The widespread use of QDs raises health and environmental concerns, and the ROS induced oxidative stress is reported as the main mechanism of QDs toxicity. Cytochrome P450 (CYP450) superfamily, the primary enzyme system for metabolizing external compounds in the liver, also generates reactive oxygen species (ROS), making it crucial for detoxification and ROS production. Therefore, we investigated whether QDs could cause liver tissue damage by affecting the activity of CYP450 isoenzymes (CYP1A2, CYP2E1, CYP2D2, and CYP3A1) in liver microsomes, thereby altering ROS generation. This mechanism has not been previously reported. Our experiments indicate that CdTe QDs exhibit a dose/time-dependent relationship with the enzymatic activities of CYP1A2 and CYP2E1, which are closely related to ROS generation. However, an inconsistency was observed between the data for CYP2E1 activity in vivo and in vitro due to the complexity of in vivo regulatory factors. More importantly, in vitro experiments have shown that CdTe QDs can significantly promote the enzymatic activity of CYP1A2. Therefore, we speculate that CdTe QDs may induce ROS generation by enhancing CYP450 enzyme activities. In addition, molecular docking experiments were conducted to illustrate the impact of CdTe QDs on the structure of CYP1A2, leading to functional change (i.e., enzyme activity). These findings suggest a novel mechanism by which CdTe QDs regulate CYP450 activities in liver microsomes, particularly CYP1A2. This may represent a crucial pathway through which CdTe QDs induce excessive ROS generation, leading to oxidative stress and liver damage.

Imidaclothiz (IMZ), an innovative neonicotinoid insecticide, has attracted significant interest due to its environmental persistence and consequent ecological implications. In this research, the white-rot fungus Phanerochaete sordida YK-624 was used to degrade IMZ, unveiling a novel fungal degradation mechanism. The results demonstrated that IMZ was efficiently degraded by P. sordida YK-624. Transcriptomic analysis revealed that IMZ-induced stress triggered a cascade of enzymatic and cellular defense responses that are instrumental in facilitating its biodegradation. Through inhibitor experiments and enzyme activity profiling, cytochrome P450 and manganese peroxidase (MnP) were identified to play crucial roles in IMZ biodegradation. Additionally, three metabolites were isolated and identified by NMR, and two innovative degradation pathways involving hydroxylation and nitro reduction were proposed. Toxicity assessment suggested the reduced environmental risk of IMZ after its degradation by P. sordida YK-624. These findings provided insights into the IMZ degradation mechanism and highlighted the potential of white-rot fungi in neonicotinoid bioremediation.

Organophosphate esters (OPEs) pose significant environmental concerns due to their widespread presence, potential toxicity, and persistence. This study investigated the degradation of the isodecyl diphenyl phosphate (IDDP) and bis-(2-ethylhexyl)-phenyl phosphate (BEHPP) using a novel enrichment culture, which could degrade 85.4 % and 78.2 % of 1 mg/L IDDP and BEHPP after 192 h and 172 h, respectively, under extremely low bacterial dosage (the initial OD600 nm= 0.0075, biomass was approximately 1 mg/L). The identification of intermediate products suggested that the degradation reactions likely included hydrolysis, hydroxylation, methylation, carboxylation, and glycosylation. Metagenomic analysis highlighted the crucial role of enzymes in degrading IDDP and BEHPP, including phosphatase, phosphodiesterase, cytochrome P450, and hydroxylase. Pure strains Burkholderia cepacia ZY1, Sphingopyxis terrae ZY2, and Amycolatopsis ZY3 were isolated, and their efficient individual degradation abilities were confirmed. These efficiencies were lower compared to the enrichment culture, emphasizing the importance of microbial interactions for effective degradation. The pathways identified for these strains illustrated their involvement in different degradation steps, reinforcing the synergy between different degraders. Molecular dynamics simulations provided insights into the interactions between alkaline phosphatase (ALP), cytochrome P450 (CYP450), and hydroxylase with OPEs. These enzymes demonstrated a strong binding capacity with both BEHPP and IDDP, exhibiting distinct binding site preferences that may contribute to varied metabolic pathways. These findings comprehensively reveal the transformation mechanisms of OPEs.

Studies have shown that the gut microbiota may affect anti-tumor immunity by regulating the host immune system and tumor microenvironment. To date, little is known about whether the gut microbiota underlies the occurrence of diffuse large B-cell lymphoma (DLBCL) and drug resistance.

In the present study, we compared the gut microbiota structure of fecal samples from 26 patients with primary DLBCL, 28 patients with relapsed and refractory (RR) DLBCL, and 30 healthy people.

Notably, Fusobacteria (from phylum to species) was enriched in the primary group. A decrease of Fusobacterium and an increase of Enterococcus were found in the RR group. PICRUSt analysis found that genes related to cytochrome P450 were upregulated in the RR group compared to the primary group, which likely contributes to the occurrence of DLBCL and the formation of drug resistance.

Our study provides further evidence for the relationship between gut microbiota and DLBCL and the formation of drug resistance, highlighting the potential significance of the bacterial variations may be used as new biomarkers of DLBCL.

Fulvic acid (FA), a humic substance, has various applications in agricultural (animal husbandry) and pharmaceutical industries. However, to the best of our knowledge, its antitumor effects remain unclear. This study aimed to elucidate the effects and underlying mechanisms of FA in ovarian cancer cells.

To determine the half-maximal inhibitory concentration (IC50) of FA, SK-OV-3 and OVCAR3 ovarian cancer cells were exposed to various concentrations of FA. The effects of FA and expression of cytochrome P450 family 1 subfamily A member 1 (CYP1A1) on cellular proliferation, migration, and invasion were evaluated using the Cell Counting Kit-8 and transwell assays for migration and invasion. Differentially expressed messenger ribonucleic acids (mRNAs) were identified via Illumina ribonucleic acid (RNA) sequencing and verified using fluorescent quantitative reverse transcription polymerase chain reaction (qRT-PCR). CYP1A1 protein levels were measured by western blotting.

The IC50 values of FA for OVCAR3 and SK-OV-3 cells were 689.9 and 752.0 µg/ml, respectively. FA treatment suppressed cell proliferation, invasion, and migration. In FA-treated SK-OV-3 cells, 117 mRNAs were upregulated, and 342 mRNAs were downregulated, as identified by Illumina RNA sequencing. The qRT-PCR results revealed that FA upregulated CYP1A1 expression in both cell lines. CYP1A1 overexpression mimicked the effects of FA treatment on cell proliferation, invasion, and migration. Furthermore, CYP1A1 knockdown alleviated these effects induced by FA treatments.

