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See detailIn vitro model to study the endocrine disrupting activity of migration products from plastic food contact materials
Simon, Coraline ULg; Onghena, Matthias; Covaci, Adrian et al

Poster (2013, November 06)

Bisphenol A (BPA) is a chemical compound mainly used for the manufacture of plastic such as polycarbonate. This transparent thermoplastic polymer is used for the fabrication of several food containers ... [more ▼]

Bisphenol A (BPA) is a chemical compound mainly used for the manufacture of plastic such as polycarbonate. This transparent thermoplastic polymer is used for the fabrication of several food containers like baby bottle … BPA can migrate into food in contact with polycarbonate. There is a worldwide concern about BPA because several studies have shown endocrine disruptor potency of BPA causing possible adverse health effects. In January 2011, the European Commission decided to ban the use of polycarbonate to manufacture baby feeding bottles. In a recent opinion, the Superior Health Council’s issued its concern regarding the currently use of alternatives to polycarbonate in these materials. This work is part of the ALTPOLYCARB project which aims to study the migration products of alternative to polycarbonate and their endocrine disruptor activities. The first part was to have an overview of the different polymers replacing polycarbonate, that are used on the Belgian market, it resulted in the conclusion that polymers used for the manufacture of baby bottles are mainly polypropylene, polyethersulfone, silicone, polyamide, polystyrene, and melamine. The second part of this work will be to evaluate the endocrine disruptor activity(ies) of global migration residues obtained from different kinds of baby bottles. This (these) activity(ies) will be explored using cell based transactivational assays also named “reporter gene assays. The MCF7 recombinant cells used here are genitically modified cells containing the firefly luciferase gene, as a reporter gene, and a DNA responsive element specific to the human oestrogen receptor. The biological activity of a chemical compound is monitored by the measurement of light emitted by the cells exposed to it (after addition of luciferin, the substrate of luciferase). In a preliminary step, we first screened pure substances which were shown to migrate from plastic baby bottle, in a recent study performed by Simoneau & al, 2012 . Human estrogen receptor agonistic and antagonistic activities of 25 pure compounds were measured using MCF7-ER cells (genetically modified MCF7 cells). After the first screening, some substances clearly show an activity such as BPA, Benzophenone, 2-Propenoic acid-2-ethylhexyl ester, Benzaldehyde-4-methylthio, Butylated hydroxytoluene and Dodecanoic acid, methyl ester whereas others ask an in-depth analysis to confirm their activity. For active substances only the study will be continued and a full dose-response curve will be performed in order to assess quantitatively the activity [less ▲]

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See detailIn vitro model to study the endocrine disrupting activity of migration products from plastic food contact materials
Simon, Coraline ULg; Onghena, Matthias; Covaci, Adrian et al

Poster (2014, May 15)

Bisphenol A (BPA) is used since 1960 as a primary raw material for the production of polycarbonate (PC) plastic and epoxy resin, which are widely used in a variety of common products including digital ... [more ▼]

Bisphenol A (BPA) is used since 1960 as a primary raw material for the production of polycarbonate (PC) plastic and epoxy resin, which are widely used in a variety of common products including digital media (e.g., CDs, DVDs), electrical and electronic equipment, automobiles, sports safety equipment, reusable food and drink containers , as well as baby bottle. During the last decades, in several studies, the migration of BPA is documented to be a well-known source of food contamination. The measurements of BPA in human fluids and tissues highlighted that its presence in food constitutes the primary route of human exposure. Some studies showed that BPA, which could disrupt normal endocrine function by mimicking estrogen hormones,, may be associated to several health problems and diseases. Recently, the European food safety authority conducted a risk assessment on BPA and concluded that though studies related to potential health hazards associated with BPA, are suggesting a potential negative effect on human body, but results are still uncertain. Following that screening assessment, the European Union took a series of measures, including a ban for the manufacture, import and sale of PC baby bottles to reduce BPA exposure of infants. Plastic alternatives to polycarbonate have massively appeared on Belgium market. Although there are several studies on BPA migration from polycarbonate into foodstuff under various conditions, there is a small amount of information about consequences on human health of the chemicals migrating from PC alternatives, including bottles commonly labelled “free BPA”. In a recent opinion (No. 8697, 11.03.2010), the Belgium Superior Health Council's issued its concern regarding the currently used alternatives to PC. Furthermore, they asked to investigate the possible risks associated with the use of these alternatives. To know if these alternatives are safe, the activity on several receptors (estrogen (ER), androgen (AR), progesterone (PR) and glucocorticoïd receptor (GR)) of chemicals migrating from PC alternatives, identified by Simoneau & al, 2012 , were evaluated using reporter gene assays. Agonistic and antagonistic activities of 38 pure compounds were measured. After the first screening, some substances clearly showed an activity on each receptor, such as BPA, 2.4- dimethyl benzaldehyde (C4), Bisphenol S (C49), while other subtances reacted on three, two or one receptor. Only 5 substances showed no activity. For active substances only, the study will be continued and a full dose-response curve will be performed in order to assess quantitatively the activity. [less ▲]

