Lipophilic toxin pectenotoxin-2 (PTX2) is oxidatively metabolized to pectenotoxin-1 (PTX1), pectenotoxin-3 (PTX3), and pectenotoxin-6 (PTX6) in the Japanese scallop Patinopecten yessoensis. This particular metabolism has been observed only in Japanese scallops, in which PTX6 is the most dominant lipophilic toxin. We investigated the cytotoxicity of PTX2 and its metabolites PTX1,3,6 in a rat cell line (L6) and a human cell line (RD). RD showed an approximately three-fold greater sensitivity than L6 upon exposure to PTXs. The cytotoxicity of PTXs decreased with degree of oxidation in the order PTX2 > PTX1 > PTX3 > PTX6. The calculated half maximal inhibitory concentration (IC₅₀) values of PTX2 obtained for the L6 and RD cell lines were 60 and 23 ng/mL, respectively, while those obtained for PTX6 for both cell lines were over 2,000 ng/mL. These results demonstrate that PTX6 has extremely low cytotoxicity or is non-toxic and that the oxidative metabolism of PTX2 in P. yessoensis is a detoxification process.

Drug-induced hepatotoxicity is a common reason for discontinuing the development of candidate clinical drugs. In the present study, we investigated the utility of three-dimensionally cultured human hepatocytes (spheroids) for prediction of hepatotoxicity, using a panel of model drugs: acetaminophen, benzbromarone, chlorpromazine, cyclosporin A, diclofenac, fialuridine, flutamide, imipramine, isoniazid, ticlopidine and troglitazone. Cultured spheroids showed a significant increase of albumin secretion from 2 to 7 days; the secretion started to decrease at 14 days, but continued from 14 days to 21 days. The morphology of the spheroids was well maintained for 21 days. Long-term exposure of spheroids to hepatotoxic drugs resulted in concentration-dependent depression of albumin secretion and elevation of aspartate aminotransferase (AST) leakage. The estimated 50% effective concentration (IC₅₀) values for decrease of albumin secretion changed from 7 days to 14 days, but similar values were obtained at 14 and 21 days, except for diclofenac. Since the IC₅₀ values and the values of drug concentration inducing 1.2-fold elevation of AST leakage (F1.2) were similar at 14 and 21 days, an incubation period of 14 days was considered sufficient. The coefficient of determination (R²) between IC₅₀ values and F1.2 values of all drugs was 0.335. When cyclosporine A and fialuridine were excluded, the value of R² became 0.887. The results indicate that the proposed human hepatocyte spheroid assay should be helpful in the evaluation of hepatotoxicity during the early development stage of clinical drug candidates.

We developed a newborn mouse behavioral testing method to evaluate the risk of neurotoxicity of environmental toxicants, based on determining a newborn’s motor activity by applying the tare function of an analytical balance. Motor activities of newborn ICR mice exposed prenatally to diethylstilbestrol (DES) at 0.005-0.5 μg/kg/day on days 5 through 18 of gestation were evaluated on postnatal day 1. The activities of male newborns in the 0.05 μg/kg/day group were significantly increased compared to those of the controls, and the increasing tendencies were observed in both sexes of the highest group. The findings indicate that prenatal exposure to low doses of DES causes hyperactivity in newborn mice.

To characterize variability of various musculoskeletal biomarkers by different blood sampling techniques in conscious rats, plasma asparate aminotransferase (AST), creatine kinase (CK) and its isoenzymes, fatty acid binding protein 3 (FABP3), myosin light chain 3 (Myl3) and microRNA (miR-133a) obtained by jugular venipuncture (C-JV) or tail venipuncture (C-TV) were compared with those obtained by jugular venipuncture (A-JV) in isoflurane-anesthetized rats. Plasma CK, FABP3 and Myl3, especially when collected by C-TV, were higher with larger variability than when collected by A-JV, whereas miR-133a displayed large variability in all techniques. Interestingly, higher CK obtained by C-JV or C-TV was largely attributable to higher CK-MM or CK-BB, respectively. Handling and restraint stress were identified as possible factors contributing to larger variability for CK, FABP3 and Myl3. A close correlation between CK and FABP3 was demonstrated both in the C-JV and C-TV techniques. Next, we evaluated the impact of C-JV and C-TV techniques for detecting skeletal myopathy in 2,3,5,6-tetramethyl-p-phenylenediamine-treated rats. In this model, CK and CK-MM obtained by C-TV were significantly increased, but those obtained by C-JV were not modified. In contrast, AST, FABP3, Myl3 and miR-133a obtained by both techniques were drastically elevated to a similar extent. The results suggest that, in conscious rats, the tail venipuncture technique may be more appropriate to detect skeletal myopathy despite the higher variability with this technique than with the jugular venipuncture technique. Furthermore, FABP3, Myl3 and miR-133a may serve as more sensitive biomarkers with large signal-to-noise ratios regardless of the blood sampling technique in conscious rats.

