Melosira nummuloides Ethanol Extract Ameliorates Alcohol-Induced Liver Injury by Affecting Metabolic Pathways.

Melosira nummuloides is a microalga with a nutritionally favorable polyunsaturated fatty acid profile. In the present study, M. nummuloides ethanol extract (MNE) was administered to chronic-binge alcohol-fed mice and alcohol-treated HepG2 cells, and its hepatoprotective effects and underlying mechanisms were investigated. MNE administration reduced triglyceride (TG), total cholesterol (T-CHO), and liver injury markers, including aspartate transaminase (AST) and alanine transaminase (ALT), in the serum of chronic-binge alcohol-fed mice. However, MNE administration increased the levels of phosphorylated adenosine monophosphate-activated protein kinase (P-AMPK/AMPK) and PPARα, which was accompanied by a decrease in SREBP-1; this indicates that MNE can inhibit adipogenesis and improve fatty acid oxidation. Moreover, MNE administration upregulated the expression of antioxidant enzymes, including SOD, NAD(P)H quinone dehydrogenase 1, and GPX, and ameliorated alcohol-induced inflammation by repressing the Akt/NFκB/COX-2 pathway. Metabolomic analysis revealed that MNE treatment modulated many lipid metabolites in alcohol-treated HepG2 cells. Our study findings provide evidence for the efficacy and mechanisms of MNE in ameliorating alcohol-induced liver injury.

Loss of AMPK activity induces organelle dysfunction and oxidative stress during oocyte aging.

BACKGROUND: Oocyte quality is critical for the mammalian reproduction due to its necessity on fertilization and early development. During aging, the declined oocytes showing with organelle dysfunction and oxidative stress lead to infertility. AMP-activated protein kinase (AMPK) is a serine/threonine protein kinase which is important for energy homeostasis for metabolism. Little is known about the potential relationship between AMPK with oocyte aging. RESULTS: In present study we reported that AMPK was related with low quality of oocytes under post ovulatory aging and the potential mechanism. We showed the altered AMPK level during aging and inhibition of AMPK activity induced mouse oocyte maturation defect. Further analysis indicated that similar with its upstream regulator PKD1, AMPK could reduce ROS level to avoid oxidative stress in oocytes, and this might be due to its regulation on mitochondria function, since loss of AMPK activity induced abnormal distribution, reduced ATP production and mtDNA copy number of mitochondria. Besides, we also found that the ER and Golgi apparatus distribution was aberrant after AMPK inhibition, and enhanced lysosome function was also observed. CONCLUSIONS: Taken together, these data indicated that AMPK is important for the organelle function to reduce oxidative stress during oocyte meiotic maturation.

Molecular Mechanisms of Fucoxanthin in Alleviating Lipid Deposition in Metabolic Associated Fatty Liver Disease.

Metabolic-associated fatty liver disease (MAFLD) is witnessing a global surge; however, it still lacks effective pharmacological interventions. Fucoxanthin, a natural bioactive metabolite derived from marine brown algae, exhibits promising pharmacological functions, particularly in ameliorating metabolic disorders. However, the mechanisms underlying its therapeutic efficacy in addressing MAFLD remain elusive. Our present findings indicated that fucoxanthin significantly alleviated palmitic acid (PA)-induced hepatic lipid deposition in vitro and obesity-induced hepatic steatosis in ob/ob mice. Moreover, at both the protein and transcriptional levels, fucoxanthin effectively increased the expression of PPARα and CPT1 (involved in fatty acid oxidation) and suppressed FASN and SREBP1c (associated with lipogenesis) in both PA-induced HepG2 cells and hepatic tissues in ob/ob mice. This modulation was accompanied by the activation of AMPK. The capacity of fucoxanthin to improve hepatic lipid deposition was significantly attenuated when utilizing the AMPK inhibitor or siRNA-mediated AMPK silencing. Mechanistically, fucoxanthin activates AMPK, subsequently regulating the KEAP1/Nrf2/ARE signaling pathway to exert antioxidative effects and stimulating the PGC1α/NRF1 axis to enhance mitochondrial biogenesis. These collective actions contribute to fucoxanthin's amelioration of hepatic steatosis induced by metabolic perturbations. These findings offer valuable insights into the prospective utilization of fucoxanthin as a therapeutic strategy for managing MAFLD.

