Sulfadimethoxine residue in broiler-chicken skin.

The disposition and elimination of sulfadimethoxine (SDMX) in the skin of broiler-chickens were investigated. The administration of SDMX, in drinking water, at a concentration of 1,000 ppm for 5 days demonstrated that the SDMX was eliminated much more slowly from the skin than from the other tissues or plasma. These results were duplicated and confirmed in another experiment, in which a single dose of 200 mg/kg BW of SDMX was administered via a stomach tube. No significant difference in the SDMX residue level was observed between the broiler-chickens that had their skin surface sealed versus the non-sealed animals and that had ingested SDMX in their drinking water. This illustrated the higher SDMX residue in the skin was not attributable to external SDMX contamination from the drinking water, feces or urine. In addition, there was no significant difference among the SDMX residue concentrations in the thoracic, dorsal and leg skin samples, following an intravenous injection of SDMX (30 mg/kg BW). This indicated that the SDMX was distributed evenly throughout the entire skin area of the broiler-chickens.     

Generation and characterization of anti-leptin antisera against synthetic peptides and recombinant protein

The objective of this study was to generate antisera against recombinant bovine leptin and synthetic oligopeptides corresponding to the amino acid sequence 21-40 and 91-110 of bovine leptin. Recombinant bovine leptin was raised in the 293 cells and purified from 10 L of conditioned medium and utilized for immunization. The synthetic peptides were conjugated with keyhole limpet hemocyanin and inoculated into rabbits for antibody generation. Antibody titer was monitored by enzymeimmunoassay, immunoblotting and sandwich binding assay techniques. Each of the antisera, against three different antigens, was found to react with bovine leptin. The titers of anti-peptide antisera were lower than that of anti-recombinant leptin antiserum. Since anti-recombinant leptin antiserum was not neutralized by the leptin peptides 21-40 and 91-110, it is suggested that each antiserum recognizes a distinct epitope. In immunoblot analyses, all antisera exhibited cross-reactivity with human and mouse leptins. However, in the sandwich binding assay, the combination of anti-peptide antisera and anti-recombinant leptin antiserum, originated from bovine leptin, did not cross-react with either human or mouse leptin. The discrepancy of antigenic recognition between the immunoblot analyses and sandwich assay is thought to be dependent on the conformational status of leptin molecules between the species. The antisera generated in this study, which recognized distinct epitopes of bovine leptin, will provide a useful tool for studies of bovine leptin functions.     

Maternal exposure to low doses of bisphenol a has no effects on development of female reproductive tract and uterine carcinogenesis in Donryu rats

Effects of maternal exposure to low doses of bisphenol A (BPA), including those comparable with human exposure levels, on growth and development of the female reproductive system and uterine carcinogenesis in Donryu rats were investigated. Dams were administered BPA (0, 0.006 and 6 mg/kg/day) daily by gavage from gestation day 2 up to the day before weaning (postnatal day 21 at offspring). The serum levels of BPA were significantly elevated in the dams receiving 6 mg/kg/day, however, BPA levels in the milk of dams, and those in the serum and liver of offspring were similar between control and treated groups. The treatment did not exert any influences on uterine development including weight, gland genesis and estrogen receptor alpha expression, vaginal opening and gonadotropin secretion in the female offspring up to puberty. After maturation, no effects were evident with regard to estrous cyclicity in female offspring treated with BPA. In addition, the treatment had no effects on age-related morphological changes of the reproductive and endocrine organs and uterine carcinogenesis until 15 months of age. The results demonstrate that maternal exposure to BPA at levels comparable to human exposure did not have any effects on the female reproductive system of offspring in rats. In addition, BPA was also found in the serum, milk and liver of control dams and pups, and low levels of BPA were detected in drinking water and pellet diet. The present study showed that the experimental animals were also exposed to environmental BPA in the animal room.     

Generation and purification of recombinant bovine pregnancy associated glycoprotein

Bovine pregnancy-associated glycoprotein-1 (bPAG-1) is predicted to play an essential role during pregnancy and is labelled as a potential biochemical marker of pregnancy in ungulates. We have compared the generation of the glycosylated form of recombinant bPAG-1 (rbPAG-1) by human embryonic kidney 293 (HEK 293) and Chinese hamster ovary (CHO) cells in attached cultures and evaluated the adaptation of the rbPAG-1 transfected cell line to suspension culture. The PAG cDNA was cloned from placental RNA obtained from a slaughtered cow on day 55 of pregnancy. The PAG-pRcRSV expression vector was transfected into HEK 293 and CHO cells. Western blot analysis showed that clonal HEK 293 cells expressed rbPAG-1 better than CHO cells in attached cultures. Transfected HEK 293 cells were adapted to suspension culture in spinner flasks and the rbPAG-1 purified to homogeneity using ion-exchange, pepstatin-sepharose affinity chromatographies and preparative SDS-PAGE. The expression of rbPAG-1 was immunocharacterised using a polyclonal antibody. Our findings indicated that 293 cells are suitable for production of glycosylated form of rbPAG-1 and that the availability of the recombinant glycoprotein will aid in further studies to elucidate the function and structure of the protein.     

