AMP-activated Protein Kinase Inhibitor Research Articles

Regulation of AMPK and GAPDH by transglutaminase 2 plays a pivotal role in microvascular leakage in diabetic retinas.

Diabetic retinopathy is the most common microvascular complication caused by chronic hyperglycemia and is a leading cause of blindness; however, the underlying molecular mechanism has not been clearly elucidated. Thus, we investigated whether regulation of AMPactivated protein kinase (AMPK) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by transglutaminase 2 (TGase2) is important for hyperglycemia-induced microvascular leakage in the diabetic retina. In HRECs and diabetic mouse retinas, we found that TGase2, activated by sequential elevation of intracellular Ca2+ and reactive oxygen species (ROS) levels, played an essential role in hyperglycemia-induced vascular leakage. ROS generation and TGsae2 activation were involved in hyperglycemia-induced AMPK dephosphorylation, which resulted in VE-cadherin disassembly and increased fluorescein isothiocyanate-dextran extravasation. Furthermore, high glucose-induced TGase2 activation suppressed GAPDH activity, determined by an on-chip activity assay, through inhibition of AMPK, which induced VE-cadherin disassembly and endothelial permeability in HRECs. Overall, our findings suggest that inhibition of AMPK and GAPDH by TGase2 plays a pivotal role in hyperglycemia-induced microvascular leakage in the retinas of diabetic mice.

Metformin induces ZFP36 by mTORC1 inhibition in cervical cancer-derived cell lines

BackgroundMetformin, a widely prescribed antidiabetic drug, has shown several promising effects for cancer treatment. These effects have been shown to be mediated by dual modulation of the AMPK-mTORC1 axis, where AMPK acts upstream of mTORC1 to decrease its activity. Nevertheless, alternative pathways have been recently discovered suggesting that metformin can act through of different targets regulation.MethodsWe performed a transcriptome screening analysis using HeLa xenograft tumors generated in NOD-SCID mice treated with or without metformin to examine genes regulated by metformin. Western Blot analysis, Immunohistochemical staining, and RT-qPCR were used to confirm alterations in gene expression. The TNMplot and GEPIA2 platform were used for in silico analysis of genes found up-regulated by metformin, in cervical cancer patients. We performed an AMPK knock-down using AMPK-targeted siRNAs and mTOR inhibition with rapamycin to investigate the molecular mechanisms underlying the effect of metformin in cervical cancer cell lines.ResultsWe shown that metformin decreases tumor growth and increased the expression of a group of antitumoral genes involved in DNA-binding transcription activator activity, hormonal response, and Dcp1-Dcp2 mRNA-decapping complex. We demonstrated that ZFP36 could act as a new molecular target increased by metformin. mTORC1 inhibition using rapamycin induces ZFP36 expression, which could suggest that metformin increases ZFP36 expression and requires mTORC1 inhibition for such effect. Surprisingly, in HeLa cells AMPK inhibition did not affect ZFP36 expression, suggesting that additional signal transducers related to suppressing mTORC1 activity, could be involved.ConclusionsThese results highlight the importance of ZFP36 activation in response to metformin treatment involving mTORC1 inhibition.Graphical

Carnosic Acid (CA) Induces a Brown Fat-like Phenotype, Increases Mitochondrial Biogenesis, and Activates AMPK in 3T3-L1 Adipocytes

Adipose tissue plays a crucial role in regulating metabolic homeostasis, and its dysfunction in obesity leads to insulin resistance and type 2 diabetes (T2D). White adipose tissue (WAT) primarily stores energy as lipids, while brown adipose tissue (BAT) regulates thermogenesis by dissipating energy as heat. The process of browning involves the transdifferentiation of WAT into brown-like or beige adipocytes, which exhibit a similar phenotype as BAT. The browning of WAT is an attractive approach against obesity and T2D, and the activation of the energy sensor AMP-activated protein kinase (AMPK) has been shown to play a role in browning. Carnosic acid (CA), a polyphenolic diterpene, found in many plants including rosemary, is reported to possess potent antioxidant, anti-inflammatory, and anti-hyperglycemic properties. The limited evidence available indicates that CA activates AMPK and may have anti-obesity and antidiabetic potential; however, the effects in adipocyte browning remain largely unexplored. This study aimed to examine the effects of CA on the markers of adipocyte browning. The treatment of 3T3L1 adipocytes with CA activated AMPK, reduced lipid accumulation, and increased the expression of browning protein markers (UCP-1, PGC-1α, PRDM16, and TFAM) and mitochondrial biogenesis. The use of compound C, an AMPK inhibitor, significantly attenuated the effects of CA, indicating AMPK involvement. These studies demonstrate that CA can activate AMPK and stimulate the browning of white adipocytes. Future animal and human studies are required to examine the effects of CA in vivo.

