Potential estrogenic and antiandrogenic effects of permethrin in rats

Many environmental chemicals including pesticides have been reported to possess hormonal activities, and thus are classified as endocrine disruptors. Permethrin, a synthetic pyrethroid insecticide, is used worldwide, which provides potential environmental exposure. However, relatively few studies have reported on hormonal activities, particularly estrogenic and androgenic activities of permethrin, and the results of these studies are in some respects contradictory. Therefore, this study investigated the potential estrogenic and androgenic activities of permethrin in vitro and in vivo. We conducted an uterine Calbindin-D9k (CaBP-9k) gene expression assay and an uterotrophic assay for estrogenic activity, and a Hershberger assay for androgenic activity. The CaBP-9k gene, one of the intracellular calcium binding proteins, is estrogen-responsive in the uterus. The rat uterotrophic and Hershberger assays are generally used as in vivo short-term screening assays for detecting the estrogenic and androgenic activities of chemicals, although these assays are still being validated by the Organization for Economic Cooperation and Development (OECD). Northern blot analysis showed the induction of uterine CaBP-9k mRNA level in response to permethrin as well as co-administration of permethrin with E2. In the uterotrophic assay using 18-day-old female rats, subcutaneous treatments with permethrin (10 to 800 mg/kg) for three days increased relative uterine wet weights, and E2-induced uterine weights. These effects were statistically significant at 800 and 200 mg/kg, respectively. Moreover, permethrin-induced uterine weights were inhibited by the co-administration of ICI 182,780, an antiestrogen. In the Hershberger assay, the administration of permethrin orally to testosterone propionate-treated castrated male rats led to statistically significant reductions in androgen-dependent sex accessory tissue (ventral prostate, seminal vesicles, levator ani and bulbocavernosus muscles, Cowper's gland and glans penis) weights at all doses tested (10, 50 and 100 mg/kg). These results suggest that permethrin might have estrogen-like effects on female rats, but antiandrogen-like effects on males.     

The Safety and efficacy of a new self-expandable intratracheal nitinol stent for the tracheal collapse in dogs

To evaluate the potential utility of a self-expandable intratracheal nitinol stent with flared ends for the treatment of tracheal collapse in dogs, endotracheal stenting therapy was performed under fluoroscopic guidance in four dogs with severe tracheal collapse. During the 4 to 7 month follow-up, after stent implantation, clinical signs, including dyspnea and respiratory distress, dramatically improved in all dogs. The radiographs showed that the implanted stents improved the tracheal collapse, and there were no side effects such as collapse, shortening or migration of the stents. In conclusion, the self-expandable intratracheal nitinol stents provided adequate stability to the trachea and were effective for attenuating the clinical signs associated with severe tracheal collapse.     

In vivo alternative testing with zebrafish in ecotoxicology

Although rodents have previously been used in ecotoxicological studies, they are expensive, time-consuming, and are limited by strict legal restrictions. The present study used a zebrafish (Danio rerio) model and generated data that was useful for extrapolating toxicant effects in this system to that of humans. Here we treated embryos of the naive-type as well as a transiently transfected zebrafish liver cell line carrying a plasmid (phAhRE-EGFP), for comparing toxicity levels with the well-known aryl hydrocarbon receptor (AhR)-binding toxicants: 3,3'',4,4'',5-pentachlorobiphenyl (PCB126), 2,3,7,8-tetrachlorodibenzo-p-dioxin, and 3-methylcholanthrene. These toxicants induced a concentration-dependent increase in morphological disruption, indicating toxicity at early life-stages. The transient transgenic zebrafish liver cell line was sensitive enough to these toxicants to express the CYP1A1 regulated enhanced green fluorescent protein. The findings of this study demonstrated that the zebrafish in vivo model might allow for extremely rapid and reproducible toxicological profiling of early life-stage embryo development. We have also shown that the transient transgenic zebrafish liver cell line can be used for research on AhR mechanism studies.     

