Time-course comparison of pulmonary inflammation induced by intratracheal instillation of four different nickel oxide nanoparticles in male Fischer rats

Occupational exposure to nickel oxide (NiO) is an important cause of respiratory tract cancer. Toxicity is known to be associated with the dissociated component, i.e. nickel (II) ions. To address the relationship between physicochemical properties, including solubility in artificial lysosomal fluid, of NiO and time-course changes in the pulmonary response, we conducted an intratracheal instillation study in male Fischer rats using four different well-characterized NiO products, US3352 (NiO A), NovaWireNi01 (NiO B), I small particle (NiO C), and 637130 (NiO D). The NiOs were suspended in purified water and instilled once intratracheally into male F344 rats (12 weeks old) at 0 (vehicle control), 0.67, 2, and 6 mg/kg body weight. The animals were euthanized on days 3, 28, or 91 after instillation, and blood analysis, bronchoalveolar lavage fluid (BALF) testing, and histopathological examination were performed. The most soluble product, NiO B, caused the most severe systemic toxicity, leading to a high mortality rate, but the response was transient and surviving animals recovered. The second-most-soluble material, NiO D, and the third, NiO A, caused evident pulmonary inflammation, and the responses persisted for at least 91 days with collagen proliferation. In contrast, NiO C induced barely detectable inflammation in the BALF examination, and no marked changes were noted on histopathology. These results indicate that the early phase toxic potential of NiO products, but not the persistence of pulmonary inflammation, is associated with their solubility.     

The palliative efficacy of modified Mohs paste for controlling canine and feline malignant skin wounds

In veterinary medicine, the management of malignant skin wounds is highly challenging. We conducted a study on seven case animals (four dogs and three cats) which presented with malignant skin wounds. All seven animals had signs and symptoms which were controlled following treatment with a modified Mohs paste. Upon obtaining informed consent from their owners, the animals requiring management of malignant wounds were enrolled in this study. The modified Mohs paste was prepared by mixing zinc chloride, zinc oxide starch powder, glycerin, and distilled water. The modified Mohs paste was topically applied to and left to remain on the malignant wounds for one hour, under controlled conditions. Once the paste was removed, the wounds were irrigated with a solution of sterile saline. At the first examination, the wounds of each animal were observed for signs of exudate, malodor, and bleeding. In every case, visible improvement was observed immediately after the modified Mohs paste treatment. Specifically, the size of the malignant wounds, and the number of times the dressing gauze required changing, significantly decreased (p < 0.05 and p < 0.01, respectively). The open malignant skin wounds caused by mammary gland tumors disappeared in two cases. The Mohs paste has been shown to be a viable option for the palliative treatment in canine and feline malignant skin wound management.     

Estrone sulfate and progesterone profiles during late gestation in recipient cows transferred embryos produced by nuclear transfer and in vitro fertilization

The aim of present experiment was to evaluate the plasma concentrations of estrone sulfate (E(1)S) and progesterone (P(4)) during late gestation in recipient cows transferred embryos produced by nuclear transfer (NT) and in vitro fertilization (IVF). Blood samples were collected from recipients transferred embryos produced by NT (n=9) and IVF (n=13) at 160, 220, 240, 260 and 270 d of gestation and then at 5 d intervals until parturition. Plasma samples were analyzed for E(1)S and P(4) by ELISA. One NT and three IVF cows aborted between days 220 and 260 of gestation. Two NT and one IVF cow had prolonged gestation (over 290 d). One IVF cow had an overweight fetus (50 kg) after abortion (257 d). The patterns of changes in the concentrations of E(1)S during late gestation in the NT and IVF cows were almost identical. The NT and IVF cows that aborted had prolonged gestation and much higher E(1)S concentrations than the average. One NT cow aborted after 220 d of gestation and had a sudden high increase in its E(1)S concentration from 160 d to 220 d of gestation. The NT and IVF cows that had prolonged gestation also had significantly higher (P<0.05) P(4) concentrations than the average. These results raise the possibility that the E(1)S and P(4) profiles can be used to monitor some late gestational problems, such as higher birth weight, abortion and prolonged gestation.     

