Search Results (1,311)

Background: The cytochrome P450 family 21 subfamily A member 2 gene (CYP21A2) encodes 21-hydroxylase, a key enzyme in adrenal steroid biosynthesis. Despite its physiological importance, the diversity of CYP21A2 transcript variants and their tissue-specific expression in domestic animals, including cattle, remains largely unexplored. This study aimed to characterize CYP21A2 transcription in adrenal glands and ovaries and assess the potential transcriptional activity of its pseudogene, CYP21A1P. Methods: CYP21A2 transcription was investigated in adrenal and ovarian tissues of 12 healthy cows using semi-quantitative PCR and Sanger sequencing. Real-time PCR was performed to confirm expression levels. Melting curve analysis and electrophoresis were used to validate distinct amplicons corresponding to different transcript variants. Extended amplicons were sequenced to identify transcripts corresponding to reference sequences and potential pseudogene products. Results: A single transcript variant (NM_001013596.1) was consistently detected in adrenal glands, whereas ovaries expressed two variants: NM_001013596.1 and XM_024983378.2. Semi-quantitative analysis showed significantly higher CYP21A2 expression in adrenal glands compared to ovaries (p < 0.01). In ovarian samples, the NM_001013596.1 variant was more abundant than the XM_024983378.2 (p < 0.01). Sanger sequencing revealed two products matching CYP21A2 reference transcripts and an additional, longer product containing sequence motifs specific to the pseudogene CYP21A1P, indicating its transcriptional activity. Conclusions: These results provide the first evidence of tissue-specific expression and differential abundance of CYP21A2 transcript variants in cattle and suggest the transcription of the CYP21A1P pseudogene. The findings reveal the complexity of CYP21A2 expression in steroidogenic tissues and suggest potential regulatory roles for transcript and pseudogene variants in bovine physiology. Full article

Aspergillus flavus is both an agricultural and clinical pathogen, notable for its ability to contaminate crops with aflatoxins and cause invasive aspergillosis. The increasing emergence of azole resistance in A. flavus poses a serious challenge to food safety and human health. Although mutations in ergosterol biosynthesis genes have been reported in resistant isolates, their functional contributions remain largely unvalidated. In this study, we investigated the role of the CYP51A Y119F mutation in azole resistance. Site-directed mutants were generated using PCR-based gene editing, and their susceptibility to antifungal agents was assessed through Clinical and Laboratory Standards Institute broth microdilution and agar diffusion assays. The Y119F mutation reduced susceptibility specifically to voriconazole and isavuconazole, while susceptibility to itraconazole and posaconazole remained unchanged. To explore the structural basis of this phenotype, molecular dynamics simulations were performed. The mutant protein exhibited greater fluctuations and reduced conformational stability compared to the wild-type enzyme. Tunnel analysis further indicated that the Y119F substitution caused narrowing and shortening of the main access tunnels to the heme-binding pocket, likely impairing azole access and binding. The combined biochemical and structural analyses suggest that Y119F represents a primary resistance-conferring mutation that modifies the structural dynamics of CYP51A. Full article

The unique cytochrome P450 BM3 from Priestia megaterium (syn. Bacillus megaterium) is renowned for its versatile high catalytic activity. The cyp102A1-LG23 gene encoding its CYP102A1-LG23 mutant variant was expressed in Escherichia coli and Mycolicibacterium smegmatis. The in vivo activity of the heterologous enzyme was assessed with respect to androstenedione (AD), androstadienedione (ADD), testosterone (TS) and dehydroepiandrosterone (DHEA). Alongside 7β-hydroxylation, the heterologous enzyme catalyzed the mono- and dihydroxylation of C19 steroids. For the first time, the formation of 7β-, 6β- and 11α-hydroxylated derivatives of ADD using a bacterial enzyme, as well as the hydroxylation of DHEA at the C7α and C7β positions, and its dihydroxylation with the formation of the 7α,15α-dihydroxylated derivative using the mutant cytochrome P450 BM3 were demonstrated. The steroid structures were confirmed using mass spectrometry and ¹H NMR spectroscopy. The advantages of using mycolicibacteria as a bacterial chassis for gene expression were also shown. The results demonstrate the unusual properties of the mutant cytochrome P450 BM3-LG23 and open up prospects for its application in the biotechnological production of valuable hydroxysteroids. Full article

