AMP-activated Protein Kinase Signaling Pathway Research Articles

Aflatoxin B1 Promotes Pyroptosis in IPEC-J2 Cells by Disrupting Mitochondrial Dynamics through the AMPK/NLRP3 Pathway.

Aflatoxin B1 (AFB1) is one of the most toxic mycotoxins in food and feed, seriously jeopardizing the intestinal health, while the effects of AFB1 on intestinal damage remain to be well understood. This study aims to evaluate the effect of AFB1 on intestinal injury by regulating AMP-activated protein kinase (AMPK)-mediated pyroptosis in vitro. The present study showed that AFB1 led to the formation of large number of bubble-like protrusions on the cell membrane, releasing lactate dehydrogenase (LDH) and interleukin-1β (IL-1β). Stimulation with AFB1 resulted in the activation of the NOD-like receptor protein 3 (NLRP3) pathway, as indicated by the increased expression of pyroptosis-associated factor mRNAs and proteins, which ultimately led to a significant upregulation of the pyroptosis rate. Meanwhile, AFB1 caused dysfunction of mitochondrial dynamics by activating the AMPK signaling pathway as mainly evidenced by upregulating dynamin-1-like protein 1 (Drp1) mRNA and protein expression. Moreover, inhibition of NLRP3 and AMPK pathways by MCC950 and compound C, respectively, significantly alleviated AFB1-induced damage in IPEC-J2 cells, evidenced by suppressed NLRP3-mediated pyroptosis, and ameliorated AMPK-mediated mitochondrial dynamics imbalance. In conclusion, these results demonstrated that AFB1 promoted pyroptosis of IPEC-J2 cells by interfering with mitochondrial dynamics by activating the AMPK/NRLP3 pathway.

Delphinidin induces a fast-to-slow muscle fiber type shift through the AMPK signaling pathway in C2C12 myotubes

Delphinidin, a plant anthocyanidin, suppresses disuse muscle atrophy in mice. However, its effect on muscle fiber type shift is unclear. To examine whether delphinidin affects skeletal muscle fiber type, differentiated C2C12 cells were treated with delphinidin. Results revealed that delphinidin upregulated the mRNA expression of myosin heavy chain type I (MyHCI), troponin C1, troponin I1, and MyHCIIx and increased slow MyHC protein level in C2C12 myotubes. Delphinidin also enhanced succinic dehydrogenase (SDH) activities and suppressed lactate dehydrogenase (LDH) activity. Adenosine monophosphate–activated protein kinase (AMPK) inhibition attenuated delphinidin-induced MyHCI upregulation and MyHCIIb downregulation. We investigated the effect of delphinidin on the upstream factors involved in AMPK activation. Delphinidin increased liver kinase B1 (LKB1) phosphorylation and nuclear respiratory factor 1 (NRF1) and calcium/calmodulin-dependent protein kinase 2 (CaMKK2) protein levels. In conclusion, delphinidin induced muscle fiber type conversion from fast-twitch to slow-twitch muscles through the AMPK signaling pathway.

Improvement of osteogenic differentiation potential of placenta-derived mesenchymal stem cells by metformin via AMPK pathway activation

