Thirteen-week Intravenous Toxicity Study of a Novel Humanized Anti-Human Death Receptor 5 Monoclonal Antibody, CS-1008, in Cynomolgus Monkeys

CS-1008, a humanized monoclonal antibody that is agonistic to human death receptor 5, was intravenously administered to cynomolgus monkeys twice a week for 13 weeks at 3 different dose levels (5, 15 and 42 mg/kg) in order to evaluate its potential toxicity. A control group received phosphate buffered saline containing 0.01% polysorbate 80. Each of the 4 groups consisted of 3 male and 3 female cynomolgus monkeys. No animal in any group died during the dosing period. No toxic changes in clinical signs, food consumption, body weight, electrocardiography, ophthalmology, urinalysis, hematology, blood chemistry, gross pathology, organ weights or histopathology were noted in any group during the dosing period. In the toxicokinetic analysis, the values for the maximum concentration of CS-1008 in plasma and the area under the curve generally increased with increasing dose. No clear differences in the toxicokinetic parameters or profiles were observed between the sexes. Development of anti-CS-1008 antibodies was not detected in any sample. The no-observed adverse-effect level (NOAEL) of CS-1008 in cynomolgus monkeys under the conditions of this study was concluded to be 42 mg/kg in both sexes, when administered intravenously twice a week for 13 weeks. This study supports the development of CS-1008 as a therapeutic biopharmaceutical.     

Basal levels and GnRH-induced responses of peripheral testosterone and estrogen in Holstein bulls with poor semen quality

The present study investigated the basal levels and GnRH-induced responses of peripheral testosterone and estrogen in Holstein bulls with poor semen quality. On the basis of semen parameters, bulls (n=5) having poor semen quality were selected as experimental bulls, and good semen quality bulls (n=4) were used as control bulls. Both groups were treated intramuscularly once with GnRH (250 μg of fertirelin acetate). Blood samples were collected at -1 day (d), -30 min and 0 h (treatment) followed by every 30 min for 5 h and 1, 3 and 5 d post-GnRH treatment (PGT), and LH, testosterone and estradiol-17β (E(2)) concentrations were measured. The pretreatment concentrations were used as basal levels. The percentage increments based on the 0-h levels were calculated per bull for each sampling time until 5 h PGT, and differences were compared between the experimental and control groups. The PGT concentrations of testosterone and basal and PGT concentrations of E(2) were significantly lower in the experimental group. The testosterone increment in the experimental group was delayed and significantly lower from 1 to 5 h PGT than those in the control group. It can be suggested that bulls with poor semen quality have delayed and lower GnRH-induced testosterone response and may also have lower estrogen levels.     

Oestradiol enhances plasma growth hormone and insulin-like growth factor-I concentrations and increased the expression of their receptors mRNAs in the liver of ovariectomized cows

Many metabolic hormones, growth hormone (GH), insulin-like growth factor-I (IGF-I) and insulin affect ovarian functions. However, whether ovarian steroid hormones affect metabolic hormones in cattle remains unknown. This study aimed to determine the effect of sex steroids on the plasma profiles of GH, IGF-I and insulin and their receptors in the liver and adipose tissues of dairy cows. Ovariectomized cows (n = 14) were randomly divided into four groups: control group (n = 3) was treated with saline on Day 0; oestradiol (E2) group (n = 3), with saline and 1 mg oestradiol benzoate (EB) on Day 0 and 5, respectively; progesterone (P4) group (n = 4) with two CIDRs (Pfizer Inc., Tokyo, Japan) from Day 0; and E2 + P4 group (n = 4) with two CIDRs on Day 0 that were removed on Day 6 and were immediately injected with 1 mg EB. The animals were euthanized after the experiment, and liver and adipose tissues samples were quantitatively analysed using real-time PCR for the expression of mRNA for the GH (GHR), IGF-I (IGFR-I) and insulin (IR) receptor mRNAs. Oestradiol benzoate significantly increased the number of peaks (p < 0.05), pulse amplitude (p < 0.05) and area under the curve (AUC; p < 0.01) for plasma GH; moreover, it increased plasma IGF-I concentration (p < 0.05), but it had no effect on the plasma insulin profile. P4 significantly decreased the AUC (p < 0.01), compared with the control group, whereas it did not affect the number of peaks and the amplitude of GH pulses. P4 + E2 did not affect the GH pulse profile. E2 increased the mRNA expression of GHR, IGFR-I and IR in the liver (p < 0.05), whereas both P4 and E2 + P4 did not change their expressions. Our results provide evidence that the metabolic and reproductive endocrine axes may regulate each other to ensure optimal reproductive and metabolic function.     

