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Experimental and Therapeutic Medicine Aug 2024Excessive alcohol consumption is considered to be a major risk factor of alcohol-induced osteonecrosis of the femoral head (AONFH). The gut microbiota (GM) has been...
Excessive alcohol consumption is considered to be a major risk factor of alcohol-induced osteonecrosis of the femoral head (AONFH). The gut microbiota (GM) has been reported to aid in the regulation of human physiology and its composition can be altered by alcohol consumption. The aim of the present study was to improve the understanding of the GM and its metabolites in patients with AONFH. Metabolomic sequencing and 16S rDNA analysis of fecal samples were performed using liquid chromatography-mass spectrometry to characterize the GM of patients with AONFH and healthy normal controls (NCs). Metagenomic sequencing of fecal samples was performed to identify whether GM changes on the species level were associated with the expression of gut bacteria genes or their associated functions in patients with AONFH. The abundance of 58 genera was found to differ between the NC group and the AONFH group. Specifically, , , and were significantly more abundant in the AONFH group compared with those in the NC group. Metagenomic sequencing demonstrated that the majority of the bacterial species that exhibited significantly different abundance in patients with AONFH belonged to the genus . Fecal metabolomic analysis demonstrated that several metabolites were present at significantly different concentrations in the AONFH group compared with those in the NC group. These metabolites were products of vitamin B6 metabolism, retinol metabolism, pentose and glucuronate interconversions and glycerophospholipid metabolism. In addition, these changes in metabolite levels were observed to be associated with the altered abundance of specific bacterial species, such as , and . According to the results of the present study, a comprehensive landscape of the GM and metabolites in patients with AONFH was revealed, suggesting the existence of interplay between the gut microbiome and metabolome in AONFH pathogenesis.
PubMed: 38873043
DOI: 10.3892/etm.2024.12599 -
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 -
The Journal of Clinical Investigation Jun 2024Neutrophil hyperactivity and neutrophil extracellular trap release (NETosis) appear to play important roles in the pathogenesis of the thromboinflammatory autoimmune...
Neutrophil hyperactivity and neutrophil extracellular trap release (NETosis) appear to play important roles in the pathogenesis of the thromboinflammatory autoimmune disease known as antiphospholipid syndrome (APS). The understanding of neutrophil metabolism has advanced tremendously in the past decade, and accumulating evidence suggests that a variety of metabolic pathways guide neutrophil activities in health and disease. Our previous work characterizing the transcriptome of APS neutrophils revealed that genes related to glycolysis, glycogenolysis, and the pentose phosphate pathway (PPP) were significantly upregulated. Here, we found that APS patient neutrophils used glycolysis more avidly than healthy control neutrophils, especially when the neutrophils were from APS patients with a history of microvascular disease. In vitro, inhibiting either glycolysis or the PPP tempered phorbol myristate acetate- and APS IgG-induced NETosis, but not NETosis triggered by a calcium ionophore. In mice, inhibiting either glycolysis or the PPP reduced neutrophil reactive oxygen species production and suppressed APS IgG-induced NETosis ex vivo. When APS-associated thrombosis was evaluated in mice, inhibiting either glycolysis or the PPP markedly suppressed thrombosis and circulating NET remnants. In summary, these data identify a potential role for restraining neutrophil glucose flux in the treatment of APS.
PubMed: 38869951
DOI: 10.1172/JCI169893 -
World Journal of Microbiology &... Jun 2024Lignocellulosic biomass is a valuable, renewable substrate for the synthesis of polyhydroxybutyrate (PHB), an ecofriendly biopolymer. In this study, bacterial strain...
