Estrogenic effects of fluorinated diiodine alkanes in MCF‐7 cells,H295R cells and zebrafish embryo assays

Fluorinated diiodine alkanes (FDIAs), important industrial intermediates in the synthesis of various perfluorinated compounds, which are distributed widely in wildlife and humans. Recent studies showed that FDIAs had in vitro estrogenic effects. However, to date, little information is available regarding the in vivo estrogenic effects of FDIAs and the mechanisms are unclear. In this study, a combination of in vitro and in vivo assays was used to investigate the estrogenic effects of FDIAs. We tested the in vitro estrogenic effects and estrogen receptor‐related gene expression via MCF‐7 cell assay. The hormone level of estradiol and the expression of estrogenic synthesis genes were measured in the H295R cell assay. Finally, the in vivo effects of FDIAs on development and estrogen‐related gene expression were assessed in the zebrafish embryos assay. The results demonstrated that FDIAs could exhibit estrogenic activity through inducing cell proliferation (1.6‐6.7‐fold of the control) and estrogen receptor alpha gene expression (1.07‐1.39‐fold of the control), altering estradiol production (1.14‐1.22‐fold of the control) and the major estrogenic synthesis gene expression of CYP19 (1.22‐1.31‐fold of the control), disrupting the estrogen‐related genes (esr1 and cyp19b) levels in zebrafish (1.52‐2.99‐fold and 2.95‐5.00‐fold of the control for esr1 and cyp19b, respectively). The current findings indicated the potential estrogenic effects of FDIAs and provided novel information for human risk assessment.     

Developmental toxicity,EROD, and CYP1A mRNA expression in zebrafish embryos exposed to dioxin‐like PCB126

Dioxin‐like PCB126 is a persistent organic pollutant that causes a range of syndromes including developmental toxicity. Dioxins have a high affinity for aryl hydrocarbon receptor (AhR) and induce cytochrome P4501A (CYP1A). However, the role of CYP1A activity in developmental toxicity is less clear. To better understand dioxin induced developmental toxicity, we exposed zebrafish (Danio rerio) embryos to PCB126 at concentrations of 0, 16, 32, 64, and 128 μg L^?1 from 3‐h post‐fertilization (hpf) to 168 hpf. The embryonic survival rate decreased at 144 and 168 hpf. The fry at 96 hpf displayed gross developmental malformations, including pericardial and yolk sac edema, spinal curvature, abnormal lower jaw growth, and non‐inflated swim bladder. The pericardial and yolk sac edema rate significantly increased and the heart rate declined from 96 hpf compared with the controls. PCB126 did not alter the hatching rate. To elucidate the mechanism of PCB126‐induced developmental toxicity, we conducted ethoxyresorufin‐O‐deethylase (EROD) in vivo assay to determine CYP1A enzyme activity, and real‐time PCR to study the induction of CYP1A mRNA gene expression in embryo/larval zebrafish at 24, 72, 96, and 132 hpf. In vivo EROD activity was induced by PCB126 at 16 μg L^?1 concentration as early as 72 hpf but significant increases were observed only in zebrafish exposed to 64 and 128 μg L^?1 doses (p < 0.005) at 72, 96, and 132 hpf. Induction of CYP1A mRNA expression was significantly upregulated in zebrafish exposed to 32 and 64 μg L^?1 at 24, 72, 96, and 132 hpf. Overall, the severe pericardial and yolk sac edema and reduced heart rate suggest that heart defects are a sensitive endpoint, and the general trend of dose‐dependent increase in EROD activity and induction of CYP1A mRNA gene expression provide evidence that the developmental toxicity of PCB126 to zebrafish embryos is mediated by activation of AhR. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 201–210, 2016.     

Cyclosporine exacerbates ketamine toxicity in zebrafish: Mechanistic studies on drug–drug interaction

Cyclosporine A (CsA) is an immunosuppressive drug commonly used in organ transplant patients to prevent allograft rejections. Ketamine is a pediatric anesthetic that noncompetitively inhibits the calcium‐permeable N‐methyl‐d ‐aspartic acid receptors. Adverse drug–drug interaction effects between ketamine and CsA have been reported in mammals and humans. However, the mechanism of such drug–drug interaction is unclear. We have previously reported adverse effects of combination drugs, such as verapamil/ketamine and shown the mechanism through intervention by other drugs in zebrafish embryos. Here, we show that ketamine and CsA in combination produce developmental toxicity even leading to lethality in zebrafish larvae when exposure began at 24 h post‐fertilization (hpf), whereas CsA did not cause any toxicity on its own. We also demonstrate that acetyl l ‐carnitine (ALCAR) completely reversed the adverse effects. Both ketamine and CsA are CYP3A4 substrates. Although ketamine and CsA independently altered the expression of the hepatic marker CYP3A65 , a zebrafish ortholog of human CYP3A4 , both drugs together induced further increase in CYP3A65 expression. In the presence of ALCAR, however, CYP3A65 expression was normalized. ALCAR has been shown to prevent ketamine toxicity in mammal and zebrafish. In conclusion, CsA exacerbated ketamine toxicity and ALCAR reversed the effects. These results, providing evidence for the first time on the reversal of the adverse effects of CsA/ketamine interaction by ALCAR, would prove useful in addressing potential occurrences of such toxicities in humans. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.     