Nanoplastics are pervasive contaminants that adversely affect male reproductive function, yet the molecular basis of polystyrene nanoplastic (PS-NP) toxicity in immature testes and effective preventive strategies remain unclear. Here, male mice (postnatal days 22–35, PND 22–35) and TM3 Leydig cells were exposed to graded PS-NPs, followed by transcriptomic profiling to identify differentially expressed genes (DEGs). Candidate therapeutics were prioritized using Connectivity Map (CMap) analysis and molecular docking, and protein interactions were examined by co-immunoprecipitation (Co-IP). PS-NPs accumulated in immature testes, eliciting excessive reactive oxygen species (ROS) and activation of NF-κB. These events coincided with the downregulation of steroidogenic enzymes (CYP11A1 and StAR) and disruption of testicular microarchitecture. In TM3 cells, PS-NPs suppressed testosterone synthesis in a concentration-dependent manner; this effect was fully reversed by pretreatment with N-acetylcysteine (NAC) or Bay 11-7082. Co-IP demonstrated p65–steroidogenic factor-1 (SF-1) binding consistent with formation of a transcriptional repressor complex targeting steroidogenic genes. CMap and docking analyses nominated parthenolide (PTL) as a candidate inhibitor of NF-κB nuclear translocation (predicted binding affinity, −6.585 kcal/mol), and PTL mitigated PS-NP-induced impairment of testosterone synthesis in vitro. Collectively, these data indicate that PS-NPs disrupt testosterone biosynthesis in immature testes through the ROS/NF-κB/p65–SF-1 axis, while PTL emerges as a candidate small molecule to counter nanoplastic-associated reproductive toxicity. These findings underscore translational relevance and support future evaluation under chronic low-dose exposure conditions, including in vivo validation of PTL efficacy, pharmacokinetics, and safety. Full article

Although traditionally contraindicated, the coadministration of tamoxifen and estradiol may hold clinical relevance in specific contexts, particularly in breast cancer survivors with premature menopause and a high risk of osteoporosis, thereby justifying the need to re-evaluate this therapeutic combination. This study presents an innovative physiologically based pharmacokinetic (PBPK) modeling approach to evaluate the coadministration of tamoxifen and estradiol in women with breast cancer and a high risk of osteoporosis. Using GastroPlus^® software, PBPK models were developed and validated for both drugs, based on physicochemical and kinetic data obtained from the literature and, where necessary, supplemented by estimates generated in ADMET Predictor^®. The simulations considered different hormonal profiles (pre and postmenopausal) and therapeutic regimens, evaluating potential interactions mediated by the CYP3A4 enzyme. Analysis of the pharmacokinetic parameters (F, C_max, T_max and AUC) revealed strong agreement between the simulated and experimental values, with prediction errors of less than twofold. The drug interaction studies, carried out in dynamic and stationary modes, indicated that estradiol does not significantly alter the pharmacokinetics of tamoxifen, even at increasing doses or in enlarged virtual populations. These results represent the first in silico evidence that, under certain conditions, the concomitant use of estradiol does not compromise the pharmacokinetic efficacy of tamoxifen. Although the study is computational, it provides a solid scientific basis for re-evaluating this therapeutic combination and proposes a pioneering model for personalized strategies in complex oncological contexts. All simulations assumed average enzyme abundance/activity without CYP polymorphism parameterization; findings are restricted to parent-tamoxifen pharmacokinetics and do not infer metabolite (e.g., endoxifen) exposure or phenotype effects. Full article