FA suppressed cell proliferation, invasion, and migration and upregulated CYP1A1 expression in SK-OV-3 and OVCAR3 cells. Our results suggest that FA demonstrates antitumor effects in ovarian cancer cells through CYP1A1 regulation.

Resistance to cancer therapies is increasingly recognized as being influenced by long non-coding RNAs (lncRNAs), which are pivotal in regulating cellular functions and gene expression. Elucidating the intricate relationship between lncRNAs and the mechanisms underlying drug resistance is critical for advancing effective therapeutic strategies. This study offers an in-depth review of the regulatory roles lncRNAs play in various signaling and immunological pathways implicated in cancer chemoresistance. lncRNA-mediated influence on drug resistance-related signaling pathways will be presented, including immune evasion mechanisms and other essential signaling cascades. Furthermore, the interplay between lncRNAs and the immune landscape will be dissected, illustrating their substantial impact on the development of chemoresistance. Overall, the potential of lncRNA-mediated signaling networks as a therapeutic strategy to combat cancer resistance has been highlighted. This review reiterates the fundamental role of lncRNAs in chemoresistance and proposes promising avenues for future research and the development of targeted therapeutic interventions.

Fomesafen is a diphenyl ether herbicide developed by Zeneca Group PLC (UK), mainly used in soybean and peanut fields to control annual and perennial broad-leaved weeds. Fomesafen has strong persistence in the soil, slow degradation rate, and is prone to harm subsequent sensitive crops. This study utilized Autodock molecular docking technology to investigate the binding and interaction between degradation enzyme CYP57A1 and small molecules of fomesafen herbicides. The CYP57A1 gene cloned from a fomesafen-resistant fungus Fusarium verticilloids, belongs to a fragment of the P450 family, contains 587 bases, encodes 190 amino acids, and has an isoelectric point of 5.16. Visualization of the active surface of the protein receptor reveals that fomesafen is located in the cavity formed by the CYP57A1 protein and the cavity is small and tightly, the proteins are connected to small molecules through hydrogen bonds, halogen atom and π - cation interactions. Molecular modification of CYP57A1 enzyme was carried out using virtual amino acid mutation technology. Four key amino acids, LEU143, MET52, PHE176, and GLU177, were subjected to site-specific mutations. This study successfully constructed mutant engineered bacteria with stable protein expression. Mutations (1) MET52 > TRP showed a a decrease in enzyme activity, and the degradation rate of fomesafen was only 7.8 % of the wild-type. It is believed that MET52 is a key active site for the binding of CYP57A1 enzyme to small molecules of fomesafen, playing a crucial role in the degradation of fomesafen by this enzyme. This provides new insights into the impact on the degradation activity of fomesafen.

Cytochrome P450 3A4 (CYP3A4) is a key mediator in xenobiotic metabolism and drug-drug interactions (DDI), developing orally active fluorogenic substrates for sensing and imaging of a target enzyme in biological systems remains challenging. Here, an artificial intelligence (AI)-driven strategy is used to construct a highly specific and orally active fluorogenic substrate for imaging CYP3A4 in complex biological systems. After the fusion of an AI-selected drug-like fragment with a CYP3A4-preferred fluorophore, three candidates are designed and synthesized. Among all evaluated candidates, NFa exhibits excellent isoform-specificity, ultra-high sensitivity, outstanding spatial resolution, favorable safety profiles, and acceptable oral bioavailability. Specifically, NFa excels at functional in situ imaging of CYP3A4 in living systems with exceptional endoplasmic reticulum (ER)-colocalization performance and high imaging resolution, while this agent can also replace hCYP3A4 drug-substrates for high-throughput screening of CYP3A4 inhibitors and for assessing DDI potential in vivo. With the help of NFa, a novel CYP3A4 inhibitor (D13) was discovered, and its anti-CYP3A4 effects are assessed in live cells, ex vivo and in vivo. Collectively, an AI-powered strategy is adapted for developing highly-specific and drug-like fluorogenic substrates, resulting in the first orally available tool (NFa) for sensing and imaging CYP3A4 activities, which facilitates CYP3A4-associated fundamental investigations and the drug discovery process.

The intestinal barrier constitutes the largest surface of the human body communicating with the external environment. Alterations affecting elements of intestinal wall may lead to increased intestinal permeability and resulting translocation of bacteria or its components to the bloodstream in the form of the "leaky gut syndrome" (LGS). One of the most common causes of LGS is the disruption of tight junctions (TJ) maintained by tight junction proteins (TJP). LGS and associated alterations in TJP are observed in numerous gastrointestinal (GI) diseases, including inflammatory bowel diseases (IBD) such as Crohn's disease (CD) and ulcerative colitis (UC). Current literature indicates the key role of LGS in many pathological processes, further emphasizing the need for effective pharmacological approaches to treat this syndrome. One of the potential pharmacological targets in LGS treatment are members of the cytochrome P450 (CYP450) superfamily. By affecting intestinal permeability, they may lead to LGS development. It was found that the expression of CYP8B1 synthesizing cholic acid and CYP26 degrading all-trans retinoic acid indirectly influence TJs. CYP2E1 responsible for the metabolism of a wide variety of chemicals, including ethanol, plays a crucial role in the impairment of the intestinal wall. Contrarily, the overexpression of CYP27B1 has a protective effect on the intestinal integrity. CYP1A1, CYP2A6, CYP2J2 and CYP3A were also suggested to influence the GI tract, through their capability to metabolize serotonin, nicotine, endocannabinoids and gemcitabine, respectively. This review summarizes the findings on the role of CYP450 isoforms in intestinal hyperpermeability and their potential involvement in the pathophysiology of LGS.