Platycodin D Ameliorates Type 2 Diabetes-Induced Myocardial Injury by Activating the AMPK Signaling Pathway.

The current study aimed to assess the effectiveness of pharmacological intervention with Platycodin D (PD), a critically active compound isolated from the roots of Platycodon grandiflorum, in mitigating cardiotoxicity in a murine model of type 2 diabetes-induced cardiac injury and in H9c2 cells in vitro. Following oral administration for 4 weeks, PD (2.5 mg/kg) significantly suppressed the elevation of fasting blood glucose (FBG) levels, improved dyslipidemia, and effectively inhibited the rise of the cardiac injury markers creatine kinase isoenzyme MB (CK-MB) and cardiac troponin T (cTnT). PD treatment could ameliorate energy metabolism disorders induced by impaired glucose uptake by activating AMPK protein expression in the DCM mouse model, thereby promoting the GLUT4 transporter and further activating autophagy-related proteins. Furthermore, in vitro experiments demonstrated that PD exerted a concentration-dependent increase in cell viability while also inhibiting palmitic acid and glucose (HG-PA)-stimulated H9c2 cytotoxicity and activating AMPK protein expression. Notably, the AMPK activator AICAR (1 mM) was observed to upregulate the expression of AMPK in H9c2 cells after high-glucose and -fat exposure. Meanwhile, we used AMPK inhibitor Compound C (20 µM) to investigate the effect of PD activation of AMPK on cells. In addition, the molecular docking approach was employed to dock PD with AMPK, revealing a binding energy of -8.2 kcal/mol and indicating a tight interaction between the components and the target. PD could reduce the expression of autophagy-related protein p62, reduce the accumulation of autophagy products, promote the flow of autophagy, and improve myocardial cell injury. In conclusion, it has been demonstrated that PD effectively inhibits cardiac injury-induced type 2 diabetes in mice and enhances energy metabolism in HG-PA-stimulated H9c2 cells by activating the AMPK signaling pathway. These findings collectively unveil the potential cardioprotective effects of PD via modulation of the AMPK signaling pathway.

NEAT1 inhibits the angiogenic activity of cerebral arterial endothelial cells by inducing the M1 polarization of microglia through the AMPK signaling pathway.

BACKGROUND: Enhancing angiogenesis may be an effective strategy to promote functional recovery after ischemic stroke. Inflammation regulates angiogenesis. Microglia are crucial cells that initiate inflammatory responses after various brain injuries. Long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) plays a role in regulating brain injury. This study aimed to explore the effects of NEAT1-regulated microglial polarization on the neovascularization capacity of cerebrovascular endothelial cells and the underlying molecular regulatory mechanisms. METHODS: Mouse cerebral arterial endothelial cells (mCAECs) were co-cultured with BV-2 cells in different groups using a Transwell system. NEAT1 expression levels were measured by fluorescence quantitative reverse transcription PCR. Levels of IL-1ß, IL-6, TNF-α, Arg-1, IL-4, and IL-10 were determined using ELISA. Expression levels of CD86 and CD163 were detected by immunofluorescence. The neovascularization capacity of mCAECs was assessed using CCK-8, Transwell, Transwell-matrigel, and tube formation assays. Label-free quantification proteomics was carried out to identify differentially expressed proteins. Protein levels were measured by Western blotting. RESULTS: NEAT1 overexpression induced M1 polarization in BV-2 cells, whereas NEAT1 knockdown blocked lipopolysaccharide-induced M1 polarization in microglia. NEAT1-overexpressing BV-2 cells suppressed the angiogenic ability of mCAECs, and NEAT1-knocking BV-2 cells promoted the angiogenic ability of mCAECs under lipopolysaccharide treatment. Label-free quantitative proteomic analysis identified 144 upregulated and 131 downregulated proteins that were induced by NEAT1 overexpression. The AMP-activated protein kinase (AMPK) signaling pathway was enriched in the Kyoto Encyclopedia of Genes and Genomes analysis of the differentially expressed proteins. Further verification showed that NEAT1 inactivated the AMPK signaling pathway. Moreover, the AMPK activator 5-aminoimidazole-4-carboxamide ribonucleotide reversed the effect of NEAT1 on BV-2 polarization and the regulatory effect of NEAT1-overexpressing BV-2 cells on the angiogenic ability of mCAECs. CONCLUSIONS: NEAT1 inhibits the angiogenic activity of mCAECs by inducing M1 polarization of BV-2 cells through the AMPK signaling pathway. This study further clarified the impact and mechanism of NEAT1 on microglia and the angiogenic ability of cerebrovascular endothelial cells.

