Transcriptome analysis during 4-vinylcyclohexene diepoxide exposure-induced premature ovarian insufficiency in mice.

The occupational chemical 4-Vinylcyclohexene diepoxide (VCD) is a reproductively toxic environmental pollutant that causes follicular failure, leading to premature ovarian insufficiency (POI), which significantly impacts a woman's physical health and fertility. Investigating VCD's pathogenic mechanisms can offer insights for the prevention of ovarian impairment and the treatment of POI. This study established a mouse model of POI through intraperitoneal injection of VCD into female C57BL/6 mice for 15 days. The results were then compared with those of the control group, including a comparison of phenotypic characteristics and transcriptome differences, at two time points: day 15 and day 30. Through a comprehensive analysis of differentially expressed genes (DEGs), key genes were identified and validated some using RT-PCR. The results revealed significant impacts on sex hormone levels, follicle number, and the estrous cycle in VCD-induced POI mice on both day 15 and day 30. The DEGs and enrichment results obtained on day 15 were not as significant as those obtained on day 30. The results of this study provide a preliminary indication that steroid hormone synthesis, DNA damage repair, and impaired oocyte mitosis are pivotal in VCD-mediated ovarian dysfunction. This dysfunction may have been caused by VCD damage to the primordial follicular pool, impairing follicular development and aggravating ovarian damage over time, making it gradually difficult for the ovaries to perform their normal functions.

4-vinylcyclohexene diepoxide induces apoptosis by excessive reactive oxygen species and DNA damage in human ovarian granulosa cells.

4-Vinylcyclohexene diepoxide (VCD) is a hazardous industrial material which is widely used in the production of fragrances, rubber tires, antioxidants, pesticides, flame retardants and plasticizers. Previous studies have shown that exposure to VCD damages the female reproductive system, but the effects and mechanisms of VCD exposure on human granulosa cells are not reported. In this study, we used a human granulosa cell line (SVOG) to explore the effects of VCD exposure and found that VCD exposure had toxic effects on SVOG cells in vitro. VCD exposure led to excessive accumulation of intracellular ROS, caused DNA damage in cells, altered the expression of some key genes related with apoptosis and oxidative stress, and ultimately inhibited the proliferative capacity of granulosa cells, resulting in increased apoptosis. Overall, our findings provide solid evidence showing that VCD exposure produces severe damage to human granulosa cells, which is helpful for understanding the reproductive toxicity of VCD and etiology of infertility.

Maternal exposure to 4-vinylcyclohexene diepoxide during pregnancy induces subfertility and birth defects of offspring in mice.

4-vinylcyclohexene diepoxide (VCD), widely used in industry, is a hazardous compound that can cause premature ovarian failure, but whether maternal VCD exposure affects the health and reproduction of offspring is unknown. Here we focused on the effects of VCD on fertility and physical health of F1 and F2 offspring in mice. The pregnant mice were injected intraperitoneally with different dosages of VCD once every day from 6.5 to 18.5 days post-coitus (dpc). We showed that maternal exposure to VCD during pregnancy significantly reduced the litter size and ovarian reserve, while increasing microtia occurrences of F1 mice. The cytospread staining showed a significant inhibition of meiotic prophase I progression from the zygotene stage to the pachytene stage. Mechanistically, the expression level of DNA damage marker (γ-H2AX) and BAX/BCL2 ratios were significantly increased, and RAD51 and DMC1 were extensively recruited to DNA double strand breaks sites in the oocytes of offspring from VCD-exposed mothers. Overall, our results provide solid evidence showing that maternal exposure to VCD during pregnancy has intergenerational deleterious effects on the offspring.

Fluorovinylsulfones and -Sulfonates as Potent Covalent Reversible Inhibitors of the Trypanosomal Cysteine Protease Rhodesain: Structure-Activity Relationship, Inhibition Mechanism, Metabolism, and In Vivo Studies.

