A dose-response study on functional and transcriptomic effects of FSH on ex vivo mouse folliculogenesis.

Follicle-stimulating hormone (FSH) binds to its membrane receptor (FSHR) in granulosa cells to activate various signal transduction pathways and drive the gonadotropin-dependent phase of folliculogenesis. Poor female reproductive outcomes can result from both FSH insufficiency owing to genetic or non-genetic factors and FSH excess as encountered with ovarian stimulation in assisted reproductive technology (ART), but the underlying molecular mechanisms remain elusive. Herein, we conducted single-follicle and single-oocyte RNA sequencing analysis along with other approaches in an ex vivo mouse folliculogenesis and oogenesis system to investigate the effects of different concentrations of FSH on key follicular events. Our study revealed that a minimum FSH threshold is required for follicle maturation into the high estradiol-secreting preovulatory stage, and such threshold is moderately variable among individual follicles between 5-10 mIU/mL. FSH at 5, 10, 20, and 30 mIU/mL induced distinct expression patterns of follicle maturation-related genes, follicular transcriptomics, and follicular cAMP levels. RNA-seq analysis identified FSH-stimulated activation of G proteins and downstream canonical and novel signaling pathways that may critically regulate follicle maturation, including the cAMP/PKA/CREB, PI3K-AKT/FOXO1, and glycolysis pathways. High FSH at 20 and 30 mIU/mL resulted in non-canonical FSH responses including premature luteinization, high production of androgen and proinflammatory factors, and reduced expression of energy metabolism-related genes in oocytes. Together, this study improves our understanding of gonadotropin-dependent folliculogenesis and provides crucial insights into how high doses of FSH used in ART may impact follicular health, oocyte quality, pregnancy outcome, and systemic health.

Sluggish and Ion-Resilient Behavior of Interfacial Aqueous Layer on Single-Layer Graphene Oxide: Insights from In Situ Atomic Force Microscopy.

Understanding interfacial interactions of graphene oxide (GO) is important to evaluate its colloidal behavior and environmental fate. Single-layer GO is the fundamental unit of GO colloids, and its interfacial aqueous layers critically dictate these interfacial interactions. However, conventional techniques like X-ray diffraction are limited to multilayer systems and are inapplicable to single-layer GO. Therefore, our study employed atomic force microscopy to precisely observe the in situ dynamic behaviors of interfacial aqueous layers on single-layer GO. The interfacial aqueous layer height was detected at the subnanometer level. In real-time monitoring, the single-layer height increased from 1.17 to 1.70 nm within 3 h immersion. This sluggish process is different from the rapid equilibration of multilayer GO in previous studies, underscoring a gradual transition in hydration kinetics. Ion strength exhibited negligible influence on the single-layer height, suggesting a resilient response of the interfacial aqueous layer to ion-related perturbations due to intricate ion interactions and electrical double-layer compression. Humic acid led to a substantial increase in the interfacial aqueous layers, improving the colloidal stability of GO and augmenting its potential for migration. These findings hold considerable significance regarding the environmental behaviors of the GO interfacial aqueous layer in ion- and organic-rich water and soil.

Bisphenols and brominated bisphenols induced endothelial dysfunction via its disruption of endothelial nitric oxide synthase.

