The effects of repeated administration of phenobarbital (PB) on blood coagulation time were examined using male Japanese white SPF rabbits, which are widely used for toxicological studies. PB was administered to the rabbits by oral gavage for 2 weeks, at dose levels of 0, 12.5, 25 and 50 mg/kg/day. Blood was collected on Days 8 and 14 after each day’s dosing to perform blood coagulation examination. The liver was excised, weighed and examined histopathologically. Activated partial thromboplastin time (APTT) was prolonged at dose levels of 12.5 mg/kg/day or more and Thrombotest® (TBT) was prolonged at 50 mg/kg/day on Day 8. APTT was prolonged at dose levels of 12.5 mg/kg/day or more, TBT was prolonged at 25 mg/kg/day or more and factor IX activity decreased at 50 mg/kg/day on Day 14. At pathological examination, liver weight increased at dose levels of 25 mg/kg/day or more, and a ground-glass appearance of the hepatocytes was observed in the central and middle parts of lobules at 12.5 mg/kg/day or more. However, changes in factor VII or X activity or prolongation of prothrombin time (PT) were not observed. Therefore, prolongation of blood coagulation time by PB administration in rabbits was considered to be due to PB’s effect on the endogenous pathway alone. Moreover, an increase in anti-thrombin III (ATIII) concentration was noted at 50 mg/kg/day; however, no change was noted at dose levels of 25 mg/kg/day or less. This suggests that the contribution of ATIII to the PB-induced prolongation of coagulation time in rabbits was small.
The use of nano-sized materials offers exciting new options in technical and medical applications. On the other hand, adverse effects on cells have been reported and may limit their use. In addition to physico-chemical parameters such as contamination with toxic elements, fibrous structure and high surface charge, the generation of radical species was identified as key mechanism for cytotoxic action of nanoparticles. The cytotoxic potential of nanoparticles in the absence of radical generation is less well investigated. This study aims to investigate the size-dependent effect of carboxyl polystyrene particles on cells to identify potential adverse effects of these particles. Particles were characterized in different solutions to assess the influence of the medium on size and surface charge. Viability, membrane integrity, apoptosis, proliferation and generation of oxidative stress were investigated. In addition the intracellular localization of the particles was recorded. 20 nm polystyrene particles induced cellular damage by induction of apoptosis and necrosis. These particles generated radicals to the same degree as larger polystyrene particles. Particles were taken up into endosomes and lysosomes in a size-dependent manner. Protein containing solutions led to increases in particle size, decreased cytotoxicity and reduced cellular uptake. It can be concluded that even in the absence of high surface reactivity and not linked to the generation of radicals nano-sized particles may cause cell damage. The mechanism of this damage includes apoptosis, necrosis and inhibition of proliferation.
Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels are widely used as sensitive markers of possible tissue damage, particularly liver toxicity. Lipid-lowering drugs, such as fibrates, slightly increase serum transaminase levels in humans, but there is little evidence that the phenomenon is related to drug-induced liver injury (DILI). Some in vitro studies have indicated that the elevations of serum transaminase activities after treatment of humans with fenofibrate, one of the fibrates, are related to increased transaminase synthesis in the hepatocytes rather than to transaminase leakage from the hepatocytes associated with cell lysis. In this study, male F344/DuCrlCrlj (Fischer) rats were treated once with fenofibrate at a dose level of 400 mg/kg and the relationships between the pharmacological effects, blood and hepatic transaminase activities and the gene expression of the transaminases in the liver were investigated. Fenofibrate treatment slightly increased plasma transaminase activities in rats with the findings directly related to the pharmacological action of the drug. The increases were in parallel with increases in hepatic transaminase activities associated with increases in the transaminase genes in the liver and were not considered to be a consequence of hepatotoxicity from the drug. The modification in transaminase gene expression is likely to be secondary to the pharmacological action of fenofibrate. The evidence obtained in our study underlines the importance of gene regulation as a possible alternative mechanism for increased blood transaminase activities.