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See detailIn vitro modelisation of prions neuroinvasion mediated by dendritic cells
Dorban, G.; Defaweux, Valérie ULg; Heinen, Ernst ULg et al

Poster (2008, October)

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See detailIn vitro models for the study of cartilage damage and repair
Henrotin, Yves ULg; Labasse, A; Zheng, SX et al

in Rheumatology in Europe (1998), 27(S2), 7

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See detailIn vitro models for the study of cartilage damage and repair
Henrotin, Yves ULg; Reginster, Jean-Yves ULg

in Reginster, Jean-Yves; Pelletier, J-P; Martel-Pelletier, J (Eds.) et al Osteoarthritis: Clinical and experimental aspects (1999)

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See detailIn vitro models of non persistent and persistent infection of human and murine neuroblastoma cell lines by the varicella zoster virus
Schlabertz, Tania; Sadzot-Delvaux, Catherine ULg; Piette, Jacques ULg et al

in Archives Internationales de Physiologie et de Biochimie (1997)

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See detailIn vitro modulation of human gingival epithelial cell attachment and migration by minocycline-HCL.
Van Heusden, Alain ULg; Nusgens, Betty ULg; Goffinet, Gerhard ULg et al

in Journal of Periodontal Research (1998), 33(6), 377-85

Although the influence of tetracyclines on periodontal connective tissue cells has been the topic of many in vitro and in vivo studies, data regarding their effects on gingival epithelial cells are scarce ... [more ▼]

Although the influence of tetracyclines on periodontal connective tissue cells has been the topic of many in vitro and in vivo studies, data regarding their effects on gingival epithelial cells are scarce. The present in vitro study was designed to examine the influence of minocycline, a semi-synthetic analog of tetracycline, on human gingival keratinocyte (HGK) attachment and migration. Attachment tests were performed with HGK prelabeled by tritiated amino-acids. Increasing concentrations of minocycline (10, 50, 100 micrograms/ml) in the medium produced no significant modification of cell adhesion kinetics compared to control conditions, except for 100 micrograms/ml which statistically significantly (p < 0.05) reduced the number of attached cells beyond 6 h. A 24-h cell preincubation in 10 micrograms/ml of minocycline did not alter the kinetics of HGK attachment. Scanning electron microscopic observations of attached HGK showed that the presence of 10 micrograms/ml of minocycline in the "attachment medium" induced the production of multiple filopodial extensions. Migration tests in Boyden chambers for 40 h demonstrated that HGK preincubation for 24 h in a 10 micrograms/ml minocycline-HCl solution increased significantly (p < 0.005) cell migration towards a gradient of fetal calf serum. The presence of 10 micrograms/ml of minocycline in contact with the keratinocytes in the upper compartment of the migration chambers also produced a significant (p < 0.005) result. In contrast, the presence of minocycline in the lower compartments did not produce any chemoattractive effect. Within the limits of their significance, these results suggest that, at concentrations not beyond 50 micrograms/ml, minocycline could fasten the periodontal wound coverage by epithelial cells and allow the normal reformation of a junctional epithelium. [less ▲]

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See detailIn vitro modulation of human gingival keratinocyte migration by minocycline-HCl
Van Heusden, Alain ULg; Nusgens, Betty ULg; Lapière, Charles et al

in Journal of Dental Research (1998), 77

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See detailIn vitro modulation of keratinocyte attachment by minocycline H-Cl
Van Heusden, Alain ULg; Rompen, Eric ULg; Lebfevre, P. et al

in Journal of Dental Research (1995), 74

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See detailIn vitro modulation of keratinocyte attachment by minocycline HCl.
Van Heusden, Alain ULg; Rompen, Eric ULg; Lebfevre, P. et al

Poster (1994)

