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International Journal of Molecular... May 2024Marine natural products constitute a great source of potential new antidiabetic drugs. The aim of this study was to evaluate the role of phosphoeleganin (PE), a...
Marine natural products constitute a great source of potential new antidiabetic drugs. The aim of this study was to evaluate the role of phosphoeleganin (PE), a polyketide purified from the Mediterranean ascidian , and its derivatives PE/2 and PE/3 on insulin sensitivity in human hepatocellular carcinoma (HepG2) cells. In our experiments, insulin stimulates the phosphorylation of its receptor (INSR) and AKT by 1.5- and 3.5-fold, respectively, whereas in the presence of PE, PE/2, and PE/3, the insulin induced INSR phosphorylation is increased by 2.1-, 2-, and 1.5-fold and AKT phosphorylation by 7.1-, 6.0-, and 5.1-fold, respectively. Interestingly, PE and PE/2 have an additive effect on insulin-mediated reduction of phosphoenolpyruvate carboxykinase (PEPCK) expression. Finally, PE and PE/2, but not PE/3, decrease interleukin 6 (IL6) secretion and expression before and after palmitic acid incubation, while in the presence of high glucose (HG), only PE reduces IL6. Levels of other cytokines are not significantly affected by PE and its derivates. All these data suggest that PE and its synthetic-derived compound, PE/2, significantly decrease IL6 and improve hepatic insulin signaling. As IL6 impairs insulin action, it could be hypothesized that PE and PE/2, by inhibiting IL6, may improve the hepatic insulin pathway.
Topics: Humans; Interleukin-6; Insulin; Liver Neoplasms; Carcinoma, Hepatocellular; Signal Transduction; Hep G2 Cells; Animals; Receptor, Insulin; Phosphorylation; Proto-Oncogene Proteins c-akt; Insulin Resistance; Antigens, CD
PubMed: 38892230
DOI: 10.3390/ijms25116039 -
Scientific Reports Jun 2024Mitochondrial phosphoenolpyruvate carboxykinase (PCK2), a mitochondrial isoenzyme, supports the growth of cancer cells under glucose deficiency conditions in vitro. This...
Mitochondrial phosphoenolpyruvate carboxykinase (PCK2), a mitochondrial isoenzyme, supports the growth of cancer cells under glucose deficiency conditions in vitro. This study investigated the role and potential mechanism of PCK2 in the occurrence and development of Hepatocellular carcinoma (HCC). The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and other databases distinguish the expression of PCK2 and verified by qRT-PCR and Western blotting. Kaplan-Meier was conducted to assess PCK2 survival in HCC. The potential biological function of PCK2 was verified by enrichment analysis and gene set enrichment analysis (GSEA). The correlation between PCK2 expression and immune invasion and checkpoint was found by utilizing Tumor Immune Estimation Resource (TIMER). Lastly, the effects of PCK2 on the proliferation and metastasis of hepatocellular carcinoma cells were evaluated by cell tests, and the expressions of Epithelial mesenchymal transformation (EMT) and apoptosis related proteins were detected. PCK2 is down-regulated in HCC, indicating a poor prognosis. PCK2 gene mutation accounted for 1.3% of HCC. Functional enrichment analysis indicated the potential of PCK2 as a metabolism-related therapeutic target. Subsequently, we identified several signaling pathways related to the biological function of PCK2. The involvement of PCK2 in immune regulation was verified and key immune checkpoints were predicted. Ultimately, after PCK2 knockdown, cell proliferation and migration were significantly increased, and N-cadherin and vimentin expression were increased. PCK2 has been implicated in immune regulation, proliferation, and metastasis of hepatocellular carcinoma, and is emerging as a novel predictive biomarker and metabolic-related clinical target.
Topics: Carcinoma, Hepatocellular; Liver Neoplasms; Humans; Prognosis; Cell Proliferation; Gene Expression Regulation, Neoplastic; Cell Line, Tumor; Phosphoenolpyruvate Carboxykinase (GTP); Epithelial-Mesenchymal Transition; Mitochondria; Male; Female; Apoptosis; Cell Movement; Biomarkers, Tumor; Middle Aged; Phosphoenolpyruvate Carboxykinase (ATP)
PubMed: 38890507
DOI: 10.1038/s41598-024-64907-7 -
Veterinary Research Jun 2024Bacteria utilize intercellular communication to orchestrate essential cellular processes, adapt to environmental changes, develop antibiotic tolerance, and enhance...