BackgroundPlacenta-derived human mesenchymal stem cells (PL-MSCs) have gained a lot of attention in the field of regenerative medicine due to their availability and bone-forming capacity. However, the osteogenic differentiation capacity of these cells remains inconsistent and could be improved to achieve greater efficiency. Although metformin, a widely used oral hypoglycemic agent, has been shown to increase bone formation in various cell types, its effect on osteogenic differentiation of PL-MSCs has not yet been investigated. Therefore, the objective of this study was to examine the effect of metformin on the osteogenic differentiation capacity of PL-MSCs and the underlying mechanisms.MethodsThe PL-MSCs were treated with 0.5 to 640 µM metformin and their osteogenic differentiation capacity was examined by an alkaline phosphatase (ALP) activity assay, Alizarin red S staining and expression levels of osteogenic genes. The role of adenosine 5′-monophosphate-activated protein kinase (AMPK) signaling in mediating the effect of metformin on the osteogenic differentiation capacity of PL-MSCs was also investigated by determining levels of phosphorylated AMPK (pAMPK)/AMPK ratio and by using compound C, an AMPK inhibitor.ResultsThe results showed that 10–160 µM metformin significantly increased the viability of PL-MSCs in a dose- and time-dependent manner. Furthermore, 80–320 µM metformin also increased ALP activity, matrix mineralization, and expression levels of osteogenic genes, runt-related transcription factor 2 (RUNX2), osterix (OSX), osteocalcin (OCN) and collagen I (COL1), in PL-MSCs. Metformin increases osteogenic differentiation of PL-MSCs, at least in part, through the AMPK signaling pathway, since the administration of compound C inhibited its enhancing effects on ALP activity, matrix mineralization, and osteogenic gene expression of PL-MSCs.ConclusionsThis study demonstrated that metformin at concentrations of 80–320 μM significantly enhanced osteogenic differentiation of PL-MSCs in a dose- and time-dependent manner, primarily through activation of the AMPK signaling pathway. This finding suggests that metformin could be used with other conventional drugs to induce bone regeneration in various bone diseases. Additionally, this study provides valuable insights for future osteoporosis treatment by highlighting the potential of modulating the AMPK pathway to improve bone regeneration.

AdipoRon improves mitochondrial homeostasis and protects dopaminergic neurons through activation of the AMPK signaling pathway in the 6-OHDA-lesioned rats

The progressive decline of dopaminergic neurons in Parkinson's disease (PD) has been linked to an imbalance in energy and the failure of mitochondrial function. AMP-activated protein kinase (AMPK), the major intracellular energy sensor, regulates energy balance, and damage to nigral dopaminergic neurons induced by 6-hydroxydopamine (6-OHDA) is exacerbated in the absence of AMPK activity. This study aimed to examine the potential therapeutic advantages of AdipoRon, an AMPK activator, on motor function and mitochondrial homeostasis in a 6-OHDA-induced PD model. Male Wistar rats were subjected to unilateral injection of 6-OHDA (10 μg) into the left medial forebrain bundle at two points, and after 7 days, they were treated with intranasal AdipoRon (0.1, 1, and 10 μg) or Levodopa (10 mg/kg, p. o.) for 21 successive days. Following the last treatment day, motor behavior was evaluated through the Murprogo's test, bar test, beam walking test, and apomorphine-induced rotation test. After euthanasia, the left substantia nigra (SN) was separated for evaluation of ATP, mitochondrial membrane potential (MMP), and protein expressions of AMPK, p-AMPK, and mitochondrial dynamics markers (Mfn-2 and Drp-1). Moreover, the number of tyrosine hydroxylase-positive (TH+) cells was quantified in the left substantia nigra. Intranasal AdipoRon effectively reversed muscle rigidity, akinesia, bradykinesia, and rotation caused by 6-OHDA. Moreover, AdipoRon increased the phospho-AMPK/AMPK ratio, mitigated mitochondrial dysfunction, and improved mitochondrial dynamics in the SN. Furthermore, AdipoRon increased the number of TH+ cells in the SN of PD animals. These findings suggest that AdipoRon could protect dopaminergic neurons by activating the AMPK pathway and improving mitochondrial dysfunction.

A polysaccharide from edible red seaweed Bangia fusco-purpurea prevents obesity in high-fat diet-induced C57BL/6 mice

The study aimed to investigate the impacts of a polysaccharide (BFP) from Bangia fusco-purpurea on high-fat diet (HFD)-induced obesity in C57BL/6 mice, as well as its underlying mechanisms. Our results showed that orally administrated BFP was more effective than inulin (INU) in reducing body weight and fat accumulation in obese mice, indicating its anti-obesity effect. BFP effectively improved the compositions and metabolites of intestinal microbiota in obese mice, leading to enhanced energy metabolism and lipid metabolism, thus contributing to its anti-obesity effect. Notably, the better anti-obesity effect of BFP compared to INU were attributed to their varying degrees of modulation of specific intestinal microbial taxa, such as Clostridium and Aerococcus, as well as the regulation of differential metabolites (including biotin, piperine, G6P, etc.) also varies. Also, both in vitro (3T3-L1 preadipocytes) and in vivo (HFD-induced obese mice) models confirmed that BFP achieved anti-obesity effect attributed to enhance energy metabolism, promote lipolysis, increase fatty acid oxidation, and inhibit adipogenesis via activating the AMP-activated protein kinase and Acetyl-CoA carboxylase signaling pathways and suppressing the peroxisome proliferator-activated receptor γ expression. Our findings suggest that BFP has the potential to be used as prebiotics, dietary agents, and nutritional supplements against obesity.