Lignocellulosic biomass is a valuable, renewable substrate for the synthesis of polyhydroxybutyrate (PHB), an ecofriendly biopolymer. In this study, bacterial strain E5-3 was isolated from soil in Japan; it was identified as Burkholderia ambifaria strain E5-3 by 16 S rRNA gene sequencing. The strain showed optimal growth at 37 °C with an initial pH of 9. It demonstrated diverse metabolic ability, processing a broad range of carbon substrates, including xylose, glucose, sucrose, glycerol, cellobiose, and, notably, palm oil. Palm oil induced the highest cellular growth, with a PHB content of 65% wt. The strain exhibited inherent tolerance to potential fermentation inhibitors derived from lignocellulosic hydrolysate, withstanding 3 g/L 5-hydroxymethylfurfural and 1.25 g/L acetic acid. Employing a fed-batch fermentation strategy with a combination of glucose, xylose, and cellobiose resulted in PHB production 2.7-times that in traditional batch fermentation. The use of oil palm trunk hydrolysate, without inhibitor pretreatment, in a fed-batch fermentation setup led to significant cell growth with a PHB content of 45% wt, equivalent to 10 g/L. The physicochemical attributes of xylose-derived PHB produced by strain E5-3 included a molecular weight of 722 kDa, a number-average molecular weight of 191 kDa, and a polydispersity index of 3.78. The amorphous structure of this PHB displayed a glass transition temperature of 4.59 °C, while its crystalline counterpart had a melting point of 171.03 °C. This research highlights the potential of lignocellulosic feedstocks, especially oil palm trunk hydrolysate, for PHB production through fed-batch fermentation by B. ambifaria strain E5-3, which has high inhibitor tolerance.
Topics: Lignin; Biomass; Palm Oil; Fermentation; Hydroxybutyrates; Burkholderia; Xylose; RNA, Ribosomal, 16S; Soil Microbiology; Glucose; Polyesters; Hydrogen-Ion Concentration; Furaldehyde; Cellobiose
PubMed: 38869634
DOI: 10.1007/s11274-024-04041-8 -
Extremophiles : Life Under Extreme... Jun 2024In this study, EPS production conditions of Geobacillus thermodenitrificans HBB 111, a thermophilic microorganism, were optimized and the amount of produced EPS (EPS...
In this study, EPS production conditions of Geobacillus thermodenitrificans HBB 111, a thermophilic microorganism, were optimized and the amount of produced EPS (EPS 111) was found to be 44.0 mg/L. EPS 111 was purified using ion exchange chromatography and gel filtration chromatography, and a single type of exopolysaccharide was obtained. The structure of the purified EPS 111 was evaluated by TLC, FTIR, NMR, and GC-MS, and it was observed that it contained hexose (glucose, fructose, galactose and mannose) and pentose sugars. From the SEM photographs, it was understood that EPS 111 had an amorphous, rough, and layered structure. It was found that purified EPS 111 had low cytotoxicity (2.3%) and exhibited high antioxidant activity and remarkable antidiabetic, prebiotic and fibrinolytic activities. It is very valuable that the purified EPS 111 in this study offers multiple biological activities compared to the thermophilic EPSs reported in the literature and has a high potential for use in biotechnological and biomedical fields.
Topics: Geobacillus; Polysaccharides, Bacterial; Antioxidants
PubMed: 38861190
DOI: 10.1007/s00792-024-01344-4 -
Environmental Science & Technology Jun 2024Microbial organic matter turnover is an important contributor to the terrestrial carbon dioxide (CO) budget. Partitioning of organic carbons into biomass relative to CO...
Microbial organic matter turnover is an important contributor to the terrestrial carbon dioxide (CO) budget. Partitioning of organic carbons into biomass relative to CO efflux, termed carbon-use efficiency (CUE), is widely used to characterize organic carbon cycling by soil microorganisms. Recent studies challenge proposals of CUE dependence on the oxidation state of the substrate carbon and implicate instead metabolic strategies. Still unknown are the metabolic mechanisms underlying variability in CUE. We performed a multiomics investigation of these mechanisms in , a versatile soil bacterium of the Gammaproteobacteria, processing a mixture of plant matter derivatives. Our C-metabolomics data captured substrate carbons into different metabolic pathways: cellulose-derived sugar carbons in glycolytic and pentose-phosphate pathways; lignin-related aromatic carbons in the tricarboxylic acid cycle. Subsequent C-metabolic flux analysis revealed a 3-fold lower investment of sugar carbons in CO efflux compared to aromatic carbons, in agreement with reported substrate-dependent CUE. Proteomics analysis revealed enzyme-level regulation only for substrate uptake and initial catabolism, which dictated downstream fluxes through CO-producing versus biomass-synthesizing reactions. Metabolic partitioning as shown here explained the substrate-dependent CUE calculated from reported metabolic flux analyses of other bacteria, further supporting a metabolism-guided perspective for predicting the microbial conversion of accessible organic matter to CO efflux.