Development and evaluation of pyramiding lines carrying early or late heading QTLs in the indica rice cultivar ‘IR64’

The heading date is an important trait for determining regional and climatic adaptability in rice. To expand the adaptability of the indica rice cultivar ‘IR64’, we pyramided multiple early or late heading quantitative trait locus (QTLs) in the ‘IR64’ genetic background by crossing previously developed near-isogenic lines (NILs) with a single QTL for early or late heading. The effects of pyramiding QTLs were observed in three different climatic zones of the Philippines, Madagascar, and Japan. The early heading pyramiding lines (PYLs) headed 6.2 to 12.8 days earlier than ‘IR64’ while the late heading PYLs headed 18.8 to 27.1 days later than ‘IR64’. The PYLs tended to produce low grain yield compared to ‘IR64’. The low yield was not improved by combining SPIKE, which is a QTL that increases the number of spikelets per panicle. Conversely, ‘IR64-PYL(7+10)’ carrying Hd5 and Hd1 headed earlier, produced more tillers, and more panicles per m² than ‘IR64’, and mitigated the yield decrease in early heading. These results suggest that the effects of pyramided QTLs on heading date were consistent across various environments and PYLs could be used to enhance the adaptation of ‘IR64’ in other rice growing environments.     

Establishment of testicular and ovarian cell lines from Honmoroko (Gnathopogon caerulescens)

We succeeded to establish cell lines from endemic fish species Honmoroko Gnathopogon caerulescens, which inhabits Lake Biwa, the third oldest lake in the world. Two cell lines designated as RMT1 and RMO1 were established from testis and ovary of G. caerulescens, respectively. These cell lines were initially cultured in Leibovitz’s L-15 medium supplemented with fetal bovine serum (FBS), fish embryo extract, epidermal growth factor, and basic fibroblast growth factor. Further addition of forskolin and β-mercaptoethanol was required to establish and maintain these cell lines for more than 60 passages. RMT1 and RMO1 cells showed fibroblast- and epithelial-like morphology, respectively. From immunocytochemical staining and gene expression patterns, RMT1 cells showed a characteristic of testicular Sertoli cells and RMO1 cells did that of ovarian theca cells. Both RMT1 and RMO1 cells multiplied well in the medium supplemented with 10 % FBS at 28 °C and their minimum population doubling times were 24.4 and 28.8 h, respectively. At the 45th passage, most of the RMT1 and RMO1 cells had a hyperploid set of chromosomes (67.3 and 96.1 %, respectively). Cells with normal diploid chromosome set were not observed. RMT1 cells were transfected with an enhanced green fluorescent protein (EGFP) expression vector and human elongation factor 1 α promoter worked efficiently to express EGFP. In addition, EGFP-expressing cell lines were also established, suggesting that the cell lines could be utilized as an in vitro monitor system (biosensor) for the evaluation of endocrine disruptors which might affect gonadal function.     

Effect of artemia nauplii enriched with vitamin A palmitate on hypermelanosis on the blind side in juvenile Japanese flounder Paralichthys olivaceus

ABSTRACT:   The effect of Artemia nauplii enriched with different level of vitamin A (VA) palmitate (1 µg = 1 IU) on the occurrence of hypermelanosis on the blind side of Japanese flounder Paralichthys olivaceus was determined. Artemia were enriched with 0, 1, 2, 5 or 10 mg VA palmitate/L (control group, and 1-, 2-, 5-, and 10-mg groups). The enriched Artemia were fed to the larvae from 27 to 31 days post hatching (dph) corresponding to the F–G stage. VA palmitate, retinol and retinoic acid (RA) contents of Artemia were correlatively elevated with increasing VA palmitate in the culture medium. RA was detected in Artemia enriched with 5 mg and 10 mg, and a significantly high frequency of hypermelanosis on the blind side was observed in these groups at 65 dph ( P  < 0.05). These results suggest that RA synthesized from VA palmitate in Artemia could induce hypermelanosis on blind side of flounder when Artemia are enriched with more than 5 mg VA palmitate/L.     