Fus3 Interacts with Gal83, Revealing the MAPK Crosstalk to Snf1/AMPK to Regulate Secondary Metabolic Substrates in Aspergillus flavus.

Aflatoxins (AFs), highly carcinogenic natural products, are produced by the secondary metabolism of fungi such as Aspergillus flavus. Essential for the fungi to respond to environmental changes and aflatoxin synthesis, the pheromone mitogen-activated protein kinase (MAPK) is a potential regulator of aflatoxin biosynthesis. However, the mechanism by which pheromone MAPK regulates aflatoxin biosynthesis is not clear. Here, we showed Gal83, a new target of Fus3, and identified the pheromone Fus3-MAPK signaling pathway as a regulator of the Snf1/AMPK energy-sensing pathway modulating aflatoxins synthesis substrates. The screening for Fus3 target proteins identified the ß subunit of Snf1/AMPK complexes using tandem affinity purification and multiomics. This subunit physically interacted with Fus3 both in vivo and in vitro and received phosphorylation from Fus3. Although the transcript levels of aflatoxin synthesis genes were not noticeably downregulated in both gal83 and fus3 deletion mutant strains, the levels of aflatoxin B1 and its synthesis substrates and gene expression levels of primary metabolizing enzymes were significantly reduced. This suggests that both the Fus3-MAPK and Snf1/AMPK pathways respond to energy signals. In conclusion, all the evidence unlocks a novel pathway of Fus3-MAPK to regulate AFs synthesis substrates by cross-talking with the Snf1/AMPK complexes.

AMP-activated protein kinase can be allosterically activated by ADP but AMP remains the key activating ligand.

The AMP-activated protein kinase (AMPK) is a sensor of cellular energy status. When activated by increases in ADP:ATP and/or AMP:ATP ratios (signalling energy deficit), AMPK acts to restore energy balance. Binding of AMP to one or more of three CBS repeats (CBS1, CBS3, CBS4) on the AMPK-γ subunit activates the kinase complex by three complementary mechanisms: (i) promoting α-subunit Thr172 phosphorylation by the upstream kinase LKB1; (ii) protecting against Thr172 dephosphorylation; (iii) allosteric activation. Surprisingly, binding of ADP has been reported to mimic the first two effects, but not the third. We now show that at physiologically relevant concentrations of Mg.ATP2- (above those used in the standard assay) ADP binding does cause allosteric activation. However, ADP causes only a modest activation because (unlike AMP), at concentrations just above those where activation becomes evident, ADP starts to cause competitive inhibition at the catalytic site. Our results cast doubt on the physiological relevance of the effects of ADP and suggest that AMP is the primary activator in vivo. We have also made mutations to hydrophobic residues involved in binding adenine nucleotides at each of the three γ subunit CBS repeats of the human α2ß2γ1 complex and examined their effects on regulation by AMP and ADP. Mutation of the CBS3 site has the largest effects on all three mechanisms of AMP activation, especially at lower ATP concentrations, while mutation of CBS4 reduces the sensitivity to AMP. All three sites appear to be required for allosteric activation by ADP.