Human cytochrome P450 (CYP) enzymes in the brain represent a crucial frontier in neuroscience, with far-reaching implications for drug detoxification, cellular metabolism, and the progression of neurodegenerative diseases. The brain's complex architecture, composed of interconnected cell types and receptors, drives unique neuronal signaling pathways, modulates enzyme functions, and leads to distinct CYP gene expression and regulation patterns compared to the liver. Despite their relatively low levels of expression, brain CYPs exert significant influence on drug responses, neurotoxin susceptibility, behavior, and neurological disease risk. These enzymes are essential for maintaining brain homeostasis, mediating cholesterol turnover, and synthesizing and metabolizing neurochemicals, neurosteroids, and neurotransmitters. Moreover, they are key participants in oxidative stress responses, neuroprotection, and the regulation of inflammation. In addition to their roles in metabolizing psychotropic drugs, substances of abuse, and endogenous compounds, brain CYPs impact drug efficacy, safety, and resistance, underscoring their importance beyond traditional drug metabolism. Their involvement in critical physiological processes also links them to neuroprotection, with significant implications for the onset and progression of neurodegenerative diseases. Understanding the roles of cerebral CYP enzymes is vital for advancing neuroprotective strategies, personalizing treatments for brain disorders, and developing CNS-targeting therapeutics. This review explores the emerging roles of CYP enzymes, particularly those within the CYP1-3 and CYP46 families, highlighting their functional diversity and the pathological consequences of their dysregulation on neurological health. It also examines the potential of cerebral CYP-based biomarkers to improve the diagnosis and treatment of neurodegenerative disorders, offering new avenues for therapeutic innovation.

NADPH-cytochrome P450 reductase (CPR) plays important roles in the metabolism of both endogenous and exogenous compounds through cytochrome P450, and is also involved in the detoxification of insecticides mediated by cytochrome P450. However, the CPR from Apis cerana cerana has not been well characterized and its function is still undescribed. This study isolated the CPR gene from Apis cerana cerana and investigated its functional role in the resistance to pesticide and heavy metal stress. Bioinformatic analysis revealed significant homology between the gene and its counterparts in other species. Functional investigations demonstrated diverse expression and localization patterns of this gene, with AccCPR primarily expressed in muscular tissues and the gut, suggesting its potential roles in flight activities and intestinal barrier function of bees. Furthermore, the expression levels of this gene were significantly modulated under pesticide and heavy metal stress. Notably, the overexpression of AccCPR led to a marked alteration the tolerance to external stressors in E. coli. Additionally, the silencing of the AccCPR gene resulted in a significant decrease in antioxidant enzyme activity and the expression levels of genes associated with antioxidant functions. Consequently, the mortality rate of Apis cerana cerana under imidacloprid stress was significantly elevated. Taken together, our findings suggest that AccCPR may play a pivotal role in the resistance of Apis cerana cerana to abiotic stresses such as pesticides and heavy metals by regulating antioxidant pathways.

C14-functionalized steroids enabled diverse biological activities in anti-gonadotropin and anticancer therapy. However, access to C14-functionalized steroids was impeded by the deficiency of chemical synthetic methods. Recently, several membrane-bound fungal cytochrome P450s (CYPs) have been identified with steroid C14α-hydroxylation activity. However, the lack of efficient heterologous overexpression strategy hampered their further characterization and molecular engineering. In the present study, sequences of fungi-derived CYP genes encoding putative 14α-hydroxylase were selected and bioinformatically analyzed. Substitution of the N-terminal hydrophobic helix by a soluble maltose binding protein tag significantly enhanced the soluble expression level in Escherichia coli. A novel CYP originated from Bipolaris oryzae was discovered with high steroidal C14α-hydroxylation activity when coupled with the redox partner CPRlun. A catalytically self-sufficient chimeric CYP-CPR was built by intramolecular fusion, and the electronic transfer rate was improved. A coenzyme NADPH regeneration system was finally constructed by the co-expression of glucose dehydrogenase. The developed soluble multi-enzyme cascade biotransformation system supported the selective C14α-hydroxylation toward progesterone with a final titer of 34.54 mg/L, the highest level achieved in E. coli-based heterologous expression system. This study provides insightful ideas on the functional expression of fungi-derived CYPs and promises an efficient C14α-hydroxylation system for steroidal drugs through protein engineering.

Calcidiol (25(OH)VD3) and calcitriol (1α,25(OH)2VD3) are active vitamin D3 with high medicinal value, which can maintain calcium and phosphorus balance and treat vitamin D deficiency. Microbial synthesis is an important method to produce high-value-added compounds. It can produce active vitamin D3 through the hydroxylation reaction of P450, which can reduce the traditional chemical synthesis steps, and greatly improve the production efficiency and economic benefits. In this work, Bacillus megaterium H-1 was screened for its ability to produce 25(OH)VD3 and 1α,25(OH)2VD3 from vitamin D3. A new highly inducible vitamin D3 hydroxylase CYP109E1-H was identified from B. megaterium H-1 through searching for transcripts with cytochrome P450 structural domains, combining the transcriptome sequencing with functional expression in Bacillus subtilis WB600. Biotransformation in recombinant B. subtilis confirmed that CYP109E1-H has C-25 hydroxylase activity towards vitamin D3. CYP109E1-H is a natural mutant of CYP109E1 with greater stereoselectivity and it is a new vitamin D3 mono-hydroxylase. The cloning and characterization of the CYP109E1-H gene provide useful information on the structural basis for improving the regional and stereoselectivity of the CYP109E gene.

Cytochrome P450 enzymes catalyze a large number of oxidative transformations that are responsible for natural product synthesis. Recent studies have revealed their unique ability to catalyze the formation of C-N and C-S bonds, broadening their biosynthetic applications. However, the enzymatic mechanisms behind these reactions are still unclear. This review focuses on theoretical insights into the mechanisms of P450-catalyzed C-N and C-S bond formation. The key roles of the conformational dynamics of substrate radicals, influenced by the enzyme environment, in modulating selectivity and reactivity are highlighted. Understanding these reaction mechanisms offers valuable guidance for P450 enzyme engineering and the design of biosynthetic applications.

The heme enzymes of the cytochrome P450 superfamily (CYPs) catalyse the selective hydroxylation of unactivated C-H bonds in organic molecules. There is great interest in applying these enzymes as biocatalysts with a focus on self-sufficient CYP 'fusion' enzymes, comprising a single polypeptide chain with the electron transfer components joined to the heme domain. Here we elucidate the function of the self-sufficient CYP116B46 fusion enzyme, from the thermophilic bacterium Tepidiphilus thermophilus. We demonstrate that it efficiently hydroxylates aromatic organic acids, exemplified by oxidation of 2-hydroxyphenylacetic acid to homogentisic acid (2,5-dihydroxyphenylacetic acid), an important metabolite in bacterial catabolism. In line with the thermophilic nature of the source bacterium, activity increased at higher temperatures, (50 °C), with a catalytic preference for NADPH over NADH. While self-sufficient fusion enzymes simplify biocatalysis; engineered peroxygenase activity is also a key advance in the application of these enzymes as biocatalysts as it eliminates the need for electron transfer partner proteins and nicotinamide cofactors. We demonstrate that a T278E mutation in the heme domain of CYP116B46, confers peroxygenase activity. This engineered peroxygenase enzyme is stable to elevated temperatures and catalytic concentrations of hydrogen peroxide, with an observed optimal activity resulting in a total turnover number of ~650.

