No Promoting Effect of Ethyl Tertiary-butyl Ether (ETBE) on Rat Urinary Bladder Carcinogenesis Initiated with N-Butyl-N-(4-hydroxybutyl)nitrosamine

The effects of ethyl tertiary-butyl ether (ETBE) on two-stage urinary bladder carcinogenesis in male F344 rats initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) were investigated at various dose levels with regard to possible promoting activity. Groups of 30 rats were given drinking water containing 500 ppm BBN, as an initiator, for 4 weeks and starting one week thereafter received ETBE by gavage (daily, 7 days/week) at dose levels of 0 (control), 100, 300, 500 or 1000 mg/kg/day until experimental week 36. No statistically significant differences in incidences of preneoplastic lesions, papillomas, and carcinomas of the urinary bladder were evident in rats treated with 100–1000 mg/kg/day ETBE as compared with control values. Furthermore, the average numbers of preneoplastic or neoplastic lesions per unit length of basement membrane in rats given 100–1000 mg/kg/day ETBE were also comparable to control values. However, papillomatosis of the urinary bladder was found in 4 out of 30 rats (13%) in the group given 1000 mg/kg/day ETBE, and soft stones in the urinary bladder were found in 3 out of these 4 rats. The results thus demonstrated that ETBE did not exert promotional activity on urinary bladder carcinogenesis. However, papillomatosis of the urinary bladder developed in small numbers of the rats given ETBE at 1000 mg/kg/day but not in rats given 500 mg/kg/day or lower doses.     

Development of a screening method for genetically modified soybean by plasmid-based quantitative competitive polymerase chain reaction

A novel type of quantitative competitive polymerase chain reaction (QC-PCR) system for the detection and quantification of the Roundup Ready soybean (RRS) was developed. This system was designed based on the advantage of a fully validated real-time PCR method used for the quantification of RRS in Japan. A plasmid was constructed as a competitor plasmid for the detection and quantification of genetically modified soy, RRS. The plasmid contained the construct-specific sequence of RRS and the taxon-specific sequence of lectin1 (Le1), and both had 21 bp oligonucleotide insertion in the sequences. The plasmid DNA was used as a reference molecule instead of ground seeds, which enabled us to precisely and stably adjust the copy number of targets. The present study demonstrated that the novel plasmid-based QC-PCR method could be a simple and feasible alternative to the real-time PCR method used for the quantification of genetically modified organism contents.     

Comparison of cholesterol levels among lipoprotein fractions separated by anion-exchange high-performance liquid chromatography in periparturient Holstein–Friesian dairy cows

Changes in lipoprotein profiles occur in dairy cows during the periparturient period and in cows with transition cow disease. Here, the lipoprotein profiles of Holstein–Friesian dairy cows during the periparturient period were obtained by anion-exchange, high-performance liquid chromatography to evaluate the usefulness of lipoprotein profile evaluation during the periparturient period and in cows with fatty liver and milk fever. Lipoprotein levels (including total and high- (HDL-C) and low-density lipoprotein (LDL-C) cholesterol) in 10 healthy cows were low 4 weeks prepartum, with the lowest values at calving or within 1 week of calving; the values increased at 8 weeks postpartum. The lipoprotein levels were measured in 16 cows diagnosed with fatty liver (n=10) or milk fever (n=6) and compared to 10 healthy dairy cows. A significant difference was observed in HDL-C between healthy cows (at calving and 1 week postpartum), and the fatty liver and milk fever cows. Cows with fatty liver and milk fever had a lower mean HDL-C than the 10 healthy dairy cows at calving and 1 week postpartum. HDL-C might be a good indicator of energy balance for differentiating healthy cows from those with transition cow disease.     

Influence of Equine Conformation on Rider Oscillation and Evaluation of Horses for Therapeutic Riding

To obtain basic knowledge about selecting horses for therapeutic riding, the influence of equine conformation on rider oscillation and relationships between these factors and the evaluation on horses as the therapeutic riding were studied. Thirty-five riding horses were used. Equine conformation was estimated by 24 indices. Rider oscillation was measured by an accelerometer fixed at the rider’s waist. The spatial position of the oscillation was estimated by a double integration of the acceleration. Horses were evaluated for therapeutic riding by a Riding for the Disabled Association instructor as a rider. Evaluations were on a scale of 1 to 5, with 5 being the highest score for 27 items. Horses were classified into 4 groups: the short and narrow (SN), short and wide (SW), tall and narrow (TN), and tall and wide (TW). The frequencies of rider oscillation both at walk and trot were higher (P<0.01), and the vertical (P<0.01) and longitudinal (P<0.05) amplitudes at trot were smaller, on short horses than on tall horses. The vertical amplitude at walk was smaller (P<0.05) and the lateral amplitude at trot was larger (P<0.01) on wide horses than on narrow horses. Short horses could be used for the rider who requires side walkers. Wide horses could be used for relieving muscular tension and for the rider who could not maintain good balance on the horse. Short and wide horses should be suitable for therapeutic riding.     