Lysine specific demethylase 1 inhibits sodium arsenite activation of HSCs by regulating SESN2/AMPK/ULK1 signaling pathway activity.

Lysine specific demethylase 1 (LSD1) is a histone demethylase that specifically catalyzes the demethylation of histone H3K4 (H3K4me1/2) and regulates gene expression. In addition, it can mediate the process of autophagy through its demethylase activity. Sestrin2 (SESN2) is a stress-induced protein and a positive regulator of autophagy. In NaAsO2-induced mouse fibrotic livers and activated hepatic stellate cells (HSCs), LSD1 expression is decreased, SESN2 expression is increased, and autophagy levels are also increased. Overexpression of LSD1 and silencing of SESN2 decreased the level of autophagy and attenuated the activation of HSCs induced by NaAsO2. LSD1 promoted SESN2 gene transcription by increasing H3K4me1/2 in the SESN2 promoter region. 3-methyladenine (3-MA) and chloroquine were used to inhibit autophagy of HSCs, and the degree of activation was also alleviated. Taken together, LSD1 positively regulates SESN2 by increasing H3K4me1/2 enrichment in the SESN2 promoter region, which in turn increases the level of autophagy and promotes the activation of HSCs. Our results may provide new evidence for the importance of LSD1 in the process of autophagy and activation of HSCs induced by arsenic poisoning. Increasing the expression and activity of LSD1 is expected to be an effective way to reverse the autophagy and activation of HSCs induced by arsenic poisoning.

Network visualization of genes involved in skeletal muscle myogenesis in livestock animals.

BACKGROUND: Muscle growth post-birth relies on muscle fiber number and size. Myofibre number, metabolic and contractile capacities are established pre-birth during prenatal myogenesis. The aim of this study was to identify genes involved in skeletal muscle development in cattle, sheep, and pigs - livestock. RESULTS: The cattle analysis showed significant differences in 5043 genes during the 135-280 dpc period. In sheep, 444 genes differed significantly during the 70-120 dpc period. Pigs had 905 significantly different genes for the 63-91 dpc period.The biological processes and KEGG pathway enrichment results in each species individually indicated that DEGs in cattle were significantly enriched in regulation of cell proliferation, cell division, focal adhesion, ECM-receptor interaction, and signaling pathways (PI3K-Akt, PPAR, MAPK, AMPK, Ras, Rap1); in sheep - positive regulation of fibroblast proliferation, negative regulation of endothelial cell proliferation, focal adhesion, ECM-receptor interaction, insulin resistance, and signaling pathways (PI3K-Akt, HIF-1, prolactin, Rap1, PPAR); in pigs - regulation of striated muscle tissue development, collagen fibril organization, positive regulation of insulin secretion, focal adhesion, ECM-receptor interaction, and signaling pathways (PPAR, FoxO, HIF-1, AMPK). Among the DEGs common for studied animal species, 45 common genes were identified. Based on these, a protein-protein interaction network was created and three significant modules critical for skeletal muscle myogenesis were found, with the most significant module A containing four recognized hub genes - EGFR, VEGFA, CDH1, and CAV1. Using the miRWALK and TF2DNA databases, miRNAs (bta-miR-2374 and bta-miR-744) and transcription factors (CEBPB, KLF15, RELA, ZNF143, ZBTB48, and REST) associated with hub genes were detected. Analysis of GO term and KEGG pathways showed that such processes are related to myogenesis and associated with module A: positive regulation of MAP kinase activity, vascular endothelial growth factor receptor, insulin-like growth factor binding, focal adhesion, and signaling pathways (PI3K-Akt, HIF-1, Rap1, Ras, MAPK). CONCLUSIONS: The identified genes, common to the prenatal developmental period of skeletal muscle in livestock, are critical for later muscle development, including its growth by hypertrophy. They regulate valuable economic characteristics. Enhancing and breeding animals according to the recognized genes seems essential for breeders to achieve superior gains in high-quality muscle mass.