Bacteria utilize intercellular communication to orchestrate essential cellular processes, adapt to environmental changes, develop antibiotic tolerance, and enhance virulence. This communication, known as quorum sensing (QS), is mediated by the exchange of small signalling molecules called autoinducers. AI-2 QS, regulated by the metabolic enzyme LuxS (S-ribosylhomocysteine lyase), acts as a universal intercellular communication mechanism across gram-positive and gram-negative bacteria and is crucial for diverse bacterial processes. In this study, we demonstrated that in Streptococcus suis (S. suis), a notable zoonotic pathogen, AI-2 QS enhances galactose utilization, upregulates the Leloir pathway for capsular polysaccharide (CPS) precursor production, and boosts CPS synthesis, leading to increased resistance to macrophage phagocytosis. Additionally, our molecular docking and dynamics simulations suggest that, similar to S. pneumoniae, FruA, a fructose-specific phosphoenolpyruvate phosphotransferase system prevalent in gram-positive pathogens, may also function as an AI-2 membrane surface receptor in S. suis. In conclusion, our study demonstrated the significance of AI-2 in the synthesis of galactose metabolism-dependent CPS in S. suis. Additionally, we conducted a preliminary analysis of the potential role of FruA as a membrane surface receptor for S. suis AI-2.
Topics: Streptococcus suis; Galactose; Quorum Sensing; Virulence; Animals; Bacterial Capsules; Lactones; Streptococcal Infections; Homoserine; Polysaccharides, Bacterial
PubMed: 38886823
DOI: 10.1186/s13567-024-01335-5 -
Plant Physiology and Biochemistry : PPB Jun 2024The phosphoenolpyruvate carboxylase kinase of Medicago sativa L. (MsPPCK1) modulates the phosphorylation status and activity of the C4 pathway phosphoenolpyruvate...
Overexpression of phosphoenolpyruvate carboxylase kinase gene MsPPCK1 from Medicago sativa L. increased alkali tolerance of alfalfa by enhancing photosynthetic efficiency and promoting nodule development.
The phosphoenolpyruvate carboxylase kinase of Medicago sativa L. (MsPPCK1) modulates the phosphorylation status and activity of the C4 pathway phosphoenolpyruvate carboxylase enzyme, which is pivotal for photosynthetic carbon assimilation in plants. This study investigated the role of MsPPCK1 in alfalfa by creating transgenic plants overexpressing MsPPCK1 under the control of the CaMV35S promoter. The enhanced alkali tolerance of transgenic plants indicated an important role of MsPPCK1 gene in regulating plant alkali tolerance. Transgenic plants exhibited heightened antioxidant activity (SOD, POD, and CAT), reduced MDA, HO, OFR and REC% content, increased activity of key photosynthetic enzymes (PEPC, PPDK, NADP-ME, and NADP-MDH), and enhanced photosynthetic parameters (Pn, E, Gs, and Ci). Moreover, MsPPCK1 overexpression increased the content of organic acids (oxaloacetic, malic, citric, and succinic acids) in the plants. The upregulation of MsPPCK1 under rhizobial inoculation showcased its other role in nodule development. In transgenic plants, MsDMI2, MsEnod12, and MsNODL4 expression increased, facilitating root nodule development and augmenting plant nodulation. Accelerated root nodule growth positively influences plant growth and yield and enhances alfalfa resistance to alkali stress. This study highlights the pivotal role of MsPPCK1 in fortifying plant alkali stress tolerance and improving yield, underscoring its potential as a key genetic target for developing alkali-tolerant and high-yielding alfalfa varieties.
PubMed: 38879983
DOI: 10.1016/j.plaphy.2024.108764 -
Journal of Dairy Science Jun 2024Nutrition and physiological state affect hepatic metabolism. Our objective was to determine if feeding flaxseed oil (∼50% C18:3n-3 cis), high oleic soybean oil (∼70%...