Personalized treatment optimization considers individual clinical, genetic, and environmental factors influencing drug efficacy and tolerability. As evidence accumulates, these approaches may become increasingly integrated into standard psychiatric care, potentially transforming the treatment landscape for mental health disorders. While personalized treatment optimization shows promise in enhancing therapeutic outcomes and minimizing adverse effects, further research is needed to establish its clinical utility and cost-effectiveness across various psychiatric disorders. This review examines the potential utility of personalized treatment optimization in psychiatry, addressing the challenge of suboptimal effectiveness and variable patient responses to psychiatric medications. It explores how therapeutic drug monitoring, pharmacogenomics, and biomarker testing can be used to individualize and optimize pharmacotherapy for mental disorders such as depression, bipolar disorder, and schizophrenia.

The landscape of psychiatric care is poised for transformation through the integration of pharmaco-multiomics, encompassing genomics, proteomics, metabolomics, transcriptomics, epigenomics, and microbiomics. This review discusses how these approaches can revolutionize personalized treatment strategies in psychiatry by providing a nuanced understanding of the molecular bases of psychiatric disorders and individual pharmacotherapy responses. With nearly one billion affected individuals globally, the shortcomings of traditional treatments, characterized by inconsistent efficacy and frequent adverse effects, are increasingly evident. Advanced computational technologies such as artificial intelligence (AI) and machine learning (ML) play crucial roles in processing and integrating complex omics data, enhancing predictive accuracy, and creating tailored therapeutic strategies. To effectively harness the potential of pharmaco-multiomics approaches in psychiatry, it is crucial to address challenges such as high costs, technological demands, and disparate healthcare systems. Additionally, navigating stringent ethical considerations, including data security, potential discrimination, and ensuring equitable access, is essential for the full realization of this approach. This process requires ongoing validation and comprehensive integration efforts. By analyzing recent advances and elucidating how different omic dimensions contribute to therapeutic customization, this review aims to highlight the promising role of pharmaco-multiomics in enhancing patient outcomes and shifting psychiatric treatments from a one-size-fits-all approach towards a more precise and patient-centered model of care.

Polychlorinated biphenyls (PCBs), a typical type of persistent organic pollutants (POPs), were previously widely employed as insulating and heat exchange fluids in transformers and capacitors. Despite knowledge of its adverse effects, the precise mechanism underlying PCB77 toxicity remains enigmatic. In this study, we utilized zebrafish as a model organism to explore the toxic effects of various concentrations of PCB77 (10, 200, and 1000 μg/L) and its molecular toxicity mechanisms. Upon exposure to dosages of PCB77 throughout embryonic and larval stages, the zebrafish exhibited adverse phenotypic manifestations, including deformities, decreased heart rates, increased distances between the bulbus arteriosus (BA) and sinus venosus (SV) and reduced locomotor ability. Transcriptome analysis revealed the common enriched pathways across all PCB77 concentration groups, such as retinol metabolism, steroid hormone biosynthesis, and metabolism of xenobiotics by cytochrome P450, which are closely related to the activity of cytochrome P450 (cyp1a) enzymes. Furthermore, Adverse Outcome Pathway (AOP) framework which integrates AOPs and dose-dependent transcriptomics to predict PCB77-induced adverse outcomes (AOs) revealed that aryl hydrocarbon receptor (AhR) associated AOPs triggered by PCB77 exposure may increase early-life stage mortality and decrease cardiac development, indicating that the primary toxic pathways of PCB77 in zebrafish may involve AhR-mediated signaling. Besides, molecular docking modeling demonstrated that PCB77 could bind to the groove within the AhR domain, suggesting that PCB77 induces embryotoxicity in zebrafish through its interaction with AhR. Collectively, these findings not only deliver a thorough examination of PCB77-induced developmental toxicity as well as the underlying mechanisms, but also validate the efficacy of the analytical approach leveraging AOP framework in unraveling toxicity mechanisms of environmental contaminants, which holds promise for risk assessment associated with novel environmental pollutants.

Insects have highly developed olfactory systems in which cytochrome P450s (CYPs) were involved as odor-degrading enzymes throughout the olfactory recognition of odor compounds by insects to avoid continuous stimulation of signaling molecules and thus damage to the olfactory nervous. To understand whether the highly expressed CYPs in the antennae play an olfactory function in Solenopsis invicta worker, in this study, we find six highly expressed antennal CYPs from the transcriptome of S. invicta. Multiple sequence alignment and phylogenetic analysis divided them into two families: the CYP3 family (SinvCYP6K1, SinvCYP6K1-1) and the CYP4 family (SinvCYP4C1, SinvCYP4C1-1, SinvCYP4C1-2, SinvCYP4V2). The expression patterns of these six CYPs were analyzed by RT-qPCR, which revealed that SinvCYP6K1 and SinvCYP4V2 were only highly expressed in the antennae of adult workers. The expression of SinvCYP6K1 and SinvCYP4V2 in workers was markedly diminished after feeding with dsRNA. The electroantennography (EAG) assay demonstrated that the silencing of either SinvCYP6K1 or SinvCYP4V2 resulted in a notable reduction in the EAG response of workers to 2-ethyl-3,6(5)-dimethylpyrazine (EDMP). Furthermore, the trajectory behavior assay showed that the worker's range and speed of movement in response to EDMP significant decreased after the silencing of SinvCYP6K1 and SinvCYP4V2. The findings indicated that both SinvCYP6K1 and SinvCYP4V2 were implicated in the recognition of EDMP by S. invicta.

The presented retrospective analysis suggests, to the best of our knowledge for the first time, a potential significant interaction between statins and niraparib in clinical settings. Nevertheless, further investigations are required to gain a better understanding of the potential clinical benefit.

