RÉSUMÉ
BACKGROUND: Bile acids (BAs) are steroid-derived molecules with important roles in digestion, the maintenance of host metabolism, and immunomodulation. Primary BAs are synthesized by the host, while secondary BAs are produced by the gut microbiome through transformation of the former. The regulation of microbial production of secondary BAs is not well understood, particularly the production of 7-dehydroxylated BAs, which are the most potent agonists for host BA receptors. The 7-dehydroxylation of cholic acid (CA) is well established and is linked to the expression of a bile acid-inducible (bai) operon responsible for this process. However, little to no 7-dehydroxylation has been reported for other host-derived BAs (e.g., chenodeoxycholic acid, CDCA or ursodeoxycholic acid, UDCA). RESULTS: Here, we demonstrate that the 7-dehydroxylation of CDCA and UDCA by the human isolate Clostridium scindens is induced when CA is present, suggesting that CA-dependent transcriptional regulation is required for substantial 7-dehydroxylation of these primary BAs. This is supported by the finding that UDCA alone does not promote expression of bai genes. CDCA upregulates expression of the bai genes but the expression is greater when CA is present. In contrast, the murine isolate Extibacter muris exhibits a distinct response; CA did not induce significant 7-dehydroxylation of primary BAs, whereas BA 7-dehydroxylation was promoted upon addition of germ-free mouse cecal content in vitro. However, E. muris was found to 7-dehydroxylate in vivo. CONCLUSIONS: The distinct expression responses amongst strains indicate that bai genes are regulated differently. CA promoted bai operon gene expression and the 7-dehydroxylating activity in C. scindens strains. Conversely, the in vitro activity of E. muris was promoted only after the addition of cecal content and the isolate did not alter bai gene expression in response to CA. The accessory gene baiJ was only upregulated in the C. scindens ATCC 35704 strain, implying mechanistic differences amongst isolates. Interestingly, the human-derived C. scindens strains were also capable of 7-dehydroxylating murine bile acids (muricholic acids) to a limited extent. This study shows novel 7-dehydroxylation activity in vitro resulting from the presence of CA and suggests distinct bai gene expression across bacterial species.
Sujet(s)
Acides et sels biliaires , Acide cholique , Acide cholique/métabolisme , Animaux , Acides et sels biliaires/métabolisme , Souris , Humains , Clostridium/métabolisme , Clostridium/génétique , Régulation de l'expression des gènes bactériens , Hydroxylation , Opéron , Chénodiol/métabolisme , Acide ursodésoxycholique/métabolisme , Microbiome gastro-intestinalRÉSUMÉ
Flavin-dependent monooxygenases (FMOs) are a valuable group of biocatalysts that can regioselectively introduce a hydroxy group for the targeted modification of biologically active compounds. Here, we present the fdeE, the FMO from Herbaspirillum seropedicae SmR1 that is a part of the naringenin degradation pathway and is active towards a wide range of flavonoids-flavanones, flavones, isoflavones, and flavonols. Bioinformatics and biochemical analysis revealed a high similarity between the analyzed enzyme and other F8H FMOs what might indicate convergent evolutionary mechanism of flavonoid degradation pathway emergence by microorganism. A simple approach with the manipulation of the reaction environment allowed the stable formation of hydroxylation products, which showed very high reactivity in both in vivo and in vitro assays. This approach resulted in an 8-hydroxyquercetin-gossypetin titer of 0.16 g/L and additionally it is a first report of production of this compound.
Sujet(s)
Flavonols , Isoflavones , Isoflavones/métabolisme , Isoflavones/composition chimique , Isoflavones/biosynthèse , Flavonols/métabolisme , Flavonoïdes/métabolisme , Flavonoïdes/composition chimique , Flavonoïdes/biosynthèse , Hydroxylation , Spécificité du substratRÉSUMÉ
Vitamin D deficiencies are linked to multiple human diseases. Optimizing its synthesis, physicochemical properties, and delivery systems while minimizing side effects is of clinical relevance and is of great medical and industrial interest. Biotechnological techniques may render new modified forms of vitamin D that may exhibit improved absorption, stability, or targeted physiological effects. Novel modified vitamin D derivatives hold promise for developing future therapeutic approaches and addressing specific health concerns related to vitamin D deficiency or impaired metabolism, such as avoiding hypercalcemic effects. Identifying and engineering key enzymes and biosynthetic pathways involved, as well as developing efficient cultures, are therefore of outmost importance and subject of intense research. Moreover, we elaborate on the critical role that microbial bioconversions might play in the a la carte design, synthesis, and production of novel, more efficient, and safer forms of vitamin D and its analogs. In summary, the novelty of this work resides in the detailed description of the physiological, medical, biochemical, and epidemiological aspects of vitamin D supplementation and the steps towards the enhanced and simplified industrial production of this family of bioactives relying on microbial enzymes. KEY POINTS: ⢠Liver or kidney pathologies may hamper vitamin D biosynthesis ⢠Actinomycetes are able to carry out 1α- or 25-hydroxylation on vitamin D precursors.