Emerging literatures have concentrated on the association between cardiovascular diseases risk of typical endocrine disruptor bisphenols, which also put forward the further studies need respect to the potential mechanism. Herein, we investigated the endothelial dysfunction effects of bisphenols and brominated bisphenols involved in aortic pathological structure, endothelial nitric oxide synthase (eNOS) protein phosphorylation, synthase activity and nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs) and C57BL/6 mice. Bisphenol A (BPA) and bisphenol S (BPS) increased NO production by 85.7% and 68.8% at 10-6 M level in vitro and 74.3%, 41.5% in vivo, respectively, while tetrabromobisphenol S (TBBPS) significantly inhibited NO by 55.7% at 10-6 M in vitro and 28.9% in vivo at dose of 20 mg/kg BW/d. Aortic transcriptome profiling revealed that the process of 'regulation of NO mediated signal transduction' was commonly induced. The mRNA and protein expression of phosphorylated eNOS at Ser1177 were promoted by BPA and BPS but decreased by TBBPA and TBBPS in HUVECs. Phosphorylation and enzymatic activity of eNOS were significantly increased by 43.4% and 13.8% with the treatment of BPA and BPS at 10-7 M, but decreased by 16.9% after exposure to TBBPS at 10-6 M in vitro. Moreover, only TBBPS was observed to increase aorta thickness significantly in mice and induce endothelial dysfunction. Our work suggests that bisphenols and brominated bisphenols may exert adverse outcome on vascular health differently in vitro and in vivo, and emphasizes areas of public health concern similar endocrine disruptors vulnerable on the vascular endothelial function.

Revolutionizing Solid-State NASICON Sodium Batteries: Enhanced Ionic Conductivity Estimation through Multivariate Experimental Parameters Leveraging Machine Learning.

Na superionic conductor (NASICON) materials hold promise as solid-state electrolytes due to their wide electrochemical stability and chemical durability. However, their limited ionic conductivity hinders their integration into sodium-ion batteries. The conventional approach to electrolyte design struggles with comprehending the intricate interactions of factors impacting conductivity, encompassing synthesis parameters, structural characteristics, and electronic descriptors. Herein, we explored the potential of machine learning in predicting ionic conductivity in NASICON. We compile a database of 211 datasets, covering 160 NASICON materials, and employ facile descriptors, including synthesis parameters, test conditions, molecular and structural attributes, and electronic properties. Random forest (RF) and neural network (NN) models were developed and optimized, with NN performing notably better, particularly with limited data (R2=0.820). Our analysis spotlighted the pivotal role of Na stoichiometric count in ionic conductivity. Furthermore, the NN algorithm highlighted the comparable significance of synthesis parameters to structural factors in determining conductivity. In contrast, the impact of electronegativity on doped elements appears less significant, underscoring the importance of dopant size and quantity. This work underscores the potential of machine learning in advancing NASICON electrolyte design for sodium-ion batteries, offering insights into conductivity drivers and a more efficient path to optimizing materials.

Long-Term Exposure of Graphene Oxide Suspension to Air Leading to Spontaneous Radical-Driven Degradation.

Understanding the environmental transformation and fate of graphene oxide (GO) is critical to estimate its engineering applications and ecological risks. While there have been numerous investigations on the physicochemical stability of GO in prolonged air-exposed solution, the potential generation of reactive radicals and their impact on the structure of GO remain unexplored. In this study, using liquid-PeakForce-mode atomic force microscopy and quadrupole time-of-flight mass spectroscopy, we report that prolonged exposure of GO to the solution leads to the generation of nanopores in the 2D network and may even cause the disintegration of its bulk structure into fragment molecules. These fragments can assemble themselves into films with the same height as the GO at the interface. Further mediated electrochemical analysis supports that the electron-donating active components of GO facilitate the conversion of O2 to •O2- radicals on the GO surface, which are subsequently converted to H2O2, ultimately leading to the formation of •OH. We experimentally confirmed that attacks from •OH radicals can break down the C-C bond network of GO, resulting in the degradation of GO into small fragment molecules. Our findings suggest that GO can exhibit chemical instability when released into aqueous solutions for prolonged periods of time, undergoing transformation into fragment molecules through self-generated •OH radicals. This finding not only sheds light on the distinctive fate of GO-based nanomaterials but also offers a guideline for their engineering applications as advanced materials.

Associations between exposure to cadmium, lead, mercury and mixtures and women's infertility and long-term amenorrhea.