Background/Objectives: Perillyl alcohol (POH), a monoterpene natural product derived from the essential oils of plants such as perilla (Perilla frutescens), is currently in phase I and II clinical trials as a chemotherapeutic agent. In this study, we investigated the effect of POH on cytochrome P450 (CYP) activity for evaluating POH-drug interaction potential. Methods: The investigation was conducted using pooled human liver microsomes (HLMs), recombinant CYP3A4 (rCYP3A4) enzymes, and human pluripotent stem cell-derived hepatic organoids (hHOs) employing liquid chromatography-tandem mass spectrometry. Results: POH inhibited the activities of CYP2A6 and CYP2B6 with Ki of 6.35 and 3.78 μM, respectively, whereas it stimulated CYP3A4 activity in pooled HLMs incubated with midazolam (MDZ). In a direct CYP inhibition assay using HLMs, activities of CYP2C9, CYP2C19, and CYP2E1 were also inhibited by POH, with IC50 values greater than 50 μM, but those of CYP1A2, CYP2C8, CYP2D6, and CYP3A4 (testosterone) were not significantly inhibited. In pooled HLMs, the Vmax/Km value of 1'-hydroxy MDZ, but not that of 4-hydroxy MDZ, was increased 2.7-fold by 100 μM POH compared with that in the absence of POH. Moreover, stimulation of MDZ 1'-hydroxylation by CYP3A4 was observed in hHOs and rCYP3A4 with cytochrome b5 but not rCYP3A4 without cytochrome b5. Furthermore, activation of CYP3A4-mediated metabolism by POH was observed in HLMs incubated with fimasartan but not atorvastatin, buspirone, donepezil, nifedipine, or tadalafil, suggesting a substrate-dependent activation of CYP3A4 by POH. Conclusions: POH inhibits CYP2A6 and CYP2B6, but it activates CYP3A4. These findings underscore the need for further evaluation of the interactions of clinical drugs with POH.

The cytochrome P450 (CYP) gene superfamily plays a significant role in various physiological processes, producing different compounds such as hormones, fatty acids, and biomolecules. However, little information is known their roles during gonad development in Pacific oyster (Crassostrea gigas). In this study, total of 116 CgCYP (Crassostrea gigas cytochrome P450) genes were identified and their expression pattern was analyzed for the first time. The relative molecular weights of these CgCYP genes ranged from 63.52 to 113.41 kDa, and the length of encoded amino acids ranged from 103 to 993. And total 26 cis-acting elements of these CgCYP genes were identified. GO and KEGG enrichment analysis showed some CgCYP genes are essential for the metabolism of male and female sex hormones. Additionally, expression anslysis showed 69 CgCYP genes were over-expressed in early gonad development and triploid infertile individuals. More importantly, expression levels of CgCYP1, CgCYP15, CgCYP34, CgCYP46, CgCYP69, CgCYP87, CgCYP88, and CgCYP103, were found to be significantly higher in female gonad, suggesting their important roles in female gonad development. The results of this study will provide a better understanding of the CgCYP genes in the gonad development of Pacific oyster.

Regulated cell death (RCD) refers to the form of cell death that can be regulated by various biomacromolecules. Each cell death modalities have their distinct morphological changes and molecular mechanisms. However, intense evidences suggest that lipid peroxidation can be the common feature that initiates and propagates the cell death. Excessive lipid peroxidation alters the property of membrane and further damage the proteins and nucleic acids, which is implicated in various human pathologies. Here, we firstly review the classical chain process of lipid peroxidation, and further clarify the current understanding of the myriad roles and molecular mechanisms of lipid peroxidation in various RCD types. We also discuss how lipid peroxidation involves in diseases and how such intimate association between lipid peroxidation-driven cell death and diseases can be leveraged to develop rational therapeutic strategies.

Cytochrome P450 (CYP450) enzymes, which are widely distributed and pivotal in various biochemical reactions, catalyze diverse processes such as hydroxylation, epoxidation, dehydrogenation, dealkylation, nitrification, and bond formation. These enzymes have been applied in drug metabolism, antibiotic production, bioremediation, and fine chemical synthesis. Recent research revealed that CYP450 catalytic kinetics deviated from the classic Michaelis-Menten model. A notable substrate inhibition phenomenon that affects the catalytic efficiency of CYP450 at high substrate concentrations was identified. However, the substrate inhibition of various reactions catalyzed by CYP450 enzymes have not been comprehensively reviewed. This review describes CYP450 substrate inhibition examples and atypical Michaelis-Menten kinetic models, and provides insight into mechanisms of these enzymes. We also reviewed 3D structure and dynamics of CYP450 with substrate binding. Outline methods for alleviating substrate inhibition in CYP450 and other enzymes, including traditional fermentation approaches and protein engineering modifications. The comprehensive analysis presented in this study lays the foundation for enhancing the catalytic efficiency of CYP450 by deregulating substrate inhibition.

Pulmonary fibrosis (PF) results from excessive extracellular matrix (ECM) deposition and tissue remodeling after activation of fibroblasts into myofibroblasts. Abnormally deposited fibrotic ECM, in turn, promotes fibroblast activation and accelerates loss of lung structure and function. However, the molecular mediators and exact mechanisms by which fibrotic ECM promotes fibroblast activation are unclear. In a bleomycin-induced PF mouse model, we found Galectin-1 (Gal-1) expression was significantly increased in lung tissue, and overexpression of Gal-1 plasmid-transfected fibroblasts were activated into myofibroblasts. Using the decellularization technique to prepare decellularized fibrotic ECM and constructing a 3D in vitro co-culture system with fibroblasts, we found that decellularized fibrotic ECM induced a high expression of Gal-1 and promoted the activation of fibroblasts into myofibroblasts. Therefore, Gal-1 has been identified as a pivotal mediator in PF. Further, we found that decellularized fibrotic ECM delivered mechanical signals to cells through the Gal-1-mediated FAK-Src-P130Cas mechanical signalling pathway, while the CYP450 enzymes (mainly involved in CYP1A1, CYP24A1, CYP3A4, and CYP2D6 isoforms) acted as a chemical signalling pathway to receive mechanical signals transmitted from upstream Gal-1, thereby promoting fibroblast activation. The Gal-1 inhibitor OTX008 or the CYP1A1 inhibitor 7-Hydroxyflavone prevented PF in mice and inhibited the role of fibrotic ECM in promoting fibroblast activation into myofibroblasts, preventing PF. These results reveal novel molecular mechanisms of lung fibrosis formation and identify Gal-1 and its downstream CYP1A1 as potential therapeutic targets for PF disease treatmnts.

Adriamycin (ADR)-induced cardiomyopathy is a common problem in many cancer survivors. Recently, specific chemokine receptors have garnered interest as therapeutic targets in cardiovascular diseases.

This study aim to report the role of C-C chemokine receptor 9 (CCR9)/C-C chemokine ligand 25 (CCL25) and its therapeutic potential in ADR-induced cardiomyopathy.