Nutrition and physiological state affect hepatic metabolism. Our objective was to determine if feeding flaxseed oil (∼50% C18:3n-3 cis), high oleic soybean oil (∼70% C18:1 cis-9), or milk fat (∼50% C16:0) alters hepatic expression of PC, PCK1, and PCK2 and the flow of carbons from propionate and pyruvate into the TCA cycle in preruminating calves. Male Holstein calves (n = 40) were assigned to a diet of skim milk with either: 3% milk fat (MF; n = 8), 3% flaxseed oil (Flax; n = 8), 3% high oleic soybean oil (HOSO; n = 8), 1.5% MF + 1.5% high oleic soybean oil (MF-HOSO; n = 8), or 1.5% MF + 1.5% flaxseed oil (MF-Flax; n = 8) from d 14 to d 21 postnatal. At d 21 postnatal, a liver biopsy was taken for gene expression and metabolic flux analysis. Liver explants were incubated in [U-C] propionate and [U-C] pyruvate to trace carbon flux through TCA cycle intermediates or with [U-C] lactate, [1-C] palmitic acid, or [2-C] propionate to quantify substrate oxidation to CO and acid soluble products. Compared with other treatments, plasma C18:3n-3 cis was 10 times higher and C18:1 cis-9 was 3 times lower in both flax (Flax and MF-Flax) treatments. PC, PCK1, and PCK2 expression and flux of [U-C] pyruvate as well as [U-C] propionate were not different between treatments. PC expression was negatively correlated with the enrichment of citrate M+5 and malate M+3, and PCK2 was negatively correlated with citrate M+5, suggesting that when expression of these enzymes is increased, carbon from pyruvate enters the TCA cycle via PC mediated carboxylation, and then OAA is converted to phosphoenolpyruvate via PCK2. Acid soluble product formation and PC expression were reduced in HOSO (MF-HOSO and HOSO) treatments compared with flax (MF-Flax and Flax), indicating that fatty acids regulate PC expression and carbon flux, but that fatty acid flux control points are not connected to PC, PCK1, or PCK2. In conclusion, fatty acids regulate hepatic expression of PC, PCK1, and PCK2, and carbon flux, but the point of control is distinct.
PubMed: 38876219
DOI: 10.3168/jds.2023-24500 -
Scientific Reports Jun 2024Cervical cancer, one of the most common gynecological cancers, is primarily caused by human papillomavirus (HPV) infection. The development of resistance to chemotherapy...
Cervical cancer, one of the most common gynecological cancers, is primarily caused by human papillomavirus (HPV) infection. The development of resistance to chemotherapy is a significant hurdle in treatment. In this study, we investigated the mechanisms underlying chemoresistance in cervical cancer by focusing on the roles of glycogen metabolism and the pentose phosphate pathway (PPP). We employed the cervical cancer cell lines HCC94 and CaSki by manipulating the expression of key enzymes PCK1, PYGL, and GYS1, which are involved in glycogen metabolism, through siRNA transfection. Our analysis included measuring glycogen levels, intermediates of PPP, NADPH/NADP ratio, and the ability of cells to clear reactive oxygen species (ROS) using biochemical assays and liquid chromatography-mass spectrometry (LC-MS). Furthermore, we assessed chemoresistance by evaluating cell viability and tumor growth in NSG mice. Our findings revealed that in drug-resistant tumor stem cells, the enzyme PCK1 enhances the phosphorylation of PYGL, leading to increased glycogen breakdown. This process shifts glucose metabolism towards PPP, generating NADPH. This, in turn, facilitates ROS clearance, promotes cell survival, and contributes to the development of chemoresistance. These insights suggest that targeting aberrant glycogen metabolism or PPP could be a promising strategy for overcoming chemoresistance in cervical cancer. Understanding these molecular mechanisms opens new avenues for the development of more effective treatments for this challenging malignancy.
Topics: Humans; Female; Uterine Cervical Neoplasms; Reactive Oxygen Species; Drug Resistance, Neoplasm; Neoplastic Stem Cells; Animals; Mice; Cell Line, Tumor; Phosphoenolpyruvate Carboxykinase (GTP); Glycogen; Intracellular Signaling Peptides and Proteins; Glycogenolysis; Pentose Phosphate Pathway; Cell Survival
PubMed: 38871968
DOI: 10.1038/s41598-024-64255-6 -
Journal of Agricultural and Food... Jun 2024β-Alanine, a valuable β-type amino acid, is experiencing increased demand due to its multifaceted applications in food flavoring, nutritional supplements,...