Immunochemical sexing of living yellowfin tuna, Thunnus albacares (Bonnaterre), using a vitellogeninlike protein

Sexing of living yellowfin tuna, Thunnus albacares (Bonnaterre), was carried out based on immunochemical detection of vitellogenin (VTG), a female-specific protein. Tissue samples were taken from swimming fish in a sea pen by a hand-made apparatus. Recently fertilized eggs were also collected before tissue sample trials. A double immunodiffusion pattern showed that the egg homogenate of T. albacares reacted with rabbit antibody against the purified VTG of greater amberjack, Seriola dumerili (Risso). This suggests that the VTG of T albacares has an antigenicity similar to that of S, dumerili and that this antibody can be used to detect T albacares VTG. Immunodot-blotting analysis revealed that, out of the two fish tested, the tissue sample from one, but not the other, reacted with this antibody, suggesting that the former was a mature female, and the latter a male or immature female. Examination of the fish gonads revealed an ovary and a testis, respectively. Immunological and morphological results corresponded completely. Therefore, the method used in the present study is available for discriminating the sexes of T albacares.     

Effect of temperature and photoperiod on the reproductive condition and performance of a tropical damselfish <Emphasis Type="Italic">Chrysiptera cyanea</Emphasis> during different phases of the reproductive season

Effects of temperature and photoperiod on the reproductive activity of a reef associated tropical damselfish Chrysiptera cyanea were evaluated under three phases with different environmental patterns, phase I (April–May; increasing water temperature and photoperiod), phase II (June–July; increasing water temperature and peak/decreasing photoperiod), and phase III (August–September; peak/decreasing water temperature and decreasing photoperiod). When the fish were reared at 20, 25, or 30°C under natural photoperiod, the reproductive conditions differed within and among the phases depending on experimental temperature and environmental patterns. From phases I through III, ovaries with vitellogenic oocytes were notable only at 25°C, whereas regressing and immature oocytes were noticed at 20 and 30°C. The fish underwent active spawning at 25°C, whereas no or few spawnings were observed at other temperatures. In phase III, there was a resultant prevention of decrease in the gonadosomatic index and disappearance of vitellogenic oocytes in the ovaries of fish under a long photoperiod (LD14:10) compared to those under a short photoperiod (LD10:14). These results indicate that a long photoperiod with a suitable range of water temperature is a principal determinant in continuity of reproductive activity and performance, and that a high temperature has a negative impact on their ovarian development.     

Assessing sexual maturity of feminized Japanese eel Anguilla japonica by measuring eye size

For efficient production of Japanese eel Anguilla japonica eggs, knowledge of the status of the sexual maturity of potential broodstock females is important because this status directly influences the time required to produce mature eggs by successive hormone doses. Here we apply an eye index (relative eye size) to evaluate the gonadal status of feminized A. japonica, which were induced by administration of estradiol-17β. Examination of gonad somatic and eye indexes of 267 feminized eels, cultivated for 12–56 months, revealed a significant correlation between these two indexes. Artificial maturation of 35 feminized eels showed that the number of hormone injections administered before ovulation was significantly lower as the eye index score increased, indicating availability of this noninvasive method of predicting sexual maturity of feminized eel. There was no correlation between eye index and fertilization rate, hatching rate, or larval survival rate, suggesting that sexual maturity before hormonal treatment does not affect egg quality.     

Effect of salinity on occurrence of notochord deformities in Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis> larvae

One of the difficulties to rear Anguilla japonica larvae is the frequent occurrence of notochord deformities. We tested the effect of salinity on the occurrence of the deformities, because we have been using 50 % diluted seawater (50 % SW) for glass eel production, on the basis of the fact that intermediate salinity saves energy due to lower cost for osmoregulation and contributes higher survival and growth rates. We reared 6-day-old larvae in 50 and 100 % SW for 85 days and observed their morphology. The occurrence rate of deformed larvae, including kyphosis and scoliosis, was significantly higher in 50 % SW (35.8 %) than in 100 % SW (25.4 %), while survival rate was significantly higher in 50 % SW (69.8 %) than in 100 % SW (32.3 %) and growth in 50 % SW (mean body depth: 7.9 ± 5.3 mm) was better than in 100 % SW (6.8 ± 4.6 mm). We speculate that the most of severely deformed larvae could not survive in the tougher condition of 100 % SW, showing the lower occurrence of deformed larvae. Eventually, the yield of normal larvae after 85 days rearing was 1.9-fold higher in 50 % SW than in 100 % SW, implying that the advantage of 50 % SW for rearing eel larvae outweighs the risk of deformities.     

Structure-Activity Relationship Study of the Neuritogenic Potential of the Glycan of Starfish Ganglioside LLG-3

LLG-3 is a ganglioside isolated from the starfish Linchia laevigata. To clarify the structure-activity relationship of the glycan of LLG-3 toward rat pheochromocytoma PC12 cells in the presence of nerve growth factor, a series of mono- to tetrasaccharide glycan derivatives were chemically synthesized and evaluated in vitro. The methyl group at C8 of the terminal sialic acid residue was crucial for neuritogenic activity, and the terminal trisaccharide moiety was the minimum active motif. Furthermore, the trisaccharide also stimulated neuritogenesis in human neuroblastoma SH-SY5Y cells via mitogen-activated protein kinase (MAPK) signaling. Phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 was rapidly induced by adding 1 or 10 nM of the trisaccharide. The ratio of phosphorylated ERK to ERK reached a maximum 5 min after stimulation, and then decreased gradually. However, the trisaccharide did not induce significant Akt phosphorylation. These effects were abolished by pretreatment with the MAPK inhibitor U0126, which inhibits enzymes MEK1 and MEK2. In addition, U0126 inhibited the phosphorylation of ERK 1/2 in response to the trisaccharide dose-dependently. Therefore, we concluded that the trisaccharide promotes neurite extension in SH-SY5Y cells via MAPK/ERK signaling, not Akt signaling.