Ubidecarenone (coenzyme Q10) has been widely used as a complementary therapy in heart failure and as a dietary supplement for over two decades. Ubidecarenone is manufactured by organic synthesis, yeast (non-Saccharomyces cerevisiae) fermentation, or bacteria fermentation. There are many reports on the safety of ubidecarenone. However, genotoxicity of ubidecarenone manufactured by bacteria fermentation has not been reported. We carried out genotoxicity evaluation of ubidecarenone manufactured by bacteria fermentation through the bacterial reverse mutation test (Ames test) and in vitro chromosome aberration test in compliance with the Japanese guidelines on genotoxicity testing of pharmaceuticals and the Organization for Economic Co-operation and Development (OECD) guidelines for testing chemicals. The results indicate neither increase of revertant colonies nor chromosome aberration, suggesting that the ubidecarenone manufactured by bacteria fermentation has no genotoxic activities under the condition of this study.
Fincoal type fluorosis has only been reported from China, but its pathogenesis is unclear. Many people believe that fluorosis is associated with oxidative stress. Oxidative stress can be reduced at higher selenium (Se) level. Heat shock protein (HSP70) is the most conserved and induced against different stressors. The aim of this study is to detect the expression of HSP70 in fluorosis patients and explore the role of Se in fluorosis protection. The subjects were divided into four groups: “High Se + F group” (n = 50), “High F group” (n = 50), “High Se group” (n = 20) and “Control group” (n = 46). Expression of HSP70 was evaluated by Western blotting and real-time PCR techniques. The concentration of fluoride, content of Se in hair, activity of antioxidant enzymes (GSH-Px, SOD, CAT) and content of malondialdehyde (MDA) were determined. The relative amount of HSP70 gene transcription was significantly higher in “High Se + F group” than the other groups. The same results were found for expression of HSP70 protein to β-actin ratio. There was a significant difference between “High Se + F group” and “High F group” regarding MDA content and glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) activity. These results suggest that oxidative stress plays an important role in the pathogenesis of the Fincoal type fluorosis and it can be reduced at higher Se level.
We recently demonstrated the incidence and multiplicity of N-methyl-N-nitrosourea (MNU)-induced mammary tumors to be increased by administration of acrylamide (AA) in post-initiation in rats. In the present study, to clarify the mechanisms of enhancement, H-ras gene mutations in mammary tumors induced in MNU-initiated rats with or without subsequent AA administration were investigated. Frequencies of mutations in codon 12 from GGA to GAA were significantly (p < 0.05) higher in rats with AA administration (82%, 23 out of 28 tumors) as compared to those without AA (50%, 9 out of 18 tumors), but the latency and volume of H-ras mutation-harboring tumors were similar to those of the mutation-lacking tumors. No mutations in codons 13 or 61 were detected in either treatment groups. The results thus indicate that H-ras gene mutations in codon 12 play a pivotal role in initiation of carcinogenesis and it appears possible that AA administration may selectively co-stimulate and/or maintain initiated cells via other genomic or non-genomic events in MNU-treated rats.
We searched a yeast genomic library for genes that conferred methylmercury resistance to yeast. Ymr258c was identified as a factor that conferred strong methylmercury resistance when overexpressed in budding yeast. Ymr258c is regarded as one of the F-box proteins, i.e., component factors of the Skp1/Cullin/F-box protein (SCF) complex that functions as a ubiquitin ligase in the ubiquitin-proteasome system. In this study, Ymr258c was demonstrated to function as an F-box protein to reduce methylmercury toxicity. The overexpression of Ymr258c might promote the ubiquitination of proteins that are involved in the enhancement of methylmercury toxicity, and thereby promote their degradation by proteasomes to reduce methylmercury toxicity.