BACKGROUND: Cadmium (Cd), lead (Pb), and mercury (Hg) have been shown to exhibit endocrine disrupting properties. Their effects on women's reproductive health, however, remain elusive. Here, we investigated associations between blood concentrations of Pb, Cd, Hg, and their mixture and infertility and long-term amenorrhea in women aged 20-49 years using the US National Health and Nutrition Examination Survey (NHANES) 2013-2018 cross-sectional survey. METHODS: A total of 1,990 women were included for the analysis of infertility and 1,919 women for long-term amenorrhea. The methods of log-transformation and use of quartiles were used to analyze blood heavy metal concentrations. Statistical differences in the covariates between the outcome groups were evaluated using a chi-squared test for categorical variables and a t-test for continuous variables. Multiple logistic regression models were used to examine the associations. RESULTS: The blood concentrations of Pb and heavy metal mixtures were significantly higher in ever-infertile women than pregnant women, but the concentrations of Cd and Hg were comparable. After full adjustment, multiple logistic regression analyses revealed a significant and dose-dependent positive association between blood Pb concentrations and women's historical infertility, a negative association between Cd and women's long-term amenorrhea, and no associations between Hg and heavy metal mixture and women's infertility or long-term amenorrhea. CONCLUSIONS: Our study suggests that exposure to heavy metals exhibit differential associations with history of infertility and amenorrhea, and Pb may adversely impact women's reproduction and heighten the risks of infertility and long-term amenorrhea.

Benzophenone-1 induced aberrant proliferation and metastasis of ovarian cancer cells via activated ERα and Wnt/ß-catenin signaling pathways.

Benzophenone-1 (BP-1) belongs to personal care product-related contaminants of emerging concern and has been recently reported to induce xenoestrogenic effects. However, the underlying mechanisms leading to the activation of target receptors and subsequent various adverse outcomes remain unclear, which is beneficial to safety and health risk assessment of benzophenone-type ultraviolet filters with their widespread occurrence. Herein, we investigated disrupting effects of BP-1 at environmentally relevant concentrations (10-9-10-6 M) on estrogen receptor (ER) α-associated signaling pathways. Molecular dynamics simulations together with yeast-based assays revealed the steady binding of BP-1 to ERα ligand binding domain (LBD) and hence the observed agonistic activity. BP-1 triggered interaction between ERα and ß-catenin in human SKOV3 ovarian cancer cells and caused translocation of ß-catenin from the cytoplasm to the nucleus, leading to aberrant activation of Wnt/ß-catenin. BP-1 consequently induced dissemination of SKOV3 via regulating epithelial-mesenchymal transitions (EMT) biomarkers including minimally downregulating ZO-1 gene to 78.0 ± 10.1% and maximally upregulating MMP9 gene to 144.1 ± 29.7% and promoted 1.03-1.83 fold proliferation, migration and invasion of SKOV3. We provide the first evidence that the BP-1 activated ERα triggers crosstalk between ERα and Wnt/ß-catenin pathway, leading to the abnormal stimulation and progression of SKOV3 cancer cells.

Endothelial barrier dysfunction induced by anthracene and its nitrated or oxygenated derivatives at environmentally relevant levels.

Polycyclic aromatic hydrocarbons (PAHs) are epidemiologically associated with cardiovascular diseases characterized by early key events involving in the disruption of endothelial barrier function. Whether PAHs can induce adverse cardiovascular outcome by directly destabilizing endothelial barrier function remains elusive. Herein, we investigated the effect of anthracene (ANT), 9-nitroanthracene (9-NANT), and 9,10-anthraquinone (9,10-AQ) on vascular endothelial barrier functions in human umbilical vein endothelial cells (HUVECs). The integrity of endothelial barrier in HUVECs was disturbed with a 1.15-1.42 fold increase in fluorescein leakage, and 21.8%-58.3% downregulated transendothelial electrical resistance. ANT, 9-NANT and 9,10-AQ promoted paracellular gap formation as revealed by transmission electron microscope. The disrupted cell junctions after 24 h exposure to ANT, 9-NANT and 9,10-AQ at 0.01 µM were indicated by the downregulated mRNA expression of vascular endothelial cadherin (VE-cadherin), zona occludens-1 (ZO-1) and occludin by 33.2%-71.4%, 19.1%-21.0%, and 31.9% respectively, and the downregulated protein expression of ZO-1 and occludin, and by the internalization of VE-cadherin. We demonstrated that ANT and its derivatives at environmentally relevant concentrations induced endothelial barrier dysfunction via the disruption of cell junctions, providing essential in vitro evidence on the association with their adverse cardiovascular outcomes.

