Adenosine Monophosphate-activated Protein Kinase Activation Research Articles

Involvement of oxidative stress-AMPK-Cx43-NLRP3 pathway in extracellular matrix remodeling of gastric smooth muscle cells in rats with diabetic gastroparesis.

This study aimed to investigate the changes in oxidative stress, adenosine monophosphate-activated protein kinase (AMPK), connexin43 (Cx43), nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) expression, and extracellular matrix (ECM) in the gastric smooth muscle tissues of rats with diabetic gastroparesis (DGP) and high glucose-cultured gastric smooth muscle cells, determine the existence of oxidative stress-AMPK-Cx43-NLRP3 pathway under high glucose condition, and the involvement of this pathway in ECM remodeling in DGP rats. The results showed that with increasing duration of diabetes, oxidation stress levels gradually increased, the AMPK activity decreased first and then increased, NLRP3, CX43 expression, and membrane/cytoplasmratio of Cx43 expression were increased in the gastric smooth muscle tissues of diabetic rats. Changes in ECM of gastric smooth muscle cells wereobserved in DGP rats. The DGP group showed higher collagen type I content, increased expression of Caspase-1, transforming growth factor-beta 3 (TGF-β3), and matrix metalloproteinase-2 (MMP-2), decreased tissue inhibitor of metalloproteinase-1 (TIMP-1) expression, and higher interleukin-1betacontent when compared with the control group. For gastric smooth muscle cells cultured under higher glucose, the MMP-2 and TGF-β3 expression wasdecreased, TGF-β1 and TIMP-1 expression was increased, the interleukin-1betacontent was decreased in cells after inhibition of NLRP3 expression; the NLRP3 and Caspase-1expression was decreased, and adenosine triphosphate content was lower after inhibition of Cx43; the expression of NLRP3, Caspase-1, P2X7, and the membrane/cytoplasm ratio of CX43 expression was decreased in cells after inhibition of AMPK and oxidative stress, the phospho-AMPK expression was also decreased after suppressing oxidative stress. Our findings suggest that high glucose induced the activation of the AMPK-Cx43-NLRP3 pathway through oxidative stress, and this pathway was involved in the ECM remodeling of gastric smooth muscles in DGP rats by regulating the biological functions of TGF-β3, TGF-β1, MMP-2, and TIMP-1.

Inhibition of lactate dehydrogenase A by diclofenac sodium induces apoptosis in HeLa cells through activation of AMPK.

Cancer cells exhibit a unique metabolic preference for the glycolytic pathway over oxidative phosphorylation for maintaining the tumor microenvironment. Lactate dehydrogenase A (LDHA) is a key enzyme that facilitates glycolysis by converting pyruvate to lactate and has been shown to be upregulated in multiple cancers due to the hypoxic tumor microenvironment. Diclofenac (DCF), a nonsteroidal anti-inflammatory drug, has been shown to exhibit anticancer effects by interfering with the glucose metabolism pathway. However, the specific targets of this drug remain unknown. Using in silico, biochemical, and biophysical studies, we show that DCF binds to LDHA adjacent to the substrate binding site and inhibits its activity in a dose-dependent and allosteric manner in HeLa cells. Thus, DCF inhibits the hypoxic microenvironment and induces apoptosis-mediated cell death. DCF failed to induce cytotoxicity in HeLa cells when LDHA was knocked down, confirming that DCF exerts its antimitotic effects via LDHA inhibition. DCF-induced LDHA inhibition alters pyruvate, lactate, NAD+, and ATP production in cells, and this could be a possible mechanism through which DCF inhibits glucose uptake in cancer cells. DCF-induced ATP deprivation leads to mitochondria-mediated oxidative stress, which results in DNA damage, lipid peroxidation, and apoptosis-mediated cell death. Reduction in intracellular ATP levels additionally activates the sensor kinase, adenosine monophosphate-activated protein kinase (AMPK), which further downregulates phosphorylated ribosomal S6 kinase (p-S6K), leading to apoptosis-mediated cell death. We find that in LDHA knocked down cells, intracellular ATP levels were depleted, resulting in the inhibition of p-S6K, suggesting the involvement of DCF-induced LDHA inhibition in the activation of the AMPK/S6K signaling pathway.

Glucagon-like peptide-1 analogs activate AMP kinase leading to reversal of the Warburg metabolic switch in breast cancer cells.