Background/Objectives: Although several studies have been reported on the modulation of Cytochrome P450 by resveratrol, inconsistencies in the results obtained require further investigation. Here, we report the results of in vivo and in vitro experiments investigating the effect of resveratrol on CYP1A2, which participates in the biotransformation of several drugs used for the treatment of human malignancies. Methods: Male Wistar rats were exposed to resveratrol through diet (1%) for 30 days, and the hepatic CYP1A2 activity and protein concentration were assayed at the end of the treatment. Additionally, the capacity of the phytochemical to interfere with the induction of CYP1A2 by benzo[a]pyrene (50 mg/kg body weight) was also studied. The inhibition of CYP1A2 activity in rat liver microsomal and recombinant human enzymes by resveratrol, as well as its inhibitory kinetics and type of inhibition, were compared. Results: No significant increase in the protein concentration of hepatic CYP1A2 was found in resveratrol-treated rats, but it induces CYP1A2 activity and enhances the induction effect of benzo[a]pyrene. In silico and in vitro experiments demonstrated that resveratrol binds to the active site of human CYP1A2 through hydrophobic interactions with PHE125, PHE226, PHE260, and ALA317, and hydrogen bonds with SER122 and ASP313. It inhibits human recombinant CYP1A2 activity as well as that in rat liver microsomes, with IC₅₀ values of 46 µM and 485 µM, respectively. Resveratrol showed a mixed type of inhibition of recombinant human protein and a competitive inhibition of rat liver microsomal CYP1A. Conclusions: We can conclude that resveratrol is an in vitro inhibitor of CYP1A2, but it increases the benzo[a]pyrene CYP induction effect in vivo. Full article

Ammonia is a key factor in the water, impacting the physiological functions of aquatic organisms. To explore the effect of ammonia stress on mature eggs, female A. baerii at the end of the fourth stage of ovarian development were subjected to varying ammonia concentrations (0 mg/L (control, C), 10 mg/L (low concentration, T1), and 50 mg/L (high concentration, T2)) for 96 h. After 96 h of stress, histological analysis revealed that the follicular membranes of group T1 remained intact and clear compared to group C, although the vacuole fusion had begun. In contrast, the T2 group exhibited ruptured follicular membranes and adhered yolk granules compared to the C group, indicating structural damage. Transcriptome analysis generated 97.89 Gb of clean data, with each sample yielding over 6.09 Gb. A total of 5576, 3719, and 9446 differentially expressed genes (DEG_S) were screened from T1 vs. C, T2 vs. C, and T2 vs. T1 comparisons, respectively. Gene Ontology analysis (GO) functional enrichment analysis showed that DEG_S were significantly enriched in multicellular organism processes (T1 vs. C), cell surface receptor signaling pathways (T2 vs. C), and immune system processes (T2 vs. T1) during biological processes. It indicates that ammonia exposure may enrich cellular components in the extracellular space, potentially disrupting the function of the extracellular matrix. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment indicated significant impacts on amino acid metabolism, particularly glutamate and arginine pathways, as well as key pathways involved in steroid biosynthesis and antioxidation. Weighted gene co-expression network analysis (WGCNA) revealed that a total of 26,369 DEGs were divided into 29 distinct modules, displaying obvious associations with their traits. In the T2 vs. C group, antioxidation-related genes such as GST and GCLM were significantly downregulated, and the expressions of key enzymes for steroid synthesis, such as CYP11A1, CYP17, and CYP19A1 were suppressed, indicating that high ammonia nitrogen concentrations impair oocyte function by inducing oxidative stress and disrupting hormone synthesis. This study provides a comprehensive repertoire of candidate genes associated with ammonia stress in the mature egg of A. baerii, which will be useful for development of sturgeon breeding and reproduction. Full article