Rhodesain is a major cysteine protease of Trypanosoma brucei rhodesiense, a pathogen causing Human African Trypanosomiasis, and a validated drug target. Recently, we reported the development of α-halovinylsulfones as a new class of covalent reversible cysteine protease inhibitors. Here, α-fluorovinylsulfones/-sulfonates were optimized for rhodesain based on molecular modeling approaches. 2d, the most potent and selective inhibitor in the series, shows a single-digit nanomolar affinity and high selectivity toward mammalian cathepsins B and L. Enzymatic dilution assays and MS experiments indicate that 2d is a slow-tight binder (Ki = 3 nM). Furthermore, the nonfluorinated 2d-(H) shows favorable metabolism and biodistribution by accumulation in mice brain tissue after intraperitoneal and oral administration. The highest antitrypanosomal activity was observed for inhibitors with an N-terminal 2,3-dihydrobenzo[b][1,4]dioxine group and a 4-Me-Phe residue in P2 (2e/4e) with nanomolar EC50 values (0.14/0.80 µM). The different mechanisms of reversible and irreversible inhibitors were explained using QM/MM calculations and MD simulations.

Effects of low intensity pulsed ultrasound on expression of B-cell lymphoma-2 and BCL2-Associated X in premature ovarian failure mice induced by 4-vinylcyclohexene diepoxide.

BACKGROUND: Premature ovarian failure (POF) is a common disease in the field of Gynecology. Low intensity pulsed ultrasound (LIPUS) can promote tissue repair and improve function. This study was performed to determine the effects of LIPUS on granulosa cells (GCs) apoptosis and protein expression of B-cell lymphoma-2 (Bcl-2) and BCL2-Associated X (Bax) in 4-vinylcyclohexene diepoxide (VCD)-induced POF mice and investigate the mechanisms of LIPUS on ovarian function and reserve capacity. METHODS: The current POF mice model was administrated with VCD (160 mg/kg) by intraperitoneal injection for 15 consecutive days. The mice were divided into the POF group, LIPUS group and control group. In the LIPUS group, the right ovary of mice was treated by LIPUS (acoustic intensity was 200 mW/cm2, frequency was 0.3 MHz, and duty cycle was 20%) for 20 min, 15 consecutive days from day 16. The mice of the POF group and control group were treated without ultrasonic output. The basic observation and body weight were recorded. Hematoxylin and eosin staining (H&E staining) and enzyme-linked immunosorbent assay (ELISA) were applied to detect ovarian follicle development, ovarian morphology and sex hormone secretion. Ovarian GCs apoptosis was detected by TUNEL assay and immunohistochemistry. RESULTS: The results showed that VCD can induce estrus cycle disorder, follicular atresia, sex hormone secretion decreased and GCs apoptosis in mice to establish POF model successfully. LIPUS significantly promoted follicular development, increased sex hormone secretion, inhibited excessive follicular atresia and GCs apoptosis. The mechanism might be achieved by increasing the protein expression of Bcl-2 and decreasing the expression of Bax in ovaries. CONCLUSIONS: LIPUS can improve the POF induced by VCD. These findings have the potential to provide novel methodological foundation for the future research, which help treat POF patients in the clinic.

N-vinyl compounds: studies on metabolism, genotoxicity, carcinogenicity.

Several N-vinyl compounds are produced in high volumes and are widely employed in the production of copolymers and polymers used in chemical, pharmaceutical, cosmetic and food industry. Hence, information on their genotoxicity and carcinogenicity is requisite. This review presents hitherto available information on the carcinogenicity and genotoxicity of N-vinyl compounds as well as their metabolism potentially generating genotoxic and carcinogenic derivatives. The genotoxicity and carcinogenicity of the investigated N-vinyl compounds vary widely from no observed carcinogenicity tested in lifetime bioassays in two rodent species (up to very high doses) to carcinogenicity in rats at very low doses in the absence of apparent genotoxicity. Despite of the presence of the vinyl group potentially metabolized to an epoxide followed by covalent binding to DNA, genotoxicity was observed for only one of the considered N-vinyl compounds, N-vinyl carbazole. Carcinogenicity was investigated only for two, of which one, N-vinyl pyrrolidone was carcinogenic (but not genotoxic) and ranitidine was neither carcinogenic nor genotoxic. As far as investigated, neither a metabolically formed epoxide nor a therefrom derived diol has been reported for any of the considered N-vinyl compounds. It is concluded that the information collected in this review will further the understanding of the carcinogenic potentials of N-vinyl compounds and may eventually allow approaching their prediction and prevention. A suggestion how to prevent genotoxicity in designing of N-vinyl compounds is presented. However, the available information is scarce and further research especially on the metabolism of N-vinyl compounds is highly desirable.