Functional gene knockout and overexpression mouse models were utilized to investigate the role of CCR9 against ADR-induced cardiomyopathy. Transcriptome sequencing was also performed to identify the downstream molecular mechanisms of CCR9.

This study revealed that CCR9 and CCL25 levels were increased in mice and HL-1 cells injured by ADR, consistent with the results of patients with heart failure. Both in vivo and in vitro, CCR9 overexpression overtly aggravated cardiac dysfunction, accompanied by decreased AMPK activity and increased mitochondrial dysfunction, fibrosis, oxidative stress, and apoptosis. However, the cardiac harmful effects of ADR were reserved by CCR9 knockdown, as well as CCR9 overexpression aggravated cardiotoxicity were reserved by AMPK agonist GSK621. By constructing different domain-missing CCR9 mutants, we suspected that the △4 region of CCR9 is important for AMPK activity. Furthermore, transcriptome sequencing  further illustrated the mechanism of CCR9 overexpression aggravated ADR-induced cardiotoxicity, which was associated with CYP1A1. Finally, lithospermic acid (LA) was screened and alleviated ADR-induced cardiotoxicity through regulation of CCR9/CCL25-AMPK signaling, bolstering CCR9-targeted potential clinical application.

These findings present a promising target and drug for treating chemotherapy-induced cardiotoxicity.

Rice kernel smut caused by the biotrophic basidiomycete fungus Tilletia horrida causes significant yield losses in hybrid rice-growing areas around the world. Cytochrome P450 (CYP) enzyme is a membrane-bound heme-containing monooxygenase. In fungi, CYPs play a role in cellular metabolism, adaptation, pathogenicity, decomposition, and biotransformation of hazardous chemicals. In this study, we identified 20 CYP genes based on complete sequence analysis and functional annotation from the T. horrida JY-521 genome. The subcellular localization, conserved motifs, and structures of these 20 CYP genes were further predicted. The ThCYP genes exhibit differences in gene structures and protein motifs. Subcellular localization showed that they were located in the plasma membrane, cytoplasm, nucleus, mitochondria, and extracellular space, indicating that they had multiple functions. Some cis-regulatory elements related to stress response and plant hormones were found in the promoter regions of these genes. Protein-protein interaction (PPI) analysis showed that several ThCYP proteins interact with multiple proteins involved in the ergosterol pathway. Moreover, the expression of 20 CYP genes had different responses to different infection time points and underwent dynamic changes during T. horrida JY-521 infection, indicating that these genes were involved in the interaction with rice and their potential role in the pathogenic mechanism. These results provided valuable resources for elucidating the structure of T. horrida CYP family proteins and laid an important foundation for further research of their roles in the pathogenesis.

This study focused on exploring the effects of SW033291, an inhibitor of 15-hydroxyprostaglandin dehydrogenase, on type 2 diabetes mellitus (T2DM) mice from a comprehensive perspective. Studies have demonstrated that SW033291 benefits tissue repair, organ function, and muscle mass in elderly mice. Our recent investigation initially reported the beneficial effect of SW033291 on T2DM progression. Herein, we used a T2DM mouse model induced by a high-fat diet and streptozotocin injection. Then, serum and liver metabolomics, as well as liver transcriptomic analyses, were performed to provide a systematic perspective of the SW033291-ameliorated T2DM. The results indicate SW033291 improved T2DM by regulating steroid hormone biosynthesis and linoleic/arachidonic acid metabolism. Furthermore, integrated transcriptomic and metabolomic analyses suggested that key genes and metabolites such as Cyp2c55, Cyp3a11, Cyp21a1, Myc, Gstm1, Gstm3, 9,10-dihydroxyoctadecenoic acid, 11-dehydrocorticosterone, and 12,13-dihydroxy-9Z-octadecenoic acid played crucial roles in these pathways. qPCR analysis validated the significant decreases in the hepatic gene expressions of Cyp2c55, Cyp3a11, Myc, Gstm1, and Gstm3 in the T2DM mice, which were reversed following SW033291 treatment. Meanwhile, the elevated mRNA level of Cyp21a1 in T2DM mice was decreased after SW033291 administration. Taken together, our findings suggest that SW033291 has promising potential in alleviating T2DM and could be a novel therapeutic candidate.

Cytochrome P450 monooxygenases in insects have been verified to implicated in insecticide and phytochemical detoxification metabolism. However, the regulation of P450s, which are modulated by signal-regulated transcription factors (TFs), is less well studied in insects. Here, we found that the Malpighian tubule specific P450 gene SlCYP9A75b in Spodoptera litura is induced by xenobiotics. The transgenic Drosophila bioassay and RNAi results indicated that this P450 gene contributes to α-cypermethrin, cyantraniliprole, and nicotine tolerance. In addition, functional analysis revealed that the MAPKs p38, PI3K/Akt, and JAK-STAT activate the transcription factor fushi tarazu factor 1 (FTZ-F1) to regulate CYP9A75b expression. These findings provide mechanistic insights into the contributions of CYP9A genes to xenobiotic detoxification and support the possible involvement of different signaling pathways and TFs in tolerance to xenobiotics in insects.

Cytochrome P450 monooxygenases are an extensive and unique class of enzymes, which can regio- and stereo-selectively functionalise hydrocarbons by way of oxidation reactions. These enzymes are naturally occurring but have also been extensively applied in a synthesis context, where they are used as efficient biocatalysts. Recently, a biosynthetic pathway where a cytochrome P450 monooxygenase catalyses a critical step of the pathway was uncovered, leading to the production of a number of products that display high antitumour potency. In this work, we use computational techniques to gain insight into the factors that determine the relative yields of the different products. We use conformational search algorithms to understand the substrate stereochemistry. On a machine-learned 3D protein structure, we use molecular docking to obtain a library of favourable poses for substrate-protein interaction. With molecular dynamics, we investigate the most favourable poses for reactivity on a molecular level, allowing us to investigate which protein-substrate interactions favour a given product and thus gain insight into the product selectivity.