Binding profiles and cytokine-inducing effects of fish rhamnose-binding lectins on Burkitt’s lymphoma Raji cells

Rhamnose-binding lectin (RBL) is one of the animal lectin categories which take part in the innate immune responses of fish. Osmerus lanceolatus lectin (OLL) from shishamo smelt eggs is an RBL composed of two tandem-repeated domains, both of which are considered to be a carbohydrate-recognition domain. SAL, catfish (Silurus asotus) egg RBL composed of three domains, binds to Burkitt’s lymphoma Raji cells through globotriaosylceramide (Gb3) carbohydrate chain and to reduce cell size and growth by altering membrane composition without causing cell death. In this experiment, we tried to compare the binding effects of these two RBLs on Raji cells. Flow cytometric and fluorescence microscopic analyses revealed that OLL also directly bound to and shrunk Raji cells with ten times less reactivity than SAL but reduced cell growth with decreasing cell viability. Anti-Gb3 antibody completely blocked the binding of SAL to Raji cells but not that of OLL. In addition, the direct bindings of OLL and SAL to Raji cells were comparably inhibited by melibiose, but lactose was more effective inhibitor for the binding of OLL than that of SAL. These results suggest that OLL has slightly different cell-binding property compared with SAL and binds not only to Gb3 but also to the other carbohydrate receptor-bearing β-galactoside chains. The quantitative RT-PCR analysis revealed that SAL induced the expression of TNF-α but not of IFN-γ, IL-1β, and IL-10. Thus, SAL-induced cytostatic effect on Raji cells might be partially caused by TNF-α-mediated signaling pathway.     

Susceptibility by amberjack (Seriola dumerili), yellowtail (S. quinqueradiata) and Japanese flounder (Paralichthys olivaceus) to Neobenedenia girellae (Monogenea) infection and their acquired protection

This study investigated: 1) susceptibility differences to infection by Neobenedenia girellae (Capsalidae) between amberjack Seriola dumerili (Carangidae), yellowtail S. quinqueradiata and Japanese flounder Paralichthys olivaceus (Paralichthyidae); 2) growth and egg production of N. girellae on each fish species; 3) acquired protection of each fish species against this parasite. The number of N. girellae on S. dumerili was significantly higher than on S. quinqueradiata and P. olivaceus when these fishes were exposed to oncomiracidia in the same aquarium. Neobenedenia girellae growth on S. dumerili was fastest and, thus the number of eggs laid by parasites on S. dumerili was greater than on the other two species. Seriola dumerili and P. olivaceus, which were previously infected with N. girellae and treated by freshwater bath, acquired partial protection against re-infection by N. girellae. The relative re-infection of three S. dumerili individuals out of eleven individuals was markedly low compared with the initial infection, and the relative initial infection and re-infection on two P. olivaceus out of eleven individuals was markedly low. The results of this study could be useful to control N. girellae infections when cultivating S. dumerili, S. quinqueradiata and P. olivaceus.     

Immunohistochemical analysis of pseudorabies virus spread through neurons innervating the eyeball

Pseudorabies virus (PRV) was inoculated intraocularly into BALB/c mice, and the distribution pattern of cells positive for several neurotransmitters and viral antigens in the eyeball, trigeminal nerve ganglia, and superior cervical ganglia was examined immunohistochemically to clarify the neural route of the virus spread. In the eyeball, substance P (SP)- and calcitonin gene-related peptide (CGRP)-positive cells were detected in the ipsilateral iris and ciliary body, neuropeptide tyrosine (NPY)-positive cells were detected in the choloid membrane, and tyrosine hydroxylase (TH)-positive cells were detected in the ipsilateral inner nuclear layer of the retina; all these cells contained viral antigens. In the superior cervical ganglia, viral antigen-positive cells containing TH or NPY were found at bilateral sites. In the trigeminal nerve ganglia, viral antigen-positive cells containing SP or CGRP were found at bilateral sites. These findings indicated that the SP- and CGRP-positive ganglion cells of the trigeminal nerve ganglia innervating the iris or ciliary body, and the NPY-positive ganglion cells of the superior cervical ganglia innervating the iris, ciliary body, and choroid membrane served as the route for the virus spread. These findings also suggested that dopaminergic neurons, such as the TH-positive retinal cells and TH-positive ganglion cells of the superior cervical ganglia, served as the route for virus spread.