APOE4 genotype exacerbates the impact of menopause on cognition and synaptic plasticity in APOE-TR mice.

The impact of sex and menopausal status in Alzheimer's disease remains understudied despite increasing evidence of greater female risk, particularly in APOE4 carriers. Utilizing female APOE-TR mice maintained on a high-fat diet background we induced ovarian failure through repeated VCD injections, to mimic human menopause. At 12 months of age, recognition memory and spatial memory were assessed using object recognition, Y-maze spontaneous alternation, and Barnes maze. A VCD*genotype interaction reduced the recognition memory (P < .05), with APOE4 VCD-treated animals unable to distinguish between novel and familiar objects. APOE4 mice displayed an additional 37% and 12% reduction in Barnes (P < .01) and Y-maze (P < .01) performance, indicative of genotype-specific spatial memory impairment. Molecular analysis indicated both VCD and genotype-related deficits in synaptic plasticity with BDNF, Akt, mTOR, and ERK signaling compromised. Subsequent reductions in the transcription factors Creb1 and Atf4 were also evident. Furthermore, the VCD*genotype interaction specifically diminished Ephb2 expression, while Fos, and Cnr1 expression reduced as a consequence of APOE4 genotype. Brain DHA levels were 13% lower in VCD-treated animals independent of genotype. Consistent with this, we detected alterations in the expression of the DHA transporters Acsl6 and Fatp4. Our results indicate that the combination of ovarian failure and APOE4 leads to an exacerbation of cognitive and neurological deficits.

Effect of Jiajian Guishen Formula on the senescence-associated heterochromatic foci in mouse ovaria after induction of premature ovarian aging by the endocrine-disrupting agent 4-vinylcyclohexene diepoxide.

ETHNOPHARMACOLOGICAL RELEVANCE: Jiajian Guishen Formula (JJGSF), which is a prescription of Traditional Chinese Medicine (TCM), has been reported to be useful in the treatment of premature ovarian insufficiency (POI). AIM OF THE STUDY: To investigate the therapeutic effects of JJGSF on the treatment of POI induced by 4-vinylcyclohexene diep-oxide (VCD), an endocrine-disrupting chemical (EDC), and to elucidate the potential mechanism. MATERIALS AND METHODS: Female 8-week-old ICR mice (N = 72) were randomized into six groups, containing the Model group, Control group, three JJGSF groups, and Progynova group which was served as a positive control. After model establishment by VCD, the Progynova group were given a daily intragastric administration of Progynova, and the three JJGSF groups (high dose group, medium dose group and low dose group) received a daily intragastric administration of JJGSF at doses of 9, 4.5 and 2.25 g/kg for four weeks. The general growth of the mice was observed and the estrous cycles were examined. The serum hormone concentrations were measured by enzyme-linked immunosorbent assay (ELISA). To explore the potential mechanism of effect, the protein expressions of H3K9me3, HP1, and HMGA1/HMGA2 related to senescence-associated heterochromatic foci (SAHF), were determined by Immunofluorescence and Western blot analysis, respectively. RESULTS: After treating with JJGSF, the estrous cycles were improved significantly. The level of estrogen (E2) and anti-müllerian hormone (AMH) was increased and the ratio of follicle-stimulating hormone (FSH) to luteinizing hormone (LH) in serum was decreased significantly. Furthermore, a significant down-regulation of HMGA1/HMGA2 on protein level, a reduction distribution of HP1 and H3K9me3 in ovarian, and a lower fraction of SAHF-positive cells were observed after the administration with JJGSF, additionally effects showed a positive correlation with dosages. CONCLUSIONS: JJGSF could treat POI by the mechanism of inhibiting SAHF.

Systemic changes in a mouse model of VCD-induced premature ovarian failure.