Breast cancer (BC) is associated with type 2 diabetes mellitus (T2DM) and obesity. Glucagon-like peptide (GLP)-1 regulates post-prandial insulin secretion, satiety, and gastric emptying. Several GLP-1 analogs have been FDA-approved for the treatment of T2DM and obesity. Moreover, GLP-1 regulates various metabolic activities across different tissues by activating metabolic signaling pathways like adenosine monophosphate (AMP) activated protein kinase (AMPK), and AKT. Rewiring metabolic pathways is a recognized hallmark of cancer, regulated by several cancer-related pathways, including AKT and AMPK. As GLP-1 regulates AKT and AMPK, we hypothesized that it alters BC cells' metabolism, thus inhibiting proliferation. The effect of the GLP-1 analogs exendin-4 (Ex4) and liraglutide on viability, AMPK signaling and metabolism of BC cell lines were assessed. Viability of BC cells was evaluated using colony formation and MTT/XTT assays. Activation of AMPK and related signaling effects were evaluated using western blot. Metabolism effects were measured for glucose, lactate and ATP. Exendin-4 and liraglutide activated AMPK in a cAMP-dependent manner. Blocking Ex4-induced activation of AMPK by inhibition of AMPK restored cell viability. Interestingly, Ex4 and liraglutide reduced the levels of glycolytic metabolites and decreased ATP production, suggesting that GLP-1 analogs impair glycolysis. Notably, inhibiting AMPK reversed the decline in ATP levels, highlighting the role of AMPK in this process. These results establish a novel signaling pathway for GLP-1 in BC cells through cAMP and AMPK modulation affecting proliferation and metabolism. This study suggests that GLP-1 analogs should be considered for diabetic patients with BC.

AMPK regulates immature boar Sertoli cell proliferation through affecting CDK4/Cyclin D3 pathway and mitochondrial function

Sertoli cell (SC) proliferation plays an important role in sperm production and quality; however, the regulatory mechanism of SC proliferation is not well understood. This study investigated the role of adenosine monophosphate-activated protein kinase (AMPK) in the regulation of immature boar SC activity. Cell counting kit-8, Seahorse XFe96, mitochondrial respiratory enzyme-related assay kits, and transmission electron microscopy were used to detect SC proliferative viability, oxygen consumption rate (OCR), mitochondrial respiratory enzyme activity, and the ultrastructure of primary cultured SCs in vitro from the testes of 21-day-old boars. A dual luciferase reporter assay was performed to determine the miRNA-mRNA target interaction. Western blotting was used to analyze cell proliferation-related protein expression of p38, p21, proliferating cell nuclear antigen (PCNA), Cyclin-dependent kinase 4 (CDK4), Cyclin D3, and phosphorylated retinoblastoma protein (Rb). Each experiment had a completely randomized design, with three replicates in each experiment. The results showed that the AMPK inhibitor (Compound C, 20 μM-24 h) increased cell proliferation viability, ATP production, and maximal respiration of SCs by 0.64-, 0.12-, and 0.08-fold (p < 0.05), respectively; increased the SC protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.13-, 0.09-, 0.88-, and 0.12-fold (p < 0.05), respectively; and decreased the SC protein expression of p38 and p21 by 0.36- and 0.27-fold (p < 0.05), respectively. The AMPK agonist AICAR (2 mM-6 h) significantly inhibited SC ultrastructure, OCR, mitochondrial respiratory enzyme activity, and cell proliferation-related protein levels. AMPK was validated to be a target gene of miR-1285 based on the result in which the miR-1285 mimic inhibited the luciferase activity of wild-type AMPK by 0.54-fold (p < 0.001). MiR-1285 mimic promoted the OCR of SCs, with 0.45-, 0.15-, 0.21-, and 0.30-fold (p < 0.01) increases in ATP production, basal and maximal respiration, and spare capacity, respectively. MiR-1285 mimic increased the mitochondrial respiratory enzyme activity of SCs, with 0.63-, 0.70-, and 0.97-fold (p < 0.01) increases in NADH-Q oxidoreductase, cytochrome c oxidase, and ATP synthase, respectively. Moreover, the miR-1285 mimic increased the protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.24-, 0.30-, 0.22-, and 0.13-fold (p < 0.05), respectively, and reduced the protein expression of p38 and p21 by 0.58- and 0.66-fold (p < 0.001). MiR-1285 inhibitor showed opposite effects on the above indicators and induced numerous autophagosomes and large lipid droplets in SCs. A high dose of estradiol (10 μM-6 h, showed a promotion of AMPK activation in a previous study) significantly inhibited SC ultrastructure, mitochondrial function, and proliferation-related pathways, while these adverse effects were weakened by Compound C treatment or miR-1285 mimic transfection. Our findings suggest that the activation and inhibition of AMPK induced by specific drugs or synthesized targeted miRNA fragments could regulate immature boar SC proliferative activity by influencing the CDK4/Cyclin D3 pathway and mitochondrial function; this helps to provide a basis for the prevention and treatment of male sterility in clinical practice.