CTRP6 alleviates endometrial fibrosis by regulating Smad3 pathway in intrauterine adhesion†.

Intrauterine adhesion (IUA) is manifestations of endometrial fibrosis and excessive extracellular matrix deposition. C1q/tumor necrosis factor-related protein-6 (CTRP6) is a newly identified adiponectin paralog which has been reported to modulate the fibrosis process of several diseases; however, the endometrial fibrosis function of CTRP6 remains unknown. Our study aimed to assess the role of CTRP6 in endometrial fibrosis and further explore the underlying mechanism. Here, we found that the expression of CTRP6 was downregulated in the endometrial tissues of IUA. In vitro experiments demonstrated the reduced level of CTRP6 in facilitated transforming growth factor-β1 (TGF-β1)-induced human endometrial stromal cells (HESCs). In addition, CTRP6 inhibited the expression of α-smooth muscle actin (α-SMA) and collagen I in TGF-β1-treated HESCs. Mechanistically, CTRP6 activated the AMP-activated protein kinase (AMPK) and protein kinase B (AKT) pathway in HESCs, and AMPK inhibitor (AraA) or PI3K inhibitor (LY294002) pretreatment abolished the protective effect of CTRP6 on TGF-β1-induced fibrosis. CTRP6 markedly decreased TGF-β1-induced Smad3 phosphorylation and nuclear translocation, and AMPK or AKT inhibition reversed these effects. Notably, CTRP6-overexpressing treatment alleviated the fibrosis of endometrium in vivo. Therefore, CTRP6 ameliorates endometrial fibrosis, among which AMPK and AKT are essential for the anti-fibrotic effect of CTRP6 via the Smad3 pathway. Taken together, CTRP6 may be a potential therapeutic target for the treatment of intrauterine adhesion.

CTRP6 alleviates endometrial fibrosis by regulating Smad3 pathway in intrauterine adhesion†.

Intrauterine adhesion (IUA) is manifestations of endometrial fibrosis and excessive extracellular matrix deposition. C1q/tumor necrosis factor-related protein-6 (CTRP6) is a newly identified adiponectin paralog which has been reported to modulate the fibrosis process of several diseases; however, the endometrial fibrosis function of CTRP6 remains unknown. Our study aimed to assess the role of CTRP6 in endometrial fibrosis and further explore the underlying mechanism. Here, we found that the expression of CTRP6 was downregulated in the endometrial tissues of IUA. In vitro experiments demonstrated the reduced level of CTRP6 in facilitated transforming growth factor-β1 (TGF-β1)-induced human endometrial stromal cells (HESCs). In addition, CTRP6 inhibited the expression of α-smooth muscle actin (α-SMA) and collagen I in TGF-β1-treated HESCs. Mechanistically, CTRP6 activated the AMP-activated protein kinase (AMPK) and protein kinase B (AKT) pathway in HESCs, and AMPK inhibitor (AraA) or PI3K inhibitor (LY294002) pretreatment abolished the protective effect of CTRP6 on TGF-β1-induced fibrosis. CTRP6 markedly decreased TGF-β1-induced Smad3 phosphorylation and nuclear translocation, and AMPK or AKT inhibition reversed these effects. Notably, CTRP6-overexpressing treatment alleviated the fibrosis of endometrium in vivo. Therefore, CTRP6 ameliorates endometrial fibrosis, among which AMPK and AKT are essential for the anti-fibrotic effect of CTRP6 via the Smad3 pathway. Taken together, CTRP6 may be a potential therapeutic target for the treatment of intrauterine adhesion.