AIMS: Premature ovarian failure (POF) is a phenomenon in which the ovaries fail before the age of 40 years. Prior research has used a wide range of mouse models designed to reflect different causes of POF, including genetic factors, iatrogenic factors, and immune factors. The current study employed a mouse model of POF induced by 4-vinylcyclohexene diepoxide (VCD). VCD can specifically kill primordial and primary ovarian follicles, which destroys the follicular reserve and causes POF. The current study sought to specify and extend the applications of this model by examining the effect of timing and VCD dose and by exploring the effect of the model on systems outside of the ovaries. MATERIALS AND METHODS: A VCD-induced mouse model of POF was constructed using established methods (VCD injected continuously at a concentration of 160 mg/kg for 15 days). Evidence for a graded effect of VCD was observed using a range of concentrations, and the best windows for examining VCD's effects on follicles and associated tissues were identified. KEY FINDINGS: The mouse model used here successfully simulated two common complications of POF - emotional changes and decreased bone density. The model's application was then extended to examine the links between disease and intestinal microorganisms, and evidence was found linking POF to the reproductively relevant composition of the gut microbiota. SIGNIFICANCE: These findings provide novel methodological guidance for future research, and they significantly extend the applications and scope of VCD-induced POF mouse models.

Impact of estrogen receptor agonists and model of menopause on enzymes involved in brain metabolism, acetyl-CoA production and cholinergic function.

Our goal is to understand how loss of circulating estrogens and estrogen replacement affect brain physiology and function, particularly in brain regions involved in cognitive processes. We recently conducted a large metabolomics study characterizing the effects of rodent models of menopause and treatment with estrogen receptor (ER) agonists on neurochemical targets in hippocampus, frontal cortex, and striatum. Here we characterize effects on levels of several key enzymes involved in glucose utilization and energy production, specifically phosphofructokinase, glyceraldehyde 3-phosphate dehydrogenase, and pyruvate dehydrogenase. We also evaluated effects on levels of ß-actin and α-tubulin, choline acetyltransferase (ChAT) activity, and levels of ATP citrate lyase. All experiments were conducted in young adult rats. Experiment 1 compared the effects of ovariectomy (OVX), a model of surgical menopause, and 4-vinylcyclohexene diepoxide (VCD)-treatments, a model of transitional menopause, with tissues collected at proestrus and at diestrus. Experiment 2 used a separate cohort of rats to evaluate the same targets in OVX and VCD-treated rats treated with estradiol or with selective ER agonists. Differences in the expression of metabolic enzymes between cycling animals and models of surgical and transitional menopause were detected. These differences were model-, region- and time- dependent, and were modulated by selective ER agonists. Collectively, the findings demonstrate that loss of ovarian function and ER agonist treatments have differing effects in OVX vs. VCD-treated rats. Differences may help to explain differences in the effects of estrogen treatments on brain function and cognition in women who have experienced surgical vs. transitional menopause.

Rescue role of hesperidin in 4-vinylcyclohexene diepoxide-induced toxicity in the brain, ovary and uterus of wistar rats.

Background The ovotoxicity of 4-vinylcyclohexene diepoxide (VCD) has been established in several experimental models. Hesperidin (HSD) is a bi-flavonoid found in citrus fruits and has been reported to be a potent antioxidant and anti-inflammatory agent. Here, we have evaluated the rescue role of hesperidin on VCD-induced toxicity in the brain, ovary, and uterus of rats. Methods Six groups of rats containing ten rats in each group were orally given corn oil (control), hesperidin (100 mg/kg), hesperidin (200 mg/kg), VCD (250 mg/kg), VCD [(250 mg/kg)+hesperidin (100 mg/kg)] and VCD [(250 mg/kg)+hesperidin (200 mg/kg)] once a day for 30 days, respectively. Thereafter, we determined the selected biomarkers of oxidative damage, inflammation, endocrine balance, and histology of the reproductive organs. Results The data showed that hesperidin rescued VCD-induced increase in oxidative stress (hydrogen peroxide and malondialdehyde) and inflammatory (nitric oxide) biomarkers. In addition, hesperidin restored the reduction in antioxidant enzymes (catalase, glutathione S-transferase, glutathione peroxidase) activities and glutathione level in the brain, ovary, and uterus of rats (p<0.05). Lastly, hesperidin preserved the histological structure of the ovary and uterus of rats exposed to VCD. Conclusions Overall, the rescue role of hesperidin on VCD-induced toxicity in the brain and reproductive organs of female rats may be due to its antioxidative and anti-inflammatory properties.