Inhibitory Effects of Metformin for Pancreatic Neuroendocrine Neoplasms: Experimental Study on Mitochondrial Function

Although pancreatic neuroendocrine neoplasms (panNENs) are much less common and have a better prognosis than exocrine pancreatic cancers, their recurrence rate is not low, even in Grade 1 (World Health Organization classification) panNEN. Recently, there have been several reports that the progression-free survival in patients with unresectable panNEN could be improved by an antidiabetic drug, metformin, with the co-treatment of everolimus or a somatostatin analog. In this study, we aimed to evaluate the effects of metformin on cell metabolism and viability using the panNEN cell line, QGP-1, and RIN-m in culture. We observed an inhibitory effect of metformin on QGP-1 cell proliferation in a dose-dependent manner. Metformin was found to decrease the oxygen consumption rate in QGP-1 and RIN-m cells after metformin 48 h treatment and immediately after exposure. Cell proliferation was suppressed after metformin treatment. Phosphorylated adenosine monophosphate-activated protein kinase (AMPK) expression was increased, and cyclin D1 expression was decreased in RIN-m cells 24 h after metformin treatment by Western blotting in a dose-dependent manner. In conclusion, suppressive mitochondrial respiration and AMPK activation by metformin are, thus, suggested to inhibit panNEN cell viability and cell survival.

Rottlerin Enhances the Autophagic Degradation of Phosphorylated Tau in Neuronal Cells.

Intra-neuronal accumulation of hyper-phosphorylated tau as neurofibrillary tangles (NFT) is a hallmark of Alzheimer's disease (AD). To prevent the aggregation of phosphorylated tau in neurons, decreasing the phosphorylated tau protein levels is important. Here, we examined the biological effects of rottlerin, a phytochemical compound extracted from the Kamala tree, Mallotus philippinensis, on phosphorylated tau levels. Notably, rottlerin decreased the levels of intracellular phosphorylated and total tau. A marked increase in the LC3-II, a hallmark of autophagy, was observed in these cells, indicating that rottlerin strongly induced autophagy. Interestingly, rottlerin induced the phosphorylation of Raptor at S792 through the activation of adenosine-monophosphate activated-protein kinase (AMPK), which likely inhibits the mammalian target of rapamycin complex 1 (mTORC1), thus resulting in the activation of transcription factor EB (TFEB), a master regulator of autophagy. In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) activity increased in the presence of rottlerin. The decrease of phosphorylated tau levels in the presence of rottlerin was ameliorated by the knockdown of TFEB andpartially attenuated by the knockout of the Nrf2 gene. Taken together, rottlerin likely enhances the degradation of phosphorylated tau through autophagy activated by TFEB and Nrf2. Thus, our results suggest that a natural compound rottlerin could be used as a preventive and therapeutic drug for AD.

Identification of kinase modulators as host-directed therapeutics against intracellular methicillin-resistant Staphylococcus aureus.

The increasing prevalence of antimicrobial-resistant Staphylococcus aureus strains, especially methicillin-resistant S. aureus (MRSA), poses a threat to successful antibiotic treatment. Unsuccessful attempts to develop a vaccine and rising resistance to last-resort antibiotics urge the need for alternative treatments. Host-directed therapy (HDT) targeting critical intracellular stages of S. aureus emerges as a promising alternative, potentially acting synergistically with antibiotics and reducing the risk of de novo drug resistance. We assessed 201 ATP-competitive kinase inhibitors from Published Kinase Inhibitor Sets (PKIS1 and PKIS2) against intracellular MRSA. Seventeen hit compounds were identified, of which the two most effective and well-tolerated hit compounds (i.e., GW633459A and GW296115X) were selected for further analysis. The compounds did not affect planktonic bacterial cultures, while they were active in a range of human cell lines of cervical, skin, lung, breast and monocyte origin, confirming their host-directed mechanisms. GW633459A, structurally related to lapatinib, exhibited an HDT effect on intracellular MRSA independently of its known human epidermal growth factor receptor (EGFR)/(HER) kinase family targets. GW296115X activated adenosine monophosphate-activated protein kinase (AMPK), thereby enhancing bacterial degradation via autophagy. Finally, GW296115X not only reduced MRSA growth in human cells but also improved the survival rates of MRSA-infected zebrafish embryos, highlighting its potential as HDT.