The Effects of Warm Acupuncture on the Expression of AMPK in High-Fat Diet-Induced MAFLD Rats.

This study investigated the effects of acupuncture and warm acupuncture on the expression and mechanism of the AMP-activated protein kinase (AMPK) signalling pathway associated with lipid accumulation in the liver tissue of rats with metabolic dysfunction-associated fatty liver disease (MAFLD) induced by a high-fat diet. Sprague-Dawley rats were categorised into four groups: control (CON), untreated MAFLD (MAFLD), and two MAFLD groups treated with acupuncture (ACU) and warm acupuncture (WA). The treatment groups underwent 16 application sessions over 8 weeks at the SP9 and BL18 acupoints. We measured the expression levels of AMPK, sterol regulatory element-binding protein1 (SREBP1), acetyl-coenzyme A carboxylase (ACC), peroxisome proliferator-activated receptorα (PPARα), carnitine palmitoyltransferase1 (CPT1), and CPT2. AMPK was activated in both ACU and WA groups. WA downregulated both SREBP1 and ACC expression at the protein level, whereas the acupuncture treatment downregulated SREBP1 expression. Additionally, WA selectively induced the activation of signalling pathways related to AMPK, PPARα, CPT1, and CPT2 at the mRNA level. Histological observations confirmed that fat accumulation was reduced in both the ACU and the WA groups compared to the MAFLD group. The WA treatment-promoted amelioration of HFD-induced MAFLD may be related to the activation of the AMPK/SREBP1/ACC pathway in the liver.

Atractylodes Lancea and Its Constituent, Atractylodin, Ameliorates Metabolic Dysfunction-Associated Steatotic Liver Disease via AMPK Activation.

Metabolic dysfunction-associated steatotic liver disease (MASLD), which encompasses a spectrum of conditions ranging from simple steatosis to hepatocellular carcinoma, is a growing global health concern associated with insulin resistance. Since there are limited treatment options for MASLD, this study investigated the therapeutic potential of Atractylodes lancea, a traditional herbal remedy for digestive disorders in East Asia, and its principal component, atractylodin, in treating MASLD. Following 8 weeks of high-fat diet (HFD) feeding, mice received oral doses of 30, 60, or 120 mg/kg of Atractylodes lancea. In HFD-fed mice, Atractylodes lancea treatment reduced the body weight; serum triglyceride, total cholesterol, and alanine aminotransferase levels; and hepatic lipid content. Furthermore, Atractylodes lancea significantly ameliorated fasting serum glucose, fasting serum insulin, and homeostatic model assessment of insulin resistance levels in response to HFD. Additionally, a glucose tolerance test demonstrated improved glucose homeostasis. Treatment with 5 or 10 mg/kg atractylodin also resulted in anti-obesity, anti-steatosis, and glucose-lowering effects. Atractylodin treatment resulted in the downregulation of key lipogenic genes (Srebf1, Fasn, Scd2, and Dgat2) and the upregulation of genes regulated by peroxisome proliferator-activated receptor-α. Notably, the molecular docking model suggested a robust binding affinity between atractylodin and AMP-activated protein kinase (AMPK). Atractylodin activated AMPK, which contributed to SREBP1c regulation. In conclusion, our results revealed that Atractylodes lancea and atractylodin activated the AMPK signaling pathway, leading to improvements in HFD-induced obesity, fatty liver, and glucose intolerance. This study suggests that the phytochemical, atractylodin, can be a treatment option for MASLD.

Leonurine Inhibits Hepatic Lipid Synthesis to Ameliorate NAFLD via the ADRA1a/AMPK/SCD1 Axis.