Kidney targeting and modulating macrophage polarization through AMPK signaling: Therapeutic mechanism of berberine in uric acid nephropathy

Uric acid nephropathy (UAN), caused by a common metabolic disorder resulting from hyperuricemia (HUA), has an increasing incidence. Previous studies have shown that berberine (BBR) has clear urate-lowering and anti-inflammatory effects in UAN mice, but its mechanism needs to be further clarified. Therefore, Potassium Oxonate (PO) combined with hypoxanthine (HX) induced UAN mice model and MSU induced THP-1 cells polarization model were adopted to investigate the mechanism of BBR on UAN in terms of tissue distribution and molecular pharmacology. Study unveiled that BBR was first found to bind to red blood cells (RBCs), which were recognized and phagocytosed by monocytes, then recruited by the injured kidney. Subsequently, BBR was enriched and functional in damaged kidney. The results of in vivo experiments revealed that, BBR reduced UA, BUN, CRE levels as well as the release of TNF-α, IL-1β, IL-18 and IL-6, and alleviated renal injury in UAN mice, as consistent with previous studies. Additionally, BBR decreased MCP-1 expression, while diminishing macrophage infiltration and decreasing M1 proportion as determined by RT-qPCR. In vitro experiments, demonstrated that MSU promoted inflammatory polarization of THP-1 cells, while BBR reduced synthesis of inflammatory factors and inhibited MSU-induced inflammatory polarization. These effects of BBR were dependent on AMPK activation along with indirect inhibition of NF-κB signaling pathway mediated. However, the anti-inflammatory and macrophage polarization regulation effects of BBR were completely reversed upon administration of Compound C, an AMPK inhibitor. Therefore, BBR ameliorated kidney injury via regulating macrophage polarization through AMPK, which has therapeutic potential for UAN patients.

Digirseophene A promotes recovery in injured developing cerebellum via AMPK/AKT/GSK3β pathway-mediated neural stem cell proliferation

Neural stem cells (NSCs) exhibit a remarkable capacity for self-renewal and have the potential to differentiate into various neural lineage cells, which makes them pivotal in the management of neurological disorders. Harnessing the inherent potential of endogenous NSCs for enhancing nerve repair and regeneration represents an optimal approach to addressing diseases of the nervous system. In this study, we explored the potential of a novel benzophenone derivative named Digirseophene A (DGA), which was isolated from the endophytic fungus Corydalis tomentella. Previous experiments have extensively identified and characterized DGA, revealing its unique properties. Our findings demonstrate the remarkable capability of DGA to stimulate neural stem cell proliferation, both in vitro and in vivo. Furthermore, we established a model of radiation-induced cerebellar injury to assess the effects of DGA on the distribution of different cell subpopulations within the damaged cerebellum, thereby suggesting its beneficial role in cerebellar repair. In addition, our observations on a primary NSCs model revealed that DGA significantly increased cellular oxygen consumption, indicating increased energy and metabolic demands. By utilizing various pathway inhibitors in combination with DGA, we successfully demonstrated its ability to counteract the suppressive impacts of AMPK and GSK3β inhibitors on NSC proliferation. Collectively, our research results strongly suggest that DGA, as an innovative compound, exerts its role in activating NSCs and promoting injury repair through the regulation of the AMPK/AKT/GSK3β pathway.

Repurposing ezetimibe as a neuroprotective agent in a rotenone-induced Parkinson’s disease model in rats: Role of AMPK/SIRT-1/PGC-1α signaling and autophagy

As a severe neurological disorder, Parkinson’s disease (PD) is distinguished by dopaminergic neuronal degeneration in the substantia nigra (SN), culminating in motor impairments. Several studies have shown that activation of the AMPK/SIRT1/PGC1α pathway contributes to an increase in mitochondrial biogenesis and is a promising candidate for the management of PD. Furthermore, turning on the AMPK/SIRT1/PGC1α pathway causes autophagy activation, which is fundamental for maintaining neuronal homeostasis. Interestingly, ezetimibe is an antihyperlipidemic agent that was recently reported to possess pleiotropic properties in neurology by triggering the phosphorylation and activation of AMPK. Thus, our study aimed to investigate the neuroprotective potential of ezetimibe in rats with rotenone-induced PD by activating AMPK. Adult male Wistar rats received rotenone (1.5 mg/kg, s.c.) every other day for 21 days to induce experimental PD. Rats were treated with ezetimibe (5 mg/kg/day, i.p.) 1 h before rotenone. Ezetimibe ameliorated the motor impairments in open field, rotarod and grip strength tests, restored striatal dopamine and tyrosine hydroxylase in the SN, up-regulated p-AMPK, SIRT1, and PGC1α striatal expression, upsurged the expression of ULK1, beclin1, and LC3II/I, reduced Bax/Bcl2 ratio, and alleviated rotenone-induced histopathological changes in striatum and SN. Our findings also verified the contribution of AMPK activation to the neuroprotective effect of ezetimibe by using the AMPK inhibitor dorsomorphin. Together, this work revealed that ezetimibe exerts a neuroprotective impact in rotenone-induced PD by activating AMPK/SIRT-1/PGC-1α signaling, enhancing autophagy, and attenuating apoptosis. Thus, ezetimibe’s activation of AMPK could hold significant therapeutic promise for PD management.