Role of monocarboxylate transporters in AMPK-mediated protection against excitotoxic injury in the rat retina

Activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway protects against N-methyl-D-aspartic acid (NMDA)-induced excitotoxic retinal injury. AMPK activation enhances fatty acid metabolism and ketone body synthesis. Ketone bodies are transported into neurons by monocarboxylate transporters (MCTs) and exert neuroprotective effects. In this study, we examined the distribution and expression levels of MCT1 and MCT2 in the retina and analyzed the effects of pharmacological inhibition of MCTs on the protective effects of metformin and 5-aminoimidazole-4-carboxamide (AICAR), activators of AMPK, against NMDA-induced retinal injury in rats. MCT1 was expressed in the blood vessels, processes of astrocytes and Müller cells, and inner segments of photoreceptors in the rat retina, whereas MCT2 was expressed in neuronal cells in the ganglion cell layer (GCL) and in astrocyte processes. The expression levels of MCT2, but not MCT1, decreased one day after intravitreal injection of NMDA (200 nmol). Intravitreal injection of NMDA decreased the number of cells in the GCL compared to the vehicle seven days after injection. Simultaneous injection of metformin (20 nmol) or AICAR (50 nmol) with NMDA attenuated NMDA-induced cell loss in the GCL, and these protective effects were attenuated by AR-C155858 (1 pmol), an inhibitor of MCTs. AR-C155858 alone had no significant effect on the retinal structure. These results suggest that AMPK-activating compounds protect against NMDA-induced excitotoxic retinal injury via mechanisms involving MCTs in rats. NMDA-induced neurotoxicity may be associated with retinal neurodegenerative changes in glaucoma and diabetic retinopathy. Therefore, AMPK-activating compounds may be effective in managing these retinal diseases.

Endogenous AMPKα2 Mediates the Inhibition of Biliary Fibroblasts Proliferation

Background: Although it has been established that activating adenosine monophosphateactivated protein kinase (AMPK) inhibits cell proliferation in several cells, it is unknown whether AMPK is involved in inhibiting biliary fibroblast growth. Objective: The objective of this study is to specifically investigate the influence of AMPK isoforms on proliferation. Methods: To further address its underlying molecular mechanisms, primary cultured rat biliary fibroblasts were transfected with sequence-specific AMPK1 or AMPK2 siRNA. Results: Our findings show that knocking down AMPK2 greatly increased the proliferation of primary cultured biliary fibroblasts, accompanied by the activation of mTOR, an increase in S-phase kinaseassociated protein 2 (Skp2) expression, and a decrease in p27 protein levels. AMPK2 inhibition-triggered Skp2 overexpression and concomitant p27 decrease, as well as biliary fibroblast proliferation, were reversed by rapamycin inhibition or previous silencing of Skp2 production by targeted small interfering RNA (siRNA) transfection. Conclusion: We concluded that AMPK2 regulates the mTOR/Skp2/p27 signaling pathway and causes endogenous suppression of primary cultured biliary fibroblast growth. The reduction of biliary fibroblast proliferation by AMPK2 could be a potential method in treating benign biliary stricture (BBS).

Hygrothermal stress increases malignant arrhythmias susceptibility by inhibiting the LKB1-AMPK-Cx43 pathway

