bims-amsmem 2022-02-27 papers

bims-amsmem

Biomed News

on AMPK signaling mechanism in energy metabolism

Issue of 2022‒02‒27
thirty papers selected by
Dipsikha Biswas, Københavns Universitet

Role of the AMPK signalling pathway in the aetiopathogenesis of ocular diseases.
AMP- activated protein kinase (AMPK) promotes breast cancer stemness and drug resistance.
Chitosan oligosaccharide attenuates hepatic steatosis in HepG2 cells via the activation of AMP-activated protein kinase.
Chronic Treatment with Metformin Has No Disrupting Effect on the Hepatic Circadian Clock in Mice.
Intestinal Epithelial AMPK Deficiency Causes Delayed Colonic Epithelial Repair in DSS-Induced Colitis.
ABCA8 inhibits breast cancer cell proliferation by regulating the AMP activated protein kinase/mammalian target of rapamycin signaling pathway.
Metformin Inhibits ROS Production by Human M2 Macrophages via the Activation of AMPK.
Anti-Obesity Effects of Polymethoxyflavone-Rich Fraction from Jinkyool (Citrus sunki Hort. ex Tanaka) Leaf on Obese Mice Induced by High-Fat Diet.
Renal tubule ectopic lipid deposition in diabetic kidney disease rat model and in vitro mechanism of leptin intervention.
Inorganic nitrate and nitrite ameliorate kidney fibrosis by restoring lipid metabolism via dual regulation of AMP-activated protein kinase and the AKT-PGC1α pathway.
Low-dose metformin targets the lysosomal AMPK pathway through PEN2.
10-Gingerol, a natural AMPK agonist, suppresses neointimal hyperplasia and inhibits vascular smooth muscle cell proliferation.
Pasteurized Akkermansia muciniphila Ameliorate the LPS-Induced Intestinal Barrier Dysfunction via Modulating AMPK and NF-κB through TLR2 in Caco-2 Cells.
Vitamin D protects against high glucose-induced pancreatic β-cell dysfunction via AMPK-NLRP3 inflammasome pathway.
Metformin attenuated sepsis-related liver injury by modulating gut microbiota.
lncRNA TUG1 protects intestinal epithelial cells from damage induced by high glucose and high fat via AMPK/SIRT1.
SIRT3-AMPK signaling pathway as a protective target in endothelial dysfunction of early sepsis.
The Natural Chemotherapeutic Capsaicin Activates AMPK through LKB1 Kinase and TRPV1 Receptors in Prostate Cancer Cells.
Heat-Killed Enterococcus faecalis EF-2001 Attenuate Lipid Accumulation in Diet-Induced Obese (DIO) Mice by Activating AMPK Signaling in Liver.
Comment on De Wendt et al. Contraction-Mediated Glucose Transport in Skeletal Muscle Is Regulated by a Framework of AMPK, TBC1D1/4, and Rac1. Diabetes 2021;70:2796-2809.
Treatments targeting autophagy ameliorate the age-related macular degeneration phenotype in mice lacking APOE (apolipoprotein E).
Response to Comment on Wendt et al. Contraction-Mediated Glucose Transport in Skeletal Muscle Is Regulated by a Framework of AMPK, TBC1D1/4, and Rac1. Diabetes 2021;70:2796-2809.
Garcinia Cambogia Improves High-Fat Diet-Induced Glucose Imbalance By Enhancing Calcium/CaMKII/AMPK/GLUT4-Mediated Glucose Uptake In Skeletal Muscle.
Metabolic Phenotyping in Mice with NASH Using Indirect Calorimetry.
Statins Aggravate the Risk of Insulin Resistance in Human Muscle.
Metformin's Mechanism of Action Is Stimulation of the Biosynthesis of the Natural Cyclic AMP Antagonist Prostaglandylinositol Cyclic Phosphate (Cyclic PIP).
MARK2/4 promotes Warburg effect and cell growth in non-small cell lung carcinoma through the AMPKα1/mTOR/HIF-1α signaling pathway.
Salt-inducible kinases inhibitor HG-9-91-01 targets RIPK3 kinase activity to alleviate necroptosis-mediated inflammatory injury.
Nrf2 activation reprograms macrophage intermediary metabolism and suppresses the type I interferon response.
Thioredoxin upregulation delays diabetes-induced photoreceptor cell degeneration via AMPK-mediated autophagy and exosome secretion.

Hum Exp Toxicol. 2022 Jan-Dec;41:41 9603271211063165

Role of the AMPK signalling pathway in the aetiopathogenesis of ocular diseases.

Dhaval K Shukal, Pooja B Malaviya, Tusha Sharma.

  BACKGROUND: AMP-activated protein kinase (AMPK) plays a precise role as a master regulator of cellular energy homeostasis. AMPK is activated in response to the signalling cues that exhaust cellular ATP levels such as hypoxia, ischaemia, glucose depletion and heat shock. As a central regulator of both lipid and glucose metabolism, AMPK is considered to be a potential therapeutic target for the treatment of various diseases, including eye disorders.OBJECTIVE: To review all the shreds of evidence concerning the role of the AMPK signalling pathway in the pathogenesis of ocular diseases.
METHOD: Scientific data search and review of available information evaluating the influence of AMPK signalling on ocular diseases.
RESULTS: Review highlights the significance of AMPK signalling in the aetiopathogenesis of ocular diseases, including cataract, glaucoma, diabetic retinopathy, retinoblastoma, age-related macular degeneration, corneal diseases, etc. The review also provides the information on the AMPK-associated pathways with reference to ocular disease, which includes mitochondrial biogenesis, autophagy and regulation of inflammatory response.
CONCLUSION: The study concludes the role of AMPK in ocular diseases. There is growing interest in the therapeutic utilization of the AMPK pathway for ocular disease treatment. Furthermore, inhibition of AMPK signalling might represent more pertinent strategy than AMPK activation for ocular disease treatment. Such information will guide the development of more effective AMPK modulators for ocular diseases.[Formula: see text].

Keywords:  AMPK; cataract; diabetic retinopathy; glaucoma; ocular disease; retinoblastoma; super kinases

DOI:  https://doi.org/10.1177/09603271211063165
Dis Model Mech. 2022 Feb 23. pii: dmm.049203. [Epub ahead of print]

AMP- activated protein kinase (AMPK) promotes breast cancer stemness and drug resistance.

Sai Balaji Andugulapati, Ananthalakshmy Sundararaman, Mohini Lahiry, Annapoorni Rangarajan.

  Breast Cancer Stem Cells (BCSCs) are a major cause of therapy resistance and tumour progression. Currently, their regulation is not entirely understood. Previous work from our lab demonstrated context-specific pro-tumorigenic role of AMP-activated protein kinase (AMPK) in breast cancer cell survival under anchorage-deprivation and mammosphere formation hallmarks of BCSCs. We therefore investigated the role of AMPK in the maintenance of BCSC state/function. AMPK depletion reduces serial sphere formation in vitro and tumour initiation in vivo. Intriguingly, tumour-derived cell analysis using stem cell markers and functional assays revealed that AMPK is required for the maintenance of BCSC population in vivo. AMPK promotes the expression of stemness genes like Nanog, Sox2 and Bmi1 through the transcriptional upregulation of Twist via promoter acetylation. Further, AMPK-driven stemness plays a critical role in resistance to doxorubicin. Significantly, we found that AMPK activity increased after chemotherapy in patient-derived tumour samples alongside an increase in stemness markers. Importantly, AMPK depletion sensitises mice tumours to doxorubicin treatment. Our work indicates that targeting AMPK in conjunction with regular chemotherapy is likely to reduce the stem cell pool and improve chemosensitivity in breast cancers.

Keywords:  AMPK; Chemotherapy; Drug resistance; Stemness; Twist

DOI:  https://doi.org/10.1242/dmm.049203
J Food Biochem. 2022 Feb 20. e14045

Chitosan oligosaccharide attenuates hepatic steatosis in HepG2 cells via the activation of AMP-activated protein kinase.

Tiange Li, Han Gong, Biyuan Zhan, Xueying Mao.