β-Alanine, a valuable β-type amino acid, is experiencing increased demand due to its multifaceted applications in food flavoring, nutritional supplements, pharmaceuticals, and the chemical industry. Nevertheless, the sustainable biosynthesis of β-alanine currently faces challenges due to the scarcity of robust strains, attributed to the complexities of modulating multiple genes and the inherent physiological constraints. Here, systems metabolic engineering was implemented in to overcome these limitations. First, an efficient l-aspartate-α-decarboxylase (ADC) was recruited for β-alanine biosynthesis. To conserve phosphoenolpyruvate flux, we subsequently modified the endogenous glucose assimilation system by inactivating the phosphotransferase system (PTS) and introducing an alternative non-PTS system, which increased β-alanine production to 1.70 g/L. The supply of key precursors, oxaloacetate and l-aspartate, was synergistically improved through comprehensive modulation, including strengthening main flux and blocking bypass metabolism, which significantly increased the β-alanine titer to 3.43 g/L. Next, the expression of ADC was optimized by promoter and untranslated region (UTR) engineering. Further transport engineering, which involved disrupting β-alanine importer CycA and heterologously expressing β-alanine exporter NCgI0580, improved β-alanine production to 8.48 g/L. Additionally, corn steep liquor was used to develop a cost-effective medium. The final strain produced 74.03 g/L β-alanine with a yield of 0.57 mol/mol glucose during fed-batch fermentation.
Topics: beta-Alanine; Escherichia coli; Metabolic Engineering; Glucose; Fermentation; Escherichia coli Proteins; Glutamate Decarboxylase
PubMed: 38867465
DOI: 10.1021/acs.jafc.4c03492 -
Nutrition Research and Practice Jun 2024Kaempferol (Ka) is one of the most widely occurring flavonoids found in large amounts in various plants. Ka has anti-obesity, antioxidant, and anti-inflammatory effects....
BACKGROUND/OBJECTIVES
Kaempferol (Ka) is one of the most widely occurring flavonoids found in large amounts in various plants. Ka has anti-obesity, antioxidant, and anti-inflammatory effects. Despite the numerous papers documenting the efficacy of Ka, some controversy remains. Therefore, this study examined the impact of Ka using 3T3-L1 and high-fat diet-induced obese mice.
MATERIALS/METHODS
3T3-L1 cells were treated with 50 μM Ka from the initiation of 3T3-L1 differentiation at D0 until the completion of differentiation on D8. Thirty male mice (C57BL/6J, 4 weeks old) were divided into 3 groups: normal diet (ND), high-fat diet (HFD), and HFD + 0.02% (w/w) Ka (Ka) group. All mice were fed their respective diets for 16 weeks. The mice were sacriced, and the plasma and hepatic lipid levels, white adipose tissue weight, hepatic glucose level, lipid level, and antioxidant enzyme activities were analyzed, and immunohistochemistry staining was performed.
RESULTS
Ka suppressed the hypertrophy of 3T3-L1 cells, and the Ka-supplemented mice showed a significant decrease in perirenal, retroperitoneal, mesenteric, and subcutaneous fat compared to the HFD group. Ka supplementation in high-fat diet-induced obese mice also improved the overall blood lipid concentration (total cholesterol, non-high-density lipoprotein-cholesterol, phospholipids, and apolipoprotein B). Ka supplementation in high-fat-induced obesity mice reduced hepatic steatosis and insulin resistance by modulating the hepatic lipid (glucose-6-phosphate dehydrogenase, fatty acid synthase, malic enzyme, phosphatidate phosphohydrolase, and β-oxidation) activities and glucose (glucokinase, phosphoenolpyruvate carboxykinase, and G6pase)-regulating enzymes. Ka supplementation ameliorated the erythrocyte and hepatic mitochondrial HO and inflammation levels (plasma tumor necrosis factor-alpha, monocyte chemoattractant protein-1, interleukin-6, and interferon-gamma and fibrosis of liver and epididymal fat).
CONCLUSION
Ka may be beneficial for preventing diet-induced obesity, inflammation, oxidative stress, and diabetes.
PubMed: 38854471
DOI: 10.4162/nrp.2024.18.3.325 -
Comparative Biochemistry and... Jun 2024Non-blood-feeding leeches, Whitmania pigra, have evolved unique digestive structures and physiological mechanisms to cope with fasting. However, the metabolic changes...