The radioprotective effects of the hexane and ethyl acetate extracts of Callophyllis japonica in mice that undergo whole body irradiation

The radioprotective activity of extracts from the red seaweed Callophyllis (C.) japonica was investigated in mice that underwent whole-body exposure to gamma radiation. A methanol extract of C. japonica and its fractions [hexane, ethyl acetate (EtOAc), butanol and the remaining H₂O] were used. Each fraction (100 mg/kg body weight) was administered intraperitoneally (i.p.) 2 times into the BALB/c mice, once at 1 and once at 24 h before exposure to 9 Gray (Gy) of gamma radiation. Pre-irradiation administration of the hexane and EtOAc fractions saved the mice, with their survival rates being greater than 80% at 30 days post-irradiation; the mice that were pretreated with the other fractions showed survival rates lower than 20% over the same time period. To examine the effect of each C. japonica fraction on the survival of intestinal and bone marrow stem cells, the number of intestinal crypts and bone marrow cells in the gamma-irradiated mice were examined. Pre-treatment of mice (i.p., 100 mg/kg body weight at 1 and 24 h before irradiation) with the hexane or EtOAc fraction prior to 6-Gy irradiation significantly protected the number of jejunal crypts and bone marrow cells at 9 days after irradiation. These findings suggest that certain extracts from C. japonica, when they are administered prior to irradiation, play an important role in the survival of irradiated mice, and this is possibly due to the extracts protecting the hematopoietic cells and intestinal stem cells against gamma irradiation.     

Eosinophilic myositis in a slaughtered Korean native cattle

Histopathological findings of eosinophilic myositis in the carcass of a slaughtered Korean native cow are presented. Lesions contained massive fibrous septae with vacuolar changes in some lesions, and the hypercontraction and rupturing of muscle bundles, with replacement by eosinophils. Necrosis and severe eosinophil infiltration were observed. Sarcoplasmic fragmentation and atrophy developed. Typical of granuloma, calcified myofibers were focally surrounded by macrophages and numerous inflammatory cells, and multinucleated giant cell formation was evident.     

Monoclonal antibodies reactive with chicken interleukin-17

Chicken interleukin-17 (chIL-17) gene was previously characterized through cloning from a chicken intestinal expressed sequence tag (EST) cDNA library. To further investigate the biological properties of chIL-17, six monoclonal antibodies (mAbs) against a bacterially expressed chIL-17 recombinant protein were produced and their binding specificities characterized. Antibodies which were initially selected on the basis of their specific binding reactivity with recombinant chIL-17 in ELISA were further characterized by Western blot analysis. Monoclonal antibodies specific for chIL-17 identified 20 and 21kDa protein bands in the culture supernatant and cell lysate of CU205 cells. These mAbs also recognized specific bands for chIL-17 in the cell lysate from conconavalin A (Con A)-activated, but not from normal splenic lymphocytes. Furthermore, these mAbs detected a 16kDa protein in the lysate of CU205 cells treated with tunicamycin and stained an intracellular protein in CU205 cells in flow cytometric analysis. Together, these results indicate that these new mAbs are specific for chIL-17 and will be a useful tool for structural and immunological studies of IL-17 in poultry.     

Dexamethasone induces apoptosis in proliferative canine tendon cells and chondrocytes

Dexamethasone (Dexa) has been commonly used in humans and domestic animals, particularly in the treatment of tendon injuries and cartilage degeneration. However, it is often associated with tendon rupture and impaired tendon and cartilage healing. In the present study, we investigated Dexa's in vitro effects on the growth of cell proliferation and the induction of apoptosis in canine Achilles tendon cells and chondrocytes. Cell proliferation after treatment with Dexa for two to six days was quantified by a 2,3-bis{2-methoxy-4-nitro-5-sulfophenyl}-2H-tetrazolium-5-carboxyanilide inner salt assay (XTT). The results showed that Dexa could inhibit the proliferation of tendon cells and chondrocytes at increasing concentrations (0.1-50 microg/ml) compared with untreated cells. Cell apoptosis was induced by Dexa, as evidenced by the typical nuclear apoptosis using Hoechst 33258 staining. Dexa increased the apoptosis of canine tendon cells and chondrocytes in a time-dependent manner. In canine tendon cells and chondrocytes that were treated with 25 and 50 microg/ml concentration of Dexa, the number of condensed apoptotic nuclei was significantly increased. In addition, culturing with Dexa and the glucocorticoid receptor blocker, mifepristone, significantly arrested apoptosis of tendon cells and chondrocytes. Based on our in vitro data, we hypothesized that in vivo treatment with glucocorticoids may diminish the proliferation of tendon and cartilage cells by increasing apoptosis and suppressing the proliferation. Our findings suggest that Dexa could be used with caution in dogs with articular or tendon problems.     