  Chitosan oligosaccharides (COSs), oligomers of decomposed chitosan possess many biological functions including immunomodulatory, antitumor, and antiinflammation. The aim of this study was to investigate the protective effects of COS against free fatty acid (FFA)-induced cellular hepatic steatosis and underlying mechanisms in HepG2 cells. Results showed that COS significantly reduced the lipid contents and elevated the activities of antioxidant enzymes including total-superoxide dismutase, glutathione peroxidase, and catalase in FFA-stimulated HepG2 cells. COS phosphorylated the acetyl-CoA carboxylase and reduced both mRNA and protein levels of lipogenesis markers including fatty acid synthase and sterol regulatory element-binding protein 1c. COS also significantly increased the expression levels of fatty acid oxidation-related factors including carnitine palmitoyltransferase 1A, acyl-coenzyme A oxidase 1, and peroxisome proliferators-activated receptor α. Besides, COS markedly phosphorylated AMP-activated protein kinase (AMPK). The inhibition of lipogenesis and the enhancement of fatty acid oxidation induced by COS were all blocked by AMPK antagonist (compound C), showing that the attenuation of hepatic steatosis by COS was dependent on AMPK activation. In conclusion, COS attenuated hepatic steatosis via suppressing lipid synthesis and enhancing fatty acid oxidation. AMPK was also involved in the alleviation of hepatic steatosis by COS. These results indicated that COS might be used as a potential ingredient to ameliorate nonalcoholic fatty liver disease. PRACTICAL APPLICATIONS: Nonalcoholic fatty liver disease (NAFLD) has been regarded as pathological fat deposition in the liver, which includes a range of pathologies, from steatosis to steatohepatitis, fibrosis, and cellular carcinoma. Our findings demonstrated that Chitosan oligosaccharides (COS) attenuated steatosis via improving lipid metabolism. COS suppressed lipogenesis and also enhanced fatty acid oxidation. Besides, the underlying molecular mechanism whereby COS elicited these beneficial effects has also been proved to be through the modulation of upstream protein kinase, AMP-activated protein kinase. This study provides new knowledge to support that COS might be used as a food supplement for the prevention of NAFLD.

Keywords:  AMP-activated protein kinase; HepG2 cells; chitosan oligosaccharide; fatty acid oxidation; lipogenesis; steatosis

DOI:  https://doi.org/10.1111/jfbc.14045
Medicina (Kaunas). 2022 Feb 15. pii: 293. [Epub ahead of print]58(2):

Chronic Treatment with Metformin Has No Disrupting Effect on the Hepatic Circadian Clock in Mice.

Nazmul Hasan, Koki Sugimoto, Koki Yamada, Jun-Ichi Morishige, Kentaro Ushijima, Akio Fujimura, Naoto Nagata, Hitoshi Ando.

  Background and Objectives: The antidiabetic agent metformin is known to activate AMP-activated protein kinase (AMPK) in various tissues. Because AMPK can modulate intracellular circadian clocks through regulating the stability of clock components, a single dose of metformin has been reported to affect circadian clocks in the peripheral tissues. In this study, therefore, we investigated whether chronic treatment with metformin causes the impairment of circadian clocks, especially if given at an inappropriate time. Materials and Methods: Non-diabetic C57BL/6J mice were allowed access to food only during 4 h at the beginning of the dark period, and repeatedly i.p. injected with a nearly maximum non-toxic dose of metformin, once daily either at 4 h after the beginning of the dark period or at the beginning of the light period. Diabetic ob/ob mice were given free access to food and treated with metformin in drinking water. Results: Under the controlled feeding regimen, 8-day treatment with metformin did not alter the mRNA expression rhythms of clock genes in both liver and adipose tissue of C57BL/6J mice, regardless of dosing time. In addition, chronic treatment with metformin for 2 weeks affected hepatic AMPK activation rhythm but did not disrupt the circadian clocks in the liver and adipose tissues of the ob/ob mice. Conclusions: These results mitigate concerns that treatment with metformin impairs peripheral circadian clocks, although confirmation is needed in humans.

Keywords:  circadian rhythm; clock genes; liver; metformin; type 2 diabetes

DOI:  https://doi.org/10.3390/medicina58020293
Cells. 2022 Feb 09. pii: 590. [Epub ahead of print]11(4):

Intestinal Epithelial AMPK Deficiency Causes Delayed Colonic Epithelial Repair in DSS-Induced Colitis.

Séverine Olivier, Hanna Diounou, Camille Pochard, Lisa Frechin, Emilie Durieu, Marc Foretz, Michel Neunlist, Malvyne Rolli-Derkinderen, Benoit Viollet.

  Dysfunctions in the intestinal barrier, associated with an altered paracellular pathway, are commonly observed in inflammatory bowel disease (IBD). The AMP-activated protein kinase (AMPK), principally known as a cellular energy sensor, has also been shown to play a key role in the stabilization and assembly of tight junctions. Here, we aimed to investigate the contribution of intestinal epithelial AMPK to the initiation, progression and resolution of acute colitis. We also tested the hypothesis that protection mediated by metformin administration on intestinal epithelium damage required AMPK activation. A dextran sodium sulfate (DSS)-induced colitis model was used to assess disease progression in WT and intestinal epithelial cell (IEC)-specific AMPK KO mice. Barrier integrity was analyzed by measuring paracellular permeability following dextran-4kDa gavage and pro-inflammatory cytokines and tight junction protein expression. The deletion of intestinal epithelial AMPK delayed intestinal injury repair after DSS exposure and was associated with a slower re-epithelization of the intestinal mucosa coupled with severe ulceration and inflammation, and altered barrier function. Following intestinal injury, IEC AMPK KO mice displayed a lower goblet cell counts with concomitant decreased Muc2 gene expression, unveiling an impaired restitution of goblet cells and contribution to wound healing process. Metformin administration during the recovery phase attenuated the severity of DSS-induced colitis through improvement in intestinal repair capacity in both WT and IEC AMPK KO mice. Taken together, these findings demonstrate a critical role for IEC-expressed AMPK in regulating mucosal repair and epithelial regenerative capacity following acute colonic injury. Our studies further underscore the therapeutic potential of metformin to support repair of the injured intestinal epithelium, but this effect is conferred independently of intestinal epithelial AMPK.

Keywords:  AMPK; epithelium repair; goblet cells; inflammatory bowel disease; intestinal barrier integrity; metformin; pro-inflammatory cytokines

DOI:  https://doi.org/10.3390/cells11040590
Environ Toxicol. 2022 Feb 22.

ABCA8 inhibits breast cancer cell proliferation by regulating the AMP activated protein kinase/mammalian target of rapamycin signaling pathway.

Chunliu Lv, Han Yang, Jinsong Yu, Xiaowei Dai.

  ATP-binding cassette (ABC) subfamily A member 8 (ABCA8) has been reported to play a vital role in cancer development. Our study aimed to explore the role and the molecular mechanism of ABCA8 in breast cancer (BC) progression. GSE65194, GSE15852, and GSE45827 datasets were used to identify differentially expressed genes (DEGs) in BC. The diagnosis and prognosis value were determined using ROC curve analysis and Kaplan-Meier plotter, respectively. The relationship between ABCA8 expression and clinicopathological features in BC was analyzed by TCGA. Co-expressed genes of ABCA8 in BC were screened out through GEPIA and subjected to KEGG pathway enrichment analysis. Cell proliferation was evaluated by CCK-8 and EdU incorporation assays. Proliferating cell nuclear antigen (PCNA) expression and the changes of the AMP activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway were measured by western blot analysis. Totally 4 overlapping DEGs were identified and all reduced in BC samples. ABCA8 with high diagnostic and prognostic values was selected for further exploration. Low ABCA8 expression was correlated with clinicopathological features in BC patients. ABCA8 overexpression inhibited BC cell proliferation. The top 20 co-expressed genes of ABCA8 were identified by GEPIA and significantly enriched in AMPK signaling pathway. Inhibition of AMPK/mTOR pathway reversed the suppressive effect of ABCA8 on BC cell growth. These results suggested that ABCA8 overexpression repressed BC cell proliferation through regulating the AMPK/mTOR signaling pathway.

