Abstract. The objective of this study was to examine whether high concentrations of epidermal growth factor (EGF) and/or insulin-like growth factor I (IGF-I) would have a beneficial effect on bovine embryo development in vitro and to obtain normal calves by using an ovum pick up method and embryo culture in a chemically defined medium. When compared with controls, EGF (100 or 200 ng/ml) or IGF-I (50 or 100 ng/ml) significantly increased the rate of embryos that developed into blastocysts during an 8-day culture after the in vitro fertilization of oocytes obtained from ovaries from a slaughterhouse. IGF-I induced a dose-dependent increase in cell number in both the inner cell mass and the trophectoderm, whereas EGF stimulated proliferation only in the inner cell mass. A combination of EGF (100 ng/ml) and IGF-I (50 ng/ml) produced an additive effect, and embryos developed into blastocysts at a comparatively high rate (27.9%) compared with controls (12.0%). A similar rate of development was achieved using a combination of EGF and IGF-I in the culture of embryos following ovum pick up by ultrasound-guided transvaginal follicular aspiration and in vitro fertilization, and 5 blastocysts that developed after the culture were transferred into uteri; two embryos implanted, and normal calves were born. These results suggest that the combined use of EGF and IGF-I makes bovine embryo culture in a chemically defined medium a practical and useful procedure for producing blastocysts, and its application to embryo culture following ovum pick up and in vitro fertilization could be useful for producing normal calves. Key words: Bovine embryo, Chemically defined medium, Epidermal growth factor (EGF), Insulin-like growth factor I (IGF-I), Ovum pick up (OPU) (J. Reprod. Dev. 58: [140][141][142][143][144][145][146] 2012) R ecently, a number of calves have been produced by the ovum pick up (OPU) method and in vitro culture (IVC) systems. Since the OPU method can be repeatedly applied to the same cow and can be used in pregnant [1] or problem cows in which embryos have not been obtained by uterine flushing after artificial insemination [2], this technology is effective for facilitating the improvement of domestic animals.An IVC system is necessary for the OPU method. Since successful pregnancy by transfer of embryos following the OPU method and in vitro fertilization has been confirmed [3], different protocols and medium conditions for oocyte maturation and embryo development have been tested [4][5][6][7]. However, there are some problems associated with the IVC system, such as a skewed sex ratio [8,9] or large offspring syndrome [10,11]. It has been suggested that factors in the culture medium such as serum used for embryo development may be involved in these problems. Fetal bovine serum (FBS) is widely employed as a supplement in the media used for culturing in vitro fertilized bovine embryos, but the subsequent rate of development of the fertilized oocytes to the blastocyst stage is known to vary among serum lots. There is thus a...

We compared the hair cortisol levels of lactating dairy cows in a cold- and a warm-temperate region out of four climatic zones in Japan. We simultaneously investigated the effects of calving number, lactation period and month of hair sampling. Hair of nine Holstein lactating cows chosen from each region (i.e. 18 cows per sampling) was sampled in March, June, September and December. Number of calvings (1, 2, ≥3) and lactation duration (early: <100, middle: 101-200, and late: >201 days) were balanced between regions. Cortisol was extracted from hair by methanol, and its level was determined with a cortisol immunoassay kit. A multi-way analysis of variance revealed that the effects of month of hair sampling (P < 0.001) and its combination with region (P < 0.05) were significant. In a multiple comparison test, significant differences (P < 0.01) in hair cortisol level (pg/mg of hair) were found between June (13.0 ± 1.0) and the other 3 months, and between September (1.6 ± 0.2) and December (4.5 ± 0.3). The rise in cortisol level from March to June was more intense in the cold-temperate region. These results demonstrate the necessity of considering seasonal variations in each climatic region when we use hair cortisol level as an indicator of stress.

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Based on our previous findings that medroxyprogesterone acetate (MPA) significantly suppressed the formation of preneoplastic mammary hyperplastic alveolar nodules in mice, and that this suppression persisted for some time, we studied the effects of different schedules of MPA treatment on spontaneous mammary tumorigenesis and uterine adenomyosis in SHN virgin mice. Mice received a subcutaneous pellet of MPA every 2 months: I) during the limited period of 1-3 months of age; II) throughout the experiment beginning at 6-8 months of age; and III) throughout the experiment beginning at 2-3 months of age. All treatments significantly enhanced mammary tumorigenesis with little difference in the effects among treatments. The progression of uterine adenomyosis was also stimulated in Experiments I and III, but not in Experiment II. These results are in good accord with our previous observations with progesterone, indicating that MPA has progesterone-like effects on mammary and uterine lesions of mice.

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This study was conducted to evaluate and compare the economic benefits of different embryo sexing methods, based on the cost per female dairy calf produced. Female calves were produced from four kinds of female embryos: (1) those collected from superstimulated donors at 7–8 days after artificial insemination (AI) with X-sorted semen; (2) those sex-determined by loop-mediated isothermal amplification assay of a biopsy sample of embryos collected from superstimulated donors after AI with conventional unsorted semen; (3) those obtained by in vitro embryo production (IVEP), using X-sorted semen and in vitro -matured oocytes collected from donors by ovum pick-up (OPU); and (4) those obtained by IVEP, using X-sorted semen and oocytes collected by OPU after dominant follicle ablation and follicle growth stimulation of the donors. The respective productivities of female calves per technical service and the total production cost per female calf of each sexing method were compared. The production cost per female calf (66,537 JPY), as calculated from the number of female calves per service (1.30), pregnancy rate of transfer (42.9%), rate of female calves obtained (92.9%), and total cost of the method (56,643 JPY plus embryo transfer fee), was less for IVEP with X-sorted semen and follicular growth-stimulated (FGS) oocytes than for the other groups (P < 0.05). The results demonstrate that embryo production with X-sorted semen and FGS oocytes provides a more efficient method for producing female calves than the other embryo sexing methods.

The mechanism for low yield of blood culture in invasive aspergillosis; the clinical importance of antigen detection tests revisited Bone Marrow Transplantation (2005) 36, 85-86.

This study examined the effects of treatment with U0126, which inhibits MAPK by inhibiting MAPK kinase, during the first 2 hr of in vitro maturation on bovine developmental competence and on gap junction (GAPJ) communication between the oocyte and cumulus cells. The percentage of oocytes developing to the blastocyst stage in the group treated with 5 μM U0126 (28%) was significantly higher than that in controls (15%, p < .05), while that in the group treated with 10 μM U0126 (18%) was not. Breakdown of the GAPJs was delayed in the group treated with 5 μM U0126 when compared to controls, as estimated by immunohistochemical examination of connexin 43, which is a primary constituent of the GAPJs. These results indicate that treatment with 5 μM U0126 during in vitro maturation delays GAPJ breakdown and improves bovine oocyte developmental competence.