Floralozone regulates MiR-7a-5p expression through AMPKα2 activation to improve cognitive dysfunction in vascular dementia.

BACKGROUND: The pathogenesis of vascular dementia (VD) is complex, and currently, no effective treatments have been recommended. Floralozone is a colorless liquid first discovered in Lagotis Gaertn. Recently, its medicinal value has been increasingly recognized. Our previous study has demonstrated that Floralozone can improve cognitive dysfunction in rats with VD by regulating the transient receptor potential melastatin 2 (TRPM2) and N-methyl-D-aspartate receptor (NMDAR) signaling pathways. However, the mechanism by which Floralozone regulates TRPM2 and NMDAR to improve VD remains unclear. AMP-activated protein kinase (AMPK) is an energy regulator in vivo; however, its role of AMPK activation in stroke remains controversial. MiR-7a-5p has been identified to be closely related to neuronal function. PURPOSE: To explore whether Floralozone can regulate the miR-7a-5p level in vivo through AMPKα2 activation, affect the TRPM2 and NR2B expression levels, and improve VD symptoms. METHODS: The VD model was established by a modified bilateral occlusion of the common carotid arteries (2-VO) of Sprague-Dawley (SD) rats and AMPKα2 KO transgenic (AMPKα2-/-) mice. Primary hippocampal neurons were modeled using oxygen and glucose deprivation (OGD). Morris water maze (MWM) test, hematoxylin-eosin staining (HE staining), and TUNEL staining were used to investigate the effects of Floralozone on behavior and hippocampal morphology in rats. Minichromosome maintenance complex component 2(MCM2) positive cells were used to investigate the effect of Floralozone on neurogenesis. Immunofluorescence staining, qRT-PCR, and western blot analysis were used to investigate the effect of Floralozone on the expression levels of AMPKα2, miR-7a-5p, TRPM2, and NR2B. RESULTS: The SD rat experiment revealed that Floralozone improved spatial learning and memory, improved the morphology and structure of hippocampal neurons, reduced apoptosis of hippocampal neurons and promoted neurogenesis in VD rats. Floralozone could increase the miR-7a-5p expression level, activate AMPKα2 and NR2B expressions, and inhibit TRPM2 expression in hippocampal neurons of VD rats. The AMPKα2 KO transgenic (AMPKα2-/-) mice experiment demonstrated that Floralozone could regulate miR-7a-5p, TRPM2, and NR2B expression levels through AMPKα2 activation. The cell experiment revealed that the TRPM2 and NR2B expression levels were regulated by miR-7a-5p, whereas the AMPKα2 expression level was not. CONCLUSION: Floralozone could regulate miR-7a-5p expression level by activating the protein expression of AMPKα2, control the protein expression of TRPM2 and NR2B, improve the morphology and structure of hippocampus neurons, reduce the apoptosis of hippocampus neurons, promote neurogenesis and improve the cognitive dysfunction.

The AMPK-related kinase NUAK1 controls cortical axons branching by locally modulating mitochondrial metabolic functions.

The cellular mechanisms underlying axonal morphogenesis are essential to the formation of functional neuronal networks. We previously identified the autism-linked kinase NUAK1 as a central regulator of axon branching through the control of mitochondria trafficking. However, (1) the relationship between mitochondrial position, function and axon branching and (2) the downstream effectors whereby NUAK1 regulates axon branching remain unknown. Here, we report that mitochondria recruitment to synaptic boutons supports collateral branches stabilization rather than formation in mouse cortical neurons. NUAK1 deficiency significantly impairs mitochondrial metabolism and axonal ATP concentration, and upregulation of mitochondrial function is sufficient to rescue axonal branching in NUAK1 null neurons in vitro and in vivo. Finally, we found that NUAK1 regulates axon branching through the mitochondria-targeted microprotein BRAWNIN. Our results demonstrate that NUAK1 exerts a dual function during axon branching through its ability to control mitochondrial distribution and metabolic activity.

Aflatoxin B1-Induced Testosterone Biosynthesis Disorder via the ROS/AMPK Signaling Pathway in Male Mice.