Carcinogenic Risk of 2,6-Di-tert-Butylphenol and Its Quinone Metabolite 2,6-DTBQ Through Their Interruption of RARß: In Vivo, In Vitro, and In Silico Investigations.

Thousands of contaminants are used worldwide and eventually released into the environment, presenting a challenge of health risk assessment. The identification of key toxic pathways and characterization of interactions with target biomacromolecules are essential for health risk assessments. The adverse outcome pathway (AOP) incorporates toxic mechanisms into health risk assessment by emphasizing the relationship among molecular initiating events (MIEs), key events (KEs), and adverse outcome (AO). Herein, we attempted the use of AOP to decipher the toxic effects of 2,6-di-tert-butylphenol (2,6-DTBP) and its para-quinone metabolite 2,6-di-tert-butyl-1,4-benzoquinone (2,6-DTBQ) based on integrated transcriptomics, molecular modeling, and cell-based assays. Through transcriptomics and quantitative real-time PCR validation, we identified retinoic acid receptor ß (RARß) as the key target biomacromolecule. The epigenetic analysis and molecular modeling revealed RARß interference as one MIE, including DNA methylation and conformational changes. In vitro assays extended subsequent KEs, including altered protein expression of p-Erk1/2 and COX-2, and promoted cancer cell H4IIE proliferation and metastasis. These toxic effects altogether led to carcinogenic risk as the AO of 2,6-DTBP and 2,6-DTBQ, in line with chemical carcinogenesis identified from transcriptome profiling. Overall, our simplified AOP network of 2,6-DTBP and 2,6-DTBQ facilitates relevant health risk assessment.

Transcriptome aberration in mice uterus associated with steroid hormone response and inflammation induced by dioxybenzone and its metabolites.

Benzophenone-type UV filters have been implicated in multiple adverse reproductive outcomes, yet the underlying processes and molecular targets on the female reproductive tract remain largely unknown. Herein, we investigated the effect of dioxybenzone, one of the widely used congeners, and its demethylated (M1) and hydroxylated (M2) metabolites on transcriptome profiles of ICR mice uterus and identified potential cellular targets in human endometrial stromal cells (HESCs) separated from normal endometrium tissues. Dioxybenzone, M1 and M2 (20 mg/kg bw/d) significantly induced transcriptome aberration with the induction of 683, 802, and 878 differentially expressed genes mainly involved in cancer, reproductive system disease and inflammatory disease. Compared to dioxybenzone, M1 and M2 exhibited a transcriptome profile more similar to estradiol in mice uterus, and subsequently promoted thicker endometrial columnar epithelial layer through upregulation of estrogen receptor target genes-Sprr2s. Dioxybenzone, M1 and M2 (0.1 or 1 µM) also exhibited estrogenic disrupting effect via increasing the mRNA expressions and production of the growth factors responsible for epithelial proliferation, including Fgfs and Igf-1 in HESCs. Additionally, the mRNA expressions of several inflammatory cytokines especially IL-1ß in mice uterus and HESCs was significantly upregulated by dioxybenzone and its metabolites. Overall, we revealed that dioxybenzone and its metabolites triggered transcriptome perturbation dually associated with abnormal steroid hormone response and inflammation, both as key determinants to reproductive health risks.

Enhanced Disrupting Effect of Benzophenone-1 Chlorination Byproducts to the Androgen Receptor: Cell-Based Assays and Gaussian Accelerated Molecular Dynamics Simulations.