Drug-Induced Liver Injury (DILI) and herb Induced Liver Injury (HILI) continues to pose a substantial challenge in both clinical practice and drug development, representing a grave threat to patient well-being. This comprehensive review introduces a novel perspective on DILI and HILI by thoroughly exploring the intricate microenvironment of the liver. The dynamic interplay among hepatocytes, sinusoidal endothelial cells, Kupffer cells, hepatic stellate cells, cholangiocytes, and the intricate vascular network assumes a central role in drug metabolism and detoxification. Significantly, this microenvironment is emerging as a critical determinant of susceptibility to DILI and HILI. The review delves into the multifaceted interactions within the liver microenvironment, providing valuable insights into the complex mechanisms that underlie DILI and HILI. Furthermore, we discuss potential strategies for mitigating drug-induced liver injury by targeting these influential factors, emphasizing their clinical relevance. By highlighting recent advances and future prospects, our aim is to shed light on the promising avenue of leveraging the liver microenvironment for the prevention and mitigation of DILI and HILI. This deeper understanding is crucial for advancing clinical practices and ensuring patient safety in the realm of DILI and HILI.

Polyhalogenated carbazoles (PHCZs) are a class of nitrogen-containing heterocyclic compounds that are widely distributed throughout the marine environment and sediment. These compounds share structural and toxicity similarities with dioxins. However, our understanding of the toxicological effects of PHCZs on marine organisms and their underlying molecular mechanisms remains limited. In this study, we employed the marine model organism Mugilogobius chulae as the experimental subject and selected 2,7-dibromocarbazole (2,7-DBCZ), a compound known for its high toxicity and detection frequency, to conduct both an acute toxicity test and transcriptome analysis on M. chulae embryos. Our findings revealed that the 96 h median lethal concentration (LC50) of 2,7-DBCZ for M. chulae embryos was 174 μg/L, with a median effective concentration (EC50) resulting in pericardial edema deformity of 88.82 μg/L. Transcriptome analysis revealed significant impacts on various systems in M. chulae embryos following exposure to 2,7-DBCZ, including the sensory, cardiovascular, immune, and endocrine systems. Furthermore, this compound perturbed signaling pathways such as phototransduction, protein folding and processing, amino acid metabolism, lipid transport, and exogenous compound metabolism. Notably, transcript abundance of the CYP1A gene associated with the activation of the AhR signaling pathway, similar to dioxin-like compounds, was 18.18 times higher than that in the control group. This observation suggests that M. chulae embryos mount a stress response when exposed to PHCZs. In summary, this study contributes to our understanding of the toxicological implications of PHCZ in marine fish and offers a theoretical foundation for risk assessment and regulatory frameworks for PHCZs in the marine environment.

In the current antiretroviral landscape, continuous efforts are still needed to search for novel chemotypes of human immunodeficiency virus type 1 (HIV-1) inhibitors with improved drug resistance profiles and favorable drug-like properties. Herein, we report the design, synthesis, biological characterization, and druggability evaluation of a class of non-nucleoside reverse transcriptase inhibitors. Guided by the available crystallographic information, a series of novel indolylarylsulfone derivatives were rationally discovered via the substituent decorating strategy to fully explore the chemical space of the entrance channel. Among them, compound 11h bearing the cyano-substituted benzyl moiety proved to be the most effective inhibitor against HIV-1 wild-type and mutant strains (EC50 = 0.0039-0.338 μM), being far more potent than or comparable to etravirine and doravirine. Besides, 11h did not exhibit cytotoxicity at the maximum test concentration. Meanwhile, the binding target of 11h was further confirmed to be reverse transcriptase (IC50 = 0.055 μM). Preliminary structure-activity relationship were discussed to guide further optimization work. Molecular docking and dynamics simulation studies were investigated in detail to rationalize the biological evaluation results. Further drug-likeness assessment indicated that 11h possessed excellent physicochemical properties. Moreover, no apparent hERG blockade liability and cytochrome P450 inhibition were observed for 11h. Notably, 11h was characterized by favorable in vitro metabolic stability with moderate clearance rates and long half-lives in human plasma and liver microsomes. Overall, 11h holds great promise as an ideal Anti-HIV-1 lead compound due to its potent antiviral efficacy, low toxicity, and favorable drug-like profiles.

Arsenite is an important heavy metal. Some Chinese traditional medicines contain significant amounts of arsenite. The aim of this study was to investigate subacute exposure of arsenite on activities of cytochrome P450 enzymes and pharmacokinetic behaviors of drugs in rats. Midazolam, tolbutamide, metoprolol, omeprazole, caffeine, and chlorzoxazone, the probe substrates for cytochrome P450 (CYP) s3A, 2C6, 2D, 2C11, 1A, and 2E, were selected as probe drugs for the pharmacokinetic study. Significant decreases in areas under the curves of probe substrates were observed in rats after consecutive 30-day exposure to As at 12 mg/kg. Microsomal incubation study showed that the subacute exposure to arsenite resulted in little change in effects on the activities of P450 enzymes examined. However, everted gut sac study demonstrated that such exposure induced significant decreases in intestinal absorption of these drugs by both passive diffusion and carrier-mediated transport. In addition, in vivo study showed that the arsenite exposure decreased the rate of peristaltic propulsion. The decreases in intestinal permeability of the probe drugs and peristaltic propulsion rate most likely resulted in the observed decreases in the internal exposure of the probe drugs. Exposure to arsenite may lead to the reduction of the efficiencies of pharmaceutical agents coadministered resulting from the observed drug-drug interactions. SIGNIFICANCE STATEMENT: Exposure to arsenite may lead to the reduction of the efficiencies of pharmaceutical agents coadministered resulting from the observed drug-drug interactions. The present study, we found that P450 enzyme probe drug exposure was reduced in arsenic-exposed animals (areas under the curve) and the intestinal absorption of the drug was reduced in the animals. Subacute arsenic exposure tends to cause damage to intestinal function, which leads to reduced drug absorption.

Sexual dimorphism in immune responses is an essential factor in environmental adaptation. However, the mechanisms involved remain obscure owing to the scarcity of data from sex-role-reversed species in stressed conditions. Benzo[a]pyrene (BaP) is one of the most pervasive and carcinogenic organic pollutants in coastal environments. In this study, we evaluated the potential effects on renal immunotoxicity of the sex-role-reversed lined seahorse (Hippocampus erectus) toward environmental concentrations BaP exposure. Our results discovered the presence of different energy-immunity trade-off strategies adopted by female and male seahorses during BaP exposure. BaP induced more severe renal damage in female seahorses in a concentration-dependent manner. BaP biotransformation and detoxification in seahorses resemble those in mammals. Benzo[a]pyrene-7,8-dihydrodiol-9,10-oxide (BPDE) and 9-hydroxybenzo[a]pyrene (9-OH-BaP) formed DNA adducts and disrupted Ca2+ homeostasis may together attribute the renal immunotoxicity. Sexual dimorphisms in detoxification of both BPDE and 9-OH-BaP, and in regulation of Ca2+, autophagy and inflammation, mainly determined the extent of renal damage. Moreover, the mechanism of sex hormones regulated sexual dimorphism in immune responses needs to be further elucidated. Collectively, these findings contribute to the understanding of sexual dimorphism in the immunotoxicity induced by BaP exposure in seahorses, which may attribute to the dramatic decline in the biodiversity of the genus.