Astragaloside IV promotes cerebral tissue restoration through activating AMPK- mediated microglia polarization in ischemic stroke rats

Ethnopharmacological relevanceAstragaloside IV (AS), a key active ingredient obtained from Chinese herb Astragalus mongholicus Bunge, exerts potent neuroprotective and anti-inflammatory effects for treating neurodegenerative diseases. However, mechanisms of AS on improvement of ischemic brain tissue repair remain unclear. Aim of the studyThis research aims at using magnetic resonance imaging (MRI) to noninvasively determine whether AS facilitates brain tissue repair, and investigating whether AS exerts brain remodeling through adenosine monophosphate-activated protein kinase (AMPK) metabolic signaling regulating key glycolytic enzymes and energy transporters, thereby impacting microglia polarization. Materials and methodsIschemic stroke model in male Sprague–Dawley rats were induced through permanent occlusion of the middle cerebral artery (MCAO). Infarct volume, the alterations of brain microstructure and nerve fibers reorganization were examined by multi-parametric MRI. The pathological damages of myelinated axons and microglia polarization surrounding infarct tissue were detected using pathological techniques. Furthermore, M1/M2 microglia polarization associated protein, glycolytic rate-limiting enzymes, energy transporters and AMPK/mammalian target of rapamycin (mTOR)/hypoxia inducible factor-1α (HIF-1α) signal were examined both in ischemic stroke rats and BV2 microglia treated with lipopolysaccharide (LPS) + interferon-γ (IFN-γ) by western blotting. ResultsMRI revealed that AS obviously decreased infarct volume, relieved brain microstructure damage and improved nerve fibers reorganization in ischemic stroke rats. Histological tests supported MRI findings. Notably, AS promoted microglia M2 and reduced M1 polarization, induced the AMPK activation accompanied with decreased levels of phosphorylated mTOR and HIF-1α. Moreover, AS suppressed the expression of glycolytic rate-limiting enzymes and energy transporters in ischemic stroke rats and BV2 microglia. In contrast, these beneficial effects were greatly blocked by AMPK inhibitor compound C. ConclusionOverall, these results collectively suggested that AS facilitated tissue remodeling that may be partially through modulating polarization of microglia in AMPK- dependent metabolic pathways after ischemic stroke.

Fisetin ameliorates polycystic ovary syndrome in rats via a mechanistic modulation of AMP-activated protein kinase and SIRT1 molecular pathway.

Fisetin, a polyphenolic flavonoid, exhibits numerous pharmacological activities against metabolic syndromes. The present research aims to explore the therapeutic efficacy of fisetin in experimental polycystic ovary syndrome (PCOS). Female Sprague-Dawley rats were administered mifepristone (20mg/kg/day) to induce PCOS. PCOS rats were treated with fisetin (20mg/kg and 40mg/kg) and further compared with metformin HCl, the conventional drug for PCOS. The mechanism of fisetin was explored using dorsomorphin (an AMPK inhibitor). Then, rats were sacrificed for further analysis of biochemical and histological parameters. PCOS rats exhibited irregular estrous cycles, increased serum testosterone (4.72 ± 0.139ng/ml), estradiol (750.2 ± 16.56pg/ml), LH (30.33 ± 1.563 mIU/ml), HOMA-IR (1.115 ± 0.049), TNF-α (86.59 ± 3.93pg/ml), IL-6 (55.34 ± 4.432pg/ml), and TBARS (3.867 ± 0.193µmol/mg) along with declined progesterone (11.67 ± 1.54ng/ml), FSH (13.33 ± 1.256 mIU/ml), GSH (33.47 ± 1.348µmol/mg) levels, and SOD (2.163 ± 0.298 U/mg) activity as compared to normal control group. Fisetin high dose significantly lowers testosterone (3.014 ± 0.234ng/ml), estradiol (533.7 ± 15.39pg/ml), LH (16.67 ± 1.62 mIU/ml), HOMA-IR (0.339 ± 0.20), TNF-α (46.02 ± 2.66pg/ml), IL-6 (31.77 ± 3.47pg/ml), and TBARS (1.747 ± 0.185µmol/mg) and enhances progesterone (33.17 ± 1.447ng/ml), FSH (27.17 ± 1.42 mIU/ml), GSH (60.35 ± 1.1.102µmol/mg) levels, and SOD (4.513 ± 0.607 U/mg) activity. The histology of ovarian tissues shows a significant increase in cystic follicles in PCOS rats compared with the normal control group. These alterations were attenuated with fisetin treatment. Administration of dorsomorphin with fisetin can reverse the beneficial effects of fisetin in PCOS rats. Altogether, these present findings highlight the potential of fisetin as a promising therapeutic intervention for the management of PCOS by modulating AMPK/SIRT1 signaling in rats.