Association between Common CYP1A2 Polymorphisms and Theophylline Metabolism in Non‐smoking Healthy Volunteers

This study was designed to investigate the impact of cytochrome P450 (CYP) 1A2 polymorphisms on theophylline metabolism in a non‐smoking healthy male Chinese population. Four polymorphisms CYP1A2*1C (G‐3860A), G‐3113A, CYP1A2*1F (C‐163A) and CYP1A2*1B (C‐5347T) were screened in 238 unrelated male volunteers. Then, a single oral 200‐mg dose of theophylline was administered to 37 volunteers, who were selected from 238 volunteers based on the CYP1A2 genotype. CYP1A2 activities were evaluated by plasma 1,7‐dimethylxanthine/caffeine ratios (17X/137X) after administration of 100‐mg caffeine. The plasma concentrations of theophylline, 17X and 137X were determined by high‐performance liquid chromatography. The activity of CYP1A2 was lower in volunteers with the ‐3113 AA genotype compared with those with the ‐3113 AG genotype (0.35 ± 0.04 versus 0.48 ± 0.07, p = 0.016) or the ‐3113 GG genotype (0.35 ± 0.04 versus 0.58 ± 0.22, p = 0.037). CYP1A2*1F polymorphisms were associated with increased CYP1A2 activity in volunteers with ‐3860G/‐3113G/5347C homozygosity (0.66 ± 0.24 versus 0.46 ± 0.05, p = 0.034). However, theophylline metabolism showed no difference among volunteers carrying different haplotype pairs. CYP1A2 genetic polymorphisms influenced CYP1A2 enzyme activity as measured by caffeine, but CYP1A2 gene polymorphisms appeared to have limited influence on theophylline metabolism in our study.     

Metabolism of the tryptamine‐derived new psychoactive substances 5‐MeO‐2‐Me‐DALT, 5‐MeO‐2‐Me‐ALCHT,and 5‐MeO‐2‐Me‐DIPT and their detectability in urine studied by GC–MS,LC–MSn,and LC‐HR‐MS/MS

Many N,N‐dialkylated tryptamines show psychoactive properties and were encountered as new psychoactive substances. The aims of the presented work were to study the phase I and II metabolism and the detectability in standard urine screening approaches (SUSA) of 5‐methoxy‐2‐methyl‐N,N‐diallyltryptamine (5‐MeO‐2‐Me‐DALT), 5‐methoxy‐2‐methyl‐N‐allyl‐N‐cyclohexyltryptamine (5‐MeO‐2‐Me‐ALCHT), and 5‐methoxy‐2‐methyl‐N,N‐diisopropyltryptamine (5‐MeO‐2‐Me‐DIPT) using gas chromatography–mass spectrometry (GC–MS), liquid chromatography coupled with multistage accurate mass spectrometry (LC–MSⁿ), and liquid chromatography‐high‐resolution tandem mass spectrometry (LC‐HR‐MS/MS). For metabolism studies, urine was collected over a 24 h period after administration of the compounds to male Wistar rats at 20 mg/kg body weight (BW). Phase I and II metabolites were identified after urine precipitation with acetonitrile by LC‐HR‐MS/MS. 5‐MeO‐2‐Me‐DALT (24 phase I and 12 phase II metabolites), 5‐MeO‐2‐Me‐ALCHT (24 phase I and 14 phase II metabolites), and 5‐MeO‐2‐Me‐DIPT (20 phase I and 11 phase II metabolites) were mainly metabolized by O‐demethylation, hydroxylation, N‐dealkylation, and combinations of them as well as by glucuronidation and sulfation of phase I metabolites. Incubations with mixtures of pooled human liver microsomes and cytosols (pHLM and pHLC) confirmed that the main metabolic reactions in humans and rats might be identical. Furthermore, initial CYP activity screenings revealed that CYP1A2, CYP2C19, CYP2D6, and CYP3A4 were involved in hydroxylation, CYP2C19 and CYP2D6 in O‐demethylation, and CYP2C19, CYP2D6, and CYP3A4 in N‐dealkylation. For SUSAs, GC–MS, LC‐MSⁿ, and LC‐HR‐MS/MS were applied to rat urine samples after 1 or 0.1 mg/kg BW doses, respectively. In contrast to the GC–MS SUSA, both LC–MS SUSAs were able to detect an intake of 5‐MeO‐2‐Me‐ALCHT and 5‐MeO‐2‐Me‐DIPT via their metabolites following 1 mg/kg BW administrations and 5‐MeO‐2‐Me‐DALT following 0.1 mg/kg BW dosage. Copyright © 2017 John Wiley & Sons, Ltd.     