Sujet(s)
Biotransformation , Vitamine D , Vitamine D/métabolisme , Humains , Voies de biosynthèse/génétique , Génie métabolique/méthodes , Actinobacteria/métabolisme , Actinobacteria/génétique , Biotechnologie/méthodes , Bactéries/métabolisme , Bactéries/génétique , HydroxylationRÉSUMÉ
Selection of chemical-resistant predatory mites is a good alternative to balance the contradiction between chemical control and biological control. Previously, a resistant strain of Neoseiulus barkeri for chlorpyrifos was obtained. In the current study, two up-regulated (NbCYP3A6, NbCYP3A16) and one down-regulated (NbCYP3A24) P450s were screened through differential expression analysis and other detoxification-related genes such as CCEs, GST, etc. were not found. 3D modelling and molecular docking indicated that the chlorine at position 5 on the pyridine ring of chlorpyrifos, as well as a methyl group, were closest to the heme iron of the enzymes (less than 5 Å). Three active recombinant P450 proteins were heterologously expressed and metabolized with chlorpyrifos in vitro. HPLC assay showed that only NbCYP3A24 could metabolize chlorpyrifos, with a metabolism rate of 21.60 %. Analysis of the m/z of metabolites by LC-MS/MS showed that chlorine at the 5C position of chlorpyrifos was stripped and hydroxylated, whereas chlorpyrifos-oxon, a common product of oxidation by P450, was not found. Knockdown of the NbCYP3A24 gene in the susceptiblestrain did reduce the susceptibility of N. barkeri to chlorpyrifos, suggesting that the biological activity of the metabolite may be similar to chlorpyrifos-oxon, thus enhancing the inhibitory effect on the target.
Sujet(s)
Chlorpyriphos , Cytochrome P-450 enzyme system , Insecticides , Mites (acariens) , Simulation de docking moléculaire , Chlorpyriphos/métabolisme , Chlorpyriphos/composition chimique , Chlorpyriphos/analogues et dérivés , Animaux , Mites (acariens)/effets des médicaments et des substances chimiques , Cytochrome P-450 enzyme system/métabolisme , Cytochrome P-450 enzyme system/génétique , Insecticides/métabolisme , Insecticides/composition chimique , Insecticides/toxicité , Régulation négative , HydroxylationRÉSUMÉ
5F-EDMB-PICA is a newly emerged synthetic cannabinoid which has been characterized in relevant literature in recent years. Although phase-I metabolites of 5F-EDMB-PICA have been partly reported, the phase-II metabolism of this synthetic cannabinoid has not been studied yet. In this study, we established a phase-I and phase-II metabolism model in vitro by using pooled human liver microsomes, NADPH regeneration system, and UGT incubation system, with 1 mg/ml 5F-EDMB-PICA added and incubated at 37 °C for 60 min. The metabolites were analyzed by Q Exactive™ Hybrid Quadrupole-Orbitrap™ Mass Spectrometer, via which we discovered and identified 14 phase-I metabolites and 4 phase-II metabolites of 5F-EDMB-PICA, involving pathways such as ester hydrolysis, dehydrogenation, hydrolytic defluorination, hydroxylation, dihydroxylation, glucuronidation, and combinations of the pathways mentioned above. We recommend considering the monohydroxylation metabolites (M9, M10) with higher content and intact ester and 5-fluoropentyl structures as potential biomarkers of 5F-EDMB-PICA.