Objectives: This study aimed to elucidate the determinants of interindividual variability in the pharmacokinetics of ibrutinib among healthy Chinese subjects, focusing on the influence of demographic characteristics, dietary conditions, and genetic polymorphisms on CYP enzymes and ABC transporters. Methods: Thirty-two participants were randomly assigned to either a fasting (n = 16) or fed (n = 16) group, each receiving a single 140 mg oral dose of ibrutinib. Plasma concentrations were quantified using a validated UPLC–MS/MS method. Genetic polymorphisms in CYP3A4, CYP3A5, CYP2D6, and ABCG2 were identified by Sanger sequencing. Pharmacokinetic parameters, including apparent clearance (CL/F), maximum plasma concentration (Cmax), area under the plasma concentration–time curve (AUC0-t), and time to maximum concentration (Tmax), were estimated by non-compartmental analysis and statistically evaluated for associations with demographic, dietary, and genetic variables. Results: Food intake significantly affected ibrutinib pharmacokinetics, with postprandial administration resulting in reduced CL/F and increased Cmax and AUC0-t (p < 0.01). Gender differences were also observed, as females exhibited higher CL/F, lower Cmax, and AUC0-t than males (p < 0.05). The CYP2D6 c.100C>T polymorphism significantly decreased CL/F and increased exposure in fasting and male subjects (p < 0.05), but this effect was absent under fed conditions. Conversely, the ABCG2 c.421C>A variant was associated with increased CL/F and decreased AUC0-t (p < 0.05), while other genotypes exerted negligible effects. Conclusions: Ibrutinib pharmacokinetics are significantly modulated by dietary status, gender, and genetic polymorphisms, particularly CYP2D6 c.100C>T and ABCG2 c.421C>A. These findings underscore the importance of integrating pharmacogenetic and physiological factors into individualized dosing strategies to optimize therapeutic efficacy and minimize adverse effects. Full article

CYP24A1, a mitochondrial cytochrome P450 enzyme, plays a critical role in the catabolism of active vitamin D metabolites and is a key regulator of local vitamin D signaling in the small intestine. While traditionally studied in the context of renal physiology, increasing evidence highlights its distinct regulatory mechanisms and functional significance within the intestinal epithelium. This review explores the molecular architecture, tissue-specific expression patterns, and multifactorial regulation of CYP24A1 in enterocytes, encompassing nuclear receptor signaling, epigenetic and post-transcriptional control, and environmental influences such as inflammation, diet, and the gut microbiota. We discuss how intestinal CYP24A1 modulates the expression of vitamin D target genes involved in transcellular calcium absorption and epithelial barrier function, and how its dysregulation contributes to gastrointestinal disorders including inflammatory bowel diseases, celiac disease, microbiota dysbiosis, and colorectal cancer. In addition, we examine preclinical and translational evidence supporting CYP24A1 as a potential therapeutic target. Emerging strategies such as selective enzyme inhibitors, microbiota modulation, RNA-based technologies, and personalized supplementation approaches are considered in the context of restoring local vitamin D bioactivity and mineral homeostasis. Together, this review underscores the clinical importance of intestinal CYP24A1 and highlights novel opportunities for targeted interventions in vitamin D-responsive gastrointestinal pathologies. Full article

Mythimna separata (Walker) (Lepidoptera: Noctuidae) is a major migratory pest causing severe damage to cereal crops such as maize, wheat, and rice across Asia, and is also found in many parts of Oceania. With increasing insecticide resistance, botanical alternatives are urgently needed. This study evaluated the insecticidal potential of hypaconitine, a C19-diterpenoid alkaloid from Aconitum coreanum, against M. separata larvae. Hypaconitine exhibited significant stomach toxicity and strong antifeedant activity. It also caused pronounced growth inhibition, prolonged larval and pupal development, reduced pupation and adult emergence, induced morphological deformities, and significantly shortened adult longevity. Crucially, biochemical assays revealed sustained, time- and concentration-dependent upregulation of key detoxification enzymes—carboxylesterase (CarE), glutathione S-transferase (GST), and cytochrome P450 (CYP450)—over 72 h, indicating that hypaconitine imposes severe metabolic fitness costs rather than being readily detoxified. These effects collectively demonstrate that hypaconitine’s insecticidal efficacy arises not only from direct toxicity but also from exploiting the physiological trade-offs inherent in xenobiotic defense. Its multi-modal action—combining larvicidal, antifeedant, growth-regulatory, and metabolism-disrupting effects—presents a novel strategy for bioinsecticide development with a lower risk of resistance evolution. These findings highlight hypaconitine as a promising candidate for sustainable, integrated management of M. separata and other resistant lepidopteran pests. Full article