High mortality due to hygrothermal stress during heat waves is mostly linked to cardiovascular malfunction, the most serious of which are malignant arrhythmias. However, the mechanism associated with hygrothermal stress leading to malignant arrhythmias remains unclear. The energy metabolism regulated by liver kinase B1 (LKB1) and adenosine monophosphate-activated protein kinase (AMPK) and the electrical signaling based on gap junction protein, connexin43 (Cx43), plays important roles in the development of cardiac arrhythmias. In order to investigate whether hygrothermal stress induces arrhythmias via the LKB1-AMPK-Cx43 pathway, Sprague–Dawley rats were exposed to high temperature and humidity for constructing the hygrothermal stress model. A final choice of 40 °C and 85% humidity was made by pre-exploration based on different gradient environmental conditions with reference to arrhythmia event-inducing stability and risk of sudden death. Then, the incidence of arrhythmic events, as well as the expression, phosphorylation at Ser368, and distribution of Cx43 in the myocardium, were examined. Meanwhile, the adenosine monophosphate-activated protein kinase activator, Acadesine, was also administered to investigate the role played by AMPK in the process. Our results showed that hygrothermal stress induced malignant arrhythmias such as ventricular tachycardia, ventricular fibrillation, and severe atrioventricular block. Besides, hygrothermal stress decreased the phosphorylation of Cx43 at Ser368, induced proarrhythmic redistribution of Cx43 from polar to lateral sides of the cardiomyocytes, and also caused LKB1 and phosphorylated-AMPK expression to be less abundant. While, pretreatment with Acadesine significantly actived the LKB1-AMPK-Cx43 pathway and thus ameliorated malignant arrhythmias, indicating that the hygrothermal stress-induced arrhythmias is associated with the redistribution of gap junctions in cardiomyocytes and the organism's energy metabolism.

Statement of Retraction: Forkhead-box C1 attenuates high glucose-induced trophoblast cell injury during gestational diabetes mellitus via activating adenosine monophosphate-activated protein kinase through regulating fibroblast growth factor 19

Statement of Retraction: Forkhead-box C1 attenuates high glucose-induced trophoblast cell injury during gestational diabetes mellitus via activating adenosine monophosphate-activated protein kinase through regulating fibroblast growth factor 19

Hypothalamic AMP-Activated Protein Kinase as a Whole-Body Energy Sensor and Regulator.

5´-Adenosine monophosphate (AMP)-activated protein kinase (AMPK), a cellular energy sensor, is an essential enzyme that helps cells maintain stable energy levels during metabolic stress. The hypothalamus is pivotal in regulating energy balance within the body. Certain neurons in the hypothalamus are sensitive to fluctuations in food availability and energy stores, triggering adaptive responses to preserve systemic energy equilibrium. AMPK, expressed in these hypothalamic neurons, is instrumental in these regulatory processes. Hypothalamic AMPK activity is modulated by key metabolic hormones. Anorexigenic hormones, including leptin, insulin, and glucagon-like peptide 1, suppress hypothalamic AMPK activity, whereas the hunger hormone ghrelin activates it. These hormonal influences on hypothalamic AMPK activity are central to their roles in controlling food consumption and energy expenditure. Additionally, hypothalamic AMPK activity responds to variations in glucose concentrations. It becomes active during hypoglycemia but is deactivated when glucose is introduced directly into the hypothalamus. These shifts in AMPK activity within hypothalamic neurons are critical for maintaining glucose balance. Considering the vital function of hypothalamic AMPK in the regulation of overall energy and glucose balance, developing chemical agents that target the hypothalamus to modulate AMPK activity presents a promising therapeutic approach for metabolic conditions such as obesity and type 2 diabetes mellitus.

Fructose Potentiates Bone Loss and Marrow Adipose Tissue Accumulation by Inhibiting Adenosine 5'-Monophosphate-Activated Protein Kinase in Mesenchymal Stem Cells.

Increased fructose consumption has been elucidated to contribute to metabolic diseases. Bone is a dynamic organ that undergoes constant remodeling. However, the effects of fructose on bone health are still in dispute. Here, we identified fructose deteriorated bone mineral density while promoting the abundance of bone marrow adipose tissue. Fructose remarkably promoted the bone marrow mesenchymal stem cells' (BMMSCs) adipogenic commitment at the expense of osteogenic commitment. Fructose boosted the glycolysis of BMMSCs and inhibited phosphorylation of adenosine 5'-monophosphate-activated protein kinase (AMPK), which played a crucial role in bone-fat alteration. Our results suggested that fructose potentiated bone loss and marrow adipose tissue accumulation by suppressing AMPK activation in BMMSCs. Understanding fructose which affected bone metabolism was thus of primary importance in order to establish preventative measures or treatments for this condition.

Schizophrenia risk proteins ZNF804A and NT5C2 interact in cortical neurons.