Bis(vinylsulfonyl)piperazines as efficient linkers for highly homogeneous antibody-drug conjugates.

Disulfide re-bridging strategy has demonstrated significant advantages in the construction of homogeneous antibody drug conjugates (ADCs). However, a major issue that disulfide scrambling at the hinge region of antibody leads to the formation of "half-antibody" has appeared for many re-bridging linkers. We present bis(vinylsulfonyl)piperazines (BVP) as efficient linkers to selectively re-bridge disulfides at the antigen-binding fragment (Fab) regions and produce highly homogeneous conjugates with a loading of two drugs without disulfide scrambling. We also found that optically active (S)-configuration linkers led to more sufficient conjugation compared with (R)-configuration. The BVP-linked ADCs demonstrated superior efficacy and antigen-selectivity in vitro cytotoxicity.

Cardiac changes during the peri-menopausal period in a VCD-induced murine model of ovarian failure.

AIM: Cardiovascular disease (CVD) risk is lower in pre-menopausal females vs age matched males. After menopause risk equals or exceeds that of males. CVD protection of pre-menopausal females is ascribed to high circulating oestrogen levels. Despite experimental evidence that oestrogen are cardioprotective, oestrogen replacement therapy trials have not shown clear benefits. One hypothesis to explain the discrepancy proposed hearts remodel during peri-menopause. Peri-menopasual myocardial changes have never been investigated, nor has the ability of oestrogen to regulate heart function during peri-menopause. METHODS: We injected female mice with 4-vinylcyclohexene diepoxide (VCD, 160 mg/kg/d IP) to cause gradual ovarian failure over 120d and act as a peri-menopausal model RESULTS: Left ventricular function assessed by Langendorff perfusion found no changes in VCD-injected mice at 60 or 120 days compared to intact mice. Cardiac myofilament activity was altered at 60 and 120 days indicating a molecular remodelling in peri-menopause. Myocardial TGF-ß1 increased at 60 days post-VCD treatment along with reduced Akt phosphorylation. Acute activation of oestrogen receptor-α (ERα) or -ß (ERß) depressed left ventricular contractility in hearts from intact mice. ER-regulation of myocardial and myofilament function, and myofilament phosphorylation, were disrupted in the peri-menopausal model. Disruption occurred without alterations in total ERα or ERß expression. CONCLUSIONS: This is the first study to demonstrate remodelling of the heart in a model of peri-menopause, along with a disruption in ER-dependent regulation of the heart. These data indicate that oestrogen replacement therapy initiated after menopause affects a heart that is profoundly different from that found in reproductively intact animals.

Radical polymerization inside living cells.

Polymerization reactions conducted inside cells must be compatible with the complex intracellular environment, which contains numerous molecules and functional groups that could potentially prevent or quench polymerization reactions. Here we report a strategy for directly synthesizing unnatural polymers in cells through free radical photopolymerization using a number of biocompatible acrylic and methacrylic monomers. This offers a platform to manipulate, track and control cellular behaviour by the in cellulo generation of macromolecules that have the ability to alter cellular motility, label cells by the generation of fluorescent polymers for long-term tracking studies, as well as generate a variety of nanostructures within cells. It is remarkable that free radical polymerization chemistry can take place within such complex cellular environments. This demonstration opens up a multitude of new possibilities for how chemists can modulate cellular function and behaviour and for understanding cellular behaviour in response to the generation of free radicals.

Comparison of Reproductive Function in Female TgMISIIR-TAg Transgenic and Wildtype C57BL/6 Mice.