Cytochrome P450 enzymes (CYPs) are versatile biocatalysts capable of performing selective oxidation reactions valuable for industrial and pharmaceutical applications. However, their catalytic efficiency is often constrained by dependence on costly electron donors, the requirement for redox partners, and uncoupling reactions that divert reducing power toward reactive oxygen species. Improving electron transfer efficiency through optimized redox partner interactions is therefore critical for developing effective CYP-based biocatalysts. In this study, we investigated the interaction between CYP119, a thermophilic CYP from Sulfolobus acidocaldarius, and putidaredoxin (Pdx), the redox partner of P450cam. Using rational design and computational modeling with PyRosetta 3, 14 CYP119 variants were modeled and analyzed by docking simulations on the Rosie Docking Server. Structural analysis identified three key mutations (N34E, D77R, and N34E/D77R) for site-directed mutagenesis. These mutations (N34E, D77R, and N34E/D77R) enhanced Pdx binding affinity by 20-, 3-, and 12-fold, respectively, without affecting substrate binding. Catalytic assays using lauric acid and indirect assays to monitor electron transfer revealed that, despite improved complex formation, the N34E variant showed reduced electron transfer efficiency compared to D77R. These findings highlight the delicate balance between redox partner binding affinity and catalytic turnover, emphasizing that fine-tuning electron transfer interfaces are essential for engineering efficient CYP biocatalysts. Full article

Hypersensitized P2X3 receptor signaling has been described to play a role in several disorders, including chronic cough. The goal of our in vitro and in vivo studies was to investigate the biotransformation and the influence of CYP3A4 inhibition on the pharmacokinetics of the selective P2X3 antagonist filapixant. Metabolic turnover of filapixant in human liver microsomes and hepatocytes was moderate to high, indicating a complex metabolic pattern with mainly oxidative biotransformation. In recombinant CYP enzymes, depletion of filapixant was observed mainly with CYP3A4 and, to a significantly lesser extent, with CYP1A1, 2D6, 2J2, and 3A5. Drug depletion of [³H]filapixant and metabolite formation in human liver microsomes was significantly inhibited in the presence of strong CYP3A4 inhibitors, whereas other CYP isoform–selective inhibitors showed no or very minor effects. Co-administration of multiple daily doses of 200 mg itraconazole with 80 mg filapixant in humans increased the AUC and C_max of filapixant to 4.01 and 1.89-fold, respectively, indicating that filapixant is a moderately sensitive CYP3A4 substrate. Co-administration of itraconazole also prolonged the half-life of filapixant from 12.1 h to 22.8 h. Overall, changes in AUC, C_max, and half-life indicate that both the bioavailability and elimination of filapixant were affected. Filapixant was well tolerated alone and in combination with itraconazole. Full article

Benzo[a]pyrene (B[a]P), a ubiquitous environmental pollutant, poses a significant threat to male reproductive health, but the underlying latent molecular mechanisms remain virtually unknown. This study investigated the effects of embryonic B[a]P exposure on testicular function and spermatogenesis in F0 and F1 adult male medaka (Oryzias latipes). Embryos were exposed to sublethal concentrations (2.5, 20, and 80 μg/L) for 8 days and then raised in clean water until they reached adulthood. Transcriptomic analysis of F0 testicular tissues revealed widespread dysregulation of critical pathways. Exposure impaired the brain–pituitary–gonadal axis by disrupting GnRH signaling and downregulating genes encoding key steroidogenic enzymes (CYP17A1, HSD3B2), indicating suppressed testosterone biosynthesis. Concurrently, pathways essential for cellular energy metabolism (AMPK signaling, insulin signaling), amino acid biosynthesis, and cytoskeletal organization (actin cytoskeleton, focal adhesion) were profoundly altered. Furthermore, B[a]P activated apoptotic pathways and disrupted the balance between cell survival (PI3K-Akt signaling) and death, compromising spermatogenic cell fate. These molecular disruptions manifested in drastic physiological impairments, including a reduced gonadosomatic index, decreased sperm motility, and compromised fertilization success in F0 males, although these effects were recovered in the F1 generation. This study provides a comprehensive molecular basis for the long-term reproductive toxicity of early-life B[a]P exposure. Full article