Benzophenone-1 (BP-1), one of the commonly used ultraviolet filters, has caused increasing public concern due to frequently detected residues in environmental and recreational waters. Its susceptibility to residual chlorine and the potential to subsequently trigger endocrine disruption remain unknown. We herein investigated the chlorination of BP-1 in swimming pool water and evaluated the endocrine disruption toward the human androgen receptor (AR). The structures of monochlorinated (P1) and dichlorinated (P2) products were separated and characterized by mass spectrometry and 1H-1H NMR correlation spectroscopy. P1 and P2 exhibited significantly higher antiandrogenic activity in yeast two-hybrid assays (EC50, 6.13 µM and 9.30 µM) than did BP-1 (12.89 µM). Our 350 ns Gaussian accelerated molecular dynamics simulations showed the protein dynamics in a long-time scale equilibrium, and further energy calculations revealed that although increased hydrophobic interactions are primarily responsible for enhanced binding affinities between chlorinated products and the AR ligand binding domain, the second chloride in P2 still hinders the complex motion because of the solvation penalty. The mixture of BP-1-P1-P2 elicited additive antiandrogenic activity, well fitted by the concentration addition model. P1 and P2 at 1 µM consequently downregulated the mRNA expression of AR-regulated genes, NKX3.1 and KLK3, by 1.7-9.1-fold in androgen-activated LNCaP cells. Because chlorination of BP-1 occurs naturally by residual chlorine in aquatic environments, our results regarding enhanced antiandrogenic activity and disturbed AR signaling provided evidence linking the use of personal care products with potential health risks.

2,6-Di-tert-butylphenol and its quinone metabolite trigger aberrant transcriptional responses in C57BL/6 mice liver.

2,6-Di-tert-butylphenol (2,6-DTBP) is used as an antioxidant with wide commercial applications and its residues have been detected in various environmental matrices. 2,6-DTBP may enter human body via ingestion, inhalation or other exposure pathways. However, its susceptibility to biotransformation and potential of the metabolic products to trigger aberrant transcriptional responses remain unclear. Here, we investigated in vitro and in vivo biotransformation of 2,6-DTBP and characterized the RNA-Seq based transcriptional profiling of C57BL/6 mice liver after the exposure to 2,6-DTBP and its metabolites. 2,6-DTBP was metabolized into hydroxylated (2,6-DTBH) and para-quinone (2,6-DTBQ) products with residues detected in serum and liver of C57BL/6 mice. 2,6-DTBP and 2,6-DTBQ induced the aberrant transcription in C57BL/6 mice liver featured with 373-2861 differentially expressed genes (DEGs). They also up-regulated 1.09-2.92 fold mRNA expression of carcinogenesis-related genes such as Ccnd1, TGFß1 and FOS in C57BL/6 mice liver. Our study indicated potential carcinogenic risk of 2,6-DTBP and its metabolites, beneficial to further evaluation of health risk of TBPs-related contaminants.

Dioxybenzone triggers enhanced estrogenic effect via metabolic activation: in silico, in vitro and in vivo investigation.

Dioxybenzone is widely used in cosmetics and personal care products and frequently detected in multiple environmental media and human samples. However, the current understanding of the metabolic susceptibility of dioxybenzone and the potential endocrine disruption through its metabolites in mimicking human estrogens remains largely unclear. Here we investigated the in vitro metabolism of dioxybenzone, detected the residue of metabolites in rats, and determined the estrogenic disrupting effects of these metabolites toward estrogen receptor α (ERα). In vitro metabolism revealed two major metabolites from dioxybenzone, i.e., M1 through the demethylation of methoxy moiety and M2 through hydroxylation of aromatic carbon. M1 and M2 were both rapidly detected in rat plasma upon exposure to dioxybenzone, which were then distributed into organs of rats in the order of livers > kidneys > uteri > ovaries. The 100 ns molecular dynamics simulation revealed that M1 and M2 formed hydrogen bond to residue Leu387 and Glu353, respectively, on ERα ligand binding domain, leading to a reduced binding free energy. M1 and M2 also significantly induced estrogenic effect in comparison to dioxybenzone as validated by the recombinant ERα yeast two-hybrid assay and uterotrophic assay. Overall, our study revealed the potential of metabolic activation of dioxybenzone to induce estrogenic disrupting effects, suggesting the need for incorporating metabolic evaluation into the health risk assessment of benzophenones and their structurally similar analogs.

