Interspecies nuclear transfer (NT) could be an alternative to replicate animals when supply of recipient oocytes is limited or in vitro embryo production systems are incomplete. In the present study, embryonic development was assessed following interspecies NT of donor cumulus cells derived from yak and dog into the recipient ooplasm of domestic cow. The percentages of fusion and subsequent embryo development to the eight-cell stage of interspecies NT embryos were comparable to those of intraspecies NT embryos (cow-cow NT embryos). The percentage of development to blastocysts was significantly lower (p < 0.05) in yak-cow NT embryos than that in cow-cow NT embryos (10.9% vs. 39.8%). In dog-cow NT embryos, only one embryo (0.4%) developed to the blastocyst stage. These results indicate that interspecies NT embryos possess equally developmental competence to the eight-cell stage as intraspecies NT embryos, but the development to blastocysts is very low when dog somatic cells are used as the donor nuclei.

Abstract. The influence of acute exposure to zearalenone (ZEN) on porcine oocyte maturation, fertilization or sperm penetration ability during both in vitro maturation and fertilization was evaluated. First, oocytes were cultured in ZENcontaining (0−1000 μg/l) maturation medium and then fertilized. The oocytes maturing in vitro without ZEN were then fertilized in ZEN-containing fertilization medium. The maturation rates of oocytes and penetration ability of sperm decreased significantly in the presence of 1000 μg/l of ZEN. However, neither increases in the rates of degeneration and DNA fragmentation of oocytes nor reductions in normal and polyspermic fertilization were observed. ZEN did not affect the sperm penetration rates; however, 1000 μg/l ZEN had positive effects on normal and polyspermic fertilization rates. Therefore, it can be suggested that an acute exposure of porcine oocytes during maturation and of oocytes and sperm during fertilization to ZEN up to 1000 μg/l may not affect the fertility of the oocytes.Key words: Oocytes, Porcine, Sperm, Zearalenone (J. Reprod. Dev. 57: [547][548][549][550] 2011) earalenone (ZEN) is a nonsteroidal estrogen-like mycotoxin produced by Fusarium species on several grains. It is an estrogen receptor agonist; its distinct estrogenic and anabolic properties in several animal species exert detrimental effects on the reproductive system resulting in reproductive disorders in domestic animals, particularly in swine [1][2][3][4]. Although in vitro culture systems do not always provide accurate predictions of toxicity in animals, they can be used to assess risks and can help to define the mechanisms by which mycotoxins act on germ cells [5]. Several in vitro culture assays have been employed to determine the effect of ZEN and its metabolites on the reproductive organs of swine. Previous in vitro experiments revealed that exposure to these mycotoxins affects oocyte maturation, pronucleus formation and embryonic development [6,7], as well as viability, motility and acrosome reactions in sperm [8,9]. Nevertheless, the acute effects of exposure to ZEN during in vitro fertilization remain unknown. This study aimed to examine the effects of exposure to ZEN on porcine oocytes and sperm by using an in vitro maturation (IVM) and in vitro fertilization (IVF) systems to assess the state of nuclear DNA damage and fertilization.As shown in Table 1, exposure to 1000 μg/l of ZEN had a negative effect on the meiotic competence of porcine oocytes (P<0.05). However, there were no significant differences among the groups with respect to the percentages of oocytes showing degeneration and DNA damage. Exposure to 100 and 1000 μg/l of ZEN during maturation culture decreased the total rate of sperm penetration (P<0.05) compared with the control group, but did not influence the rates of normal or polyspermic fertilization of oocytes.As shown in Table 2, exposure to ZEN during IVF did not affect total rates of sperm penetration irrespective of the ZEN concentration. However, exposure to 1000 μg/l of...

The chemical toxicity of cryoprotectants to porcine embryos was examined by the evaluation of survival and DNA damage after exposure to cryoprotectants. Porcine blastocysts were exposed to 10% of ethylene glycol (EG), 1,2-propanediol (PD) or glycerol (GLY) for 1 h at room temperature (23-25 degrees C) and then cultured in vitro for 24 h. The survival rates of blastocysts exposed to PD and GLY were significantly lower than those of control blastocysts in which the embryos were exposed to carrier solution without cryoprotectants. Significantly more DNA-fragmented nuclei occurred in the cryoprotectant-exposed blastocysts, compared with the control blastocysts. Moreover, the indices of DNA-fragmented nuclei in the blastocysts without blastocoele re-formation after culture were significantly higher than those with blastocoele re-formation, irrespective of the exposure treatment. These results indicate that the exposure of porcine blastocysts to cryoprotectant decreases the survival rates and increases the DNA-fragmented nuclei in embryos.

Abstract. Zearalenone (ZEN) and its metabolites are important nonsteroidal estrogenic mycotoxins that cause reproductive disorders in domestic animals, especially pigs. We aimed to simultaneously detect ZEN and its metabolites α-zearalenol (α-ZOL) and β-zearalenol (β-ZOL) in porcine follicular fluid (FF) by liquid chromatographytandem mass spectrometry. ZEN and α-ZOL, but not β-ZOL, were detected in all pooled FF samples collected from coexisting follicles (diameter ≥ 6 mm) within 10 ovaries. Furthermore, ZEN and α-ZOL were detected in samples pretreated with β-glucuronidase/arylsulfatase, but not in those left untreated, suggesting that the FF samples contained glucuronide-conjugated forms of the mycotoxins that may be less harmful to porcine oocytes due to glucuronidation affecting the receptor binding. Nonetheless, the effects of the glucuronide-conjugated forms should be studied, both in vitro and in vivo. earalenone (ZEN) is a nonsteroidal estrogenic mycotoxin that is produced by Fusarium species on several grains. Despite its low acute toxicity and carcinogenicity, ZEN and its metabolites exhibit distinct estrogenic and anabolic properties in several animal species because of their agonistic effect on the estrogenic receptor. Thus, they affect the reproductive system and play important roles in reproductive disorders in domestic animals, particularly swine [1][2][3][4].The presence of environmental pollutants with potential reproductive toxicity in the follicular fluid (FF) of livestock may be of particular importance because the oocyte completes maturation before ovulation within the FF [5]. Recently, we reported that the FF was naturally contaminated with ZEN and its metabolites and showed the in vitro effects of ZEN on oocyte maturation in cattle [6]. However, no reports are available on the contamination of porcine FF by ZEN and its metabolites.The objectives of this preliminary investigation were to determine the concentrations of ZEN and its metabolites, including α-zearalenol (α-ZOL) and β-zearalenol (β-ZOL), in porcine FF by using liquid chromatography-negative ion electrospray tandem mass spectrometry with electrospray ionization (LC/MS/MS).As shown in Table 1, ZEN and α-ZOL were detected in all 10 FF samples supplemented with β-glucuronidase/arylsulfatase solution during preincubation, while the 10 FF samples that were not treated with β-glucuronidase/arylsulfatase solution did not contain ZEN or α-ZOL, even at trace levels. Additionally, β-ZOL was not detected even at trace levels, irrespective of β-glucuronidase/arylsulfatase treatment. Figure 1 shows representative LC/MS/MS chromatograms of the porcine FF samples contaminated with ZEN and α-ZOL. The mean (± SEM) concentrations of ZEN and α-ZOL were 38.9 ± 4.0 pg/ml (max and min: 54.8 and 15.2 pg/ml, respectively) and 17.6 ± 1.7 pg/ml (max and min: 26.4 and 10.0 pg/ ml, respectively), respectively.In the present study, we detected ZEN and α-ZOL but not β-ZOL in porcine FF only after the FF was treated with β-glucuronidase/arylsulfatase solution...