Keywords:  ABCA8; AMPK/mTOR signaling pathway; breast cancer; cell proliferation; prognosis

DOI:  https://doi.org/10.1002/tox.23495
Biomedicines. 2022 Jan 29. pii: 319. [Epub ahead of print]10(2):

Metformin Inhibits ROS Production by Human M2 Macrophages via the Activation of AMPK.

Rana M Nassif, Elias Chalhoub, Pia Chedid, Margarita Hurtado-Nedelec, Elia Raya, Pham My-Chan Dang, Jean-Claude Marie, Jamel El-Benna.

  Metformin (1,1-dimethylbiguanide hydrochloride) is the most commonly used drug to treat type II diabetic patients. It is believed that this drug has several other beneficial effects, such as anti-inflammatory and anticancer effects. Here, we wanted to evaluate the effect of metformin on the production of reactive oxygen species (ROS) by human macrophages. Macrophages are generated in vivo from circulating monocytes depending on the local tissue environment. In vitro proinflammatory macrophages (M1) and anti-inflammatory macrophages (M2) can be generated by culturing monocytes in the presence of different cytokines, such as GM-CSF or M-CSF, respectively. We show that metformin selectively inhibited human monocyte differentiation into proinflammatory macrophages (M1) without inhibiting their differentiation into anti-inflammatory macrophages (M2). Moreover, we demonstrate that, in response to LPS, M2 macrophages produced ROS, which could be very harmful for nearby tissues, and metformin inhibited this process. Interestingly, metformin with LPS induced activation of the adenosine-monophosphate-activated protein kinase (AMPK) and pharmacological activation of AMPK by AICAR, a known AMPK activator, decreased ROS production, whereas the deletion of AMPK in mice dramatically enhanced ROS production in different types of immune cells. These results suggest that metformin exhibits anti-inflammatory effects by inhibiting the differentiation of human monocytes into M1 macrophages and by limiting ROS production by macrophages via the activation of AMPK.

Keywords:  AMPK; NADPH oxidase; NOX2; ROS; inflammation; macrophage; metformin

DOI:  https://doi.org/10.3390/biomedicines10020319
Nutrients. 2022 Feb 18. pii: 865. [Epub ahead of print]14(4):

Anti-Obesity Effects of Polymethoxyflavone-Rich Fraction from Jinkyool (Citrus sunki Hort. ex Tanaka) Leaf on Obese Mice Induced by High-Fat Diet.

Yeong-Jun Jin, Mi-Gyeong Jang, Jae-Won Kim, Songyee Baek, Hee-Chul Ko, Sung-Pyo Hur, Se-Jae Kim.

  Polymethoxyflavones (PMFs) are flavonoids exclusively found in certain citrus fruits and have been reported to be beneficial to human health. Most studies have been conducted with PMFs isolated from citrus peels, while there is no study on PMFs isolated from leaves. In this study, we prepared a PMF-rich fraction (PRF) from the leaves of Citrus sunki Hort ex. Tanaka (Jinkyool) and investigated whether the PRF could improve metabolic decline in obese mice induced by a high-fat diet (HFD) for 5 weeks. The HFD-induced obese mice were assigned into HFD, OR (HFD + orlistat at 15.6 mg/kg of body weight/day), and PRF (HFD + 50, 100, and 200 mg/kg of body weight/day) groups. Orlistat and PRF were orally administered for 5 weeks. At the end of the experiment, the serum biochemical parameters, histology, and gene expression profiles in the tissues of each group were analyzed. The body weight gain of the obese mice was significantly reduced after orlistat and PRF administration for 5 weeks. PRF effectively improved HFD-induced insulin resistance and dyslipidemia. Histological analysis in the liver demonstrated that PRF decreased adipocyte size and potentially improved the liver function, as it inhibited the incidence of fatty liver. PRF activated AMP-activated protein kinase (AMPK), acetyl-CoA carboxylase (ACC), and hormone-sensitive lipase (HSL) in HFD-induced obese mice. Moreover, liver transcriptome analysis revealed that PRF administration enriched genes mainly related to fatty-acid metabolism and immune responses. Overall, these results suggest that the PRF exerted an anti-obesity effect via the modulation of lipid metabolism.

Keywords:  C57BL/6; Citrus sunki; anti-obesity; fatty-acid oxidation; high-fat diet; lipolysis; polymethoxyflavones

DOI:  https://doi.org/10.3390/nu14040865
J Physiol Biochem. 2022 Feb 22.

Renal tubule ectopic lipid deposition in diabetic kidney disease rat model and in vitro mechanism of leptin intervention.

Shasha Liu, Jingjing Da, Jiali Yu, Rong Dong, Jing Yuan, Fuxun Yu, Yan Zha.

  Diabetic kidney disease (DKD) is a major health burden closely related to lipid metabolism disorders. Leptin has lipid-lowering efficacy, but the specific mechanism of its local effects on kidney is still unclear. This study aims to investigate the role of ectopic lipid deposition (ELD) in DKD and evaluate the lipid-lowering efficacy of leptin in the palmitic acid (PA)-induced renal tubular epithelial cells (NRK-52E). DKD model was established in Sprague-Dawley (SD) rats by giving single intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) after high-fat diet for 8 weeks. Then, the expression changes of lipid metabolism-related markers were observed. At week 12, the protein expression level of lipid-deposited marker adipose differentiation-related protein (ADRP) was significantly increased. Besides, the lipid synthesis marker sterol regulatory element-binding protein 1c (SREBP 1c) was highly expressed while the expression of insulin-induced gene 1 (Insig-1), a key molecular of inhibiting SREBP 1c, was decreased. Leptin and compound c were incubated with the PA-induced NRK-52E cells to investigate the lipid-lowering effects and whether this effect was mediated by the AMPK/Insig-1/SREBP 1c signaling pathways. mRNA and protein of ADRP and SREBP 1c were reduced after leptin treatment, while Insig-1 and phosphorylated AMP-activated protein kinase (AMPK) were increased. Conversely, inhibition of AMPK phosphorylation by compound c mostly eliminated lipid-lowering efficacy of leptin in PA-induced cells. Collectively, these results suggested that there was ELD of renal tubular epithelial cells in DKD rats. Leptin upregulated the expression level of Insig-1 by activating AMPK to attenuate ELD in PA-induced NRK-52E cells.

Keywords:  AMP-activated protein kinase; Diabetic kidney disease; Ectopic lipid deposition; Insulin-induced gene 1; Leptin

DOI:  https://doi.org/10.1007/s13105-022-00874-9
Redox Biol. 2022 Feb 17. pii: S2213-2317(22)00038-6. [Epub ahead of print]51 102266

Inorganic nitrate and nitrite ameliorate kidney fibrosis by restoring lipid metabolism via dual regulation of AMP-activated protein kinase and the AKT-PGC1α pathway.

Xuechen Li, Zhengbing Zhuge, Lucas Rannier R A Carvalho, Valdir A Braga, Ricardo Barbosa Lucena, Shuijie Li, Tomas A Schiffer, Huirong Han, Eddie Weitzberg, Jon O Lundberg, Mattias Carlström.