Pentabromoethylbenzene Exposure Induces Transcriptome Aberration and Thyroid Dysfunction: In Vitro, in Silico, and in Vivo Investigations.

Pentabromoethylbenzene (PBEB), as one of the novel brominated flame retardants (NFBRs), has caused increasing public concern for health risks. Till now, information regarding potential effects of PBEB on thyroid function remains unclear. Herein, we investigated thyroid disruption of PBEB in vitro and in silico and evaluated thyroid dysfunction induced by PBEB using Sprague-Dawley rats. PBEB showed thyroid receptor (TR) ß antagonistic activity with IC50 of 9.82 × 10-7 M in the dual-luciferase reporter gene assay and induced relative reorientation of helix 11 (H11) and H12 of the TR ligand binding domain as revealed by molecular dynamics simulations. PBEB (0.2, 2, 20 mg/kg BW/d) markedly altered the transcriptome profile of thyroid with induction of 17, 42, and 119 differentially expressed genes (DEGs) involved in thyroid hormone signaling and synthesis pathway, of which transthyretin and albumin are common DEGs. The 28-d exposure to PBEB significantly decreased the triiodothyronine level (from 7.23 to 5.67 ng/mL) and increased the thyrotropin level (from 7.88 to 12.86 mU/L) for female rats. PBEB consequently reduced thyroid weight and altered its morphology with more depleted follicles. Overall, our study provides the first account of evidence on PBEB exerted thyroid disruption, transcriptome aberration, and morphological alteration, facilitating health risk assessment of PBEB and structurally related NBFRs.

Metabolic Susceptibility of 2-Chlorothioxanthone and Its Toxic Effects on mRNA and Protein Expression and Activities of Human CYP1A2 and CYP3A4 Enzymes.

Thioxanthones (TXs) are photoinitiators widely used in UV curable resins and food packaging, and their residues have been frequently detected in human bodies. Our current understanding of the susceptibility of residual TXs to metabolism and their effects on human health is very limited. The in vitro metabolism of TXs and its toxic effects on cytochrome P450 (CYP) (the key xenobiotic metabolizing enzymes) were examined in this study. 2-Chlorothioxanthone (2-Cl-TX) significantly inhibited the enzymatic activities of CYP1A2 and CYP3A4 with IC50 of 8.36 and 0.86 µM, respectively. The exposure to 2-Cl-TX at 2.5 µM up-regulated the mRNA expression of CYP1A2 and CYP3A4 in human hepatocellular carcinoma cells to 3.03-fold and 2.02-fold, respectively. 2-Cl-TX at 2.5 µM caused 2.19-fold and 1.98-fold overexpression of CYP1A2 and CYP3A4, respectively. In vitro studies revealed that 2-Cl-TX was biotransformed into two metabolites through the sulfoxidation of the sulfur atom, or via the hydroxylation of aromatic carbon. Results from this study, including the metabolic susceptibility of residual 2-Cl-TX, the proposed metabolites and the significant toxic effect on the activities, mRNA, and protein expression of CYP1A2 and CYP3A4, are vital to the human health and safety risk assessment from this ubiquitous xenobiotic.

Thyroid Disruption by Bisphenol S Analogues via Thyroid Hormone Receptor ß: in Vitro, in Vivo, and Molecular Dynamics Simulation Study.