Sujet(s)
Cannabinoïdes , Microsomes du foie , Humains , Microsomes du foie/métabolisme , Cannabinoïdes/métabolisme , Glucuronosyltransferase/métabolisme , Voies et réseaux métaboliques , Détoxication de phase I , Détoxication de phase II , NADP/métabolisme , HydroxylationRÉSUMÉ
Second near-infrared (NIR-II) laser-mediated photothermal therapy and sonothermal therapy using low-intensity focused ultrasound exposure for tumors have attracted increasing attention owing to their ability to penetrate deep tissues and provide noninvasive ablation with high therapeutic efficacy. However, their applications were limited by the shortness of optimal NIR-II photothermal agents and sonothermal agents. In this study, we discovered that the edge-selectively hydroxylated graphene nanosheets (EHG NSs) with excellent water dispersibility and an "intact conjugated plane" were not only an outstanding NIR-II photothermal agent but also an effective sonothermal agent for tumor therapy. EHG NSs were incorporated into an injectable adhesive thermosensitive hydrogel with a characteristic sol-gel phase transition behavior. EHG NSs endowed the injectable hydrogel with an exceptional photothermal effect under the laser irradiation (1064 nm, 1.0 W cm-2) as well as an effective sonothermal effect under ultrasonic exposure (3.0 MHz, 2.1 W cm-2), effectively killing tumor cells in vitro and inhibiting tumor growth after intratumoral injection. Especially, the NIR-II photothermal therapy based on the hybrid hydrogel completely ablated the primary tumors and effectively activated systemic anti-tumor immune responses benefiting from the protein adsorption capacity of the injectable hydrogel, significantly inhibiting the growth of the distal tumors. Collectively, EHG nanosheets loaded in the injectable hydrogel will be a promising "all-rounder" for noninvasive deep penetrating thermotherapy and a potent platform that integrates various therapies.
Sujet(s)
Graphite , Rayons infrarouges , Graphite/composition chimique , Animaux , Souris , Humains , Nanostructures/composition chimique , Hydroxylation , Antinéoplasiques/composition chimique , Antinéoplasiques/pharmacologie , Thérapie photothermique , Souris de lignée BALB C , Prolifération cellulaire/effets des médicaments et des substances chimiques , Survie cellulaire/effets des médicaments et des substances chimiques , Hydrogels/composition chimique , Hydrogels/pharmacologie , Photothérapie , Taille de particuleRÉSUMÉ
Piceatannol, resveratrol's derivative, and a valuable polyphenol has managed to become one of the most remarkable candidate molecules for drug development research, with its high bioactive properties and higher stability. On the other hand, the very low amount of piceatannol in plants which are its natural source increases the cost and limits the commercialization possibilities of the product. To overcome this bottleneck, a limited number of studies have recently shown that it is possible to produce piceatannol from the resveratrol precursor much cheaper by regioselective hydroxylation catalyzed by bacteria isolated from the soil, and the search for new bacteria of similar nature in new ecosystems has gained popularity. The aim of our study, which was prepared within this framework, is the bacterial isolate with regioselective hydroxylation potential obtained as a result of selective isolation steps; determination of resveratrol hydroxylation potentials and piceatannol product yields, investigation of possibilities to increase piceatannol yield with optimization trials and identification of isolates with the highest yield. For this purpose, 200 bacterial isolates capable of resveratrol hydroxylation were obtained from soil samples taken from Erzurum (Turkey) and its surroundings by using selective media. In the continuation of the study; resveratrol hydroxylation trials were carried out with these isolates and 55 active isolates capable of producing piceatannol by regioselective hydroxylation were selected. Then, yield improvement studies of active isolates were carried out by using different carbon sources and optimizing the culture conditions. As a result, a culture collection was created by identifying the 6 most active bacterial isolates with commercialization potential using conventional and molecular methods. These are 4 Gram-positive (Rhodococcus sp., Rhodococcus erythropolis, Paeniglutamicibacter sp., Arthrobacter sp.) and 2 Gram-negative (Shinella sp., Ensifer adhaerens) bacterial isolates. As a result of the optimization studies, three of these isolates used phenol as a biocatalyst, while the other three increased the production yield of piceatannol by using 4-hydroxyphenylacetic acid.
Sujet(s)
Bactéries , Resvératrol , Microbiologie du sol , Stilbènes , Stilbènes/métabolisme , Stilbènes/isolement et purification , Bactéries/métabolisme , Bactéries/isolement et purification , Bactéries/classification , Resvératrol/métabolisme , Turquie , HydroxylationRÉSUMÉ
Major constituents of the plant cell walls are structural proteins that belong to the hydroxyproline-rich glycoprotein (HRGP) family. Leucine-rich repeat extensin (LRX) proteins contain a leucine-rich domain and a C-terminal domain with repetitive Ser-Pro3-5 motifs that are potentially to be O-glycosylated. It has been demonstrated that pollen-specific LRX8-LRX11 from Arabidopsis thaliana are necessary to maintain the integrity of the pollen tube cell wall during polarized growth. In HRGPs, including classical extensins (EXTs), and probably in LRXs, proline residues are converted to hydroxyproline by prolyl-4-hydroxylases (P4Hs), thus defining novel O-glycosylation sites. In this context, we aimed to determine whether hydroxylation and subsequent O-glycosylation of Arabidopsis pollen LRXs are necessary for their proper function and cell wall localization in pollen tubes. We hypothesized that pollen-expressed P4H4 and P4H6 catalyze the hydroxylation of the proline units present in Ser-Pro3-5 motifs of LRX8-LRX11. Here, we show that the p4h4-1 p4h6-1 double mutant exhibits a reduction in pollen germination rates and a slight reduction in pollen tube length. Pollen germination is also inhibited by P4H inhibitors, suggesting that prolyl hydroxylation is required for pollen tube development. Plants expressing pLRX11::LRX11-GFP in the p4h4-1 p4h6-1 background show partial re-localization of LRX11-green fluorescent protein (GFP) from the pollen tube tip apoplast to the cytoplasm. Finally, immunoprecipitation-tandem mass spectrometry analysis revealed a decrease in oxidized prolines (hydroxyprolines) in LRX11-GFP in the p4h4-1 p4h6-1 background compared with lrx11 plants expressing pLRX11::LRX11-GFP. Taken together, these results suggest that P4H4 and P4H6 are required for pollen germination and for proper hydroxylation of LRX11 necessary for its localization in the cell wall of pollen tubes.