Non-blood-feeding leeches, Whitmania pigra, have evolved unique digestive structures and physiological mechanisms to cope with fasting. However, the metabolic changes and molecular mechanisms induced by fasting remain unclear. Therefore, this study recorded the weights of leeches during the fasting process. The weight changes were divided into two stages: a rapid decline period (1-9 weeks) and a fluctuating decline period (9-24 weeks). Leeches fasted for 4 (H4), 11 (H11), and 24 (H24) weeks were selected for transcriptome sequencing. Compared to the control group (H0), 436, 1157, and 337 differentially expressed genes (DEGs) were identified, which were mainly related to glycolysis/gluconeogenesis, amino acid metabolism, and the lipid metabolism pathway. The 6-phosphofructokinase (Pfk), pyruvate kinase (PK), and phosphoenolpyruvate carboxykinase (Pck) transcription levels revealed glycolysis/gluconeogenesis activation during the early stage of fasting and peaked at 11 weeks. Decreased expression of the rate-limiting enzyme acetyl-CoA carboxylase (ACC) in fatty acid synthesis during fasting may impede fatty acid synthesis. These results indicated that the nutrient storage and energy-supplying pathways in W. pigra were modified to improve fasting resistance. The findings of this study provided guidance for exploring the mechanism underlying fasting metabolism and laid a foundation for artificial breeding to improve the resistance of leeches.
PubMed: 38852903
DOI: 10.1016/j.cbpb.2024.110999 -
Biochemical and Biophysical Research... Sep 2024This study aimed to explore the involvement of phosphoenolpyruvate carboxykinase 2 (PCK2) in gefitinib-resistant non-small cell lung cancer (NSCLC) cells and assess its...
OBJECTIVES
This study aimed to explore the involvement of phosphoenolpyruvate carboxykinase 2 (PCK2) in gefitinib-resistant non-small cell lung cancer (NSCLC) cells and assess its feasibility as a therapeutic target against gefitinib resistance.
METHODS
Gefitinib-resistant cell lines, PC9GR and HCC827GR, were generated through progressive exposure of parental cells to escalating concentrations of gefitinib. Transcriptomic analysis encompassed the treatment of PC9 and PC9GR cells with gefitinib or vehicle, followed by RNA extraction, sequencing, and subsequent bioinformatic analysis. Cell viability was determined via CCK-8 assay, while clonogenic assays assessed colony formation. Apoptosis was detected utilizing the Annexin V-FITC/7AAD kit. Iron ion concentrations were quantified using FerroOrange. mRNA analysis was conducted through quantitative RT-PCR. Western blotting was employed for protein analysis. H&E and immunohistochemical staining were performed on tumor tissue sections.
RESULTS
The results revealed that depletion or inhibition of PCK2 significantly enhanced gefitinib's efficacy in inducing cell growth arrest, apoptosis, and ferroptosis in resistant NSCLC. Moreover, PCK2 knockdown led to the downregulation of key ferroptosis-related proteins, GPX4 and SLC7A11, while upregulating ASCL4. Conversely, overexpression of PCK2 in gefitinib-sensitive cells rendered resistance to gefitinib. In vivo experiments using a gefitinib-resistant xenograft model demonstrated that PCK2 silencing not only reduced tumor growth but also considerably increased the anti-tumor effect of gefitinib.
CONCLUSIONS
In conclusion, our study presents compelling evidence indicating that PCK2 plays a pivotal role in gefitinib resistance in NSCLC. The modulation of ferroptosis-related proteins and the involvement of Akt activation further elucidate the mechanisms underlying this resistance. Consequently, PCK2 emerges as a promising therapeutic target for overcoming gefitinib resistance in NSCLC, offering a new avenue for the development of more effective treatment strategies.
Topics: Ferroptosis; Gefitinib; Carcinoma, Non-Small-Cell Lung; Humans; Drug Resistance, Neoplasm; Lung Neoplasms; Cell Line, Tumor; Animals; Phosphoenolpyruvate Carboxykinase (ATP); Antineoplastic Agents; Mice; Mice, Nude; Apoptosis
PubMed: 38850814
DOI: 10.1016/j.bbrc.2024.150200