The isotope 210Po was suspected of being involved in the death of a former Russian intelligence agent in 2006 in the UK. Although human exposure to this natural radionuclide in foods is estimated to be high, few studies are available. UNSCEAR Report 2000 does not contain data on 210Po concentrations of foodstuffs in Japan. We analyzed samples of the everyday Japanese diet cooked with foodstuffs purchased at supermarkets in 7 major domestic cities in 2007-2008. 210Po was quantified by alpha spectrometry and natural radionuclides such as 40K by gamma spectrometry. The daily intake and committed effective dose of 210Po, 40K, and other natural radionuclides for Japanese adults were calculated. Daily intake was 0.34-1.84 (mean ± σ : 0.66 ± 0.53) and 68.5-94.2 (81.5 ± 8.5) Bq/d and the committed effective dose was 0.15-0.81 (0.29 ± 0.24) and 0.16-0.21 (0.18 ± 0.02) mSv for 210Po and 40K, respectively, comprising a high percentage of the total exposure. The total of the mean committed effective dose for the two nuclides (0.47 mSv) was higher than the annual effective dose from ingestion of foods reported by UNSCEAR 2000 (0.29 mSv). The mean committed effective dose of 40K in the 7 major Japanese cities was comparable to the global average (0.17 mSv). The dietary exposure of Japanese adults can be characterized by a higher 210Po contribution than in other countries. Of the total daily dietary 210Po exposure (13 food categories excluding water) for adults in Yokohama, about 70% was from fish/shellfish and 20% from vegetables/mushrooms/seaweeds, reflecting preferences of Japanese to eat a considerable amount of fish/shellfish containing high 210Po concentrations.
Many medicines exist which can cause pruritus (itching) as "serious adverse events." Many severe pruritic conditions respond poorly to histamine H1 receptor antagonists; there is no generally accepted antipruritic treatment. Recently described histamine H4 receptors are expressed in haematopoietic cells and have been linked to the pathology of allergy and asthma. We previously reported their expression in human dermal fibroblasts; in this study we have investigated H4 receptor expression in human epidermal tissue and found it to be greater in keratinocytes in the epidermal upper layer than in the lower layer. We have also investigated the effect of histamine H4 receptor antagonists on histamine H1 receptor antagonist-resistant pruritus using a mouse model. Scratching behavior was induced by histamine (300 nmol) or substance P (100 nmol) injected intradermally into the rostral part of the back of each mouse. Fexofenadine, a histamine H1 receptor antagonist, reduced scratching induced by histamine but not by substance P, whereas JNJ7777120, a histamine H4 receptor antagonist, significantly reduced both histamine- and substance P-induced scratching. These results suggest that H4 receptor antagonists may be useful for treatment of H1 receptor antagonist-resistant pruritus.
Vascular toxicity is an important feature of the neuropathy induced by methylmercury. Methylmercury does not cause nonspecific cell damage, but rather retards the repair of wounded monolayers of cultured human brain microvascular endothelial cells by inhibiting their proliferation. Since vascular endothelial cell proliferation during the repair process strongly depends on the fibroblast growth factor-2 (FGF-2) system, we investigated the effects of methylmercury on the expression of FGF-2 and related proteins (i.e., FGF receptor 1 and perlecan) in cultured human brain microvascular endothelial cells. Of the mRNAs examined, FGF-2 mRNA expression was significantly lowered by methylmercury in not only wounded monolayers but also dense and sparse cultures of endothelial cells; a lower expression of FGF-2 protein in the cells was confirmed. In addition, exogenous FGF-2 partially abrogated the proliferation-inhibitory effect of methylmercury. The results of this study suggest that suppression of FGF-2 expression is one of the mechanisms underlying the inhibitory effect of methylmercury in damaged endothelial cell monolayers. The FGF-2 system may be one of the important biological systems behind the vascular toxicity of methylmercury.
We introduced a yeast open reading frame library into Saccharomyces cerevisiae strain BY4742 to search for genes whose overexpression conferred cadmium resistance to yeast, toward the goal of elucidating the mechanism of cadmium toxicity. As a result, we found that the overexpression of two newly identified genes, Ycg1 and Ydr520c, each conferred strong cadmium resistance to yeast.
We present herein transcriptional changes in mouse spinal cords in response to physical exercise on a treadmill using a DNA microarray. By 30-min exercise, the expression of 3 genes was enhanced and expression of 29 genes reduced. By continuous 2-week exercise (30-min exercise per day), the expression of 1 gene was enhanced and expression of 13 genes reduced.