Cytochrome P450 (CYP450) is a group of enzymes that play an essential role in Phase I metabolism, with 57 functional genes classified into 18 families in the human genome, of which the CYP1, CYP2, and CYP3 families are prominent. Beyond drug metabolism, CYP enzymes metabolize endogenous compounds such as lipids, proteins, and hormones to maintain physiological homeostasis. Thus, dysregulation of CYP450 enzymes can lead to different endocrine disorders. Moreover, CYP450 enzymes significantly contribute to fatty acid metabolism, cholesterol synthesis, and bile acid biosynthesis, impacting cellular physiology and disease pathogenesis. Their diverse functions emphasize their therapeutic potential in managing hypercholesterolemia and neurodegenerative diseases. Additionally, CYP450 enzymes are implicated in the onset and development of illnesses such as cancer, influencing chemotherapy outcomes. Assessment of CYP450 enzyme expression and activity aids in evaluating liver health state and differentiating between liver diseases, guiding therapeutic decisions, and optimizing drug efficacy. Understanding the roles of CYP450 enzymes and the clinical effect of their genetic polymorphisms is crucial for developing personalized therapeutic strategies and enhancing drug responses in diverse patient populations.

Insect cytochromes P450 (CYP450s) are key enzymes responsible for a wide array of oxidative transformations of both endogenous and exogenous substrates. However, there is currently no a universal guideline established for heterologous expression of membrane-bound CYP450s, which hampers their downstream biochemical and structural studies. In this study, we conducted large-scale screening of protein overexpression in Escherichia coli using 71 insect CYP450 sequences and optimized the expression of a difficult-to-express CYP450 (CYP6HX3) using eight different optimizations, including selection of host strains and expression vectors, alternative of leader signal peptides, and N-terminal modifications. We confirmed that 1) Only insect CYP450s belonging to the CYP347 family could be expressed with N-terminal fusion of ompA2+ signal peptide in E. coli expression system. 2) E. coli Lemo 21 (DE3) effectively improved the expression of CYP6HX3 in the plasma membrane. 3) A brick-red appearance occurred frequently in the expressed thallus or membrane proteins, but this phenomenon could not necessarily indicate successful overexpression of target CYP450s. These findings provide new insights into the recombinant expression of insect CYP450s in E. coli systems and will facilitate the theoretical approaches for functional expression and production of eukaryotic CYP450s.

Cytochrome P450 (CYP) enzymes play a central role in the elimination of approximately 80% of all clinically used drugs. Differences in CYP enzyme activity between individuals can contribute to interindividual variability in exposure and, therefore, treatment outcome. In vivo CYP enzyme activity could be determined with phenotyping. Currently, (sub)therapeutic doses are used for in vivo phenotyping, which can lead to side effects. The use of microdoses (100 µg) for in vivo phenotyping for CYP enzymes could overcome the limitations associated with the use of (sub)therapeutic doses of substrates. The aim of this review is to provide a critical overview of the application of microdosing for in vivo phenotyping of CYP enzymes. A literature search was performed to find drug-drug interaction studies of CYP enzyme substrates that used microdoses of the respective substrates. A substrate was deemed sensitive to changes in CYP enzyme activity when the pharmacokinetics of the substrate significantly changed during inhibition and induction of the enzyme. On the basis of the currently available evidence, the use of microdosing for in vivo phenotyping for subtypes CYP1A2, CYP2C9, CYP2D6, and CYP2E1 is not recommended. Microdosing can be used for the in vivo phenotyping of CYP2C19 and CYP3A. The recommended microdose phenotyping test for CYP2C19 is measuring the omeprazole area-under-the-concentration-time curve over 24 h (AUC0-24) after administration of a single 100 µg dose. CYP3A activity could be best determined with a 0.1-75 µg dose of midazolam, and subsequently measuring AUC extrapolated to infinity (AUC∞) or clearance. Moreover, there are two metrics available for midazolam using a limited sampling strategy: AUC over 10 h (AUC0-10) and AUC from 2 to 4 h (AUC2-4).

The study of adrenal disorders is a key component of scientific research, driven by the complex innervation, unique structure, and essential functions of the adrenal glands. This review explores the use of non-traditional animal models for studying congenital adrenal hyperplasia. It highlights the advantages, limitations, and relevance of these models, including domestic ferrets, dogs, guinea pigs, golden hamsters, pigs, and spiny mice. We provide a detailed analysis of the histological structure, steroidogenesis pathways, and genetic characteristics of these animal models. The morphological and functional similarities between the adrenal glands of spiny mice and humans highlight their potential as an important avenue for future research.

Late-onset hypogonadism (LOH) is an age-related disease in men characterized by decreased testosterone levels with symptoms such as decreased libido, erectile dysfunction, and depression. Thymus quinquecostatus Celakovski (TQC) is a plant used as a volatile oil in traditional medicine, and its bioactive compounds have anti-inflammatory potential. Based on this knowledge, the present study aimed to investigate the effects of TQC extract (TE) on LOH in TM3 Leydig cells and in an in vivo aging mouse model. The aqueous extract of T. quinquecostatus Celakovski (12.5, 25, and 50 µg/mL concentrations) was used to measure parameters such as cell viability, testosterone level, body weight, and gene expression, via in vivo studies. Interestingly, TE increased testosterone levels in TM3 cells in a dose-dependent manner without affecting cell viability. Furthermore, TE significantly increased the expression of genes involved in the cytochrome P450 family (Cyp11a1, Cyp17a1, Cyp19a1, and Srd5a2), which regulate testosterone biosynthesis. In aging mouse models, TE increased testosterone levels without affecting body weight and testicular tissue weight tissue of an aging animal group. In addition, the high-dose TE-treated group (50 mg/kg) showed significantly increased expression of the cytochrome p450 enzymes, similar to the in vitro results. Furthermore, HPLC-MS analysis confirmed the presence of caffeic acid and rosmarinic acid as bioactive compounds in TE. Thus, the results obtained in the present study confirmed that TQC and its bioactive compounds can be used for LOH treatment to enhance testosterone production.