Sujet(s)
Protéines d'Arabidopsis , Arabidopsis , Tube pollinique , Prolyl hydroxylases , Arabidopsis/métabolisme , Arabidopsis/génétique , Hydroxylation , Tube pollinique/croissance et développement , Tube pollinique/métabolisme , Tube pollinique/génétique , Protéines d'Arabidopsis/métabolisme , Protéines d'Arabidopsis/génétique , Prolyl hydroxylases/métabolisme , Prolyl hydroxylases/génétique , Paroi cellulaire/métabolismeRÉSUMÉ
Aberrations in the Hedgehog (Hh) signaling pathway are significantly prevailed in various cancers, including B-cell lymphoma. A critical facet of Hh signal transduction involves the dynamic regulation of the suppressor of fused homolog (SUFU)-glioma-associated oncogene homolog (GLI) complex within the kinesin family member 7 (KIF7)-supported ciliary tip compartment. However, the specific post-translational modifications of SUFU-GLI complex within this context have remained largely unexplored. Our study reveals a novel regulatory mechanism involving prolyl 4-hydroxylase 2 (P4HA2), which forms a complex with KIF7 and is essential for signal transduction of Hh pathway. We demonstrate that, upon Hh pathway activation, P4HA2 relocates alongside KIF7 to the ciliary tip. Here, it hydroxylates SUFU to inhibit its function, thus amplifying the Hh signaling. Moreover, the absence of P4HA2 significantly impedes B lymphoma progression. This effect can be attributed to the suppression of Hh signaling in stromal fibroblasts, resulting in decreased growth factors essential for malignant proliferation of B lymphoma cells. Our findings highlight the role of P4HA2-mediated hydroxylation in modulating Hh signaling and propose a novel stromal-targeted therapeutic strategy for B-cell lymphoma.
Sujet(s)
Évolution de la maladie , Protéines Hedgehog , Lymphome B , Procollagen-Proline Dioxygenase , Protéines de répression , Transduction du signal , Protéines Hedgehog/métabolisme , Humains , Lymphome B/métabolisme , Lymphome B/anatomopathologie , Souris , Animaux , Protéines de répression/métabolisme , Protéines de répression/génétique , Procollagen-Proline Dioxygenase/métabolisme , Hydroxylation , Communication paracrine , Prolifération cellulaire , Kinésine/métabolisme , Kinésine/génétique , Lignée cellulaire tumorale , Prolyl hydroxylasesRÉSUMÉ
In drug discovery, metabolite profiling unveils biotransformation pathways and potential toxicant formation, guiding selection of candidates with optimal pharmacokinetics and safety profiles. Tazemetostat (TAZ) is employed in treating locally advanced or metastatic epithelioid sarcoma. Identification of drug metabolites are of significant importance in improving safety, efficacy and reduced toxicity of drugs. The current study aimed to investigate the comprehensive metabolic fate of TAZ using different in vivo (rat) and in vitro (RLM, HLM, HS9) models. For in vivo studies, drug was orally administered to Sprague-Dawley rats with subsequent analysis of plasma, feces and urine samples. A total of 21 new metabolites were detected across various matrices and were separated on Phenomenex kinetex C18 (2.5 µm; 150 × 4.6 mm) column using acetonitrile and 0.1% formic acid in water as mobile phase. LC-QTOF-MS/MS and NMR techniques were employed to identify and characterize the metabolites from extracted samples. The major metabolic routes found in biotransformation of TAZ were hydroxylation, N-dealkylation, N-oxidation, hydrogenation, hydrolysis and N-acetylation. In silico toxicity revealed potential immunotoxicity for TAZ and few of its metabolites. This research article is the first time to discuss the complete metabolite profiling including identification and characterization of TAZ metabolites as well as its biotransformation mechanism.