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See detailIn vitro modulation of keratinocyte attachment to dentin by minocycline
Van Heusden, Alain ULg; Rompen, Eric ULg; Lapière, Charles

in Journal of Dental Research (1997), 76

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See detailIn vitro modulation of radiosenitizing effect of FMdC. The importance of simultaneous alteration of the novo and salvage pathways to deoxyribonucleosides.
COUCKE, Philippe ULg; Cottin, E; Ciernick, I-F et al

in International Journal of Radiation, Oncology, Biology, Physics (2000), 46(3),

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See detailIn vitro multiplication of an industrial fiber plant: kenaf (Hibiscus cannabinus L.)
Arbaoui, Sarra; Campanella, Bruno ULg; Paul, Roger ULg et al

Poster (2012, April 23)

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See detailIn vitro neuroblastoma cell infection by varicella-zoster virus
Bourdon-Wouters, C.; Merville, Marie-Paule ULg; Piette, Jacques ULg et al

Poster (1988)

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See detailIN VITRO NUCLEAR AND CYTOPLASMIC MATURATION OF THE EQUINE OOCYTE: INFLUENCE OF CYSTEAMINE
Deleuze, Stefan ULg

Doctoral thesis (2009)

Research on in vitro embryo production (IVP) in the equine is impeded by the limited availability of mature oocytes as the mare is mono ovulating and superovulation is still difficult (Dippert and Squires ... [more ▼]