The zinc finger protein 804A (ZNF804A) and the 5'-nucleotidase cytosolic II (NT5C2) genes are amongst the first schizophrenia susceptibility genes to have been identified in large-scale genome-wide association studies. ZNF804A has been implicated in the regulation of neuronal morphology and is required for activity-dependent changes to dendritic spines. Conversely, NT5C2 has been shown to regulate 5' adenosine monophosphate-activated protein kinase activity and has been implicated in protein synthesis in human neural progenitor cells. Schizophrenia risk genotype is associated with reduced levels of both NT5C2 and ZNF804A in the developing brain, and a yeast two-hybrid screening suggests that their encoded proteins physically interact. However, it remains unknown whether this interaction also occurs in cortical neurons and whether they could jointly regulate neuronal function. Here, we show that ZNF804A and NT5C2 colocalise and interact in HEK293T cells and that their rodent homologues, ZFP804A and NT5C2, colocalise and form a protein complex in cortical neurons. Knockdown of the Zfp804a or Nt5c2 genes resulted in a redistribution of both proteins, suggesting that both proteins influence the subcellular targeting of each other. The identified interaction between ZNF804A/ZFP804A and NT5C2 suggests a shared biological pathway pertinent to schizophrenia susceptibility within a neuronal cell type thought to be central to the neurobiology of the disorder, providing a better understanding of its genetic landscape.

Pharmacological Activation of AMPK Prevents Drp1-mediated Mitochondrial Fission and Alleviates Hepatic Steatosis In vitro.

The incidence of non-alcoholic fatty liver disease (NAFLD) is increasing worldwide. Adenosine monophosphate-activated protein kinase (AMPK) activation is beneficial for NAFLD treatment. Recent studies show the excessive fission of mitochondria during NAFLD progression, so targeting mitochondria dynamics may be a possible target for NAFLD. Still, little is known about whether AMPK regulates mitochondrial dynamics in hepar. This study investigated whether AMPK activation alleviates hepatic steatosis by regulating mitochondrial dynamics mediated by GTPase dynamin-related protein 1 (Drp1). Human hepatocyte line L-02 cells were cultured and subjected to palmitic acid (PA) treatment for 24 h to establish a hepatic steatosis model in vitro, which was pre-treated with different tool drugs. Hepatocyte function, hepatocyte lipid content, mitochondrial reactive oxygen species (ROS) production, and mitochondrial membrane potential (MMP) were examined. The expression levels of genes and proteins associated with mitochondrial dynamics were assessed using reverse transcription-quantitative PCR and western blotting. The results indicated that 5-Aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR), an AMPK activator, improved hepatocyte function, as demonstrated by decreased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity (P＜0.05 or P＜0.01). In addition, AICAR decreased total cholesterol (TC) and triglyceride (TG) content and lipid deposition in hepatocytes (P＜0.01); decreased ROS production; improved MMP (P＜0.01); reduced fission-1 (Fis1) and mitochondrial fission factor (Mff) mRNA expression; and downregulated p-Drp1 (Ser 616) protein expression. In contrast, AICAR increased mitochondrial fusion factor mitofusin-1 (Mfn1) and mitofusin-2 (Mfn2) mRNA expression and upregulated p-Drp1 (Ser 637) protein expression. Mdivi-1, a Drp-1 inhibitor, was used to confirm whether mitochondrial dynamics regulated by Drp1-mediated the role of AICAR. Similar to AICAR, Mdivi-1 improved hepatocyte function and MMP significantly, decreased ROS production and lipid deposition, downregulated Fis1 and Mff mRNA expression, downregulated p-Drp1 (Ser 616) protein expression, and enhanced Mfn1 and Mfn2 mRNA and p-Drp1 (Ser 637) protein expression. However, Compound C, an AMPKspecific inhibitor, had less impact on the protective effect of Mdivi-1. The results demonstrated that AMPK activation has a protective effect on hepatic steatosis in vitro, largely dependent on the inhibition of Drp1-mediated mitochondrial fission.