IL-10 induces activated phenotypes of monocytes observed in virally-suppressed HIV-1-infected individuals

Despite viral suppression by effective combined antiretroviral therapy, HIV-1-infected individuals have an increased risk of non-AIDS-related overall morbidity, which is due to the persistent chronic inflammation exemplified by the activation of monocytes, such as increased CD16high subset, and elevated plasma level of soluble CD163 (sCD163) and soluble CD14 (sCD14). Here, we show that IL-10, which has been recognized as anti-inflammatory, induces these activated phenotypes of monocytes in vitro. IL-10 increased CD16high monocytes, which was due to the upregulation of CD16 mRNA expression and completely canceled by an inhibitor of Stat3. Moreover, IL-10 increased the production of sCD163 and sCD14 by monocytes, which was consistent with the upregulation of cell surface expression of CD163 and CD14, and mRNA expression of CD163. However, unlike the IL-10-indeuced upregulation of CD16, that of CD14 was minimally affected by the Stat3 inhibitor. Furthermore, the IL-10-induced upregulation of CD163 protein and mRNA was partially inhibited by the Stat3 inhibitor, but completely canceled by an inhibitor of AMPK, an upstream kinase of Stat3 and PI3K/Akt/mTORC1 pathways. In this study, we also found that HIV-1 pathogenic protein Nef, which is known to persist in plasma of virally-suppressed individuals, induced IL-10 production in monocyte-derived macrophages. Our results may suggest that IL-10, which is inducible by Nef-activated macrophages, is one of drivers for activated phenotypes of monocytes in virally-suppressed individuals, and that IL-10 induces the increased CD16high monocytes and elevated level of sCD163 and sCD14 through the activation of different signaling pathways.

The role of the AMPK/ERK1/2 signaling pathway in neuronal oxidative stress damage following cerebral ischemia-reperfusion

Cerebral ischemia-reperfusion injury involves a series of pathophysiological processes that occur when blood supply is restored after cerebral vascular obstruction, leading to neuronal damage. The AMPK/ERK1/2 signaling pathway has been identified as crucial in this process, although the exact mechanisms underlying the induction of ischemia-reperfusion injury remain unclear. In this study, we investigated the involvement of the AMPK/ERK1/2 signaling pathway in neuronal oxidative stress damage following cerebral ischemia-reperfusion by establishing animal and cell models. Our experimental results demonstrated that cerebral ischemia-reperfusion leads to oxidative stress damage, including cell apoptosis and mitochondrial dysfunction. Moreover, further experiments showed that inhibition of AMPK and ERK1/2 activity, using U0126 and Compound C respectively, could alleviate oxidative stress-induced cellular injury, improve mitochondrial morphology and function, reduce reactive oxygen species levels, increase superoxide dismutase levels, and suppress apoptosis. These findings clearly indicate the critical role of the AMPK/ERK1/2 signaling pathway in regulating oxidative stress damage and cerebral ischemia-reperfusion injury. The discoveries in this study provide a theoretical basis for further research and development of neuroprotective therapeutic strategies targeting the AMPK/ERK1/2 signaling pathway.