Neuro‐ and hepatic toxicological profile of (S)‐2,4‐diaminobutanoic acid in embryonic,adolescent and adult zebrafish

(S)‐2,4‐Diaminobutanoic acid (DABA) is a noncanonical amino acid often co‐produced by cyanobacteria along with β‐N‐methylamino‐l ‐alanine (BMAA) in algal blooms. Although BMAA is a well‐established neurotoxin, the toxicity of DABA remains unclear. As part of our development of biocompatible materials, we wish to make use of DABA as both a building block and as the end‐product of enzymatically induced depolymerization; however, if it is toxic at very low concentrations, this would not be possible. We examined the toxicity of DABA using both in vivo embryonic and adult zebrafish models. At higher sublethal concentrations (700 μm ), the fish demonstrated early signs of cardiotoxicity. Adolescent zebrafish were able to tolerate a higher concentration. Post‐mortem histological analysis of juvenile zebrafish showed no liver or brain abnormalities associated with hepato‐ or neurotoxicity. Combined, these results show that DABA exhibits no overt toxicity at concentrations (100‐300 μm ) within an order of magnitude of those envisioned for its application. This study further highlights the low cost and ease of using zebrafish as an early‐stage toxicological screening tool.     

Age‐related Differences in CYP3A Abundance and Activity in the Liver of the Göttingen Minipig

In view of paediatric drug development, regulatory authorities often request safety studies in juvenile animals, including minipigs. Unfortunately, knowledge on the ontogeny of the biotransformation processes in animal models remains scarce and impedes a correct interpretation of the toxicity findings. CYP3A4 is one of the most important drug‐metabolizing enzymes in human beings and shows important similarities with CYP3A in the minipig. Therefore, the aim of this study was to assess the abundance and activity of CYP3A in liver microsomes from foetal, juvenile (days 1, 3, 7 and 28) and adult male and female Göttingen minipigs. CYP3A abundance was studied by an indirect enzyme‐linked immunosorbent assay (ELISA), whereas CYP3A activity was assessed by a biotransformation assay with Luciferin‐IPA. CYP3A abundance could not be detected until day 3. From day 7 onwards, a gradual increase in expression was noted, leading to the highest abundance in adult animals. CYP3A activity was not detectable in foetuses and 1‐day‐old animals. The CYP3A activity was detectable, but below the LLOQ in day 3 animals and increased gradually with age to reach the highest level in adults. The CYP3cide and ketoconazole inhibition, and testosterone and midazolam reduction of Luciferin‐IPA metabolism in minipig liver microsomes substantiate that Luciferin‐IPA is metabolized by CYP3A in minipigs. A positive correlation was found between CYP3A abundance and biotransformation of Luciferin‐IPA (Pearson r = 0.863; p < 0.0001). In conclusion, both abundance and activity of CYP3A increased gradually in juvenile minipigs, but remained below the levels observed in adult animals.     

Effects of allicin on CYP2C19 and CYP3A4 activity in healthy volunteers with different <Emphasis Type="Italic">CYP2C19</Emphasis> genotypes