Comparative effects of canagliflozin and sitagliptin in chronically ischemic myocardium

Aim: Recent studies demonstrate that sodium-glucose cotransporter 2 inhibitors (SGLT2i) and dipeptidyl peptidase-4 inhibitors (DPP4i), two classes of antidiabetic drugs, are cardioprotective. However, the mechanisms of these benefits and their comparative efficacy remain unclear. We aimed to compare the effects of these antidiabetic agents on cardiac function, perfusion, and microvascular density using a swine model of chronic myocardial ischemia. Methods: Chronic myocardial ischemia was induced in Yorkshire swine by ameroid constrictor placement to the left circumflex artery. Two weeks later, pigs were administered vehicle (“CON”, 8 pigs), 300 mg SGLT2i canagliflozin, (“CANA”, 8 pigs), or 100 mg DPP4i sitagliptin (“SIT”, 5 pigs) daily. Five weeks later, pigs were euthanized. Cardiac function, perfusion, collateralization, and protein expression were determined by pressure-volume catheter, microsphere analysis, immunofluorescence, and immunoblotting, respectively. Results: Compared with SIT, CANA was associated with improved stroke volume and cardiac output, with a trend towards reduced left ventricular stiffness. Both CANA and SIT trended towards improved perfusion compared to CON, but there were no differences between the two treatment groups. SIT was associated with improved capillary density with a trend towards improved arteriolar density compared to CANA. Both CANA and SIT were associated with increased expression of vascular endothelial cadherin compared to CON, without differences in treatment groups. SIT pigs had decreased 5′ adenosine monophosphate-activated protein kinase activation compared to CON and CANA. There was a trend towards increased endothelial nitric oxide synthase activation in the SIT group compared to CON. There were no differences in activation of extracellular signal-regulated kinase 1/2 across groups. Conclusions: In the setting of chronic myocardial ischemia, canagliflozin is associated with improved cardiac function compared to sitagliptin, with similar effects on perfusion despite differences in microvascular collateralization.

Involvement of gut microbiota recovery and autophagy induction in Youhua Kuijie Formula's protection against experimental ulcerative colitis.

Ulcerative colitis (UC) is characterized by overactive inflammatory response, impaired intestinal mucosal barrier and disrupted gut microbiota. Youhua Kuijie formula is a classic empirical prescription based on the pathogenesis of UC. The present study was designed to verify the protective effect of Youhua Kuijie Formula on DSS-induced UC in mice and uncover the related mechanism. Youhua Kuijie Formula were orally administrated to UC mice induced by DSS dissolved in drinking water for ten days. The protective effect of Youhua Kuijie Formula was evidenced by reduced pathological symptoms accompanied by palliative inflammatory response and relatively intact intestinal barrier. The data from 16S rRNA gene sequencing and GC-MS untargeted metabolomics indicated that the supplement of Youhua Kuijie Formula restructured gut microbiota community structure, and thereby modulated the metabolic profiles in UC mice. The analysis of pathway enrichment analysis suggested the major alterations in metabolic pathway were related to protein digestion and absorption. Besides, the results of the following experiments suggested that Youhua Kuijie Formula treatment increased adenosine monophosphate-activated protein kinase (AMPK) activation, decreased mechanistic target of rapamycin (mTOR) phosphorylation, and thereby reversing autophagy deficiency in the intestinal tract of UC mice. Collectively, our results demonstrated that the regulation of AMPK/mTOR was involved in Youhua Kuijie Formula administration mediated protective effect on UC.

Hippocampal and gut AMPK activation attenuates enterocolitis-like symptoms and co-occurring depressive-like behavior in ulcerative colitis model mice: Involvement of brain-gut autophagy

Patients with inflammatory bowel disease, including ulcerative colitis (UC) and Crohn's disease, have a high incidence of psychiatric disorders, including depression and anxiety. However, the underlying pathogenic mechanism remains unknown. Dextran sulfate sodium (DSS)-treated mice, a model of UC, exhibit depressive-like behavior and reduced adenosine monophosphate-activated protein kinase (AMPK) activity, which regulates various physiological functions in the brain and gut. However, comprehensive studies on UC pathophysiology with co-occurring depression focused on brain-gut AMPK activity are lacking. Therefore, we aimed to investigate whether resveratrol (RES), an AMPK activator, prevented DSS-induced UC-like symptoms and depressive-like behavior. DSS treatment induced UC-like pathology and depressive-like behavior, as assessed via the tail suspension test. Moreover, western blotting and immunohistochemical studies revealed that DSS increased p-p70S6 kinase (Thr389), p62, tumor necrosis factor-α, interleukin (IL)-1β, IL-18, NLR family pyrin domain containing 3 (NLRP3), cleaved caspase-1, cleaved Gasdermin-D (GSDMD), and cleaved caspase-3 expression levels in the rectum and hippocampus, and increased CD40, iNOS, and Kelch-like ECH-associated protein 1 expression levels, and the number of Iba1-positive cells in the hippocampus, and decreased p-AMPK and LC3II/I expression levels, and the number of NF-E2-related factor 2 (Nrf2)-positive cells, and reduced neurogenesis in the hippocampus. These changes were reversed by the RES administration. RES also enhanced PGC1α and SOD1 expression in the hippocampus of DSS-treated male mice. Moreover, NLRP3 staining was observed in the neurons and microglia, and cleaved GSDMD staining in neurons in the hippocampus of DSS-treated mice. Notably, RES prevented UC-like pathology and depressive-like behavior and enhancement of autophagy, decreased rectal and hippocampal inflammatory cytokines and inflammasome, and induced the Nrf2-PGC1α-SOD1 pathway in the hippocampus, resulting in neurogenesis in the hippocampal dentate gyrus. Our findings suggest that brain-gut AMPK activation may be an important therapeutic strategy in patients with UC and depression.