Ginsenoside Rg1 inhibits multiple myeloma and overcomes bortezomib resistance through AMPK-mTOR pathway

BackgroundThe resistance of multiple myeloma (MM) to bortezomib (BTZ) has brought multiple challenges to its clinical use. Numerous ginsenosides have potential anti-tumor effects, however, the research on the role of Rg1 in MM has not been reported. ObjectiveTo examine the inhibitory impact of Rg1 on the growth of MM and reduce the drug resistance of MM to BTZ through in vivo and in vitro experiments, and to explore their potential mechanism. MethodsBTZ drug-resistant cell line RPMI8226R was constructed. Mouse tumor-bearing model was developed by abdominal subcutaneous injection of MM cells. MM cells were treated with AMPK inhibitor Compound C or autophagy inhibitor Chloroquine together with Rg1. RPMI8226R cells were treated with BTZ and Rg1. Cell multiplication was detected using Methylthiazolyldiphenyl-tetrazolium bromide assay. Apoptosis was assessed using flow cytometry. Immunofluorescence assay was employed to assess the autophagy markers LC3. Western blot was utilized to assess the protein expression. Immunohistochemistry was used to detect cell proliferation and apoptosis in tumor tissues. ResultsIn vitro experiments demonstrated that Rg1 could hinder the proliferation of MM cells, promote apoptosis and enhance autophagy. Rg1 could also increase the sensitivity of RPMI8226R to BTZ. In vivo experiments illustrated that Rg1 could hinder the development of MM cells in mice, weaken the proliferation of tumor cells and enhance their apoptosis. Further study found that the anti-MM impact of Rg1 was linked to AMPK-mTOR pathway, the autophagy degree of RPMI8226R was higher than that of RPMI8226, and that Rg1 could inhibit MM and overcome drug resistance through autophagy induced by AMPK-mTOR pathway. ConclusionRg1 has significant anti-MM effect and can overcome BTZ resistance, and its potential mechanism is related to the regulation of autophagy induced by AMPK-mTOR pathway. Rg1 is a promising adjuvant drug for the treatment of MM.

Reprogramming of Energy Metabolism in Human PKD1 Polycystic Kidney Disease: A Systems Biology Analysis.

Multiple alterations of cellular metabolism have been documented in experimental studies of autosomal dominant polycystic kidney disease (ADPKD) and are thought to contribute to its pathogenesis. To elucidate the molecular pathways and transcriptional regulators associated with the metabolic changes of renal cysts in ADPKD, we compared global gene expression data from human PKD1 renal cysts, minimally cystic tissues (MCT) from the same patients, and healthy human kidney cortical tissue samples. We found gene expression profiles of PKD1 renal cysts were consistent with the Warburg effect with gene pathway changes favoring increased cellular glucose uptake and lactate production, instead of pyruvate oxidation. Additionally, mitochondrial energy metabolism was globally depressed, associated with downregulation of gene pathways related to fatty acid oxidation (FAO), branched-chain amino acid (BCAA) degradation, the Krebs cycle, and oxidative phosphorylation (OXPHOS) in renal cysts. Activation of mTORC1 and its two target proto-oncogenes, HIF-1α and MYC, was predicted to drive the expression of multiple genes involved in the observed metabolic reprogramming (e.g., GLUT3, HK1/HK2, ALDOA, ENO2, PKM, LDHA/LDHB, MCT4, PDHA1, PDK1/3, MPC1/2, CPT2, BCAT1, NAMPT); indeed, their predicted expression patterns were confirmed by our data. Conversely, we found AMPK inhibition was predicted in renal cysts. AMPK inhibition was associated with decreased expression of PGC-1α, a transcriptional coactivator for transcription factors PPARα, ERRα, and ERRγ, all of which play a critical role in regulating oxidative metabolism and mitochondrial biogenesis. These data provide a comprehensive map of metabolic pathway reprogramming in ADPKD and highlight nodes of regulation that may serve as targets for therapeutic intervention.

Ecklonia cava Polyphenols Have a Preventive Effect on Parkinson's Disease through the Activation of the Nrf2-ARE Pathway.

Parkinson's disease (PD) is a degenerative neurological disorder defined by the deterioration and loss of dopamine-producing neurons in the substantia nigra, leading to a range of motor impairments and non-motor symptoms. The underlying mechanism of this neurodegeneration remains unclear. This research examined the neuroprotective properties of Ecklonia cava polyphenols (ECPs) in mitigating neuronal damage induced by rotenone via the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway. Using human neuroblastoma SH-SY5Y cells and PD model mice, we found that ECP, rich in the antioxidant polyphenol phlorotannin, boosted the gene expression and functionality of the antioxidant enzyme NAD(P)H quinone oxidoreductase-1. ECP also promoted Nrf2 nuclear translocation and increased p62 expression, suggesting that p62 helps sustain Nrf2 activation via a positive feedback loop. The neuroprotective effect of ECP was significantly reduced by Compound C (CC), an AMP-activated protein kinase (AMPK) inhibitor, which also suppressed Nrf2 nuclear translocation. In PD model mice, ECPs improved motor functions impaired by rotenone, as assessed by the pole test and wire-hanging test, and restored intestinal motor function and colon tissue morphology. Additionally, ECPs increased tyrosine hydroxylase expression in the substantia nigra, indicating a protective effect on dopaminergic neurons. These findings suggest that ECP has a preventative effect on PD.