  BACKGROUND: Renal fibrosis, associated with oxidative stress and nitric oxide (NO) deficiency, contributes to the development of chronic kidney disease and renal failure. As major energy source in maintaining renal physiological functions, tubular epithelial cells with decreased fatty acid oxidation play a key role in renal fibrosis development. Inorganic nitrate, found in high levels in certain vegetables, can increase the formation and signaling by bioactive nitrogen species, including NO, and dampen oxidative stress. In this study, we evaluated the therapeutic value of inorganic nitrate treatment on development of kidney fibrosis and investigated underlying mechanisms including regulation of lipid metabolism in tubular epithelial cells.METHODS: Inorganic nitrate was supplemented in a mouse model of complete unilateral ureteral obstruction (UUO)-induced fibrosis. Inorganic nitrite was applied in transforming growth factor β-induced pro-fibrotic cells in vitro. Metformin was administrated as a positive control. Fibrosis, oxidative stress and lipid metabolism were evaluated.
RESULTS: Nitrate treatment boosted the nitrate-nitrite-NO pathway, which ameliorated UUO-induced renal dysfunction and fibrosis in mice, represented by improved glomerular filtration and morphological structure and decreased renal collagen deposition, pro-fibrotic marker expression, and inflammation. In human proximal tubule epithelial cells (HK-2), inorganic nitrite treatment prevented transforming growth factor β-induced pro-fibrotic changes. Mechanistically, boosting the nitrate-nitrite-NO pathway promoted AMP-activated protein kinase (AMPK) phosphorylation, improved AKT-mediated peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC1α) activity and restored mitochondrial function. Accordingly, treatment with nitrate (in vivo) or nitrite (in vitro) decreased lipid accumulation, which was associated with dampened NADPH oxidase activity and mitochondria-derived oxidative stress.
CONCLUSIONS: Our findings indicate that inorganic nitrate and nitrite treatment attenuates the development of kidney fibrosis by targeting oxidative stress and lipid metabolism. Underlying mechanisms include modulation of AMPK and AKT-PGC1α pathways.

Keywords:  AMPK; Inorganic nitrate; Nitrite; Oxidative stress; PGC1α; Renal fibrosis

DOI:  https://doi.org/10.1016/j.redox.2022.102266
Nature. 2022 Feb 23.

Low-dose metformin targets the lysosomal AMPK pathway through PEN2.

Teng Ma, Xiao Tian, Baoding Zhang, Mengqi Li, Yu Wang, Chunyan Yang, Jianfeng Wu, Xiaoyan Wei, Qi Qu, Yaxin Yu, Shating Long, Jin-Wei Feng, Chun Li, Cixiong Zhang, Changchuan Xie, Yaying Wu, Zheni Xu, Junjie Chen, Yong Yu, Xi Huang, Ying He, Luming Yao, Lei Zhang, Mingxia Zhu, Wen Wang, Zhi-Chao Wang, Mingliang Zhang, Yuqian Bao, Weiping Jia, Shu-Yong Lin, Zhiyun Ye, Hai-Long Piao, Xianming Deng, Chen-Song Zhang, Sheng-Cai Lin.

Metformin, the most prescribed antidiabetic medicine, has shown other benefits such as anti-ageing and anticancer effects1-4. For clinical doses of metformin, AMP-activated protein kinase (AMPK) has a major role in its mechanism of action4,5; however, the direct molecular target of metformin remains unknown. Here we show that clinically relevant concentrations of metformin inhibit the lysosomal proton pump v-ATPase, which is a central node for AMPK activation following glucose starvation6. We synthesize a photoactive metformin probe and identify PEN2, a subunit of γ-secretase7, as a binding partner of metformin with a dissociation constant at micromolar levels. Metformin-bound PEN2 forms a complex with ATP6AP1, a subunit of the v-ATPase8, which leads to the inhibition of v-ATPase and the activation of AMPK without effects on cellular AMP levels. Knockout of PEN2 or re-introduction of a PEN2 mutant that does not bind ATP6AP1 blunts AMPK activation. In vivo, liver-specific knockout of Pen2 abolishes metformin-mediated reduction of hepatic fat content, whereas intestine-specific knockout of Pen2 impairs its glucose-lowering effects. Furthermore, knockdown of pen-2 in Caenorhabditis elegans abrogates metformin-induced extension of lifespan. Together, these findings reveal that metformin binds PEN2 and initiates a signalling route that intersects, through ATP6AP1, the lysosomal glucose-sensing pathway for AMPK activation. This ensures that metformin exerts its therapeutic benefits in patients without substantial adverse effects.

DOI: https://doi.org/10.1038/s41586-022-04431-8
Food Funct. 2022 Feb 25.

10-Gingerol, a natural AMPK agonist, suppresses neointimal hyperplasia and inhibits vascular smooth muscle cell proliferation.

Bo Deng, Xiao-Li Jiang, You-Cai Xu, Si Chen, Min Cai, Sui-Hui Deng, Wen-Jun Ding, Hong-Lin Xu, Shuang-Wei Zhang, Zhang-Bin Tan, Rui-Xue Chen, Bin Liu, Jing-Zhi Zhang.

Background: Abnormal proliferation of vascular smooth muscle cells (VSMCs) in the intimal region is a key event in the development of neointimal hyperplasia. 10-G, a bioactive compound found in ginger, exerted inhibitory effects on the proliferation of several cancer cells. However, the effect and mechanism of 10-G on neointimal hyperplasia are not clear. Purpose: To explore the suppressive effects of 10-G on the proliferation and migration of VSMCs, and investigate the underlying mechanisms. Methods: In vivo, a left common carotid artery ligation mouse model was used to observe the effects of neointimal formation through immunohistochemistry and hematoxylin-eosin staining. In vitro, the cell proliferation and migration of HASMCs and A7r5 cells were detected by MTS assay, EdU staining, wound healing assay, Transwell assay, and western blotting as well. Molecular docking, molecular dynamics simulations and surface plasmon resonance imaging were collectively used to evaluate the interaction of 10-G with AMP-activated protein kinase (AMPK). Compound C and si-AMPK were used to inhibit the expression of AMPK. Results: Treatment with 10-G significantly reduced neointimal hyperplasia in the left common carotid artery ligation mouse model. MST and EdU staining showed that 10-G inhibited the proliferation of VSMC cells A7r5 and HASMC. We also found that 10-G altered the expression of proliferation-related proteins, including CyclinD1, CyclinD2, CyclinD3, and CDK4. Molecular docking revealed that the binding energy between AMPK and 10-G is -7.4 kcal mol-1. Molecular simulations suggested that the binding between 10-G and AMPK is stable. Surface plasmon resonance imaging analysis also showed that 10-G has a strong binding affinity to AMPK (KD = 6.81 × 10-8 M). 10-G promoted AMPKα phosphorylation both in vivo and in vitro. Blocking AMPK by an siRNA or AMPK inhibitor pathway partly abolished the anti-proliferation effects of 10-G on VSMCs. Conclusion: These data showed that 10-G might inhibit neointimal hyperplasia and suppress VSMC proliferation by the activation of AMPK as a natural AMPK agonist.

DOI: https://doi.org/10.1039/d1fo03610f
Nutrients. 2022 Feb 11. pii: 764. [Epub ahead of print]14(4):

Pasteurized Akkermansia muciniphila Ameliorate the LPS-Induced Intestinal Barrier Dysfunction via Modulating AMPK and NF-κB through TLR2 in Caco-2 Cells.

Mengxuan Shi, Yunshuang Yue, Chen Ma, Li Dong, Fang Chen.

  Akkermansia muciniphila is well known for the amelioration of inflammatory responses and restoration of intestinal barrier function. The beneficial effect of A. muciniphila occurred through contacting Toll-like receptor 2 (TLR2) on intestinal epithelial cells by wall components. In this case, the downstream mechanism of pasteurized A. muciniphila stimulating TLR2 for ameliorated intestinal barrier function is worth investigating. In this study, we evaluated the effect of live and pasteurized A. muciniphila on protecting the barrier dysfunction of Caco-2 intestinal epithelial cells induced by lipopolysaccharide (LPS). We discovered that both live and pasteurized A. muciniphila could attenuate an inflammatory response and improve intestinal barrier integrity in Caco-2 monolayers. We demonstrated that A. muciniphila enhances AMP-activated protein kinase (AMPK) activation and inhibits Nuclear Factor-Kappa B (NF-κB) activation through the stimulation of TLR2. Overall, we provided a specific mechanism for the probiotic effect of A. muciniphila on the intestinal barrier function of Caco-2 cells.

Keywords:  AMPK; Akkermansia muciniphila; Caco-2; NF-κB; TLR2; intestinal barrier function

DOI:  https://doi.org/10.3390/nu14040764
Mol Cell Endocrinol. 2022 Feb 17. pii: S0303-7207(22)00043-0. [Epub ahead of print] 111596

Vitamin D protects against high glucose-induced pancreatic β-cell dysfunction via AMPK-NLRP3 inflammasome pathway.