The worldwide prevalence of Aflatoxin B1 (AFB1), which contaminates feedstock and food, is on the rise. AFB1 inhibits testosterone (T) biosynthesis, but the mechanism is not yet clear. By establishing in vivo and in vitro models, this study found the number of Leydig cells (LCs), T content, and the expression of T biosynthesis key enzymes were suppressed after AFB1 treatment. AFB1 exposure also increased reactive oxygen species (ROS) and promoted mitochondrial injury and mitochondrial pathway apoptosis. Moreover, the AMPK signaling pathway was activated, and using an AMPK inhibitor relieved apoptosis and the suppressed T biosynthesis key enzymes of LCs caused by AFB1 through regulating downstream p53 and Nur77. Additionally, adding ROS intervention could inhibit AMPK activation and alleviate the decreased T content caused by AFB1. In summary, AFB1 promotes the apoptosis of LCs and inhibits T biosynthesis key enzyme expression via activating the ROS/AMPK signaling pathway, which eventually leads to T synthesis disorder.

Sirt3 Protects Retinal Pigment Epithelial Cells From High Glucose-Induced Injury by Promoting Mitophagy Through the AMPK/mTOR/ULK1 Pathway.

Purpose: The regulation of mitophagy by Sirt3 has rarely been studied in ocular diseases. In the present study, we determined the effects of Sirt3 on AMPK/mTOR/ULK1 signaling pathway-mediated mitophagy in retinal pigment epithelial (RPE) cells in a high glucose environment. Methods: The mRNA expression levels of Sirt3, AMPK, mTOR, ULK1, and LC3B in RPE cells under varying glucose conditions were measured by real-time polymerase chain reaction (RT-PCR). The expressions of Sirt3, mitophagy protein, and AMPK/mTOR/ULK1 signaling pathway-related proteins were detected by Western blotting. Lentivirus (LV) transfection mediated the stable overexpression of Sirt3 in cell lines. The experimental groups were NG (5.5 mM glucose), hypertonic, HG (30 mM glucose), HG + LV-GFP, and HG + LV-Sirt3. Western blotting was performed to detect the expressions of mitophagy proteins and AMPK/mTOR/ULK1-related proteins in a high glucose environment during the overexpression of Sirt3. Reactive oxygen species (ROS) production in a high glucose environment was measured by DCFH-DA staining. Mitophagy was detected by labeling mitochondria and lysosomes with MitoTracker and LysoTracker probes, respectively. Apoptosis was detected by flow cytometry. Results: Sirt3 expression was reduced in the high glucose group, inhibiting the AMPK/mTOR/ULK1 pathway, with diminished mitophagy and increased intracellular ROS production. The overexpression of Sirt3, increased expression of p-AMPK/AMPK and p-ULK1/ULK1, and decreased expression of p-mTOR/mTOR inhibited cell apoptosis and enhanced mitophagy. Conclusions: Sirt3 protected RPE cells from high glucose-induced injury by activating the AMPK/mTOR/ULK1 signaling pathway. Translational Relevance: By identifying new targets of action, we aimed to establish effective therapeutic targets for diabetic retinopathy treatment.

Ginsenoside Rk1 induces autophagy-dependent apoptosis in hepatocellular carcinoma by AMPK/mTOR signaling pathway.