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.

AMPK-mediated autophagy pathway activation promotes ΔFosB degradation to improve levodopa-induced dyskinesia

BackgroundParkinson's disease patients on chronic levodopa often suffer from motor complications, which tend to reduce their quality of life. Levodopa-induced dyskinesia (LID) is one of the most prevalent motor complications, often characterized by abnormal involuntary movements, and the pathogenesis of LID is still unclear but recent studies have suggested the involvement of autophagy. MethodsThe onset of LID was mimicked by chronic levodopa treatment in a unilateral 6-hydroxydopamine (6-OHDA) -lesion rat model. Overexpression of ΔFosB in HEK293 cells to mimic the state of ΔFosB accumulation. The modulation of the AMP-activated protein kinase (AMPK)-mediated autophagy pathway using by metformin, AICAR (an AMPK activator), Compound C (an AMPK inhibitor) and chloroquine (an autophagy pathway inhibitor). The severity of LID was assessed by axial, limb, and orofacial (ALO) abnormal involuntary movements (AIMs) score and in vivo electrophysiology. The activity of AMPK pathway as well as autophagy markers and FosB-ΔFosB levels were detected by western blotting. RT-qPCR was performed to detect the transcription level of FosB-ΔFosB. The mechanism of autophagy dysfunction was further explored by immunofluorescence and transmission electron microscopy. ResultsIn vivo experiments demonstrated that chronic levodopa treatment reduced AMPK phosphorylation, impaired autophagosome-lysosomal fusion and caused FosB-ΔFosB accumulation in the striatum of PD rats. Long-term metformin intervention improved ALO AIMs scores as well as reduced the mean power of high gamma (hγ) oscillations and the proportion of striatal projection neurons unstable in response to dopamine for LID rats. Moreover, the intervention of metformin promoted AMPK phosphorylation, ameliorated the impairment of autophagosome-lysosomal fusion, thus, promoting FosB-ΔFosB degradation to attenuate its accumulation in the striatum of LID rats. However, the aforementioned roles of metformin were reversed by Compound C and chloroquine. The results of in vitro studies demonstrated the ability of metformin and AICAR to attenuate ΔFosB levels by promoting its degradation, while Compound C and chloroquine could block this effect. ConclusionsIn conclusion, our results suggest that long-term metformin treatment could promote ΔFosB degradation and thus attenuate the development of LID through activating the AMPK-mediated autophagy pathway. Overall, our results support the AMPK-mediated autophagy pathway as a novel therapeutic target for LID and also indicate that metformin is a promising therapeutic candidate for LID.

Naringenin enhances the efficacy of ferroptosis inducers by attenuating aerobic glycolysis by activating the AMPK-PGC1α signalling axis in liver cancer

Liver cancer is a heterogeneous disease characterized by poor responses to standard therapies and therefore unfavourable clinical outcomes. Understanding the characteristics of liver cancer and developing novel therapeutic strategies are imperative. Ferroptosis, a type of programmed cell death induced by lipid peroxidation, has emerged as a potential target for treatment. Naringenin, a natural compound that modulates lipid metabolism by targeting AMPK, shows promise in enhancing the efficacy of ferroptosis inducers. In this study, we utilized liver cancer cell lines and xenograft mice to explore the synergistic effects of naringenin in combination with ferroptosis inducers, examining both phenotypic outcomes and molecular mechanisms. Our study results indicate that the use of naringenin at non-toxic doses to hepatocytes can significantly enhance the anticancer effects of ferroptosis inducers (erastin, RSL3, and sorafenib). The combination index method confirmed a synergistic effect between naringenin and ferroptosis inducers. In comparison to naringenin or ferroptosis inducers alone, the combined therapy caused more robust lipid peroxidation and hence more severe ferroptotic damage to cancer cells. The inhibition of aerobic glycolysis mediated by the AMPK-PGC1α signalling axis is the key to naringenin's effect on reducing ferroptosis resistance in liver cancer, and the synergistic cytotoxic effect of naringenin and ferroptosis inducers on cancer cells was reversed after pretreatment with an AMPK inhibitor or a PGC1α inhibitor. Taken together, these findings suggest that naringenin could boost cancer cell sensitivity to ferroptosis inducers, which has potential clinical translational value.