Research on in vitro embryo production (IVP) in the equine is impeded by the limited availability of mature oocytes as the mare is mono ovulating and superovulation is still difficult (Dippert and Squires, 1994; Bezard et al., 1995; Alvarenga et al., 2001b). Despite recent improvement in IVM of equine oocytes, success rates of IVM in that species remain low in all culture media tested compared to other species (Goudet et al., 2000b). However, most studies have focused on the percentage of oocytes reaching the metaphase II stage (nuclear maturation) but few concentrated on the final oocyte competence as measured by its ability to develop into a blastocyst and further establish a pregnancy. Blastocyst production rate is influenced not only by culture environment but also by oocyte maturation conditions. Under in vitro culture conditions, oxidative modifications of cell components via increased ROS represent a major culture induced stress (Johnson and Nasr-Esfahani, 1994). Anti-oxidant systems can attenuate the deleterious effects of oxidative stress by scavenging ROS (Del Corso et al., 1994). Glutathione, a tripeptide thiol, is the major non-protein sulfydryl compound in mammalian cells that plays an important role in protecting the cell from oxidative damage (Meister and Tate, 1976; Meister and Anderson, 1983). It has been suggested that GSH content in oocytes may serve as a reservoir protecting the zygote and the early embryos from oxidative damage before genomic activation and de novo GSH synthesis occur (Furnus et al., 1998; de Matos and Furnus, 2000). The addition of GSH synthesis precursors, such as cysteamine, a thiol compound, to IVM media has been shown to improve IVP in various species (Takahashi et al., 1993; de Matos et al., 1995; Grupen et al., 1995; de Matos et al., 2002a; de Matos et al., 2002b; de Matos et al., 2003; Gasparrini et al., 2003; Oyamada and Fukui, 2004; Balasubramanian and Rho, 2007; Anand et al., 2008; Singhal et al., 2008; Zhou et al., 2008). Very little information on the use of thiol compounds in the equine is available. Conventional in vitro fertilization (IVF) has not been successful in the mare, and a repeatable IVF technique has not yet been developed (Alm et al., 2001). To overcome the limitation of conventional IVF procedures, other methods to produce embryos from oocytes, either in vivo or in vitro, have been investigated. Among these, intra cytoplasmic sperm injection (ICSI) has permitted efficient equine in vitro blastocyst production (Galli et al., 2002; Lazzari et al., 2002; Choi et al., 2006a; Choi et al., 2006c). However, ICSI requires specific equipment and skills. Transfer of an immature oocyte into the preovulatory follicle of an inseminated recipient mare (Intra-Follicular Oocyte Transfer, IFOT) has produced embryos but the success rate was low (Hinrichs and Digiorgio, 1991). Similarly, oocyte transfer (OT) into the oviduct of an inseminated recipient mare was investigated (McKinnon et al., 1988; Carnevale, 1996; Hinrichs et al., 1997; Carnevale et al., 2001; Carnevale et al., 2003; Carnevale, 2004), and commercial programs using OT for mares with reproductive abnormalities are now available (Carnevale et al., 2001). Unfortunately, IFOT is poorly documented in the literature and reports of OT have been published by various laboratories and under various conditions, making comparisons between results and choosing among these as substitutive techniques to ICSI or embryo transfer difficult. The first aim of the present work was to investigate if there is an influence of supplementation with 100 µM of cysteamine on conventional IVF success rate. Cumulus oocytes complexes (COCs) retrieved by transvaginal ultrasound guided aspiration were matured in vitro with or without cysteamine supplementation and were then submitted to conventional IVF using either calcium ionophore or heparin as capacitation treatment for spermatozoa. A total of 131 oocytes were evaluated for evidence of sperm penetration. Both techniques (ionophore or heparin) yielded 6% of IVF and results were similar both for the cysteamine and the control group. This success rate of IVF is low compared to some published data (Palmer et al., 1991; Dell'Aquila et al., 1996; McPartlin et al., 2009) but similar to what others reported in the literature (Choi et al., 1994; Dell'Aquila et al., 1997a). Although, it seems likely that cysteamine did not significantly improve IVF rates under our conditions, our general success rates for IVF procedures may be too low for us to conclude definitely about the effect of cysteamine. As ICSI was not available to us, the second aim of this work was to determine what in vivo technique could best bypass the lack of an efficient conventional IVF procedure. We compared embryo production following transfer of in vivo recovered oocytes (1) into a recipient’s oviduct or (2) into her preovulatory follicle either immediately after ovum pick up or (3) after in vitro maturation. Recipients were inseminated with fresh semen of a stallion with a known normal fertility. Ten days after transfer, rates of embryos collected in excess to the number of ovulations were calculated and compared for each group. Embryo collection rates were 32.5% (13/40), 5.5% (3/55) and 12.8% (6/47) for OT, post-IVM and immediate IFOT respectively. OT significantly yielded more embryos than immediate and post-IVM IFOT did. These results show that, when ICSI is not an option, intra-oviductal oocyte transfer is to be preferred to IFOT, as an in vivo alternative, to bypass the inadequacy of conventional in vitro fertilization and to assess oocyte developmental competence. After it was established that in comparison to IFOT, OT is the most reliable in vivo alternative to in vitro fertilization where ICSI technology is not available, this technique was used to assess the effect of cysteamine supplementation on nuclear maturation and oocyte competence. The third aim of this work was to investigate the influence of supplementation with 100 µM of cysteamine on in vitro nuclear and cytoplasmic maturation by specific DNA staining and the ability of oocytes to undergo in vivo fertilization after OT. Oocytes were collected by transvaginal ultrasound guided aspiration and matured in vitro with (cysteamine group) or without (control group) cysteamine. The nuclear stage after DNA Hoechst staining and the embryo yield following OT were used as a criterion for assessing nuclear and cytoplasmic maturation, respectively. Overall maturation rate was 52%, which is rates reported in the literature ranging from 40 to 70% in the equine (Goudet et al., 1997a; Bogh et al., 2002; Hinrichs et al., 2005; Galli et al., 2007). Nuclear maturation was not statistically different (p>0.05) between oocytes cultured with or without cysteamine (55% and 47% respectively). From 57 oocytes transferred to the oviduct in each group, the number of embryos collected was 10 (17%) in the control group and 5 in the cysteamine group (9%). Those two percentages were not statistically different (p>0.05). Contrary to the data described in other domestic species, there was no effect of cysteamine on in vitro nuclear maturation, or in vivo embryonic development under our conditions. Under our conditions, the addition of 100 µM of cysteamine to a classic culture medium does not improve equine oocyte maturation or embryonic development after OT. The same dose failed to increase GSH content in the equine (Luciano et al., 2006). However, the effect of cysteamine supplementation is highly species and concentration dependant. The inadequacy of the chosen concentration may explain that equine embryo production has not been increased by the cysteamine under our conditions as opposed to what has been observed in many other species. Alternatively, we can hypothesize that some substances present in the IVM medium can interfere with GSH synthesis. This has been suggested for FSH and estradiol (Bing et al., 2001) and, although our maturation medium is not supplemented with gonadotropins or estradiol, factors contained in fetal calf serum or EGF might also have an effect on GSH synthesis. Considering its beneficial effects in many other species, supplementation with cysteamine to different IVM media should be further investigated in the equine. Ideally combining different concentrations and ICSI or OT in order to determine an optimal concentration and its effects on oocyte developmental competence. [less ▲]

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See detailIn vitro oocyte IGF-I priming increases inner cell mass proliferation of in vitro-formed bovine blastocysts
Velazquez, MA; Hadeler, KG; Herrmann, D et al

in Theriogenology (2012), 78(3), 517-527

Studies addressing the effects of supraphysiological levels of IGF-1 on oocyte developmental competence are relevant for unravelling conditions resulting in high bioavailability of IGF-1, such as the ... [more ▼]