Hepatocellular carcinoma (HCC) is the third most lethal cancer in the world. Recent studies have shown that suppression of autophagy plays an important role in the development of HCC. Ginsenoside Rk1 is a protopanaxadiol saponin isolated from ginseng and has a significant anti-tumor effect, but its role and mechanism in HCC are still unclear. In this study, a mouse liver cancer model induced by diethylnitrosamine and carbon tetrachloride (DEN + CCl4) was employed to investigate the inhibitory effect of Rk1 on HCC. The results demonstrate that ginsenoside Rk1 effectively inhibits liver injury, liver fibrosis, and cirrhosis during HCC progression. Transcriptome data analysis of mouse liver tissue reveals that ginsenoside Rk1 significantly regulates the AMPK/mTOR signaling pathway, autophagy pathway, and apoptosis pathway. Subsequent studies show that ginsenoside Rk1 induces AMPK protein activation, upregulates the expression of autophagy marker LC3-II protein to promote autophagy, and then downregulates the expression of Bcl2 protein to trigger a caspase cascade reaction, activating AMPK/mTOR-induced toxic autophagy to promote cells death. Importantly, co-treatment of ginsenoside Rk1 with autophagy inhibitors can inhibit apoptosis of HCC cells, once again demonstrating the ability of ginsenoside Rk1 to promote autophagy-dependent apoptosis. In conclusion, our study demonstrates that ginsenoside Rk1 inhibits the development of primary HCC by activating toxic autophagy to promote apoptosis through the AMPK/mTOR pathway. These findings confirm that ginsenoside Rk1 is a promising new strategy for the treatment of HCC.

Arsenic disturbs neural tube closure involving AMPK/PKB-mTORC1-mediated autophagy in mice.

Arsenic exposure is a significant risk factor for folate-resistant neural tube defects (NTDs), but the potential mechanism is unclear. In this study, a mouse model of arsenic-induced NTDs was established to investigate how arsenic affects early neurogenesis leading to malformations. The results showed that in utero exposure to arsenic caused a decline in the normal embryos, an elevated embryo resorption, and a higher incidence of malformed embryos. Cranial and spinal deformities were the main malformation phenotypes observed. Meanwhile, arsenic-induced NTDs were accompanied by an oxidant/antioxidant imbalance manifested by elevated levels of reactive oxygen species (ROS) and decreased antioxidant activities. In addition, changes in the expression of autophagy-related genes and proteins (ULK1, Atg5, LC3B, p62) as well as an increase in autophagosomes were observed in arsenic-induced aberrant brain vesicles. Also, the components of the upstream pathway regulating autophagy (AMPK, PKB, mTOR, Raptor) were altered accordingly after arsenic exposure. Collectively, our findings propose a mechanism for arsenic-induced NTDs involving AMPK/PKB-mTORC1-mediated autophagy. Blocking autophagic cell death due to excessive autophagy provides a novel strategy for the prevention of folate-resistant NTDs, especially for arsenic-exposed populations.

Time-series analysis of hematopoietic stem cells.

This study aimed to investigate the molecular mechanisms underlying the aging of hematopoietic stem cells (HSCs). Gene expression profile GSE32719 was downloaded from the Gene Expression Omnibus database, including 14 young, 5 middle, and 8 old HSCs. Differential expression analysis, short time-series expression miner analysis, and weighted co-expression network analysis were conducted to screen for hub genes whose expression changed over time during HSC aging. Subsequently, functional enrichment and multiple regulatory network analyses of the hub genes were performed. A total of 124 intersecting time-dependent differentially expressed and module genes were obtained, which were considered hub genes whose expression changed over time during HSC aging. Hub genes were significantly enriched in pathways such as the Hippo and AMP-activated protein kinase (AMPK) signaling pathways. Moreover, AP-1 Transcription Factor Subunit (FOS) and sirtuin 1 (SIRT1) had higher degrees in the protein-protein interaction network, were regulated by more transcription factors (TFs), such as Sp1 transcription factor (SP1) and BRCA1 DNA repair-associated (BRCA1), in the TF-mRNA-miRNA network, were associated with more diseases in the disease-gene network, and could be targeted by more drugs in the drug-gene network. Furthermore, SIRT1 was targeted by miR-9-5p in the TF-mRNA-miRNA network. Hub genes such as FOS and SIRT1 and key pathways such as the Hippo and AMPK signaling pathways may play crucial roles in HSC aging. Moreover, FOS and SIRT1 were regulated by SP1 and BRCA1, respectively, during HSC aging. Furthermore, miR-9-5p may modulate HSC aging by targeting SIRT1. Thus, FOS and SIRT1 may be potential therapeutic targets for age-related hematopoietic dysfunction.