Objective  To investigate the interaction between allicin and omeprazole and to observe the effects of allicin on CYP2C19 and CYP3A4 activity in healthy Chinese male volunteers with different CYP2C19 genotypes. Methods  Eighteen subjects (six CYP2C19*1/CYP2C19*1, four CYP2C19*1/CYP2C19*2, two CYP2C19*1/ CYP2C19*3, and six CYP2C19*2/ CYP2C19*2) were enrolled in a two-phase randomized crossover trial. In each phase, all subjects received placebo or a 180 mg allicin capsule once daily for 14 consecutive days. The pharmacokinetics of omeprazole (20 mg orally on day 15) was determined for up to 12 h following administration by high-performance liquid chromatography. Results  In carriers of the CYP2C19*1/CYP2C19*1 and CYP2C19*1/CYP2C19*2 or *3 genotype, allicin treatment increased the peak plasma concentration (C_max) of omeprazole by 49.7 ± 7.2 (p < 0.001) and 54.2 ± 9.2% (p < 0.001), and increased the area under the plasma time–concentration curve ( ) of omeprazole by 48.1 ± 9.0 (p = 0.001) and 73.6 ± 26.7% (p < 0.001), respectively. The ratio of of 5-hydroxyomeprazole to omeprazole (a marker for CYP2C19 activity) decreased significantly (p < 0.001 and p = 0.001, respectively). However, no pharmacokinetic parameters were significantly changed by allicin in CYP2C19*2/CYP2C19*2. The C_max and of omeprazole sulfone were unchanged in all three genotypes. Conclusions  Allicin reduced the metabolism of omeprazole by inhibiting CYP2C19 activity in individuals with the CYP2C19*1/CYP2C19*1 and CYP2C19*1/CYP2C19*2 or *3 genotypes, but not in those with the CYP2C19*2/ CYP2C19*2 genotype. Allicin did not significantly affect the activity of CYP3A4 in all subjects.     

Remote per‐conditioning reduces oxidative stress,downregulates cyclo‐oxygenase‐2 expression and attenuates ischaemia–reperfusion‐induced acute kidney injury

相似文献   

Synthesis,biological evaluation,and molecular docking studies of novel 3‐aryl‐5‐(alkyl‐thio)‐1H‐1,2,4‐triazoles derivatives targeting Mycobacterium tuberculosis

A small library of new 3‐aryl‐5‐(alkyl‐thio)‐1H‐1,2,4‐triazoles was synthesized and screened for the antimycobacterial potency against Mycobacterium tuberculosis H₃₇Ra strain and Mycobacterium bovis BCG both in active and dormant stage. Among the synthesized library, 25 compounds exhibited promising anti‐TB activity in the range of IC₅₀0.03–5.88 μg/ml for dormant stage and 20 compounds in the range of 0.03–6.96 μg/ml for active stage. Their lower toxicity (>100 μg/ml) and higher selectivity (SI = >10) against all cancer cell lines screened make them interesting compounds with potential antimycobacterial effects. Furthermore, to rationalize the observed biological activity data and to establish a structural basis for inhibition of M. tuberculosis, the molecular docking study was carried out against a potential target MTB CYP121 which revealed a significant correlation between the binding score and biological activity for these compounds. Cytotoxicity and in vivo pharmacokinetic studies suggested that 1,2,4‐triazole analogues have an acceptable safety index, in vivo stability and bio‐availability.     

The effect of perazine on the CYP2D6 and CYP2C19 phenotypes as measured by the dextromethorphan and mephenytoin tests in psychiatric patients

There is evidence that the antipsychotic drug perazine is an inhibitor of CYP2D6. This study aimed at evaluating its effect on CYP2D6 and CYP2C19 activities in submitting psychiatric patients to phenotyping with dextromethorphan and mephenytoin, respectively, substrates of these enzymes, before and during a treatment with perazine. A total of 31 patients were phenotyped with dextromethorphan (CYP2D6) and mephenytoin (CYP2C19) before and after a 2‐week treatment with 450 ± 51 mg/day (mean ± sd) perazine. At baseline, five patients appeared to be poor metabolizers (PM) of dextromethorphan and two patients of mephenytoin. The metabolic ratio (MR) of dextromethorphan/dextrorphan as determined in collected urine increased significantly (Wilcoxon; P < .0001) from baseline (0.39 ± 1.38 [mean ± sd]) till day 14 (1.46 ± 2.22). In 19 out of 26 extensive metabolizers (EM) of dextromethorphan, the phenotype changed from EM to PM. This suggests an almost complete inhibition of CYP2D6 by perazine and/or its metabolites. On the other hand, perazine (or some of its metabolites) did seemingly not inhibit CYP2C19. In conclusion, this study suggests that in patients treated with perazine and co‐medicated with CYP2D6 substrates, there could be an increased risk of adverse effects as a consequence of a pharmacokinetic interaction.     