Sujet(s)
Chromatographie en phase liquide , Spectrométrie de masse en tandem , Animaux , Humains , Mâle , Rats , Biotransformation , Fèces/composition chimique , Hydroxylation , Microsomes du foie/métabolisme , Rat Sprague-DawleyRÉSUMÉ
Bromophenols has been proven to synthesize hydroxylated polybrominated diphenyl ethers (OH-PBDEs), which may pose additional environmental and health risks in the process of bioremediation. In this study, the removal of 2,4-dibromophenol (2,4-DBP) and 2,4,6-tribromophenol (2,4,6-TBP) and the biosynthetic of OH-PBDEs by Prorocentrum donghaiense were explored. The removal efficiencies of 2,4-DBP and 2,4,6-TBP ranged from 32.71% to 76.89% and 31.15%-78.12%, respectively. Low concentrations of 2,4-DBP stimulated algal growth, while high concentrations were inhibitory. Furthermore, exposure to 10.00 mg L-1 2,4-DBP resulted in the detection of 2'-hydroxy-2,3',4,5'-tetrabromodiphenyl ether (2'-OH-BDE-68) within P. donghaiense. In contrast, increasing concentrations of 2,4,6-TBP considerably inhibited P. donghaiense growth, with 4'-hydroxy-2,3',4,5',6-pentabromodiphenyl ether (4'-OH-BDE-121) detected within P. donghaiense under 5.00 mg L-1 2,4,6-TBP. Metabolomic analysis further revealed that the synthesized OH-PBDEs exhibited higher toxicity than their precursors and identified the oxidative coupling as a key biosynthetic mechanism. These findings confirm the capacity of P. donghaiense to remove bromophenols and biosynthesize OH-PBDEs from bromophenols, offering valuable insights into formulating algal bioremediation to mitigate bromophenol contamination.
Sujet(s)
Dépollution biologique de l'environnement , Éthers de polyhalogénophényle , Phénols , Éthers de polyhalogénophényle/métabolisme , Éthers de polyhalogénophényle/composition chimique , Phénols/métabolisme , Hydroxylation , Ignifuges/métabolismeRÉSUMÉ
Extracellular matrix (ECM) is a major component of the tumor environment, promoting the establishment of a pro-invasive behavior. Such environment is supported by both tumor- and stromal-derived metabolites, particularly lactate. In prostate cancer (PCa), cancer-associated fibroblasts (CAFs) are major contributors of secreted lactate, able to impact on metabolic and transcriptional regulation in cancer cells. Here, we describe a mechanism by which CAF-secreted lactate promotes in PCa cells the expression of genes coding for the collagen family. Lactate-exploiting PCa cells rely on increased α-ketoglutarate (α-KG) which activates the α-KG-dependent collagen prolyl-4-hydroxylase (P4HA1) to support collagen hydroxylation. De novo synthetized collagen plays a signaling role by activating discoidin domain receptor 1 (DDR1), supporting stem-like and invasive features of PCa cells. Inhibition of lactate-induced collagen hydroxylation and DDR1 activation reduces the metastatic colonization of PCa cells. Overall, these results provide a new understanding of the link between collagen remodeling/signaling and the nutrient environment exploited by PCa.
Sujet(s)
Collagène , Matrice extracellulaire , Acide lactique , Métastase tumorale , Tumeurs de la prostate , Mâle , Tumeurs de la prostate/métabolisme , Tumeurs de la prostate/anatomopathologie , Tumeurs de la prostate/génétique , Humains , Matrice extracellulaire/métabolisme , Collagène/métabolisme , Acide lactique/métabolisme , Lignée cellulaire tumorale , Fibroblastes associés au cancer/métabolisme , Fibroblastes associés au cancer/anatomopathologie , Transduction du signal , Régulation de l'expression des gènes tumoraux , Animaux , Souris , Microenvironnement tumoral , Acides cétoglutariques/métabolisme , HydroxylationRÉSUMÉ
N-Acetylnorloline synthase (LolO) is one of several iron(II)- and 2-oxoglutarate-dependent (Fe/2OG) oxygenases that catalyze sequential reactions of different types in the biosynthesis of valuable natural products. LolO hydroxylates C2 of 1-exo-acetamidopyrrolizidine before coupling the C2-bonded oxygen to C7 to form the tricyclic loline core. Each reaction requires cleavage of a C-H bond by an oxoiron(IV) (ferryl) intermediate; however, different carbons are targeted, and the carbon radicals have different fates. Prior studies indicated that the substrate-cofactor disposition (SCD) controls the site of H· abstraction and can affect the reaction outcome. These indications led us to determine whether a change in SCD from the first to the second LolO reaction might contribute to the observed reactivity switch. Whereas the single ferryl complex in the C2 hydroxylation reaction was previously shown to have typical Mössbauer parameters, one of two ferryl complexes to accumulate during the oxacyclization reaction has the highest isomer shift seen to date for such a complex and abstracts H· from C7 â¼ 20 times faster than does the first ferryl complex in its previously reported off-pathway hydroxylation of C7. The detectable hydroxylation of C7 in competition with cyclization by the second ferryl complex is not enhanced in 2H2O solvent, suggesting that the C2 hydroxyl is deprotonated prior to C7-H cleavage. These observations are consistent with the coordination of the C2 oxygen to the ferryl complex, which may reorient its oxo ligand, the substrate, or both to positions more favorable for C7-H cleavage and oxacyclization.