Leonurine is a natural product unique to the Lamiaceae plant Leonurus japonicus Houtt., and it has attracted attention due to its anti-oxidative stress, anti-apoptosis, anti-fibrosis, and metabolic regulation properties. Also, it plays an important role in the prevention and treatment of nonalcoholic fatty liver disease (NAFLD) through a variety of biological mechanisms, but its mechanism of action remains to be elucidated. Therefore, this study aims to preliminarily explore the mechanisms of action of leonurine in NAFLD. Mice were randomly divided into four groups: the normal control (NC) group, the Model (M) group, the leonurine treatment (LH) group, and the fenofibrate treatment (FB) group. The NAFLD model was induced by a high-fat high-sugar diet (HFHSD) for 12 weeks, and liver pathological changes and biochemical indices were observed after 12 weeks. Transcriptomic analysis results indicated that leonurine intervention reversed the high-fat high-sugar diet-induced changes in lipid metabolism-related genes such as stearoyl-CoA desaturase 1 (Scd1), Spermine Synthase (Sms), AP-1 Transcription Factor Subunit (Fos), Oxysterol Binding Protein Like 5 (Osbpl5), and FK506 binding protein 5 (Fkbp5) in liver tissues. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis results suggest that leonurine may exert its lipid-lowering effects through the AMP-activated protein kinase (AMPK) signaling pathway. Liver lipidomic analysis showed that leonurine could alter the abundance of lipid molecules related to fatty acyl (FAs) and glycerophospholipids (GPs) such as TxB3, carnitine C12-OH, carnitine C18:1-OH, and LPC (20:3/0:0). Molecular biology experiments and molecular docking techniques verified that leonurine might improve hepatic lipid metabolism through the alpha-1A adrenergic receptor (ADRA1a)/AMPK/SCD1 axis. In summary, the present study explored the mechanism by which leonurine ameliorated NAFLD by inhibiting hepatic lipid synthesis via the ADRA1a/AMPK/SCD1 axis.

Canola Oil Ameliorates Obesity by Suppressing Lipogenesis and Reprogramming the Gut Microbiota in Mice via the AMPK Pathway.

obesity is a worldwide problem that seriously endangers human health. Canola oil (Col) has been reported to regulate hepatic steatosis by influencing oxidative stress and lipid metabolism in Kunming mice. However, whether Col exhibits an anti-obesity effect by altering the gut microbiota remains unknown. in this study, we observed that a high-fat diet increased lipogenesis and gut microbiota disorder in C57BL/6J male mice, while the administration of Col suppressed lipogenesis and improved gut microbiota disorder. the results show that Col markedly reduced the final body weight and subcutaneous adipose tissue of C57BL/6J male mice fed a high-fat diet (HFD) after 6 weeks of administration. However, although Col did not effectively increase the serum concentration of HDL, we found that treatment with Col notably inhibited the low-density lipoprotein (LDL), total cholesterol (TC), and triglycerides (TGs) in HFD mice. Furthermore, Col ameliorated obesity in the liver compared to mice that were only fed a high-fat diet. We also found that Col significantly inhibited the relative expression of sterol regulatory element binding protein (SREBP1/2), peroxisome proliferator-activated receptor γ (PPARγ), and insulin-induced genes (Insig1/2) that proved to be closely associated with lipogenesis in HFD mice. In addition, the concentration of acetic acid was significantly increased in Col-treatment HFD mice. Further, we noted that Col contributed to the reprogramming of the intestinal microbiota. The relative abundances of Akkermansia, Dubosiella, and Alistipes were enhanced under treatment with Col in HFD mice. The results also imply that Col markedly elevated the phosphorylation level of the AMP-activated protein kinase (AMPK) pathway in HFD mice. the results of our study show that Col ameliorates obesity and suppresses lipogenesis in HFD mice. The underlying mechanisms are possibly associated with the reprogramming of the gut microbiota, in particular, the acetic acid-mediated increased expression of Alistipes via the AMPK signaling pathway.