Antiproliferative Activity of Gibbosic Acid H through Induction of G0/G1 Cell Cycle Arrest and Apoptosis in Human Lung Cancer Cells.

Lung cancer is one of the most common causative cancers worldwide. Particularly, non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancer cases. NSCLC is a serious form of lung cancer that requires prompt diagnosis, and the 5-year survival rate for patients with this disease is only 24%. Gibbosic acid H (GaH), a natural lanostanoid obtained from the Ganoderma species (Ganodermataceae), has antiproliferative activities against colon and lung cancer cells. The aim of the present study was to evaluate the antiproliferative activity of GaH in NSCLC cells and to elucidate the underlying molecular mechanisms. GaH was found to induce G0/G1 cell cycle arrest and autophagy by activating adenosine monophosphate-activated protein kinase in A549 and H1299 cells. The induction of this cell cycle arrest was associated with the downregulation of cyclin E1 and CDK2. Additionally, the induction of autophagy by GaH was correlated with the upregulation of LC3B, beclin-1, and p53 expression. GaH also induced apoptosis by upregulating cleaved caspase-3 and Bax in the lung cancer cells. These findings suggest that GaH has a potential in the growth inhibition of human lung cancer cells.

Ezetimibe ameliorates cisplatin-induced nephrotoxicity: A novel therapeutic approach via modulating AMPK/Nrf2/TXNIP signaling.

Cisplatin (Cis) is among the most powerful antineoplastic medications, nevertheless, its serious side effects; particularly nephrotoxicity designates a major concern. Previous studies reported that ezetimibe (Eze), a well-known antihyperlipidemic drug, exerts additional trivial pharmacological effects. In this work, we displayed Eze as an intriguing protective candidate in a cisplatin-induced nephrotoxicity rat model through AMPK activation. Eze (10 mg/kg, p.o.) was administered for two weeks and Cis (10 mg/kg, i.p.) was administered on the 10th day to induce nephrotoxicity in male Wistar rats. Treatment with Eze greatly augmented the phosphorylation of adenosine 5'-monophosphate-activated protein kinase (AMPK) and the antioxidant regulator; nuclear factor erythroid 2-related factor 2 (Nrf2), thus, mitigating oxidative injury through induction of the antioxidant enzymes, such as heme oxygenase-1 (HO-1) and glutathione reductase (GR). As well, Eze relieved inflammation by reducing protein expression of thioredoxin-interacting protein (TXNIP) and nucleotide-binding domain-like receptor protein 3 (NLRP3), which led to a decrease in the release of caspase-1, in addition to, the inflammatory markers IL-18and IL-1 β. Besides, Eze ameliorated apoptosis in the renal cells through inhibiting the phosphorylated Apoptosis signal-regulating kinase-1(p-ASK1), caspase-3 and reducing Bax/Bcl2ratio. Correspondingly, histopathological examination corroborated the previous biochemical findings. Collectively, Eze exerts significant renal protection against Cis-induced nephrotoxicity via antioxidant, anti-inflammatory and anti-apoptotic pathways that are probably mediated, at least partly, via activating AMPK/Nrf2/HO-1 pathway and conquering both TXNIP/NLRP3 inflammasome and TXNIP/ASK1 signaling pathways. To confirm the protective effect of Eze via AMPK-activation, an AMPK-inhibitor, dorsomorphin (Dors), when co-administered with Eze abolished its protective effect.