Mian Wu, Li Lu, Kaifeng Guo, Junxi Lu, Haibing Chen.

  Vitamin D deficiency is considered a pandemic and has been postulated to increase the risk of type 2 diabetes mellitus (T2DM). Activation of nod-like receptor protein 3 (NLRP3) signaling induced by hyperglycemia stress has been recognized as a key priming stage for pancreatic β-cell inflammation in T2DM pathogenesis. AMP-activated protein kinase (AMPK) activation attenuates NLRP3 inflammasome upregulation in diabetes. This study investigated whether vitamin D3 could protect cells against high glucose-induced inflammation by modulating this critical step. A human cross-sectional study of 78 healthy, glucose-tolerant volunteers and 399 patients with type 2 diabetes was undertaken. The relationship between serum 25(OH)D3 levels and β-cell function was assessed using Pearson correlation analysis and multiple linear regression, and a high-glucose diet-induced rat model of impaired glucose tolerance was used to evaluate the effects of cholecalciferol. Intraperitoneal glucose tolerance tests and an ELISA were performed to detect the function of pancreatic islets. Glucose-stimulated insulin secretion, pyroptosis, reactive oxygen species (ROS) production, and NLRP3 pathway were evaluated together to determine the role of vitamin D in high glucose-induced pancreatic β-cell dysfunction in INS-1E cells. The clinical results showed a positive association between serum 25(OH)D3 levels and β-cell function in male patients with type 2 diabetes. In vivo, cholecalciferol significantly reduced blood glucose levels and improved insulin secretion in response to glucose loading in the high glucose diet-induced rat model. In vitro studies have demonstrated that 1,25(OH)2D3 promotes insulin release in both islets and INS-1E cells. Mechanistically, our results demonstrated that vitamin D3 can activate AMPK, inhibiting the mTOR pathway, thus inhibiting NLRP3 inflammasome activation and alleviating pyroptosis in β-cell dysfunction. This study showed that vitamin D protects against high-glucose-induced β-cell dysfunction by enhancing the AMPK pathway, thereby suppressing NLRP3 inflammasome activation.

Keywords:  AMPK; NLRP3; Pancreatic β-cell function; Vitamin D

DOI:  https://doi.org/10.1016/j.mce.2022.111596
Emerg Microbes Infect. 2022 Feb 22. 1-34

Metformin attenuated sepsis-related liver injury by modulating gut microbiota.

Huoyan Liang, Heng Song, Xiaojuan Zhang, Gaofei Song, Yuze Wang, Xianfei Ding, Xiaoguang Duan, Lifeng Li, Tongwen Sun, Quancheng Kan.

  Increased evidence shows that gut microbiota acts as the primary regulator of the liver; however, their role in sepsis-related liver injury (SLI) in the elderly is unclear. This study assessed whether metformin could attenuate SLI by modulating gut microbiota in septic aged rats. Cecal ligation and puncture (CLP) was used to induce SLI in aged rats. Fecal microbiota transplantation (FMT) was used to validate the roles of gut microbiota in these pathologies. The composition of gut microbiota was analyzed by 16S rRNA sequencing. Moreover, the liver and colon tissues were analyzed by histopathology, immunofluorescence, immunohistochemistry,and reverse transcription polymerase chain reaction (RT-PCR). Metformin improved liver damage, colon barrier dysfunction in aged SLI rats. Moreover, metformin improved sepsis-induced liver inflammation and damage under the presence of gut microbiota. Importantly, FMT assay showed that rats gavaged with feces from metformin-treated SLI rats displayed fewer-severe liver damage and colon barrier dysfunction than did rats gavaged with feces from SLI rats.The gut microbiota composition among the sham-operated, CLP-operated and metformin-treated SLI rats were different. In particular, the proportion of Klebsiella and Escherichia_Shigella was higher in SLI rats than sham-operated and metformin-treated SLI rats;while metformin could increased the proportion of Bifidobacterium, Muribaculaceae, Parabacteroides_distasonis and Alloprevitella in aged SLI rats. Additionally, Klebsiella and Escherichia_Shigella correlated positively with the inflammatory factors in the liver. Our findings suggest that metformin may improve the liver injury by regulating the gut microbiota and alleviating colon barrier dysfunction in septic aged rats, which may be an effective therapy for SLI.

Keywords:  aged rats; gut microbiota; metformin; sepsis-related liver injury

DOI:  https://doi.org/10.1080/22221751.2022.2045876
Mol Med Rep. 2022 Apr;pii: 139. [Epub ahead of print]25(4):

lncRNA TUG1 protects intestinal epithelial cells from damage induced by high glucose and high fat via AMPK/SIRT1.

Weiwei Wei, Xingquan Wang, Yaqing Wei, Shilin Liu, Shengyu Gao, Hao Tian, Dewang Su.

  he incidence of obesity and type 2 diabetes mellitus (T2DM) is increasing year by year and shows a trend towards younger age groups worldwide. It has become a disease that endangers the health of individuals all over the world. Among numerous weight loss surgeries, sleeve gastrectomy (SG) has become one of the most common surgical strategies for the treatment of T2DM. However, SG‑mediated alterations to the molecular mechanism of metabolism require further investigation. Thus, reverse transcription‑quantitative PCR was used to detect the expression levels of long non‑coding (lnc)RNA taurine‑upregulated gene 1 (TUG1), Sirtuin 1 (SIRT1), AMP‑activated protein kinase (AMPK) and uncoupling protein 2 (UCP2) in the serum of T2DM patients, as well as in HIEC‑6 and SW480 cells following treatment with high glucose and high fat (HGHF). Protein expression was detected by western blotting. Cell Counting Kit‑8 assays were performed to analyze cell viability, and flow cytometry and a TUNEL assay was performed to evaluate cell apoptosis. The secretion of ILs in the culture medium was detected by conducting ELISAs. The results showed that lncRNA TUG1 and UCP2 expression was upregulated, SIRT1 and AMPK expression levels were decreased by SG. Under HGHF conditions, HIEC‑6 and SW480 cell viability was inhibited, apoptosis was promoted, TUG1 expression was downregulated, and SIRT1 and AMPK expression levels were upregulated. The secretory levels of IL‑1β, IL‑6 and IL‑8 were increased, whereas the secretion of IL‑10 was decreased under HGHF conditions. lncRNA TUG1 overexpression significantly reversed the effects of HGHF on cell viability, apoptosis and SIRT1, AMPK, UCP2 and Bcl‑2 expression levels. Together, the findings of the present study demonstrated that lncRNA TUG1 alleviated the damage induced by HGHF in intestinal epithelial cells by downregulating SIRT1 and AMPK expression, and upregulating UCP2 expression. Thus, the lncRNA TUG1/AMPK/SIRT1/UCP2 axis may serve an important role in the treatment of T2DM.

Keywords:  long non‑coding RNA taurine‑upregulated gene 1; obesity; sleeve gastrectomy; type 2 diabetes

DOI:  https://doi.org/10.3892/mmr.2022.12655
Int Immunopharmacol. 2022 Feb 22. pii: S1567-5769(22)00084-4. [Epub ahead of print]106 108600

SIRT3-AMPK signaling pathway as a protective target in endothelial dysfunction of early sepsis.

Huilin Yu, Qian Liu, Guodong Chen, Longxiang Huang, Minghao Luo, Dingyi Lv, Suxin Luo.