Occurrence of Corynespora cassiicola isolates resistant to boscalid on cucumber in Ibaraki Prefecture, Japan

A total of 651 isolates of cucumber corynespora leaf spot fungus ( Corynespora cassiicola ) collected from cucumber in Japan, either with (438 isolates) or without (213 isolates) a prior history of boscalid use, were tested for their sensitivity to boscalid by using a mycelial growth inhibition method on YBA agar medium. Additionally, seven isolates of C. cassiicola obtained from tomato, soybean, eggplant (aubergine) and cowpea in different locations in Japan were tested before boscalid registration. Minimum inhibitory concentration (MIC) and 50% effective concentration (EC₅₀) values for 220 isolates from crops without a prior history of boscalid use ranged from 0·5 to 7·5 μg mL⁻¹ and from 0·04 to 0·59 μg mL⁻¹, respectively. Two hundred and fourteen out of 438 isolates collected from ten cucumber greenhouses in Ibaraki Prefecture, Japan, which received boscalid spray applications showed boscalid resistance, with MIC values higher than 30 μg mL⁻¹. Moreover, resistant isolates were divided into two groups: a moderately resistant (MR) group consisting of 189 isolates with EC₅₀ values ranging from 1·1 to 6·3 μg mL⁻¹, and a very highly resistant (VHR) group consisting of 25 isolates with EC₅₀ values higher than 24·8 μg mL⁻¹. MR isolates were detected from all ten greenhouses, but VHR isolates were detected from only three. As a result of fungus inoculation tests which used potted cucumber plants, control failures of boscalid were observed against resistant isolates. Efficacy of boscalid was remarkably low against VHR isolates in particular. This is the first known report on boscalid resistance in Japan.     

Inhibition by pulp juice and enhancement by ingestion on germination of bird-dispersedPrunus seeds

To evaluate what prevents seed germination and why ingestion by birds enhances germination, we carried out germination experiments using bird-dispersed seeds of twoPrunus species (Prunus sargentii andP. ssiori) in the laboratory. Germination of seeds with following four treatments was compared: “Ingested seeds” excreted after feeding of fruits to birds; “Extracted seeds” deliberately extracted from the fruit pulp; “Juiced seeds” which were supplemented with the juice of the pulp after seeds had been deliberately extracted from the pulp; and “Intact seeds” in untreated whole fruits. Many ingested (27%, 23% inP. sargentii, 18% inP. ssiori) and extracted seeds (24% inP. sargentii, 17% inP. ssiori) germinated, and difference in germination percentages between ingested and extracted seeds were not significant, for eitherPrunus species. The Juiced seeds rarely germinated (2% inP. sargentii, 6% inP. ssiori), and seeds within intact fruits did not germinate. These results suggest that the fruit pulp and the juice of the pulp inhibit germination and the seeds could germinate without ingestion by birds once they had been manually extracted from pulp. Therefore, it is considered that bird ingestion enahnces germination by removing pulp, especially juice of the pulp, in these twoPrunus species.     

Morphological changes of epiphyseal plate in the long bone of chondrodysplastic dwarfism in Japanese brown cattle.

A total of 20 cases of disproportionate calves on Japanese Brown cattle were examined morphologically. Investigation of 5 affected calves revealed that the insufficiency of endochondral ossification was confined only to the long bones of the limbs and was not accompanied by other associated changes. On the basis of the histological changes of epiphyseal plate and the affected site, this condition may be called chondrodysplasia of the rhizomelic type. Therefore, this disorder was defined as a bovine dwarfism. From three dimensional image analysis and histological changes of the tibial proximal portion in the other 15 cases, deformity and shortness of the tibia were related to the state of distribution and the degree of damages of the epiphyseal plate. In the affected cartilage matrix, cystoid degeneration, fibrous striation, bone spicule, necrotic foci and rent were found. Inadequate metachromasia suggests the abnormal staining of sulfated glycosaminoglycans and alcian blue stainability may be attributable to the abnormal interactions between proteoglycan and other matrix components. At the interface of cartilage and bone, irregular calcification, fibrotic scar and sealing by osseous tissue with relation to vascularity were found. These changes in this study indicated the failure of modeling on the epiphyseal plate, showing disturbance of chondrocytic differentiation and abnormal formation of the matrix.