Liang, Yidong Zhu, Yan He, Long Liao, Dingkang Wang, Yanming Yang, Xingfeng Yin, Ang Li, Qingyu He, Bin Li.
      As one of the hallmarks of cancer, metabolic reprogramming leads to cancer progression, and targeting glycolytic enzymes could be useful strategies for cancer therapy. By screening a small molecule library consisting of 1320 FDA-approved drugs, we found that penfluridol, an antipsychotic drug used to treat schizophrenia, could inhibit glycolysis and induce apoptosis in esophageal squamous cell carcinoma (ESCC). Gene profiling and Ingenuity Pathway Analysis suggested the important role of AMPK in action mechanism of penfluridol. By using drug affinity responsive target stability (DARTS) technology and proteomics, we identified phosphofructokinase, liver type (PFKL), a key enzyme in glycolysis, as a direct target of penfluridol. Penfluridol could not exhibit its anticancer property in PFKL-deficient cancer cells, illustrating that PFKL is essential for the bioactivity of penfluridol. High PFKL expression is correlated with advanced stages and poor survival of ESCC patients, and silencing of PFKL significantly suppressed tumor growth. Mechanistically, direct binding of penfluridol and PFKL inhibits glucose consumption, lactate and ATP production, leads to nuclear translocation of FOXO3a and subsequent transcriptional activation of BIM in an AMPK-dependent manner. Taken together, PFKL is a potential prognostic biomarker and therapeutic target in ESCC, and penfluridol may be a new therapeutic option for management of this lethal disease.
    Keywords:  DARTS technology; Drug repurposing; Esophageal cancer; Glycolysis; Metabolic reprogramming; PFKL; Penfluridol
    DOI:  https://doi.org/10.1016/j.apsb.2021.09.007
  64. Indian J Pharmacol. 2022 Mar-Apr;54(2):54(2): 118-125
    Tert-Butylhydroquinone alleviates insulin resistance and liver steatosis in diabetes.
    Tian-Tian Zhu, Chao-Nan Zhu, Ning Huang, Xin Yu, Guang-Rui Wan, Shuang-Xi Wang, Ping Song, Jian Xu, Peng Li, Ya-Ling Yin.
       OBJECTIVES: This work aimed to determine tert-Butylhydroquinone (TBHQ)'s effects on insulin resistance (IR) and liver steatosis in diabetic animals and to explore the underpinning mechanisms.
    MATERIALS AND METHODS: Male ApoE-/-mice underwent streptozocin (STZ) administration while receiving a sucrose/fat-rich diet for type 2 diabetes mellitus (T2DM) establishment. This was followed by a 6-week TBHQ administration. Body weight, fasting (FBG) and postprandial (PBG) blood glucose amounts, and insulin concentrations were measured, and the oral glucose tolerance test (OGTT) was carried out. Hematoxylin and eosin (H and E) staining and immunoblot were carried out for assessing histology and protein amounts in the liver tissue samples. In addition, cultured HepG2 cells were administered HClO and insulin for IR induction, and immunoblot was carried out for protein evaluation. Finally, the cells were stained with the Hoechst dye for apoptosis evaluation.
    RESULTS: The model animals showed T2DM signs, and TBHQ decreased FBG, ameliorated glucose tolerance and reduced liver steatosis in these animals. In addition, TBHQ markedly upregulated AMPKα2, GLUT4 and GSK3 β, as well as phosphorylated PI3K and AKT in the liver of mice with T2DM. In agreement, TBHQ decreased HClO-and insulin-related IR in cells and suppressed apoptosis through AMPKα2/PI3K/AKT signaling.
    CONCLUSIONS: TBHQ alleviates IR and liver steatosis in a mouse model of T2DM likely through AMPKα2/PI3K/AKT signaling.
    Keywords:  AMPKα2; insulin resistance; liver steatosis; tert-Butylhydroquinone; type 2 diabetes mellitus
    DOI:  https://doi.org/10.4103/ijp.ijp_440_21
This page is being maintained by Thomas Krichel. It was last updated on 2024‒11‒22 at 4:11.