  Extensive vascular endothelial dysfunction usually occurs in sepsis, resulting in high mortality. The purpose of this study was therefore to investigate the role of AMP-dependent protein kinase (AMPK) in the aortic endothelial dysfunction of early sepsis in mice, and the relationship between AMPK and Sirtuin3 (SIRT3). Cecal ligation and puncture (CLP) surgery was performed to establish a mouse sepsis model, and human umbilical vein endothelial cells (HUVECs) were treated with lipopolysaccharide (LPS) to mimic a sepsis model in vitro. We suppressed and increased the activities of AMPK with Dorsomorphin (CC) and Acadesine (AICAR), respectively. 3-TYP (SIRT3 inhibitor) and Honokiol (SIRT3 agonist) were used to alter SIRT3 activity. Then, the inflammatory and endothelial function parameters of the vascular tissue and survival rate were determined. In vivo, the expression of Ser1177 phosphorylation of endothelial nitric oxide synthase (p-eNOS), endothelium-dependent relaxation function, and survival decreased (P < 0.05), while NF-κB and NLRP3 pathways were activated in CLP-induced early sepsis (P < 0.05). Moreover, activation of AMPK significantly reversed the reduction of p-eNOS expression (P < 0.05), prevented endothelial dysfunction (P < 0.05), deactivated NF-κB and NLRP3 pathways (P < 0.05), and improved survival (P < 0.05) in septic mice. However, AMPK inhibition led to opposite effects (P < 0.05). In addition, changing the activity of AMPK had little effect on SIRT3 expression (P > 0.05), while the expression of p-AMPK varied with the inhibition or activation of SIRT3 (P < 0.05), which was further demonstrated using in vitro experiments. Together, the results showed that the SIRT3-AMPK signaling pathway played an important role in inhibiting vascular inflammation and endothelial dysfunction during early sepsis.

Keywords:  AMPK; Endothelial dysfunction; SIRT3; Sepsis; eNOS

DOI:  https://doi.org/10.1016/j.intimp.2022.108600
Pharmaceutics. 2022 Jan 29. pii: 329. [Epub ahead of print]14(2):

The Natural Chemotherapeutic Capsaicin Activates AMPK through LKB1 Kinase and TRPV1 Receptors in Prostate Cancer Cells.

Belén G Sánchez, Alicia Bort, José M Mora-Rodríguez, Inés Díaz-Laviada.

  The natural bioactive compound capsaicin has been reported to have anticancer activity, although the underlying mechanism of action has not been completely clarified. Herein, we investigated the mechanism whereby capsaicin exerts antitumor effects on prostate cancer cells. We found that capsaicin activated AMP-activated kinase (AMPK) and promoted cell death in the LKB1-expressing prostate cancer cell lines LNCaP and PC3, but not in the liver kinase B1 (LKB1)-null cell line DU-145. Capsaicin treatment stimulated LKB1 phosphorylation and activated AMPK in LKB1-expressing cells. In addition, LKB1 silencing in LNCaP and PC3 cells abrogated capsaicin-induced AMPK activation, while the overexpression of LKB1 by lentiviral infection in DU-145 cells induced capsaicin-triggered AMPK phosphorylation. Moreover, the calcium/calmodulin-dependent kinase kinase 2 (CaMKK2) inhibitor STO-609 did not modify the activation of AMPK induced by capsaicin, suggesting a CaMKK2-independent mechanism. Capsaicin-induced LKB1 phosphorylation was dependent on the transient receptor potential cation channel subfamily V member 1 (TRPV1), since TRPV1 knocked down by shRNA abolished LKB1 and AMPK phosphorylation in LKB1-expressing cells. Altogether, our results showed that capsaicin affected AMPK activity in an LKB1- and TRPV1-dependent fashion, linking TRPV1 with cell fate. These data also suggest that capsaicin may be a rational chemotherapeutic option for prostate tumors.

Keywords:  AMPK; DU-145; LKB1; LNCaP; PC3; TRPV1; capsaicin; prostate cancer

DOI:  https://doi.org/10.3390/pharmaceutics14020329
Foods. 2022 Feb 16. pii: 575. [Epub ahead of print]11(4):

Heat-Killed Enterococcus faecalis EF-2001 Attenuate Lipid Accumulation in Diet-Induced Obese (DIO) Mice by Activating AMPK Signaling in Liver.

Meiqi Fan, Young-Jin Choi, Nishala Erandi Wedamulla, Yujiao Tang, Kwon Il Han, Ji-Young Hwang, Eun-Kyung Kim.

  To explore the inhibitory mechanism of heat-killed Enterococcus faecalis, EF-2001 on hepatic lipid deposition, a diet-induced obese (DIO) animal model was established by high-fat diet (HFD). The DIO C57BL/6 mice were divided into four groups: the normal group without HFD (ND, n = 8), obesity group (HFD, n = 8), experimental group (HFD + EF-2001, 200 mg/kg, n = 8), and positive control group (HFD + Orlistat, 60 mg/kg, n = 8). After 4 weeks, liver and adipose tissue were fixed in 10% paraformaldehyde, followed by embedding in paraffin for tissue sectioning. The differences in body mass, body fat ratio, fatty cell area, and lipid profiling of the liver (TC, LDL, and HDL) were also determined. Moreover, Western blot was performed to analyze the expression of lipid accumulation-related proteins, including AMPK, PPARγ, SREBP-1, ACC, and FAS. Compared with the HFD group, the HFD + EF-2001 group exhibited decreased fat mass, liver index, adipocyte area, TC, and LDL, and an increased level of HDL. The results of liver hematoxylin and eosin (H&E), and oil red O staining showed that the mice in each intervention group were improved on hepatic lipid accumulation, and the mice in the HFD + EF-2001 group were the most similar to those in the normal group when compared with the HFD group. From the Western blot results, we proved that EF-2001 activated the AMPK signaling pathway. EF-2001 significantly upregulated the expressions of p-AMPK and p-ACC and downregulated PPARγ, SREBP-1, and FAS in murine liver. Taken together, these results suggest that EF-2001 decrease lipid accumulation in the DIO model mice through the AMPK pathway and ameliorate liver damage by HFD.

Keywords:  EF-2001; Enterococcus faecalis; lipid accumulation; liver damage; obese

DOI:  https://doi.org/10.3390/foods11040575
Diabetes. 2022 03 01. 71(3): e3-e4

Comment on De Wendt et al. Contraction-Mediated Glucose Transport in Skeletal Muscle Is Regulated by a Framework of AMPK, TBC1D1/4, and Rac1. Diabetes 2021;70:2796-2809.

Rasmus Kjøbsted, Jørgen F P Wojtaszewski.

DOI: https://doi.org/10.2337/db21-1036
Autophagy. 2022 Feb 23. 1-17

Treatments targeting autophagy ameliorate the age-related macular degeneration phenotype in mice lacking APOE (apolipoprotein E).

Kirstan A Vessey, Andrew I Jobling, Mai X Tran, Anna Y Wang, Ursula Greferath, Erica L Fletcher.

  Age-related macular degeneration (AMD) is a leading cause of vision loss with recent evidence indicating an important role for macroautophagy/autophagy in disease progression. In this study we investigate the efficacy of targeting autophagy for slowing dysfunction in a mouse model with features of early AMD. Mice lacking APOE (apolipoprotein E; B6.129P2-Apoetm1UncJ/Arc) and C57BL/6 J- (wild-type, WT) mice were treated with metformin or trehalose in the drinking water from 5 months of age and the ocular phenotype investigated at 13 months. Control mice received normal drinking water. APOE-control mice had reduced retinal function and thickening of Bruch's membrane consistent with an early AMD phenotype. Immunohistochemical labeling showed reductions in MAP1LC3B/LC3 (microtubule-associated protein 1 light chain 3 beta) and LAMP1 (lysosomal-associated membrane protein 1) labeling in the photoreceptors and retinal pigment epithelium (RPE). This correlated with increased LC3-II:LC3-I ratio and alterations in protein expression in multiple autophagy pathways measured by reverse phase protein array, suggesting autophagy was slowed. Treatment of APOE-mice with metformin or trehalose ameliorated the loss of retinal function and reduced Bruch's membrane thickening, enhancing LC3 and LAMP1 labeling in the ocular tissues and restoring LC3-II:LC3-I ratio to WT levels. Protein analysis indicated that both treatments boost ATM-AMPK driven autophagy. Additionally, trehalose increased p-MAPK14/p38 to enhance autophagy. Our study shows that treatments targeting pathways to enhance autophagy have the potential for treating early AMD and provide support for the use of metformin, which has been found to reduce the risk of AMD development in human patients.Abbreviations:AMD: age-related macular degeneration; AMPK: 5' adenosine monophosphate-activated protein kinase APOE: apolipoprotein E; ATM: ataxia telangiectasia mutated; BCL2L1/Bcl-xL: BCL2-like 1; DAPI: 4'-6-diamidino-2-phenylindole; ERG: electroretinogram; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GCL: ganglion cell layer; INL: inner nuclear layer; IPL: inner plexiform layer; IS/OS: inner and outer photoreceptor segments; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3B/LC3: microtubule-associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; OCT: optical coherence tomography; ONL: outer nuclear layer; OPs: oscillatory potentials; p-EIF4EBP1: phosphorylated eukaryotic translation initiation factor 4E binding protein 1; p-MAPK14/p38: phosphorylated mitogen-activated protein kinase 14; RPE: retinal pigment epithelium; RPS6KB/p70 S6 kinase: ribosomal protein S6 kinase; SQSTM1/p62: sequestosome 1; TP53/TRP53/p53: tumor related protein 53; TSC2: TSC complex subunit 2; WT: wild type.

