34 research outputs found
Диалог культур – парадигма современного социокультурного процесса
To improve the predictability of the zebrafish embryotoxicity test (ZET) for developmental (neuro)toxicity screening, we used a multiple-endpoints strategy, including morphology, motor activity (MA), histopathology and kinetics. The model compounds used were antiepileptic drugs (AEDs): valproic acid (VPA), carbamazepine (CBZ), ethosuximide (ETH) and levetiracetam (LEV). For VPA, histopathology was the most sensitive parameter, showing effects already at 60. μM. For CBZ, morphology and MA were the most sensitive parameters, showing effects at 180. μM. For ETH, all endpoints showed similar sensitivity (6.6. mM), whereas MA was the most sensitive parameter for LEV (40. mM). Inclusion of kinetics did not alter the absolute ranking of the compounds, but the relative potency was changed considerably. Taking all together, this demo-case study showed that inclusion of multiple-endpoints in ZET may increase the sensitivity of the assay, contribute to the elucidation of the mode of toxic action and to a better definition of the applicability domain of ZET
Fundamental aspects of bovine oocyte maturation : the role of estradiol, VIP and GHRH
Chapter 1 presents an overview on the aspects of oocyte maturation.
Growth hormone (GH), released from the pituitary by the stimulus of GHRH, increases cumulus expansion and improves cytoplasmic maturation in bovine oocytes. GHRH is also expressed in extraneural tissues suggesting that GHRH also plays a regulatory role in peripheral tissues. Therefore in Chapter 2 we studied the role of GHRH and its related peptide VIP on the nuclear maturation and cumulus expansion. We observed that exposure of bovine cumulus oocyte complexes (COCs) to GHRH or VIP did not affect nuclear maturation or cumulus expansion, but it retarded cytoplasmic maturation.
At the time of the LH surge, follicular fluid of preovulatory follicles has a high concentration of estradiol (E2), but 6h after the LH peak a sharp decline of E2 occurs, coinciding with the germinal vesicle breakdown (GVBD). Although most in vivo effects of E2 are well identified, the role of E2 during in vitro maturation (IVM) remains contradictory. In Chapter 3 we investigated the effects of E2 on bovine oocyte IVM and subsequent embryo development. Both COCs and denuded oocytes (DO) were used to study whether effects of E2 are exerted via cumulus cells or directly on the oocyte. We observed that presence of E2 during IVM decreased the percentage of metaphase II (MII) stage oocytes and increased the percentage of nuclear aberrations. This effect of E2 was stronger in DO compared with COCs. Concomitant presence of FSH and E2 during IVM did not influence the proportion of MII oocytes, although nuclear aberrations were still observed. Presence of E2 during IVM also decreased the blastocyst rate. However, when FSH was present, E2 had no effect on the cleavage rate and blastocyst formation. We concluded that supplementation of E2 to maturation medium negatively affects bovine oocyte nuclear maturation and subsequent embryo development. Although these effects are attenuated in the presence of FSH, we suggest omission of E2 in maturation protocols of bovine oocytes.
To investigate whether the effects of E2 were mediated via genomic or nongenomic pathways, we cultured oocytes in the presence of E2 either before or after GVBD (the period when transcription ceased) (Chapter 4). Additionally, an estradiol membrane-impermeable conjugate (E2-BSA) was used to investigate the presence of a putative membrane receptor for estradiol. By using RT-PCR we observed that oocytes expressed estrogen receptor (ER)β mRNA, while cumulus cells expressed both ERα and ERβ mRNA. Exposure to E2 during the first 8h of culture (before GVBD) induced a block at the metaphase I stage. However, the presence of E2 after GVBD induced an increase of oocytes exhibiting nuclear aberrations. Exposure of oocytes to E2-BSA did not affect nuclear maturation, suggesting that the effects of E2 on in vitro nuclear maturation are not exerted via a plasma membrane receptor.
Although oocytes from small-sized antral follicles can resume meiosis, the blastocyst yield obtained with such oocytes is low. It was hypothesized that chemically preventing GVBD by roscovitine (ROS), until the initiation of IVM might allow the oocyte more time to undergo cytoplasmic maturation and consequently acquire a higher developmental competence. In Chapter 5 we demonstrated that exposure to E2, during temporary inhibition of the GVBD with ROS, affected neither nuclear nor cytoplasmic maturation of oocytes originating from small-sized follicles. Possibly, the increase of E2 during follicular growth in vivo is more important for selection of the dominant follicle than for the cytoplasmic maturation of the oocyte.