Bisphenol S (4-hydroxyphenyl sulfone, BPS) is increasingly used as a bisphenol A (BPA) alternative. The global usage of BPS and its analogues (BPSs) resulted in the frequent detection of their residues in multiple environmental media. We investigated their potential endocrine-disrupting effects toward thyroid hormone receptor (TR) ß. The molecular interaction of BPSs toward TRß ligand binding domain (LBD) was probed by fluorescence spectroscopy and molecular dynamics (MD) simulations. BPSs caused the static fluorescence quenching of TRß LBD. The 100 ns MD simulations revealed that the binding of BPSs caused significant changes in the distance between residue His435 at helix 11(H11) and residue Phe459 at H12 in comparison to no ligand-bound TRß LBD, indicating relative repositioning of H12. The recombinant two-hybrid yeast assay showed that tetrabromobisphenol S (TBBPS) and tetrabromobisphenol A (TBBPA) have potent antagonistic activity toward TRß, with an IC10 of 10.1 and 21.1 nM, respectively. BPS and BPA have the antagonistic activity with IC10 of 312 and 884 nM, respectively. BPSs significantly altered the expression level of mRNA of TRß gene in zebrafish embryos. BPS and TBBPS at environmentally relevant concentrations have antagonistic activity toward TRß, implying that BPSs are not safe BPA alternatives in many BPA-free products. Future health risk assessments for TR disruption and other adverse effects should focus more on the structure-activity relationship in the design of environmentally benign BPA alternatives.

Binding Specificity Determines the Cytochrome P450 3A4 Mediated Enantioselective Metabolism of Metconazole.

Cytochrome P450 3A4 (CYP3A4) is a promiscuous enzyme, mediating the biotransformations of ∼50% of clinically used drugs, many of which are chiral molecules. Probing the interactions between CYP3A4 and chiral chemicals is thus essential for the elucidation of molecular mechanisms of enantioselective metabolism. We developed a stepwise-restrained-molecular-dynamics (MD) method to model human CYP3A4 in a complex with cis-metconazole (MEZ) isomers and performed conventional MD simulations with a total simulation time of 2.2 µs to probe the molecular interactions. Our current study, which employs a combined experimental and theoretical approach, reports for the first time on the distinct conformational changes of CYP3A4 that are induced by the enantioselective binding of cis-MEZ enantiomers. CYP3A4 preferably metabolizes cis-RS MEZ over the cis-SR isomer, with the resultant enantiomer fraction for cis-MEZ increasing rapidly from 0.5 to 0.82. cis-RS MEZ adopts a more extended structure in the active pocket with its Cl atom exposed to the solvent, whereas cis-SR MEZ sits within the hydrophobic core of the active pocket. Free-energy-perturbation calculations indicate that unfavorable van der Waals interactions between the cis-MEZ isomers and the CYP3A4 binding pocket predominantly contribute to their binding-affinity differences. These results demonstrate that binding specificity determines the cytochrome P450 3A4 mediated enantioselective metabolism of cis-MEZ.

Side Chains of Parabens Modulate Antiandrogenic Activity: In Vitro and Molecular Docking Studies.

Parabens have been widely used in packaged foods, pharmaceuticals, and personal-care products. Considering their potential hydrolysis, we herein investigated structural features leading to the disruption of human androgen receptor (AR) and whether hydrolysis could alleviate such effects using the recombinant yeast two-hybrid assay. Parabens with an aryloxy side chain such as benzyl paraben and phenyl paraben have the strongest antiandrogenic activity. The antiandrogenic activity of parabens with alkyloxyl side chains decreases as the side chain length increases from 1 to 4, and no antiandrogenic effect occurred for heptyl, octyl, and dodecyl parabens with the number of alkoxyl carbon atoms longer than 7. The antiandrogenic activity of parabens correlates significantly with their binding energies (R2 = 0.84, p = 0.01) and were completely diminished after the hydrolysis, particularly for parabens with aryloxy side chains. The Km for the hydrolysis of parabens with aromatic moiety side chain is 1 order of magnitude higher than that of the parabens with alkyl side chains. Both in vitro and in silico data, for the first time, suggest parabens with aromatic side chains are less prone to hydrolysis. Our results provide an insight into risk of various paraben and considerations for design of new paraben-related substitutes.