Keywords:  B6.129P2-Apoetm1UncJ; bruch’s membrane; metformin; retina; retinal pigment epithelium; trehalose

DOI:  https://doi.org/10.1080/15548627.2022.2034131
Diabetes. 2022 03 01. 71(3): e5

Response to Comment on Wendt et al. Contraction-Mediated Glucose Transport in Skeletal Muscle Is Regulated by a Framework of AMPK, TBC1D1/4, and Rac1. Diabetes 2021;70:2796-2809.

Alexandra Chadt, Hadi Al-Hasani.

DOI: https://doi.org/10.2337/dbi21-0038
Mol Nutr Food Res. 2022 Feb 25. e2100669

Garcinia Cambogia Improves High-Fat Diet-Induced Glucose Imbalance By Enhancing Calcium/CaMKII/AMPK/GLUT4-Mediated Glucose Uptake In Skeletal Muscle.

Joo-Hui Han, Min-Tae Kim, Chang-Seon Myung.

  SCOPE: Garcinia cambogia (G. cambogia) is known to have antiobesity effects. In this study, the therapeutic effects of G. cambogia on glucose homeostasis in obesity-induced diabetes were explored and the underlying mechanisms were investigated.METHODS AND RESULTS: C2C12 myotubes were treated with G. cambogia; glucose uptake, intracellular Ca2+ levels, and related alterations in signaling pathways were examined. High-fat diet (HFD)-fed mice were administered G. cambogia for 8 weeks; oral glucose tolerance was evaluated, and the regulation of identified targets of signaling pathways in quadriceps skeletal muscle were examined in vivo. G. cambogia increased glucose uptake in C2C12 myotubes and induced the upregulation of AMPK, ACC and p38 MAPK phosphorylation. Notably, G. cambogia markedly elevated both intracellular Ca2+ levels, activating CaMKII, a Ca2+ -sensing protein, and TBC1D4-mediated GLUT4 translocation, to facilitate glucose uptake. Furthermore, high-glucose-induced inhibition of glucose uptake and signal transduction was reverted by G. cambogia. In an HFD-induced diabetes mouse model, G. cambogia administration resulted in significant blood glucose-lowering effects, which were attributed to the regulation of targets that had been identified in vitro, in quadricep skeletal muscle.
CONCLUSION: These findings provide new insights into the mechanism by which G. cambogia regulates glucose homeostasis in obesity-induced diabetes. This article is protected by copyright. All rights reserved.

Keywords:  AMPK; CaMKII; Diabetes; Garcinia cambogia; calcium; glucose uptake

DOI:  https://doi.org/10.1002/mnfr.202100669
Methods Mol Biol. 2022 ;2455 223-232

Metabolic Phenotyping in Mice with NASH Using Indirect Calorimetry.

Bin Ni, Shanshan Chen, Jared S Farrar, Francesco S Celi.

  Obesity caused by caloric overload has assumed epidemic proportions. Obesity is frequently associated with metabolic dysfunctions, such as type 2 diabetes, non-alcoholic steatohepatitis (NASH), cardiovascular diseases, and cancer. Metabolic phenotyping is a set of techniques for studying metabolic dysfunction and behavior information including energy expenditure, body weight gain, glucose homeostasis, and lipid profile. Among different metabolic phenotyping methods, indirect calorimetry is an indispensable tool for quantifying the energy balance/imbalance in various mouse models, which enables researchers to probe the development of disease and to evaluate the therapeutic benefit from different interventions. In this chapter, we will describe the procedures of metabolic phenotyping using indirect calorimetry in db/db mouse, a metabolic disorder mouse model which develops NASH.

Keywords:  Diabetes; Energy expenditure; Indirect calorimetry; NASH; Obesity; Oxygen consumption; Respiratory exchange ratio; db/db

DOI:  https://doi.org/10.1007/978-1-0716-2128-8_17
Int J Mol Sci. 2022 Feb 21. pii: 2398. [Epub ahead of print]23(4):

Statins Aggravate the Risk of Insulin Resistance in Human Muscle.

Stefanie A Grunwald, Stefanie Haafke, Ulrike Grieben, Ursula Kassner, Elisabeth Steinhagen-Thiessen, Simone Spuler.

  Beside their beneficial effects on cardiovascular events, statins are thought to contribute to insulin resistance and type-2 diabetes. It is not known whether these effects are long-term events from statin-treatment or already triggered with the first statin-intake. Skeletal muscle is considered the main site for insulin-stimulated glucose uptake and therefore, a primary target for insulin resistance in the human body. We analyzed localization and expression of proteins related to GLUT4 mediated glucose uptake via AMPKα or AKT in human skeletal muscle tissue from patients with statin-intake >6 months and in primary human myotubes after 96 h statin treatment. The ratio for AMPKα activity significantly increased in human skeletal muscle cells treated with statins for long- and short-term. Furthermore, the insulin-stimulated counterpart, AKT, significantly decreased in activity and protein level, while GSK3ß and mTOR protein expression reduced in statin-treated primary human myotubes, only. However, GLUT4 was normally distributed whereas CAV3 was internalized from plasma membrane around the nucleus in statin-treated primary human myotubes. Statin-treatment activates AMPKα-dependent glucose uptake and remains active after long-term statin treatment. Permanent blocking of its insulin-dependent counterpart AKT activation may lead to metabolic inflexibility and insulin resistance in the long run and may be a direct consequence of statin-treatment.

Keywords:  AKT; AMPK; human skeletal muscle; insulin resistance; primary human muscle cells; statins

DOI:  https://doi.org/10.3390/ijms23042398
Int J Mol Sci. 2022 Feb 16. pii: 2200. [Epub ahead of print]23(4):

Metformin's Mechanism of Action Is Stimulation of the Biosynthesis of the Natural Cyclic AMP Antagonist Prostaglandylinositol Cyclic Phosphate (Cyclic PIP).

Heinrich K Wasner.

  Metformin is the leading drug for treating type 2 diabetics, but the mechanism of action of metformin, despite some suggested mechanisms such as the activation of the AMP-kinase, is largely unknown. Among its many positive effects are the reduction of blood glucose levels, the inhibition of cyclic AMP synthesis, gluconeogenesis and an increase in sensitivity to insulin. Recent studies have described the natural antagonist of cyclic AMP, prostaglandylinositol cyclic phosphate. Synthesis of cyclic PIP is stimulated in all organs by hormones such as insulin and also by drugs such as metformin. Its primary action is to trigger the dephosphorylation of proteins/enzymes, phosphorylated on serine/threonine residues. Cyclic PIP triggers many of the regulations requested by insulin. The parallels between the beneficial effects of metformin and the regulations triggered by cyclic PIP suggest that the mechanism of action of this key drug may well be explained by its stimulation of the synthesis of cyclic PIP.

Keywords:  biguanides; cyclic AMP antagonist; cyclic PIP; mechanism of insulin action; metformin; prostaglandylinositol cyclic phosphate; protein serine/threonine phosphatase; protein tyrosine kinase; protein tyrosine phosphatase

DOI:  https://doi.org/10.3390/ijms23042200
Biochim Biophys Acta Mol Cell Res. 2022 Feb 19. pii: S0167-4889(22)00033-7. [Epub ahead of print] 119242

MARK2/4 promotes Warburg effect and cell growth in non-small cell lung carcinoma through the AMPKα1/mTOR/HIF-1α signaling pathway.