Studies addressing the effects of supraphysiological levels of IGF-1 on oocyte developmental competence are relevant for unravelling conditions resulting in high bioavailability of IGF-1, such as the polycystic ovary syndrome (PCOS). This study investigated the effects of supraphysiological levels of IGF-1 during in vivo folliculogenesis on the morula-blastocyst transition in bovine embryos. Compacted morulae were non-surgically collected and frozen for subsequent mRNA expression analysis (IGF1R, IGBP3, TP53, AKT1, SLC2A1, SLC2A3, and SLC2A8), or underwent confocalmicroscopy analysis for protein localization (IGF1R and TP53), or were cultured in vitro for 24 h. In vitro-formed blastocysts were subjected to differential cell staining. The mRNA expression of SLC2A8 was higher in morulae collected from cows treated with IGF-1. Both IGF1R and TP53 protein were present in the plasma membrane and cytoplasm. IGF-1 treatment did not affect protein localization of both IGF1R and TP53. In vitro-formed blastocysts derived from morulae recovered from IGF-1-treated cows displayed a higher number of cells in the inner cell mass (ICM). Total cell number (TCN) of in vitro-formed blastocysts was not affected. A higher mean ICM/TCN proportion was observed in in vitro-formed blastocysts derived from morulae collected from cows treated with IGF-1. The percentage of in vitro-formed blastocysts displaying a low ICM/TCN proportion was decreased by IGF-1 treatment. In vitro-formed blastocysts with a high ICM/TCN proportion were only detected in IGF-1 treated cows. Results show that even a short in vivo exposure of oocytes to a supraphysiological IGF-1 microenvironment can increase ICM cell proliferation in vitro during the morula to blastocyst transition [less ▲]

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See detailIn vitro ototoxicity of aminoglycosides and platin derivatives. A semi-automatic assay for sensory hair cell damage in explanted rat organ of corti.
Malgrange, B.; LEFEBVRE, Philippe ULg; van de Water, T.R. et al

in Toxicology in Vitro (1998), 12 (6)

The ototoxic damage that drugs such as neomycin, kanamycin, colistin, cisplatin, transplatin and carboplatin cause on outer and inner hair cells in postnatal day 3 rat cochlear explants was investigated ... [more ▼]

The ototoxic damage that drugs such as neomycin, kanamycin, colistin, cisplatin, transplatin and carboplatin cause on outer and inner hair cells in postnatal day 3 rat cochlear explants was investigated. Phalloidin-fluorescein conjugate-stained stereocilia bundles of sensory hair cells were quantified by video image analysis as a measurement of ototoxic effect. The video image quantification system established dose-response curves for ototoxic drugs (e.g. calculation of an IC50) and allowed comparisons between several ototoxins from the same family. This methodology provided the means to assess the efficacy of otoprotectant agents in preventing ototoxicity. Poly-l-aspartate (10-5M) and poly-l-glutamate (10-5M) protected auditory hair cells from neomycin (10-3M) toxicity while reduced glutathione (10-3M) provided protection against cisplatin (10-4M)-induced hair cell damage. [less ▲]

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See detailIn vitro paradigms for the study of GnRH neuron function and estrogen effects.
Matagne, Valerie; LEBRETHON, Marie-Christine ULg; Gerard, Arlette et al

in Annals of the New York Academy of Sciences (2003), 1007

The elaboration of in vitro paradigms has enabled direct study of GnRH secretion and the regulation of this process. Common findings using different models are the pulsatile nature and calcium-dependency ... [more ▼]

The elaboration of in vitro paradigms has enabled direct study of GnRH secretion and the regulation of this process. Common findings using different models are the pulsatile nature and calcium-dependency of GnRH secretion, the excitatory effect of glutamate, and the inhibitory or excitatory effect of GABA. Among the different paradigms, the fetal olfactory placode cultures exhibit the unique property of migration in vitro and may retain the capacity to undergo maturational changes in vitro. The short-term incubation of hypothalamic explants obtained at different ages enables one to study developmental changes as well. Estrogens may have important roles in the regulation of GnRH function and can act indirectly via the neighboring neuronal/glial apparatus and directly on GnRH neurons at the cell body and terminal levels. A direct effect is supported by the recent localization of ERalpha and ERbeta transcripts in GnRH neurons using most paradigms. Discrepant effects of estrogens on GnRH neurons were observed since GnRH biosynthesis is inhibited while GnRH secretion can be either stimulated, unaffected, or reduced. It is likely that the regulatory role of sex steroids including estradiol is very complex since it could involve direct and indirect effects on GnRH neurons through genomic and/or non-genomic mechanisms. [less ▲]

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