Protective properties of milk-derived peptide QEPV in attenuating oxidative stress through AMPK/PPARα signaling pathways

Peptides derived from fermented milk have demonstrated diverse bioactivities. In this study, we investigated the protective effects of Gln-Glu-Pro-Val (QEPV) against hydrogen peroxide (H2O2) induced oxidative stress in RAW 264.7 cells and further evaluated its potential in Drosophila melanogaster (D. melanogaster). Pre-incubation with QEPV in vitro dose-dependently promoted cell viability, suppressed oxidative damage, and increased antioxidant enzyme activities in a low-level oxidative stress model. QEPV remarkably extended lifespan under the low-level oxidative stress, validating its practical applicability. These effects were attributed to the upregulation of 5′ adenosine monophosphate-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor alpha (PPARα) signaling pathways, as revealed by label-free proteomic analysis. To confirm the roles of these two signaling pathways, antagonists of AMPK (Dorsomorphin) and PPARα (GW6471) were applied in vitro and in vivo. The protective effects of QEPV were reversed by both antagonists. Overall, QEPV exhibits protective properties by selectively upregulating the AMPK and PPARα signaling pathways, indicating its potential application as an antioxidant food ingredient.

Research progress of probiotics regulating intestinal micro-ecological environment in obese patients after bariatric surgery

Bariatric surgery may cause intestinal microecological environment imbalance due to changes in gastrointestinal anatomy. Some patients may have compli-cations, even regain weight. Probiotics can act on intestinal mucosa, epithelium and gut-associated lymphoid tissue to improve the intestinal microecological environment of obese patients after bariatric surgery. Probiotics can promote the production of short-chain fatty acids, stimulate intestinal cells to release glucagon-like peptide-1, peptide tyrosine-tyrosine, insulin and other endocrine hormones, affect the function of the central nervous system through the gut-brain axis, make patients after bariatric surgery feel full, and reduce blood sugar at the same time. Probiotics can produce lactic acid, acetic acid and lactase, to inhibit the growth of harmful bacteria and to improve gastrointestinal symptoms of patients after bariatric surgery. Probiotics can activate the AMP-activated protein kinase signaling pathway, improve lipid metabolism, and promote the recovery of symptom indicators of nonalcoholic fatty liver disease after bariatric surgery. Probiotics can regulate the release of neurotransmitters or metabolites by the microbiota through the gut-brain axis to affect brain activity and behavior, thus helping patients improve negative emotions after bariatric surgery. This article describes the intestinal microecological environment of obese patients and mechanism of the change after bariatric surgery and summarizes the effects and possible mechanisms of probiotics in improving the intestinal microecological environment of obese patients after bariatric surgery, to provide references for promoting the clinical application of probiotics.

UGT1A7 altered HER2-positive breast cancer response to trastuzumab by affecting epithelial-to-mesenchymal transition: A potential biomarker to identify patients resistant to trastuzumab treatment.

HER2-positive (HER2+) breast cancer accounts for 20-30% of all breast cancers. Although trastuzumab has significantly improved the survival of patients with HER2+ breast cancer, more than 70% of patients develop drug resistance within one year of treatment. Differential-gene-expression analysis of trastuzumab-sensitive and resistant HER2+ breast cancer cell lines from GSE15043 was performed to identify the biomarkers associated with trastuzumab resistance. Differential biomarker expression was confirmed in FFPE tissues collected from clinical HER2+ breast cancer tumor samples that were sensitive or resistant to trastuzumab treatment. UGT1A7, a member of the uronic acid transferase family, was associated with trastuzumab resistance. UGT1A7 expression was downregulated in trastuzumab-resistant tumor tissues and in a cell line that developed trastuzumab resistance (BT474TR). Overexpressing UGT1A7 in BT474TR restored their sensitivity to trastuzumab treatment, whereas downregulating UGT1A7 expression in parental cells led to trastuzumab resistance. Importantly, UGT1A7 localized to the endoplasmic reticulum and altered stress responses. Furthermore, downregulating UGT1A7 expression promoted epithelial-to-mesenchymal transition (EMT) by affecting TWIST, SNAIL, and GRP78 expression and the AMP-activated protein kinase signaling pathway, thus contributing to trastuzumab resistance. This study demonstrated the important role and novel mechanisms of UGT1A7 in tumor responses to trastuzumab. Low UGT1A7 expression plays an important role in EMT and contributes to trastuzumab resistance. UGT1A7 has the potential to be developed as a biomarker for identifying patients who are resistant to trastuzumab treatment.

Metabolomics Dysfunction in Replicative Senescence of Periodontal Ligament Stem Cells Regulated by AMPK Signaling Pathway.