Nanomaterials have been proposed as novel substrates for medical and commercial applications. However, such materials also may have novel toxicities, thus posing environmental and health concerns. We previously reported hepatic injury in mice following the intravenous administration of unmodified silica particles with diameters of 70 nm (SP70); this toxicity was not observed following administration by the same route of micro-size particles with diameters of 300 nm (SP300) or 1,000 nm (SP1000). In the present study, we used electron microscopy to investigate the dynamics of silica nanoparticles administered in mice. SP70 was observed in hepatocytes and in lung epithelial cells. Inclusion within hepatocytes was associated with accumulation of SP70 in the liver sinusoidal endothelial cells and passage through the space of Disse. In contrast, SP300 and SP1000 were not observed within the hepatocytes. To our knowledge, our report represents the first demonstration that silica nanoparticles accumulate in hepatocytes, liver sinusoidal endothelial cells, Kupffer cells, and lung tissue; accumulation of SP70 in liver sinusoidal endothelial cells correlated with the induction of liver injury.

The purpose of this study was to evaluate the effects of sub-chronic exposure to deltamethrin in lower doses on the acquisition of a two-way avoidance task and the levels of amino acid neurotransmitters in hippocampus of mice measured using shuttle-box and LC-MS/MS system. Deltamethrin was given to mice respectively at doses of 0.46, 0.92, 1.80 mg/kg BW daily for 60 days by gavage. Deltamethrin was found to decrease the number of avoidance responses, increase response latency, and increase glutamate levels in the 0.92 and 1.80 mg/kg BW-dose group. As revealed by electron microscopy, in 0.92 and 1.8 mg/kg-dose group mice, morphology of cells were changed and degeneration and necrosis morphological characteristics obviously were appeared. Collectively, results from this study suggest that deltamethrin may have cumulative effects in mice following repeated dosing of deltamethrin using moderately effective doses, beside Na⁺ and Ca²⁺ channels as well as Na⁺ and Ca²⁺-dependent glutamate release, may be involved with neurotoxic action of deltamethrin.

Lepidopteran species fly freely in the environment and their larvae feed on the leaves of host plants which may be exposed to nanomaterials. As an ecological model of nanoparticle exposure, 5th instar larvae of the sweet potato hornworm (Agrius convolvuli) were subcutaneously injected with suspensions of 10 μL (100 μg/mL) titanium dioxide nanoparticles (TiO₂-NPs), 10 μL (100 μg/mL) zinc oxide nanoparticles (ZnO-NPs) or saline (control) and the effects on spermatogenesis were examined in the pupal testis. During the larval wandering stage, larval tissues (except the testis) underwent extremely rapid histolytic changes. Pupation and emergence were not affected by the injection. On pupal day 4, there was a significant decrease in testis weight and the number of sperm bundles in the ZnO-NPs group. Electron microscopic observation revealed that cyst cells surrounding the spermatogenic cells took on small agglomerates of TiO₂-NPs or ZnO-NPs by phagocytosis. As spermatogenesis advanced in the nanoparticle-injected groups, vacuoles of various sizes were found in the nuclei of spermatocytes, the nuclear chromatin of spermatids was uncondensed and some vacuoles were found in the nuclei of sperm bundles. A possible mechanism for this is that abnormal vacuoles disturbed the chromatin condensation, resulting in the decrease of sperm bundles. Toxicity of manufactured TiO₂-NPs and ZnO-NPs were demonstrated detriment to insect spermatogenesis.