Topics: Carbon Dioxide; Carbon; Metabolic Networks and Pathways; Pseudomonas putida; Biomass
PubMed: 38860668
DOI: 10.1021/acs.est.4c01328 -
Circulation. Genomic and Precision... Jun 2024Hypertrophic cardiomyopathy (HCM) is caused by sarcomere gene mutations (genotype-positive HCM) in ≈50% of patients and occurs in the absence of mutations...
Integrating Clinical Phenotype With Multiomics Analyses of Human Cardiac Tissue Unveils Divergent Metabolic Remodeling in Genotype-Positive and Genotype-Negative Patients With Hypertrophic Cardiomyopathy.
BACKGROUND
Hypertrophic cardiomyopathy (HCM) is caused by sarcomere gene mutations (genotype-positive HCM) in ≈50% of patients and occurs in the absence of mutations (genotype-negative HCM) in the other half of patients. We explored how alterations in the metabolomic and lipidomic landscape are involved in cardiac remodeling in both patient groups.
METHODS
We performed proteomics, metabolomics, and lipidomics on myectomy samples (genotype-positive N=19; genotype-negative N=22; and genotype unknown N=6) from clinically well-phenotyped patients with HCM and on cardiac tissue samples from sex- and age-matched and body mass index-matched nonfailing donors (N=20). These data sets were integrated to comprehensively map changes in lipid-handling and energy metabolism pathways. By linking metabolomic and lipidomic data to variability in clinical data, we explored patient group-specific associations between cardiac and metabolic remodeling.
RESULTS
HCM myectomy samples exhibited (1) increased glucose and glycogen metabolism, (2) downregulation of fatty acid oxidation, and (3) reduced ceramide formation and lipid storage. In genotype-negative patients, septal hypertrophy and diastolic dysfunction correlated with lowering of acylcarnitines, redox metabolites, amino acids, pentose phosphate pathway intermediates, purines, and pyrimidines. In contrast, redox metabolites, amino acids, pentose phosphate pathway intermediates, purines, and pyrimidines were positively associated with septal hypertrophy and diastolic impairment in genotype-positive patients.
CONCLUSIONS
We provide novel insights into both general and genotype-specific metabolic changes in HCM. Distinct metabolic alterations underlie cardiac disease progression in genotype-negative and genotype-positive patients with HCM.
Topics: Humans; Cardiomyopathy, Hypertrophic; Male; Female; Middle Aged; Genotype; Adult; Phenotype; Myocardium; Metabolomics; Proteomics; Lipidomics; Lipid Metabolism; Sarcomeres; Energy Metabolism; Aged; Multiomics
PubMed: 38853772
DOI: 10.1161/CIRCGEN.123.004369 -
Water Research Aug 2024Overuse of antibiotics has led to their existence in nitrogen-containing water. The impacts of antibiotics on bio-denitrification and the metabolic response of...
Overuse of antibiotics has led to their existence in nitrogen-containing water. The impacts of antibiotics on bio-denitrification and the metabolic response of denitrifiers to antibiotics are unclear. We systematically analyzed the effect of ciprofloxacin (CIP) on bio-denitrification and found that 5 mg/L CIP greatly inhibited denitrification with a model denitrifier (Paracoccus denitrificans). Nitrate reduction decreased by 32.89 % and nitrous oxide emission increased by 75.53 %. The balance analysis of carbon and nitrogen metabolism during denitrification showed that CIP exposure blocked electron transfer and reduced the flow of substrate metabolism used for denitrification. Proteomics results showed that CIP exposure induced denitrifiers to use the pentose phosphate pathway more for substrate metabolism. This caused a substrate preference to generate NADPH to prevent cellular damage rather than NADH for denitrification. Notably, despite denitrifiers having antioxidant defenses, they could not completely prevent oxidative damage caused by CIP exposure. The effect of CIP exposure on denitrifiers after removal of extracellular polymeric substances (EPS) demonstrated that EPS around denitrifiers formed a barrier against CIP. Fluorescence and infrared spectroscopy revealed that the binding effect of proteins in EPS to CIP prevented damage. This study shows that denitrifiers resist antibiotic stress through different intracellular and extracellular defense strategies.
Topics: Ciprofloxacin; Denitrification; Anti-Bacterial Agents; Paracoccus denitrificans
PubMed: 38851110
DOI: 10.1016/j.watres.2024.121851 -
BMC Plant Biology Jun 2024The phosphorylation of the Light-Harvesting Complex of photosystem II (LHCII) driven by STATE TRANSITION 7 (STN7) kinase is a part of one of the crucial regulatory...