Periodontal ligament mesenchymal stem cells (PDLSCs) are a promising cell resource for stem cell-based regenerative medicine in dentistry, but they inevitably acquire a senescent phenotype after prolonged in vitro expansion. The key regulators of PDLSCs during replicative senescence remain unclear. Here, we sought to elucidate the role of metabolomic changes in determining the cellular senescence of PDLSCs. PDLSCs were cultured to passages 4, 10, and 20. The senescent phenotypes of PDLSCs were detected, and metabolomics analysis was performed. We found that PDLSCs manifested senescence phenotype during passaging. Metabolomics analysis showed that the metabolism of replicative senescence in PDLSCs varied significantly. The AMP-activated protein kinase (AMPK) signaling pathway was closely related to adenosine monophosphate (AMP) levels. The AMP:ATP ratio increased in senescent PDLSCs; however, the levels of p-AMPK, FOXO1 and FOXO3a decreased with senescence. We treated PDLSCs with an activator of the AMPK pathway (AICAR) and observed that the phosphorylated AMPK level at P20 PDLSCs was partially restored. These data delineate that the metabolic process of PDLSCs is active in the early stage of senescence and attenuated in the later stages of senescence; however, the sensitivity of AMPK phosphorylation sites is impaired, causing senescent PDLSCs to fail to respond to changes in energy metabolism.

AMPK and O-GlcNAcylation: interplay in cardiac pathologies and heart failure.

Heart failure (HF) represents a multifaceted clinical syndrome characterized by the heart's inability to pump blood efficiently to meet the body's metabolic demands. Despite advances in medical management, HF remains a major cause of morbidity and mortality worldwide. In recent years, considerable attention has been directed toward understanding the molecular mechanisms underlying HF pathogenesis, with a particular focus on the role of AMP-activated protein kinase (AMPK) and protein O-GlcNAcylation. This review comprehensively examines the current understanding of AMPK and O-GlcNAcylation signalling pathways in HF, emphasizing their interplay and dysregulation. We delve into the intricate molecular mechanisms by which AMPK and O-GlcNAcylation contribute to cardiac energetics, metabolism, and remodelling, highlighting recent preclinical and clinical studies that have explored novel therapeutic interventions targeting these pathways.

Theoretical investigation of complexes of uranyl(II), vanadyl(II) and zirconyl(II) with vitamin B13 as candidate compounds as anti-nonalcoholic fatty liver disease and Diabetes mellitus targets

Density functional theory (DFT) calculations were carried out utilizing the B3LYP functional in conjunction with the 6-311G++ and LanL2DZ basis sets to explore the molecular properties and geometries of Vitamin (Vit) B13 and its complexes with Uranyl (II), Vanadyl (II), and Zirconyl (II). The optimized structures, molecular electrostatic potential maps, key molecular properties, and HOMO-LUMO energy gaps of these compounds were systematically examined to gain insights into their electronic characteristics and stability. Bioactivity screenings of the synthesized complexes were performed using molecular docking studies to investigate their interactions with diabetes mellitus (DM) target proteins, specifically the Insulin Receptor (IR) (PDB ID: 1IR3) and the peroxisome proliferator-activated receptor-γ (PPARγ) (PDB ID: 3K8S). These receptors play pivotal roles in the PPAR and AMP-activated protein kinase (AMPK) signaling pathways, which are essential for protecting against nonalcoholic fatty liver disease (NAFLD). The docking results highlighted the binding affinities and potential bioactivity of Vit B13 and its metal complexes, suggesting their promise as therapeutic agents against DM and NAFLD. This comprehensive computational study provides valuable insights into the molecular interactions and electronic properties of these compounds, paving the way for further experimental validation and potential pharmaceutical applications. KEY WORDS: Vitamin B13, Metal complexes, Diabetes mellitus, Molecular docking, DFT Bull. Chem. Soc. Ethiop. 2024, 38(6), 1743-1757. DOI: https://dx.doi.org/10.4314/bcse.v38i6.19

Activation of Follicle-Stimulating Hormone Receptor in Adrenal Zona Fasciculata Cells Promotes Cortisol Secretion: Implications for the Development of Menopause-Associated Diseases.

