Inhibitory Effects of Prior Low-dose X-irradiation on Ischemia-reperfusion Injury in Mouse Paw

We have reported that low-dose, unlike high-dose, irradiation enhanced antioxidation function and reduced oxidative damage. On the other hand, ischemia-reperfusion injury is induced by reactive oxygen species. In this study, we examined the inhibitory effects of prior low-dose X-irradiation on ischemia-reperfusion injury in mouse paw. BALB/c mice were irradiated by sham or 0.5 Gy of X-ray. At 4 hrs after irradiation, the left hind leg was bound 10 times with a rubber ring for 0.5, 1, or 2 hrs and the paw thickness was measured. Results show that the paw swelling thickness by ischemia for 0.5 hr was lower than that for 2 hrs. At 1 hr after reperfusion from ischemia for 1 hr, superoxide dismutase activity in serum was increased in those mice which received 0.5 Gy irradiation and in the case of the ischemia for 0.5 or 1 hr, the paw swelling thicknesses were inhibited by 0.5 Gy irradiation. In addition, interstitial edema in those mice which received 0.5 Gy irradiation was less than that in the mice which underwent by sham irradiation. These findings suggest that the ischemia-reperfusion injury is inhibited by the enhancement of antioxidation function by 0.5 Gy irradiation

Evaluating the Toxicity of Silver Nanoparticles by Detecting Phosphorylation of Histone H3 in Combination with Flow Cytometry Side-Scattered Light

Post-translational modification of histones is linked to a variety of biological processes and disease states. This paper focuses on phosphorylation of histone H3 at serine 10 (p-H3S10), induced by silver nanoparticles (AgNPs) and discusses the usefulness of p-H3S10 as a marker to evaluate the toxicity of AgNPs. Cultured human cells showed remarkable p-H3S10 immediately after treatment with AgNPs but not with Ag microparticles. p-H3S10 lasts up to 24 h and strongly depends upon the cellular uptake of AgNPs. Removal of Ag ions suppressed p-H3S10, while adding an excess of Ag ions augmented p-H3S10. We expected that p-H3S10 requires two events: cellular uptake of AgNPs and continuous release of Ag ions from intracellular AgNPs. AgNPs enhanced the expression of the proto-oncogene <i>c-jun</i>, and p-H3S10 increased in the promoter sites of the gene, indicating that p-H3S10 might indicate a biological reaction related to carcinogenesis. We previously showed that side-scattered light from flow cytometry could be used to measure the uptake potential of nanoparticles [Suzuki, H.; Toyooka, T.; Ibuki, Y. Simple and easy method to evaluate uptake potential of nanoparticles in mammalian cells using a flow cytometric light scatter analysis. Environ. Sci. Technol. 2007, 41 (8), 3018−3024]. Our current findings suggest that p-H3S10 can be used to evaluate the toxicity of AgNPs and Ag ion release in combination with detection of side-scattered light from flow cytometry

Phosphatidylserine induces apoptosis in CHO cells without mitochondrial dysfunction in a manner dependent on caspases other than caspases-1, -3, -8 and -9.

Treatment of Chinese hamster ovary K1 cells with phosphatidylserine (PS) caused typical　apoptosis with distinct morphological and biochemical features in a dose- and time-dependent manner. However, unlike camptothecin-induced apoptosis, changes in mitochondrial transmembranepotential were not observed. In addition, cytochrome c release did not occur in PS-induced apoptosis. A pan caspase inhibitor, Z-VAD, significantly inhibited the apoptosis, but inhibitors of caspase-1, -3, -8 and -9 did not. Activities of caspase-1, -3, -8 and -9 were increased bytreatment of the cells with camptothecin, but not with PS. These results suggest that PS-induced apoptosis occurs without the collapse of mitochondrial transmembrane potential and without the release of cytochrome c, in a manner independent of caspase-1, -3, -8 and -9

Dynamics of Ku80 in Living Hamster Cells with DNA Double-Strand Breaks Induced by Chemotherapeutic Drugs.

A variety of chemotherapeutic drugs, e.g., etoposide and bleomycin, are widely used in clinical practice to treat many types of animal malignancies. In the clinical situation, cellular resistance to chemotherapy is a significant component of tumor treatment failure. A variety of DNA repair factors, e.g., Ku80, might be a key contributor to chemoresistance to anticancer agents. In both cancer and normal cells, Ku80 plays a key role as a sensor of DNA double-strand break (DSB) induced by treatment with some chemotherapeutic drugs. Although the localization and mobility of Ku80 play a key role in regulating the physiological function of Ku80, it is not clear whether those of Ku80 are affected after treatment with chemotherapeutic drugs. We examined the localization and mobility of Ku80 in living hamster cells with or without DSBs, which were induced by treatment with chemotherapeutic drugs. Our data showed that Ku80, in contrast to H2AX, is highly mobile in the nuclei. We found that before and after the induction of DNA damage by treatment with etoposide or bleomycin, a major portion of Ku80 is exchanged by the same kinetics in the nuclei of interphase cells. These results suggest that the mobility of a major portion of Ku80 is not affected by DNA DSBs in order to find other DSBs. In addition, the information would be worthy to develop some new chemotherapeutic drugs to treat many types of animal malignancie

Flow Cytometric Evaluation of Nanoparticles Using Side-Scattered Light and Reactive Oxygen Species-Mediated Fluorescence–Correlation with Genotoxicity

We recently clarified that the side-scatter­(ed) light (SSC) of flow cytometry (FCM) could be used as a guide to measure the uptake potential of nanoparticles [Suzuki et al. Environ. Sci. Technol. 2007, 41, 3018−3024]. In this paper, the method was improved so as to be able to determine simultaneously the uptake potential of nanoparticles and the production of reactive oxygen species (ROS), and correlations with genotoxicity were evaluated. In the FCM analysis, SSC and fluorescence of 6-carboxy-2,7′-diclorodihydrofluorescein diacetate, di­(acetoxy ester) based on ROS production were concurrently detected after treatments with ZnO, CuO, Fe₃O₄, TiO₂, and Ag nanoparticles. The ZnO and CuO nanoparticles caused high ROS production, which was more significant in the cells with higher SSC intensity. The increase of SSC intensity was more significant for TiO₂ than ZnO and CuO, whereas ROS production was higher for ZnO and CuO than TiO₂, suggesting that the extent of ROS production based on the uptake of nanoparticles differed with each kind of nanoparticle. ROS production was correlated with generation of the phosphorylated histone H2AX (γ-H2AX), a marker of DNA damage, and an antioxidant, <i>n</i>-acetylcysteine, could partially suppress the γ-H2AX. This method makes it possible to predict not only uptake potential but also genotoxicity