AMPK Drives both Glycolytic and Oxidative Metabolism in Murine and Human T Cells During Graft-versus-host Disease

Allogeneic T cells reprogram their metabolism during acute graft-versus-host disease (GVHD) in a process involving the cellular energy sensor AMP-activated protein kinase (AMPK). Deletion of AMPK in donor T cells limits GVHD but still preserves homeostatic reconstitution and graft-versus-leukemia (GVL) effects. In the current studies, murine AMPK KO T cells decreased oxidative metabolism at early timepoints post-transplant and lacked a compensatory increase in glycolysis following inhibition of the electron transport chain. Immunoprecipitation using an antibody specific to phosphorylated targets of AMPK determined that AMPK modified interactions of several glycolytic enzymes including aldolase, enolase, pyruvate kinase M (PKM), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and enzyme assays indicated impaired aldolase and GAPDH activity in AMPK KO T cells. Importantly, these changes in glycolysis correlated with both an impaired ability of AMPK KO T cells to produce significant amounts of interferon gamma (IFNγ) upon antigenic re-stimulation and a decrease in the total number of donor CD4 T cells recovered at later time points post-transplant. Human T cells lacking AMPK gave similar results, with glycolytic compensation impaired both in vitro and following expansion in vivo. GVHD results also mirrored those of the murine model, with reduced CD4/CD8 ratios and a significant improvement in disease severity. Together these data highlight a significant role for AMPK in controlling oxidative and glycolytic metabolism in both murine and human T cells and endorse further study of AMPK inhibition as a potential clinical target for future GVHD therapies.

GCTRP3 inhibits oophorectomy‑induced osteoporosis by activating the AMPK/SIRT1/Nrf2 signaling pathway in mice.

C1q/tumor necrosis factor‑related protein 3 (CTRP3) expression is markedly reduced in the serum of patients with osteoporosis. The present study aimed to investigate whether CTRP3 reduces bone loss in oophorectomy (OVX)‑induced mice via the AMP‑activated protein kinase (AMPK)/sirtuin 1 (SIRT1)/nuclear factor E2‑related factor 2 (Nrf2) signaling pathway. Female C57BL/6J mice and MC3T3‑E1 cells were used to construct in vivo and in vitro models of osteoporosis, respectively. The left femurs of mice were examined using micro‑computed tomography scans and bone‑related quantitative morphological evaluation was performed. Pathological changes and the number of osteoclasts in the left femurs of mice were detected using hematoxylin and eosin, and tartrate‑resistant acid phosphatase (TRAP) staining. Runt‑related transcription factor‑2 (RUNX2) expression in the left femurs was detected using immunofluorescence analysis, and the serum levels of bone resorption markers (C‑telopeptide of type I collagen and TRAP) and bone formation markers [osteocalcin (OCN) and procollagen type 1 N‑terminal propeptide] were detected. In addition, osteoblast differentiation and calcium deposits were examined in MC3T3‑E1 cells using alkaline phosphatase (ALP) and Alizarin red staining, respectively. Moreover, RUNX2, ALP and OCN expression levels were detected using reverse transcription‑quantitative PCR, and the expression levels of proteins associated with the AMPK/SIRT1/Nrf2 signaling pathway were detected using western blot analysis. The results revealed that globular CTRP3 (gCTRP3) alleviated bone loss and promoted bone formation in OVX‑induced mice. gCTRP3 also facilitated the osteogenic differentiation of MC3T3‑E1 cells through the AMPK/SIRT1/Nrf2 signaling pathway. The addition of an AMPK inhibitor (Compound C), SIRT1 inhibitor (EX527) or Nrf2 inhibitor (ML385) reduced the osteogenic differentiation of MC3T3‑E1 cells via inhibition of gCTRP3. In conclusion, gCTRP3 inhibits OVX‑induced osteoporosis by activating the AMPK/SIRT1/Nrf2 signaling pathway.