Changes in postmenopausal hormone levels are associated with a variety of disorders. This study elucidated the mechanism by which follicle-stimulating hormone (FSH) increases cortisol production involved in development of menopause-related diseases. The expression of FSH receptors (FSHRs) in murine adrenal zona fasciculata (AZF) cells and ATC7 cells was verified by immunofluorescence, western blotting and RT-PCR. The function of FSHR in promoting cortisol production was analyzed by cell culture and molecular biological methods. FSHR signaling pathways in ATC7 cells were analyzed by ELISA, qRT-PCR, and western blotting. Further, a mouse model was established by ovariectomy. Ovariectomized mice were treated with GnRHa. Ovariectomized mice initially received physiological doses of estrogen and were then injected with recombinant FSH. Then serum FSH, luteinizing hormone (LH), estradiol, and cortisol, and bone mineral density (BMD), blood pressure (BP) and heart rate (HR) were determined. FSHRs were expressed in murine AZF cells and ATC7 cells. FSH accelerated cortisol production through activated protein kinase A (PKA), cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB), protein kinase B (PKB/AKT) and 5' AMP-activated protein kinase (MAPK) signaling pathways by Gsα-coupled FSHRs in ATC7 cells. Serum FSH levels (P<0.001) were elevated in ovariectomized mice with concurrent increases in cortisol (P<0.01), areal BMD (aBMD) (P<0.05), volumetric BMD (vBMD) (P<0.05), systolic BP (SBP) (P<0.05), diastolic BP (DBP) (P<0.05), and HR (P<0.05). However, the administration of GnRHa suppressed the increase in FSH levels and the elevation of cortisol, aBMD, vBMD, SBP, DBP, and HR induced by ovariectomy, even in the presence of normal serum estradiol levels. The study findings indicate that elevated FSH levels stimulate cortisol secretion, through a mechanism related to FSHRs expression in AZF cells.

Structural Optimization and Structure-Activity Relationship of 1H-Pyrazole-4-carboxylic Acid Derivatives as DNA 6mA Demethylase ALKBH1 Inhibitors and Their Antigastric Cancer Activity.

DNA N6-methyladenine (6mA) demethylase ALKBH1 plays an important role in various cellular processes. Dysregulation of ALKBH1 is associated with the development of some cancer types, including gastric cancer, implicating a potential therapeutic target. However, there is still a lack of potent ALKBH1 inhibitors. Herein, we report the discovery of a highly potent ALKBH1 inhibitor, 1H-pyrazole-4-carboxylic acid derivative 29. The structure-activity relationship of this series of compounds was also discussed. Because of the poor cell membrane permeability of 29, we prepared a prodrug of 29 (29E), which showed excellent cellular activities. In gastric cancer cell lines HGC27 and AGS, 29E treatment significantly increased the abundance of 6mA, inhibited cell viability, and upregulated the AMP-activated protein kinase (AMPK) signaling pathway. In addition, the hydrolysis product 29 showed high exposure in mice after administration of 29E. Collectively, this research provides a new potent ALKBH1 inhibitor, which could serve as a lead compound for subsequent drug development.

Farrerol Alleviates Diabetic Cardiomyopathy by Regulating AMPK-Mediated Cardiac Lipid Metabolic Pathways in Type 2 Diabetic Rats.

Diabetic cardiomyopathy (DCM) is a prevalent complication of diabetes mellitus characterized by cardiac dysfunction and myocardial remodeling. Farrerol (FA), an active ingredient in Rhododendron with various pharmacological activities, has an unclear specific role in DCM. Therefore, this study aims to investigate the effects of FA on DCM rats and elucidate its mechanism. The type 2 diabetes mellitus (T2DM) model was induced in adult male Sprague-Dawley rats by administering a high-fat diet for 8 weeks along with STZ injection. Subsequent to successful modeling, FA and the positive drug Dapagliflozin (Dapa) were orally administered via gavage for an additional 8-week period. After administration, the rats' body weight, fasting blood glucose, fasting insulin, and blood lipid profiles were quantified. Cardiac function was assessed through evaluation of cardiac function parameters, histopathological examination and measurement of myocardial enzyme markers were conducted to assess myocardial injury and fibrosis, Oil red O staining was utilized to evaluate myocardial lipid accumulation, wheat germ agglutinin (WGA) staining was used for assessing cardiomyocyte hypertrophy, and Western blot analysis was used to detect the proteins expression level of AMP-activated protein kinase (AMPK) pathway. The rat cardiomyocyte H9c2 were induced with palmitic acid to establish an in vitro cell model of myocardial lipid toxicity. Subsequently, the cells were subjected to treatment with FA and AMPK inhibitor Compound C, followed by assessment of lipid formation and expression levels of proteins related to the AMPK signaling pathway. The findings demonstrated that both FA and Dapa exhibited efficacy in ameliorating diabetic symptoms, cardiac dysfunction, myocardial fibrosis, cardiomyocyte hypertrophy, and lipid accumulation in T2DM rats. Additionally, they were found to enhance AMPK phosphorylation and PPARα expression while down-regulating CD36. Similarly, FA was observed to inhibit lipid formation in H9c2 and activate the AMPK signaling pathway. However, the improved effect of FA on lipotoxic cardiomyocytes induced by palmitic acid was partially reversed by Compound C. Therefore, the activation of the AMPK signaling pathway by FA may enhance cardiac lipid metabolism, thereby improving cardiac dysfunction and myocardial fibrosis in DCM rats.