Sujet(s)
Fer , Acides cétoglutariques , Acides cétoglutariques/métabolisme , Acides cétoglutariques/composition chimique , Fer/métabolisme , Fer/composition chimique , Hydroxylation , Cyclisation , Oxygénases/métabolisme , Oxygénases/composition chimique , Protéines bactériennes/métabolisme , Protéines bactériennes/composition chimiqueRÉSUMÉ
The oxygen-labile transcription factor called hypoxia-inducible factor (HIF) is responsible for the cellular and organismal adaptive response to reduced oxygen availability. Deregulation of HIF is associated with the pathogenesis of major human diseases including cardiovascular disease and cancer. Under normoxia, the HIFα subunit is hydroxylated on conserved proline residues within the oxygen-dependent degradation domain (ODD) that labels HIFα for proteasome-mediated degradation. Despite similar oxygen-dependent degradation machinery acting on HIF1α and HIF2α, these two paralogs have been shown to exhibit unique kinetics under hypoxia, which suggests that other regulatory processes may be at play. Here, we characterize the protease activity found in rabbit reticulocytes that specifically cleaves the ODD of HIF1α but not HIF2α. Notably, the cleavage product is observed irrespective of the oxygen-dependent prolyl-hydroxylation potential of HIF1α, suggesting independence from oxygen. HIF1α M561T substitution, which mimics an evolutionary substitution that occurred during the duplication and divergence of HIF1α and HIF2α, diminished the cleavage of HIF1α. Protease inhibitor screening suggests that cysteine proteases cathepsins L and B preferentially cleave HIF1αODD, thereby revealing an additional layer of differential HIF regulation.
Sujet(s)
Cathepsine L , Sous-unité alpha du facteur-1 induit par l'hypoxie , Oxygène , Protéolyse , Sous-unité alpha du facteur-1 induit par l'hypoxie/métabolisme , Sous-unité alpha du facteur-1 induit par l'hypoxie/génétique , Animaux , Cathepsine L/métabolisme , Cathepsine L/génétique , Lapins , Oxygène/métabolisme , Humains , Réticulocytes/métabolisme , Facteurs de transcription à motif basique hélice-boucle-hélice/métabolisme , Facteurs de transcription à motif basique hélice-boucle-hélice/génétique , HydroxylationRÉSUMÉ
BACKGROUND: Thyroid hormones are primarily responsible for the brain development in perinatal mammals. However, this process can be inhibited by external factors such as environmental chemicals. Perinatal mammals are viviparous, which makes direct fetal examination difficult. METHODS: We used metamorphic amphibians, which exhibit many similarities to perinatal mammals, as an experimental system. Therefore, using metamorphic amphibians, we characterized the gene expression of matrix metalloproteinases, which play an important role in brain development. RESULTS: The expression of many matrix metalloproteinases (mmps) was characteristically induced during metamorphosis. We also found that the expression of many mmps was induced by T3 and markedly inhibited by hydroxylated polychlorinated biphenyls (PCBs). CONCLUSION: Overall, our findings suggest that hydroxylated PCBs disrupt normal brain development by disturbing the gene expression of mmps.