Polymorphisms in the canine glutamate transporter-1 gene: identification and variation among five dog breeds

Excitatory amino acid transporters (EAATs) are important for terminating glutamatergic neurotransmission and protect central nervous system (CNS) neurons from glutamatergic excitotoxicity. We selected these genes as targets that may relate to canine behavioral traits. After screening four EAAT genes (glutamate transporter-1; GLT-1, excitatory amino acid transporter 4; EAAT4, excitatory amino acid carrier; EAAC1, glutamate/aspartate transporter; GLAST) for single nucleotide polymorphisms (SNPs), we identified two silent SNPs (C129T and T471C) in the GLT-1 gene. We genotyped 193 dogs of 5 breeds and found significant variation among breeds in these two SNPs in GLT-1. The C129T polymorphism was not observed in Malteses and Miniature Schnauzers. These results suggest that polymorphisms in the GLT-1 gene may be useful markers for examining how the genetic background relates to the behavioral traits of dogs.     

Expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and its expected roles in the bovine endometrium during gestation

Extracellular matrix metalloproteinase inducer (EMMPRIN) and its induced matrix metalloproteinases (MMPs) play a crucial role in tissue remodeling during the peri-implantation period. However, the role of EMMPRIN in the bovine placenta is still unclear. We have postulated that EMMPRIN might play a regulatory role in trophoblastic cell functions during gestation by itself or through the regulation of MMP expression. In this study, EMMPRIN mRNA was detected in the bovine placentome and interplacentome throughout gestation, and its expression was significantly higher in the cotyledon during late gestation. In situ hybridization showed that EMMPRIN mRNA was expressed in the caruncular epithelium and the cotyledonary epithelium, including binucleate cells. Western blot analysis detected a band representing a protein of approximately 65 kDa in the caruncular and cotyledonary tissues, and the intensity of its expression was increased in both of these tissues during late gestation. The expression levels of MMP-2 and MMP-14 in the bovine placenta were higher during late gestation, as was observed for EMMPRIN. Therefore, EMMPRIN might regulate trophoblastic cell functions, especially those of binucleate cells, through MMP expression in the bovine placenta.     

Effects of photoperiod on secretory patterns of growth hormone in adult male goats

The aim of the present study was to clarify the effect of photoperiod on secretory patterns of growth hormone (GH) in male goats. Adult male goats were kept at 20°C with an 8‐h or 16‐h light photoperiod, and secretory patterns of GH secretion were compared. In addition, plasma profiles of prolactin (PRL), insulin‐like growth factor‐I (IGF‐I) and testosterone (T) were also examined to characterize GH secretion. GH was secreted in a pulsatile manner. There was no significant difference in pulse frequency between the 8‐h and 16‐h photoperiods. However, GH pulse amplitude tended to be greater in the group with the 16‐h photoperiod (P = 0.1), and mean GH concentrations were significantly greater in the 16‐h photoperiod (P < 0.05). The GH‐releasing response to GH releasing hormone was greater in the 16‐h than 8‐h photoperiod (P < 0.05). Plasma PRL and IGF‐I levels were higher in the 16‐h than 8‐h photoperiod (P < 0.05). In contrast, plasma T levels were lower in the 16‐h photoperiod (P < 0.05). These results show that a long light photoperiod enhances the secretion of GH as well as PRL and IGF‐I, but reduces plasma T concentrations in male goats.     

Oral administration of peptidergic growth hormone (GH) secretagogue KP102 stimulates GH release in goats

To assess the oral activity of KP102 (also known GHRP-2) on growth hormone (GH) release in ruminant animals, 5 or 10 mg/kg body weight (BW) of KP102 dissolved in saline was orally administered twice at 2 hr-intervals to either 1- or 3-mo-old goats (n = 5-6). Plasma GH concentrations in the 1-mo-old goats were elevated at 15 min after the first administration of both 5 and 10 mg/kg BW of KP102. Significant elevation of GH concentrations continued until 180 min after 10 mg/kg BW of KP102, whereas the elevated GH levels after the administrations of 5 mg/kg BW of KP102 subsided to basal concentrations within 90 min. The second administration of 10 mg/kg BW of KP102 failed to elevate the GH concentration, but 5 mg/kg BW of KP102 abruptly stimulated GH release. Plasma GH concentrations in the 3-mo-old goats were also significantly elevated after the administration of both 5 and 10 mg/kg BW of KP102. The plasma GH responses to 5 and 10 mg/kg BW of KP102 were almost identical. The elevated GH levels after the first administration of KP102 tended to be maintained throughout the experiment, and a transient increase in plasma GH levels was observed after the second administration. However, the stimulatory effect of KP102 on GH release in the 3-mo-old goats was small and less abrupt than that in the 1-mo-old goats. The concentrations of insulin-like growth factor-I were not increased by KP102 during the brief sampling periods used in this experiment. These results show that the oral administration of the peptidergic GH secretogogue KP102 stimulates GH release in a ruminant species, and that the oral activity of KP102 on GH release is modified by the age.