Transgenic TgMISIIR-TAg (TAg) mice express the oncogenic virus SV40 in Mullerian epithelial cells. Female TAg mice spontaneously develop epithelial ovarian carcinoma, the most common type of ovarian cancer in women. Female TAg mice are infertile, but the reason has not been determined. We therefore investigated whether female TAg mice undergo puberty, demonstrate follicular development, maintain regular cycles, and ovulate. Ovarian cancers in women commonly develop after menopause. The occupational chemical 4-vinylcyclohexene diepoxide (VCD) accelerates follicle degeneration in the ovaries of rats and mice, causing early ovarian failure. We therefore used VCD dosing of mice to develop an animal model for menopause. The purpose of this study was to characterize reproductive parameters in female TAg mice and to investigate whether the onset of ovarian failure due VCD dosing differed between female TAg and WT C57BL/6 mice. As in WT female mice, TAg female mice underwent puberty (vaginal opening) and developed cyclicity in patterns that were similar between the groups. Vehicle-only TAg mice had fewer ovulations (numbers of corpora lutea) than WT animals. VCD exposure delayed the onset of puberty (day of first estrus) in TAg as compared with WT mice. Morphologic evaluation of ovaries revealed many more degenerating follicles in TAg mice than WT mice, and more VCD-dosed TAg mice were in ovarian failure than VCD-dosed WT mice. These results suggest that despite showing similar onset of sexual maturation, TAg mice have increased follicular degeneration and fewer ovulations than WT. These features may contribute to the inability of female TAg mice to reproduce.

Depletion of follicles accelerated by combined exposure to phthalates and 4-vinylcyclohexene diepoxide, leading to premature ovarian failure in rats.

Humans are at daily risk by simultaneous exposures to a broad spectrum of man-made chemicals in the commercial products. Several classes of chemicals have been shown to alter follicle development and reduce fertility, leading to premature ovarian failure (POF) in mammals. We investigate the synergistic effects of 4-vinylcyclohexene diepoxide (VCD) and phthalate, including di(2-ethylhexyl) phthalate (DEHP), butyl benzyl phthalate (BBP) and di-n-butyl phthalate (DBP) on POF. Combination exposure with VCD and phthalate significantly reduced the numbers of primary follicles. The expressions of Amh and Sohlh2 were significantly decreased in the combination groups. Serum Amh levels were significantly lower in the combination groups. Additionally, serum levels of follicle-stimulating hormone were significantly increased in combination groups. Taken together, exposure to phthalates promotes the depletion of follicular follicles and consequently increases the risk of premature menopause, and combined exposure of phthalates and VCD to early menopausal women is likely to aggravate the POF syndrome.

Peroxiredoxin 2 deficiency accelerates age-related ovarian failure through the reactive oxygen species-mediated JNK pathway in mice.

Reactive oxygen species (ROS) produced in biological reactions have been shown to contribute to ovarian aging. Peroxiredoxin 2 (Prx2) is an antioxidant enzyme that protects cells by scavenging ROS; however, its effect on age-related, oxidative stress-associated ovarian failure has not been reported. Here, we investigated its role in age-related ovarian dysfunction and 4-vinylcyclohexene diepoxide (VCD)-induced premature ovarian failure using Prx2-deficient mice. Compared to those in wildtype (WT) mice, serum levels of anti-Müllerian hormone, 17ß-estradiol, and progesterone and numbers of follicles and corpora lutea were significantly lower in 18-month-old Prx2-/- mice. Moreover, levels of Bax, cytochrome c, cleaved caspase-3, and phosphorylated JNK proteins were higher and numbers of apoptotic (terminal deoxynucleotidyl transferase dUTP nick end labeling-positive) cells were considerably greater in 18-month-old Prx2-/- ovaries than WT ovaries. Furthermore, the effects of the ovarian toxicant VCD in significantly enhancing ROS levels and apoptosis through activation of JNK-mediated apoptotic signaling were more pronounced in Prx2-/- than WT mouse embryonic fibroblasts. Expression of the steroidogenic proteins StAR, CYP11A1, and 3ß-HSD and serum levels of 17ß-estradiol and progesterone were also reduced to a greater extent in Prx2-/- mice than WT mice after VCD injection. This reduced steroidogenesis was rescued by addition of the Prx mimic ebselen or JNK inhibitor SP600125. This constitutes the first report that Prx2 deficiency leads to acceleration of age-related or VCD-induced ovarian failure by activation of the ROS-induced JNK pathway. These findings suggest that Prx2 plays an important role in preventing accelerated ovarian failure by inhibiting ROS-induced JNK activation.

In vitro toxicity evaluation of new silane-modified clays and the migration extract from a derived polymer-clay nanocomposite intended to food packaging applications.