BACKGROUND
The phosphorylation of the Light-Harvesting Complex of photosystem II (LHCII) driven by STATE TRANSITION 7 (STN7) kinase is a part of one of the crucial regulatory mechanisms of photosynthetic light reactions operating in fluctuating environmental conditions, light in particular. There are evidenced that STN7 can also be activated without light as well as in dark-chilling conditions. However, the biochemical mechanism standing behind this complex metabolic pathway has not been deciphered yet.
RESULTS
In this work, we showed that dark-chilling induces light-independent LHCII phosphorylation in runner bean (Phaseolus coccineus L.). In dark-chilling conditions, we registered an increased reduction of the PQ pool which led to activation of STN7 kinase, subsequent LHCII phosphorylation, and possible LHCII relocation inside the thylakoid membrane. We also presented the formation of a complex composed of phosphorylated LHCII and photosystem I typically formed upon light-induced phosphorylation. Moreover, we indicated that the observed steps were preceded by the activation of the oxidative pentose phosphate pathway (OPPP) enzymes and starch accumulation.
CONCLUSIONS
Our results suggest a direct connection between photosynthetic complexes reorganization and dark-chilling-induced activation of the thioredoxin system. The proposed possible pathway starts from the activation of OPPP enzymes and further NADPH-dependent thioredoxin reductase C (NTRC) activation. In the next steps, NTRC simultaneously activates ADP-glucose pyrophosphorylase and thylakoid membrane-located NAD(P)H dehydrogenase-like complex. These results in starch synthesis and electron transfer to the plastoquinone (PQ) pool, respectively. Reduced PQ pool activates STN7 kinase which phosphorylates LHCII. In this work, we present a new perspective on the mechanisms involving photosynthetic complexes while efficiently operating in the darkness. Although we describe the studied pathway in detail, taking into account also the time course of the following steps, the biological significance of this phenomenon remains puzzling.
Topics: Phaseolus; Phosphorylation; Light; Thylakoids; Photosystem I Protein Complex; Cold Temperature; Light-Harvesting Protein Complexes; Photosystem II Protein Complex; Plant Proteins; Starch; Pentose Phosphate Pathway; Enzyme Activation; Photosynthesis; Stress, Physiological; Protein Serine-Threonine Kinases
PubMed: 38849759
DOI: 10.1186/s12870-024-05169-3 -
Free Radical Biology & Medicine Jun 2024The oxidative phase of the pentose phosphate pathway (PPP) involving the enzymes glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconolactonase (6PGL), and...
The oxidative phase of the pentose phosphate pathway (PPP) involving the enzymes glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconolactonase (6PGL), and 6-phosphogluconate dehydrogenase (6PGDH), is critical to NADPH generation within cells, with these enzymes catalyzing the conversion of glucose-6-phosphate (G6P) into ribulose-5-phosphate (Ribu5-P). We have previously studied peroxyl radical (ROO•) mediated oxidative inactivation of E. coli G6PDH, 6PGL, and 6PGDH. However, these data were obtained from experiments where each enzyme was independently exposed to ROO•, a condition not reflecting biological reality. In this work we investigated how NADPH production is modulated when these enzymes are jointly exposed to ROO•. Enzyme mixtures (1:1:1 ratio) were exposed to ROO• produced from thermolysis of 100 mM 2,2'-azobis(2-methylpropionamidine) dihydrochloride (AAPH). NADPH was quantified at 340 nm, and protein oxidation analyzed by liquid chromatography with mass spectrometric detection (LC-MS). The data obtained were rationalized using a mathematical model. The mixture of non-oxidized enzymes, G6P and NADP generated ∼175 μM NADPH. Computational simulations showed a constant decrease of G6P associated with NADPH formation, consistent with experimental data. When the enzyme mixture was exposed to AAPH (3 h, 37 ºC), lower levels of NADPH were detected (∼100 μM) which also fitted with computational simulations. LC-MS analyses indicated modifications at Tyr, Trp, and Met residues but at lower concentrations than detected for the isolated enzymes. Quantification of NADPH generation showed that the pathway activity was not altered during the initial stages of the oxidations, consistent with a buffering role of G6PDH towards inactivation of the oxidative phase of the pathway.
PubMed: 38848786
DOI: 10.1016/j.freeradbiomed.2024.05.050