Sathan Raj Natarajan, Lavanya Ponnusamy, Ravi Manoharan.

  MARKs kinase belongs to an AMPK-related family kinase plays a critical role in tumor progression, but its exact role and contribution of four different isoforms remain largely ambiguous. In this study, we used a clinical dataset compiled by The Cancer Genome Atlas (TCGA) and GEO revealed that MARK2 and MARK4 expressions were significantly upregulated in non-small cell lung cancer (NSCLC) compared with normal tissues. Furthermore, expressions of MARK2/4 were highly appeared in advanced stages and associated with the low survival rate of NSCLC patients. Functional assays demonstrated that MARK2/4 deletion or MARKs inhibition significantly suppressed aerobic glycolysis and cell growth in NSCLC cells. Mechanistically, MARK2/4 stimulates the mTOR/HIF-1α pathway and subsequently alleviates AMPK activity via physically associate with Raptor and AMPKα1, thereby facilitating aerobic glycolysis and cell growth in NSCLC cells. However, these effects were markedly reversed by MARKs inhibitor 39,621, or MARK2/4 deletion, mTOR inhibitor rapamycin, or AMPK activator AICAR. Together, the data demonstrated that MARK2/4 exerts its oncogenic effects by facilitating metabolic reprogramming in NSCLC cells. Therefore, MARK2/4 might be a potential therapeutic target for lung cancer.

Keywords:  AMPK; Aerobic glycolysis; Cell growth; MARK2; MARK4; mTOR

DOI:  https://doi.org/10.1016/j.bbamcr.2022.119242
Cell Death Dis. 2022 Feb 25. 13(2): 188

Salt-inducible kinases inhibitor HG-9-91-01 targets RIPK3 kinase activity to alleviate necroptosis-mediated inflammatory injury.

Dongxuan Huang, Pengfei Chen, Guoqing Huang, Huimin Sun, Xiaohua Luo, Chaowen He, Fei Chen, Yong Wang, Changchun Zeng, Lianhui Su, Xiaobin Zeng, Jiachun Lu, Shiyue Li, Dongsheng Huang, Hanchao Gao, Mengtao Cao.

Receptor-interacting protein kinase 3 (RIPK3) functions as a central regulator of necroptosis, mediating signaling transduction to activate pseudokinase mixed lineage kinase domain-like protein (MLKL) phosphorylation. Increasing evidences show that RIPK3 contributes to the pathologies of inflammatory diseases including multiple sclerosis, infection and colitis. Here, we identified a novel small molecular compound Salt-inducible Kinases (SIKs) inhibitor HG-9-91-01 inhibiting necroptosis by targeting RIPK3 kinase activity. We found that SIKs inhibitor HG-9-91-01 could block TNF- or Toll-like receptors (TLRs)-mediated necroptosis independent of SIKs. We revealed that HG-9-91-01 dramatically decreased cellular activation of RIPK3 and MLKL. Meanwhile, HG-9-91-01 inhibited the association of RIPK3 with MLKL and oligomerization of downstream MLKL. Interestingly, we found that HG-9-91-01 also trigger RIPK3-RIPK1-caspase 1-caspase 8-dependent apoptosis, which activated cleavage of GSDME leading to its dependent pyroptosis. Mechanistic studies revealed that SIKs inhibitor HG-9-91-01 directly inhibited RIPK3 kinase activity to block necroptosis and interacted with RIPK3 and recruited RIPK1 to activate caspases leading to cleave GSDME. Importantly, mice pretreated with HG-9-91-01 showed resistance to TNF-induced systemic inflammatory response syndrome. Consistently, HG-9-91-01 treatment protected mice against Staphylococcus aureus-mediated lung damage through targeting RIPK3 kinase activity. Overall, our results revealed that SIKs inhibitor HG-9-91-01 is a novel inhibitor of RIPK3 kinase and a potential therapeutic target for the treatment of necroptosis-mediated inflammatory diseases.

DOI: https://doi.org/10.1038/s41419-022-04633-y
iScience. 2022 Feb 18. 25(2): 103827

Nrf2 activation reprograms macrophage intermediary metabolism and suppresses the type I interferon response.

Dylan G Ryan, Elena V Knatko, Alva M Casey, Jens L Hukelmann, Sharadha Dayalan Naidu, Alejandro J Brenes, Thanapon Ekkunagul, Christa Baker, Maureen Higgins, Laura Tronci, Efterpi Nikitopolou, Tadashi Honda, Richard C Hartley, Luke A J O'Neill, Christian Frezza, Angus I Lamond, Andrey Y Abramov, J Simon C Arthur, Doreen A Cantrell, Michael P Murphy, Albena T Dinkova-Kostova.

  To overcome oxidative, inflammatory, and metabolic stress, cells have evolved cytoprotective protein networks controlled by nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) and its negative regulator, Kelch-like ECH associated protein 1 (Keap1). Here, using high-resolution mass spectrometry we characterize the proteomes of macrophages with altered Nrf2 status revealing significant differences among the genotypes in metabolism and redox homeostasis, which were validated with respirometry and metabolomics. Nrf2 affected the proteome following lipopolysaccharide (LPS) stimulation, with alterations in redox, carbohydrate and lipid metabolism, and innate immunity. Notably, Nrf2 activation promoted mitochondrial fusion. The Keap1 inhibitor, 4-octyl itaconate remodeled the inflammatory macrophage proteome, increasing redox and suppressing type I interferon (IFN) response. Similarly, pharmacologic or genetic Nrf2 activation inhibited the transcription of IFN-β and its downstream effector IFIT2 during LPS stimulation. These data suggest that Nrf2 activation facilitates metabolic reprogramming and mitochondrial adaptation, and finetunes the innate immune response in macrophages.

Keywords:  Biochemistry; Immunology; Proteomics

DOI:  https://doi.org/10.1016/j.isci.2022.103827
Diabetes Res Clin Pract. 2022 Feb 16. pii: S0168-8227(22)00600-3. [Epub ahead of print] 109788

Thioredoxin upregulation delays diabetes-induced photoreceptor cell degeneration via AMPK-mediated autophagy and exosome secretion.

Xiang Ren, Jinjuan Lv, Nina Wang, Jiasu Liu, Chuanzhou Gao, Xiaoli Wu, Yang Yu, Qiufeng Teng, Wenkang Dong, Hui Kong, Li Kong.

  AIMS: Autophagy and exosome secretion in photoreceptor and RPE cells play an important role during diabetic retinopathy (DR). Thioredoxin (Trx) upregulation delays diabetes-induced photoreceptor cell degeneration, which the effect of autophagy and exosome secretion on it is unclear. Therefore, we investigated the effect of them on Trx upregulation to delay diabetes-induced photoreceptor cell degeneration and to identify the potential therapy for DR in the future.METHODS: Trx-transgenic mice and 661w cell were as models. Retinal function and morphology were evaluated by electroretinography and H&E staining. TUNEL was used to evaluate apoptosis. The protein expression was detected by Western blotting. TEM and mRFP-GFP-LC3 method were used to analyze autophagy.
RESULTS: In vitro and in vivo, Trx upregulation can delay diabetes-induced photoreceptor cell degeneration. Moreover, the expression of LC3 and p62 was decreasing and the expression of Alix and CD63 was increasing after Trx overexpression. However, it was inhibited after AMPK inhibitor treatment. Additionally, secreted exosomes from photoreceptor were phagocytosed by RPE cells to regulate its physiological function.
CONCLUSIONS: Trx upregulation can delay diabetes-induced photoreceptor cell degeneration via AMPK-mediated autophagy and exosome secretion. Secreted exosomes from photoreceptor cells could be phagocytosed and degraded by RPE cells in DR.

Keywords:  Autophagy; Diabetic retinopathy; Exosomes; Photoreceptor; Thioredoxin

DOI:  https://doi.org/10.1016/j.diabres.2022.109788