Finally, Chapter 6 presents a summarizing discussion
The bovine oocyte in vitro maturation model: A potential tool for reproductive toxicology screening
Reproductive toxicity testing according to the present guidelines requires a high number of animals. Therefore, the development of alternative in vitro methods is urgently required. The aim of the present study was to investigate the applicability domain of the bovine oocyte in vitro maturation assay (bIVM) to study female reproductive toxicology. Therefore, bovine oocytes were exposed to a broad set of chemicals of two distinct biological function groups: (a) affecting female fertility and (b) affecting embryonic development and having a broad range of physical and chemical properties. The endpoints evaluated were the oocyte nuclear maturation (progression of meiosis) and general cytotoxicity. The oocyte nuclear maturation was negatively affected by all compounds tested and the effect was observed at concentrations lower than the cytotoxic ones. The bIVM assay correctly predicted the classification of compounds between those predefined groups. Additionally, the bIVM model contributes significantly for the 3R principle, since no test animals are used in this assay. In conclusion, the bIVM is a sensitive and valuable alternative assay to identify potential chemical hazard on female fertility. © 2012 Elsevier Inc
A category approach to predicting the developmental (neuro) toxicity of organotin compounds: The value of the zebrafish (Danio rerio) embryotoxicity test (ZET).
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Zebrafish embryotoxicity test for developmental (neuro)toxicity: Demo case of an integrated screening approach system using anti-epileptic drugs
To improve the predictability of the zebrafish embryotoxicity test (ZET) for developmental (neuro)toxicity screening, we used a multiple-endpoints strategy, including morphology, motor activity (MA), histopathology and kinetics. The model compounds used were antiepileptic drugs (AEDs): valproic acid (VPA), carbamazepine (CBZ), ethosuximide (ETH) and levetiracetam (LEV). For VPA, histopathology was the most sensitive parameter, showing effects already at 60. μM. For CBZ, morphology and MA were the most sensitive parameters, showing effects at 180. μM. For ETH, all endpoints showed similar sensitivity (6.6. mM), whereas MA was the most sensitive parameter for LEV (40. mM). Inclusion of kinetics did not alter the absolute ranking of the compounds, but the relative potency was changed considerably. Taking all together, this demo-case study showed that inclusion of multiple-endpoints in ZET may increase the sensitivity of the assay, contribute to the elucidation of the mode of toxic action and to a better definition of the applicability domain of ZET. © 2014 Elsevier Inc. Molecular Sequence Numbers: GENBANK: NM_131146, NM_131850; Chemicals/CAS: carbamazepine, 298-46-4, 8047-84-5; ethosuximide, 77-67-8; etiracetam, 102767-28-2, 33996-58-6; valproic acid, 1069-66-5, 99-66-1 Manufacturers: Sigma Aldrich, United State
Effects of Superovulation with oFSH and Norgestomet/GnRH-Controlled Release of the LH Surge on Hormone Concentrations, and Yield of Oocytes and Embryos at Specific Developmental Stages
Preovulatory estradiol and the establishment and maintenance of pregnancy in suckled beef cows1
Effects of hypoxanthine at low concentrations on in vitro maturation and fertilization of bovine oocytes
Bisphenol A exposure in utero disrupts early oogenesis in the mouse
Estrogen plays an essential role in the growth and maturation of the mammalian oocyte, and recent studies suggest that it also influences follicle formation in the neonatal ovary. In the course of studies designed to assess the effect of the estrogenic chemical bisphenol A (BPA) on mammalian oogenesis, we uncovered an estrogenic effect at an even earlier stage of oocyte development--at the onset of meiosis in the fetal ovary. Pregnant mice were treated with low, environmentally relevant doses of BPA during mid-gestation to assess the effect of BPA on the developing ovary. Oocytes from exposed female fetuses displayed gross aberrations in meiotic prophase, including synaptic defects and increased levels of recombination. In the mature female, these aberrations were translated into an increase in aneuploid eggs and embryos. Surprisingly, we observed the same constellation of meiotic defects in fetal ovaries of mice homozygous for a targeted disruption of ERbeta, one of the two known estrogen receptors. This, coupled with the finding that BPA exposure elicited no additional effects in ERbeta null females, suggests that BPA exerts its effect on the early oocyte by interfering with the actions of ERbeta. Together, our results show that BPA can influence early meiotic events and, importantly, indicate that the oocyte itself may be directly responsive to estrogen during early oogenesis. This raises concern that brief exposures during fetal development to substances that mimic or antagonize the effects of estrogen may adversely influence oocyte development in the exposed female fetus