Synthesis,molecular modeling,and biological evaluation of 4‐[5‐aryl‐3‐(thiophen‐2‐yl)‐4,5‐dihydro‐1H‐pyrazol‐1‐yl] benzenesulfonamides toward acetylcholinesterase,carbonic anhydrase I and II enzymes

In this study, 4‐[5‐aryl‐3‐(thiophen‐2‐yl)‐4,5‐dihydro‐1H‐pyrazol‐1‐yl] benzenesulfonamides were synthesized, and inhibition effects on AChE, hCA I, and hCA II were evaluated. K_i values of the compounds toward hCA I were in the range of 24.2 ± 4.6‐49.8 ± 12.8 nm , while they were in the range of 37.3 ± 9.0‐65.3 ± 16.7 nm toward hCA II. K_i values of the acetazolamide were 282.1 ± 19.7 nm and 103.60 ± 27.6 nm toward both isoenzymes, respectively. The compounds inhibited AChE with K_i in the range of 22.7 ± 10.3‐109.1 ± 27.0 nm , whereas the tacrine had K_i value of 66.5 ± 13.8 nm . Electronic structure calculations at M06‐L/6‐31 + G(d,p)//AM1 level and molecular docking studies were also performed to enlighten inhibition mechanism and to support experimental findings. Results obtained from calculations of molecular properties showed that the compounds obey drug‐likeness properties. The experimental and computational findings obtained in this study might be useful in the design of novel inhibitors against hCA I, hCA II, and AChE.     

Pharmacokinetics of the Experimental Non‐Nucleosidic DNA Methyl Transferase Inhibitor N‐Phthalyl‐l‐Tryptophan (RG 108) in Rats

DNA methyl transferase (DNMT) inhibitors can re‐establish the expression of tumour suppressor genes in malignant diseases, but might also be useful in other diseases. Inhibitors in clinical use are nucleosidic cytotoxic agents that need to be integrated into the DNA of dividing cells. Here, we assessed the in vivo kinetics of a non‐nucleosidic inhibitor that is potentially free of cytotoxic effects and does not require cell division. The non‐specific DNMT inhibitor N‐phthalyl‐l ‐tryptophan (RG 108) was injected subcutaneously in rats. Blood was drawn 0, 0.5, 1, 2, 4, 6, 8 and 24 hr after injection and RG 108 in plasma was measured by high‐performance liquid chromatography coupled to mass spectrometry. Trough levels and area under the curve (AUC) were significantly higher with multiple‐dose administration and cytochrome inhibition. In this group, time to maximal plasma concentration (t_max, mean ± S.D.) was 37.5 ± 15 min., terminal plasma half‐life was approximately 3.7 h (60% CI: 2.1–15.6 h), maximal plasma concentration (C_max) was 61.3 ± 7.6 μM, and AUC was 200 ± 54 μmol·h/l. RG 108 peak levels were not influenced by cytochrome inhibition or multiple‐dose administration regimens. Maximal tissue levels (C_max in μmol/kg) were 6.9 ± 6.7, 1.6 ± 0.4 and 3.4 ± 1.1 in liver, skeletal and heart muscle, respectively. We conclude that despite its high lipophilicity, RG 108 can be used for in vivo experiments, appears safe and yields plasma and tissue levels in the range of the described 50% inhibitory concentration of around 1 to 5 μM. RG 108 can therefore be a useful tool for in vivo DNMT inhibition.     

Protective effects of puerarin against tetrabromobisphenol a‐induced apoptosis and cardiac developmental toxicity in zebrafish embryo‐larvae

Tetrabromobisphenol A (TBBPA), a brominated flame retardant, is detected commonly in aquatic environments, where it is thought to be highly toxic to the development of aquatic life. In this study, zebrafish embryos and larvae were used to investigate the protective effects of puerarin after exposure to TBBPA. Malformation, blood flow disorders, pericardial edema, and spawn coagulation rates increased, whereas survival decreased significantly after exposure to 0.5 and 1.0 mg L^?1 TBBPA. The measured indices of morphological toxicity improved after treatment with puerarin. TBBPA also induced reactive oxygen species (ROS) production in a dose‐dependent manner. Acridine orange staining results revealed that TBBPA exposure caused cardiomyocyte apoptosis and induced the expression of three proapoptotic genes: P53, Bax, and Caspase9. In contrast, the expression of the antiapoptotic gene Bcl2 was down‐regulated. When genes related to cardiac development were assessed, the expression of Tbx1, Raldh2, and Bmp2b changed after exposure to the combination of TBBPA and puerarin. These results suggest that TBBPA induces cardiomyocyte apoptosis and ROS production, resulting in cardiac developmental toxicity in zebrafish embryos or larvae. Therefore, puerarin regulates the expression of cardiac developmental genes, such as Tbx1, Bmp2b, and Raldh2 by inhibiting ROS production, and subsequently modulates cardiac development after the exposure of zebrafish larvae to TBBPA. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1014–1023, 2015.