The clay montmorillonite (Mt) is among the nanofillers more frequently used for food packaging applications. The organomodification of clays with different modifiers, such as silanes, is an important step in the preparation of improved polymer/clay materials known as nanocomposites. However, the toxicological data about these nanofillers is still scarce. In the present study, an in vitro toxicological evaluation in Caco-2 cells of two silane-modified clays based on Mt, Clay3 and Clay4 (0-250µg/ml), was performed. The cytotoxicity, cell death, genotoxicity and oxidative stress produced by both organoclays were evaluated after 24 and 48h of exposure. Moreover, the migration extracts obtained from nanocomposites of polypropylene (PP) + Clay3 and only PP were also investigated. Only Clay4 induced cytotoxicity, showing a reduction of cell viability to 63% of the control, as well as oxidative stress in a concentration-dependent manner. Regarding the PP-Clay3 migration extract, no cytotoxic effects were observed after exposure to the tested concentrations (0-100%). Moreover, significant differences in the presence of Ca, Mg and Si compared to the PP extract were obtained, although migration levels were in accordance with the food contact materials regulation. These findings indicate that a case-by-case toxicological assessment of organoclays should be performed.

Organic Solvent-Tolerant Marine Microorganisms as Catalysts for Kinetic Resolution of Cyclic ß-Hydroxy Ketones.

Chiral cyclic ß-hydroxy ketones represent key motifs in the production of natural products of biological interest. Although the molecules are structurally simple, they require cumbersome synthetic steps to get access to them and their synthesis remains a challenge in organic chemistry. In this report, we describe a straightforward approach to enantiomerically enriched (R)- and (S)-3-hydroxycyclopentanone 2a, (R)- and (S)-3-hydroxycyclohexanone 2b, and (R)- and (S)-3-hydroxycycloheptanone 2c involving a transesterification resolution of the racemates using whole cells of marine microorganisms as catalysts and vinyl acetate the acyl donor and solvent. Twenty-six strains from a wide collection of isolates from marine sediments were screened, and seven strains were found to markedly catalyze the resolution in an asymmetric fashion. Using the strain Serratia sp., (R)-2a was isolated in 27% yield with 92% ee and (S)-2a in 65% yield with 43% ee, corresponding to an E-value of 37; (R)-2b was isolated in 25% yield with 91% ee and (S)-2b in 67% yield with 39% ee, corresponding to an E-value of 40; and (R)-2c was isolated in 30% yield with 96% ee and (S)-2c in 63% yield with 63% ee, corresponding to an E-value of 75.

Parabens Accelerate Ovarian Dysfunction in a 4-Vinylcyclohexene Diepoxide-Induced Ovarian Failure Model.

Parabens are widely used preservatives in basic necessities such as cosmetic and pharmaceutical products. In previous studies, xenoestrogenic actions of parabens were reported in an immature rat model and a rat pituitary cell line (GH3 cells). The relationship between parabens and ovarian failure has not been described. In the present study, the influence of parabens on ovarian folliculogenesis and steroidogenesis was investigated. A disruptor of ovarian small pre-antral follicles, 4-vinylcyclohexene diepoxide (VCD, 40 mg/kg), was used to induce premature ovarian failure (POF). Methylparaben (MP, 100 mg/kg), propylparaben (PP, 100 mg/kg), and butylparaben (BP, 100 mg/kg) dissolved in corn oil were treated in female 8-week-old Sprague-Dawley rat for 5 weeks. Estrus cycle status was checked daily by vaginal smear test. Ovarian follicle development and steroid synthesis were investigated through real-time PCR and histological analyses. Diestrus phases in the VCD, PP, and BP groups were longer than that in the vehicle group. VCD significantly decreased mRNA level of folliculogenesis-related genes (Foxl2, Kitl and Amh). All parabens significantly increased the Amh mRNA level but unchanged Foxl2 and Kitlg acting in primordial follicles. VCD and MP slightly increased Star and Cyp11a1 levels, which are related to an initial step in steroidogenesis. VCD and parabens induced an increase in FSH levels in serum and significantly decreased the total number of follicles. Increased FSH implies impairment in ovarian function due to VCD or parabens. These results suggest that VCD may suppress both formation and development of follicles. In particular, combined administration of VCD and parabens accelerated inhibition of the follicle-developmental process through elevated AMH level in small antral follicles.