Transient increases of glutamic acid decarboxylase 67 immunoreactivity and its protein levels in the somatosensory cortex after transient cerebral ischemia in gerbils

In this study, we investigated changes in glutamic acid decarboxylase 67 (GAD67) immunoreactivity and its protein levels in the gerbil somatosensory cortex after ischemia/reperfusion. GAD67 immunoreactivity was significantly increased in layers III and V of the somatosensory cortex 12 hr after ischemia/reperfusion. Thereafter, GAD67 immunoreactivity was decreased with time after ischemia/reperfusion. GAD67 immunoreactivity in the somatosensory cortex 4 days after ischemia/reperfusion was similar to that in the sham-operated group. In addition, GAD67 protein levels were also significantly increased 12 hr after transient forebrain ischemia. These results suggest that the transient increase of GAD67 immunoreactivity in layers III and V may be associated with responses to transient ischemia-induced neuronal damage.     

Time-course changes in the expression levels of miR-122, -155, and -21 as markers of liver cell damage,inflammation, and regeneration in acetaminophen-induced liver injury in rats

Drug-induced liver injury (DILI) is a significant threat to patient health and a major concern during drug development. Recently, multiple circulating microRNAs (miRNAs) have been reported to be potential biomarkers for DILI. To adapt and validate miRNAs for clinical use, we investigated the time-course changes in miR-122 expression levels in an acetaminophen-induced liver injury model in rats. In addition, miR-155 and miR-21 were evaluated as makers of inflammation and regeneration, respectively, to characterize liver status. Our results revealed that miR-122 is an early and sensitive biomarker of hepatocellular injury at a stage when alanine transaminase, aspartate transaminase, and total bilirubin were not detectable. However, no significant differences in the expression levels of other miRNAs (miR-155 and -21) were observed between treatment and vehicle groups. Collectively, these time-course changes in the expression levels of miRNAs may be useful as markers for clinical decision-making, in the diagnosis and treatment of DILI.     

Sequential disruption of ALV host receptor genes reveals no sharing of receptors between ALV subgroups A,B, and J

Background: Previously, we showed that targeted disruption of viral receptor genes in avian leukosis virus(ALV)subgroups using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9))-based genome editing confers resistance to ALV subgroups B and J. Here, we used the same strategy to target the receptor expressed by ALV subgroup A(TVA) and generate chicken cells resistant to infection by this virus.Results: CRISPR/Cas9-based disruption of exon 2 within the tva gene of DF-1 fibroblasts conferred resistance to infection by ALV subgroup A regardless of whether frameshift mutations were introduced during editing. Conversely,overexpression of the wild-type TVA receptor(wtTVA) by tva-modified DF-1 clones restored susceptibility to ALV subgroup A. The results confirm that exon 2, which contains the low-density lipoprotein receptor class A domain of TVA, is critical for virus entry. Furthermore, we sequentially modified DF-1 cells by editing the tva, tvb, and Na~+/H~+ exchange 1(chNHE1) genes, which are the specific receptors for ALV subgroups A, B, and J, respectively.Conclusions: Simultaneous editing of multiple receptors to block infection by different subgroups of ALV confirmed that ALV subgroups A, B, and J do not share host receptors. This strategy could be used to generate cells resistant to multiple viral pathogens that use distinct receptors for cell entry.