Sujet(s)
Encéphale , Matrix metalloproteinases , Métamorphose biologique , Polychlorobiphényles , Hormones thyroïdiennes , Xenopus laevis , Animaux , Encéphale/métabolisme , Encéphale/effets des médicaments et des substances chimiques , Encéphale/croissance et développement , Xenopus laevis/métabolisme , Xenopus laevis/génétique , Matrix metalloproteinases/métabolisme , Matrix metalloproteinases/génétique , Polychlorobiphényles/toxicité , Métamorphose biologique/effets des médicaments et des substances chimiques , Métamorphose biologique/génétique , Hormones thyroïdiennes/métabolisme , Régulation de l'expression des gènes au cours du développement/effets des médicaments et des substances chimiques , HydroxylationRÉSUMÉ
Amaryllidaceae alkaloids are structurally diverse natural products with a wide range biological properties, and based on the partial identification of the biosynthetic enzymes, norbelladine would be a common intermediate in the biosynthetic pathways. Previous studies suggested that norbelladine synthase (NBS) catalyzed the condensation reaction of 3,4-dihydroxybenzaldehyde and tyramine to form norcraugsodine, and subsequently, noroxomaritidine/norcraugsodine reductase (NR) catalyzed the nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reduction of norcraugsodine to generate norbelladine. However, recent studies have highlighted possible alternative Amaryllidaceae alkaloid biosynthetic pathways via the formation of isovanillin and vanillin from the 4-O- and 3-O-methylation reactions of 3,4-dihydroxybenzaldehyde, respectively. Herein, we focused on NpsNBS and NpsNR, which were initially identified from Narcissus pseudonarcissus, and explored their substrate recognition tolerance by performing condensation reactions of tyramine with various benzaldehyde derivatives, to shed light on the Amaryllidaceae alkaloid biosynthetic pathway from the viewpoint of the enzymatic properties. The assays revealed that both NpsNBS and NpsNR lacked the abilities to produce 4'-O- and 3'-O-methylnorbelladine from isovanillin and vanillin with tyramine, respectively. These observations thus suggested that Amaryllidaceae alkaloids are biosynthesized from norbelladine, formed through the condensation/reduction reaction of 3,4-dihydroxybenzaldehyde with tyramine.
Sujet(s)
Aldéhydes , Aldéhydes/composition chimique , Aldéhydes/métabolisme , Hydroxylation , Structure moléculaire , Spécificité du substrat , Nitrate reductase/composition chimique , Nitrate reductase/métabolismeRÉSUMÉ
A new detection platform based on a hydroxylated covalent organic framework (COF) integrated with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was constructed and used for detecting adrenergic receptor agonists (ARAs) residues in milk. The hydroxylated COF was prepared by polymerization of tris(4-aminophenyl)amine and 1,3,5-tris(4-formyl-3-hydroxyphenyl)benzene and applied to solid-phase extraction (SPE) of ARAs. This hydroxylated COF was featured with hierarchical flower-like morphology, easy preparation, and copious active adsorption sites. The adsorption model fittings and molecular simulation were applied to explore the potential adsorption mechanism. This detection platform was suitable for detecting four α2- and five ß2-ARAs residues in milk. The linear ranges of the ARAs were from 0.25 to 50 µg·kg-1; the intra-day and the inter-day repeatability were in the range 2.9-7.9% and 2.0-10.1%, respectively. This work demonstrates this hydroxylated COF has great potential as SPE cartridge packing, and provides a new way to determine ARAs residues in milk.
Sujet(s)
Lait , Extraction en phase solide , Spectrométrie de masse en tandem , Extraction en phase solide/méthodes , Lait/composition chimique , Animaux , Spectrométrie de masse en tandem/méthodes , Hydroxylation , Réseaux organométalliques/composition chimique , Adsorption , Agonistes adrénergiques/composition chimique , Agonistes adrénergiques/analyse , Limite de détection , BovinsRÉSUMÉ
Furan-containing compounds distribute widely in food, herbal medicines, industrial synthetic products, and environmental media. These compounds can undergo oxidative metabolism catalyzed by cytochrome P450 enzymes (CYP450) within organisms, which may produce reactive products, possibly reacting with biomolecules to induce toxic effects. In this work, we performed DFT calculations to investigate the CYP450-mediated metabolic mechanism of furan-ring oxidation using 2-methylfuran as a model substrate, meanwhile, we studied the regioselective competition of another hydroxylation reaction involving methyl group of 2-methylfuran. As a result, we found the toxicological-relevant cis-enedione product can be produced from O-addition directly via a concerted manner without formation of an epoxide intermediate as traditionally believed. Moreover, our calculations demonstrate the kinetic and thermodynamic feasibility of both furan-ring oxidation and methyl hydroxylation pathways, although the former pathway is a bit more favorable. We then constructed a linear model to predict the rate-limiting activation energies (ΔE*) of O-addition with 11 diverse furan substates based on their adiabatic ionization potentials (AIPs) and condensation Fukui functions (CFFs). The results show a good predictive ability (R2=0.94, Q2CV=0.87). Therefore, AIP and CFF with clear physichem meanings relevant to the mechanism, emerge as pivotal molecular descriptors to enable the fast prediction of furan-ring oxidation reactivities for quick insight into the toxicological risk of furans, using just ground-state calculations.