Ginsenoside Rh4 prevents endothelial dysfunction as a novel AMPK activator.

The AMP-activated protein kinase (AMPK) signalling pathway is a desirable target for various cardiovascular diseases (CVD), while the involvement of AMPK-mediated specific downstream pathways and effective interventions in hyperlipidaemia-induced endothelial dysfunction remain largely unknown. Herein, we aim to identify an effective AMPK activator and to explore its efficacy and mechanism against endothelial dysfunction. Molecular docking technique was adopted to screen for the potent AMPK activator among 11 most common rare ginsenosides. In vivo, poloxamer 407 (P407) was used to induce acute hyperlipidaemia in C57BL/6J mice. In vitro, palmitic acid (PA) was used to induce lipid toxicity in HAEC cells. We discovered the strongest binding of ginsenoside Rh4 to AMPKα1 and confirmed the action of Rh4 on AMPK activation. Rh4 effectively attenuated hyperlipidaemia-related endothelial injury and oxidative stress both in vivo and in vitro and restored cell viability, mitochondrial membrane potential and mitochondrial oxygen consumption rate in HAEC cells. Mechanistically, Rh4 bound to AMPKα1 and simultaneously up-regulated AKT/eNOS-mediated NO release, promoted PGC-1α-mediated mitochondrial biogenesis and inhibited P38 MAPK/NFκB-mediated inflammatory responses in both P407-treated mice and PA-treated HAEC cells. The AMPK inhibitor Compound C treatment completely abrogated the regulation of Rh4 on the above pathways and weakened the lowering effect of Rh4 on endothelial impairment markers, suggesting that the beneficial effects of Rh4 are AMPK dependent. Rh4 may serve as a novel AMPK activator to protect against hyperlipidaemia-induced endothelial dysfunction, providing new insights into the prevention and treatment of endothelial injury-associated CVD.

Jaboticaba (Plinia jaboticaba (Vell.) Berg) polyphenols alleviate skeletal muscle insulin resistance by modulating PI3K/Akt/GLUT-4 and AMPK signaling pathways in diet-induced obese mice

Skeletal muscle responds for most of the insulin-stimulated glucose disposal at postprandial state, impacting glucose homeostasis. Polyphenols were shown to prevent obesity-associated glucose intolerance and peripheral insulin resistance in animal models, but the implication of skeletal muscle to these effects is unclear. We investigated the role of polyphenolic extracts from jaboticaba (Plinia jaboticaba (Vell.) Berg) (PEJ), a Brazilian native species, on skeletal muscle insulin resistance in diet-induced obese mice. PEJ administration was associated with an increase in skeletal muscle protein content of glucose transporter-4 (GLUT-4) and AMP-activated protein kinase (AMPK) phosphorylated at Thr172. PEJ also reduced skeletal muscle mRNA levels of inflammatory genes nuclear factor-ҡB (NF-κB), tumoral necrose factor-α (TNF-α), interleukin 1β (IL-1β), and c-Jun N-terminal kinase (JNK). This study demonstrates that polyphenols from jaboticaba may be a valuable therapeutic agent in the management and prevention of obesity-associated metabolic disorders by reducing skeletal muscle obesity-associated insulin resistance and inflammation.Graphical