Propranolol is a non-selective β-adrenergic receptor antagonist widely used in cardiovascular and neurological therapy. Its naphthalene-based structure contributes to its high lipophilicityand central nervous system penetration. Clinically, propranolol is indicated for hypertension, arrhythmias, anxiety, migraine, and other conditions. It undergoes extensive hepatic metabolism via cytochrome P450 enzymes, notably CYP2D6, with a significant first-pass effect limiting oral bioavailability. This review integrates pharmacological profiling with crystallographic analysis to explore propranolol’s molecular interactions and therapeutic versatility. High-resolution crystal structures of the human β₂-adrenergic receptor (hβ₂-AR), particularly PDB ID: 6PS5 obtained via serial femtosecond crystallography (SFX), reveal key binding determinants responsible for receptor affinity and antagonism. Comparative structural analysis with other β-blockers—alprenolol, timolol, and carvedilol—highlights how variations in aromatic and heterocyclic frameworks influence pharmacokinetics and receptor selectivity. Superimposition results (RMSD: 0.032 for propranolol–alprenolol, 0.078 for propranolol–carvedilol, and 1.078 for propranolol–timolol) quantitatively illustrate molecular similarity and divergence. The enantioselective behavior of propranolol is also discussed, with the S-enantiomer showing greater receptor affinity and pharmacological potency than the R-form. Beyond canonical β-adrenergic targets, propranolol interacts with non-canonical proteins such as the cellulase enzyme Cel7A and lactoferrin, suggesting off-target effects and novel therapeutic potential. These findings underscore the importance of propranolol’s amphiphilic character, stereochemistry, and electrostatic properties in shaping its pharmacological profile. Overall, the integration of crystallographic data with pharmacological insights supports the rational design of next-generation β-adrenergic ligands with enhanced selectivity, bioavailability, and clinical efficacy. Full article

Traumatic optic neuropathy (TON) causes vision loss through compression and contusion, yet there is no consensus on the most effective treatment. Polyunsaturated fatty acid (PUFA)-derived bioactive lipids metabolized by lipoxygenase (LOX), cytochrome P450 (CYP), and cyclooxygenase (COX) enzymes are known mediators of inflammation and neurodegeneration. However, their role in TON-related retinal pathology remains unclear. Controlled orbital impact (COI) was used to induce unilateral TON in mice with controlled velocity (2–3 m/s), with the fellow eye serving as an internal control. Retina tissues were collected three days post-injury and analyzed by LC/MS to quantify bioactive lipid metabolites from ω−6 and ω−3 PUFAs. Statistical analysis was performed using paired, nonparametric Wilcoxon signed-rank tests with Benjamini–Hochberg false discovery rate (FDR) correction. Results showed that among 38 reliably detected metabolites, no individual lipid showed a statistically significant difference between TON and control eyes after FDR correction (q < 0.05). However, both individual and pathway-level analysis revealed consistent trends toward increased expression of LOX- and CYP-derived metabolites across FDA PUFA substrates, including arachidonic acid (AA), linoleic acid (LA), and docosahexaenoic acid (DHA). These findings support further investigation into lipid-mediated inflammation in TON and its potential as a therapeutic target, particularly through expanding both the sample size and the post-TON time periods. Full article