Sujet(s)
Cytochrome P-450 enzyme system , Théorie de la fonctionnelle de la densité , Furanes , Oxydoréduction , Furanes/composition chimique , Cytochrome P-450 enzyme system/métabolisme , Cytochrome P-450 enzyme system/composition chimique , Hydroxylation , Cinétique , ThermodynamiqueRÉSUMÉ
Brepocitinib is an oral once-daily Janus kinase 1 and Tyrosine kinase 2 selective inhibitor currently in development for the treatment of several autoimmune disorders. Mass balance and metabolic profiles were determined using accelerator mass spectrometry in six healthy male participants following a single oral 60 mg dose of 14C-brepocitinib (â¼300 nCi). The average mass balance recovery was 96.7% ± 6.3%, with the majority of dose (88.0% ± 8.0%) recovered in urine and 8.7% ± 2.1% of the dose recovered in feces. Absorption of brepocitinib was rapid, with maximal plasma concentrations of total radioactivity and brepocitinib achieved within 0.5 hours after dosing. Circulating radioactivity consisted primarily of brepocitinib (47.8%) and metabolite M1 (37.1%) derived from hydroxylation at the C5' position of the pyrazole ring. Fractional contributions to metabolism via cytochrome P450 enzymes were determined to be 0.77 for CYP3A4/5 and 0.14 for CYP1A2 based on phenotyping studies in human liver microsomes. However, additional clinical studies are required to understand the potential contribution of CYP1A1. Approximately 83% of the dose was eliminated as N-methylpyrazolyl oxidative metabolites, with 52.1% of the dose excreted as M1 alone. Notably, M1 was not observed as a circulating metabolite in earlier metabolic profiling of human plasma from a multiple ascending dose study with unlabeled brepocitinib. Mechanistic studies revealed that M1 was highly unstable in human plasma and phosphate buffer, undergoing chemical oxidation leading to loss of the 5-hydroxy-1-methylpyrazole moiety and formation of aminopyrimidine cleavage product M2. Time-dependent inhibition and trapping studies with M1 yielded insights into the mechanism of this unusual and unexpected instability. SIGNIFICANCE STATEMENT: This study provides a detailed understanding of the disposition and metabolism of brepocitinib, a JAK1/TYK2 inhibitor for atopic dermatitis, in humans as well as characterization of clearance pathways and pharmacokinetics of brepocitinib and its metabolites.
Sujet(s)
Inhibiteurs de protéines kinases , Humains , Mâle , Adulte , Inhibiteurs de protéines kinases/pharmacocinétique , Inhibiteurs de protéines kinases/administration et posologie , Inhibiteurs de protéines kinases/métabolisme , Jeune adulte , Pyrazoles/pharmacocinétique , Pyrazoles/métabolisme , Pyrazoles/sang , Pyrazoles/administration et posologie , Janus kinase 1/antagonistes et inhibiteurs , Janus kinase 1/métabolisme , Administration par voie orale , Cytochrome P-450 CYP3A/métabolisme , Volontaires sains , Microsomes du foie/métabolisme , Kinase Janus-2/antagonistes et inhibiteurs , Kinase Janus-2/métabolisme , Fèces/composition chimique , Hydroxylation , Cytochrome P-450 CYP1A2/métabolisme , Adulte d'âge moyenRÉSUMÉ
The antioxidant N-(1,3-Dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) and its oxidized quinone product 6PPD-quinone (6PPD-Q) in rubber have attracted attention due to the ecological risk that they pose. Both 6PPD and 6PPD-Q have been detected in various environments that humans cohabit. However, to date, a clear understanding of the biotransformation of 6PPD-Q and a potential biomarker for exposure in humans are lacking. To address this issue, this study presents a comprehensive analysis of the extensive biotransformation of 6PPD-Q across species, encompassing both in vitro and in vivo models. We have tentatively identified 17 biotransformation metabolites in vitro, 15 in mice in vivo, and confirmed the presence of two metabolites in human urine samples. Interestingly, different biotransformation patterns were observed across species. Through semiquantitative analysis based on peak areas, we found that almost all 6PPD-Q underwent biotransformation within 24 h of exposure in mice, primarily via hydroxylation and subsequent glucuronidation. This suggests a rapid metabolic processing of 6PPD-Q in mammals, underscoring the importance of identifying effective biomarkers for exposure. Notably, monohydroxy 6PPD-Q and 6PPD-Q-O-glucuronide were consistently the most predominant metabolites across our studies, highlighting monohydroxy 6PPD-Q as a potential key biomarker for epidemiological research. These findings represent the first comprehensive data set on 6PPD-Q biotransformation in mammalian systems, offering insights into the metabolic pathways involved and possible exposure biomarkers.