Is the Comet Assay a Sensitive Procedure for Detecting Genotoxicity?

Although the Comet assay, a procedure for quantitating DNA damage in mammalian cells, is considered sensitive, it has never been ascertained that its sensitivity is higher than the sensitivity of other genotoxicity assays in mammalian cells. To determine whether the power of the Comet assay to detect a low level of genotoxic potential is superior to those of other genotoxicity assays in mammalian cells, we compared the results of Comet assay with those of micronucleus test (MN test). WTK1 human lymphoblastoid cells were exposed to methyl nitrosourea (MNU), ethyl nitrosourea (ENU), methyl methanesulfonate (MMS), ethyl methanesulfonate (EMS), bleomycin (BLM), or UVC. In Comet assay, cells were exposed to each mutagen with (Comet assay/araC) and without (Comet assay) DNA repair inhibitors (araC and hydroxyurea). Furthermore, acellular Comet assay (acellular assay) was performed to determine how single-strand breaks (SSBs) as the initial damage contributes to DNA migration and/or to micronucleus formation. The lowest genotoxic dose (LGD), which is defined as the lowest dose at which each mutagen causes a positive response on each genotoxicity assay, was used to compare the power of the Comet assay to detect a low level of genotoxic potential and that of MN test; that is, a low LGD indicates a high power. Results are summarized as follows: (1) for all mutagens studied, LGDs were MN test ≦ Comet assay; (2) except for BLM, LGDs were Comet assay/araC ≦ MN test; (3) except for UVC and MNU, LGDs were acellular assay ≦ Comet assay/araC ≦ MN test ≦ Comet assay. The following is suggested by the present findings: (1) LGD in the Comet assay is higher than that in MN test, which suggests that the power of the MN test to detect a low level of genotoxic potential is superior to that of the Comet assay; (2) for the studied mutagens, all assays were able to detect all mutagens correctly, which suggests that the sensitivity of the Comet assay and that of the MN test were exactly identical; (3) the power of the Comet assay to detect a low level of genotoxic potential can be elevated to a level higher than that of MN test by using DNA resynthesis inhibitors, such as araC and HU

Epidemiological studies on stomach cancer mortality and its correlation with dietary factor in Japan

The aim of this study is to assess the effects of dietary factors on the death from stomach cancer in Japan. We analyzed the correlation between socioeconomic factors and age-adjusted death rate (ADR) from stomach cancer for every five year and that between mean intake of food or nutrient during two periods (former period : 1966-1970, latter period : 1976-1980) and the reduction rate in ADR from stomach cancer for two periods (former period : 1966-1980, latter period : 1976-1990). Moreover, principal component analyses of food intake for two periods were performed to examine the changes in the structure of food intake and its correlation with the reduction rate in ADR from stomach cancer. There were significant positive correlations between the reduction rate in ADR from stomach cancer and some socioeconomic factors (total expense for food, consumpion of alcoholic beverages and consumption of salted and dried fish). There were significant negative correlations between ADR from stomach cancer and several food or nutrient intakes (wheat and egg intake for former period, milk and animal fat intake for latter period) (p<0.01). The reduction rate in ADR from stomach cancer were greater in districts where these food or nutrient intakes were larger. On the other hand, there were positive correlations between the reduction rate in ADR from stomach cancer and some food or nutrient intakes (rice intake for former period, beans intake and the P/S ratio for latter period) (p<0.05). The reduction rate in ADR from stomach cancer were smaller in districts where these food or nutrient intakes were larger. Principal component analyses revealed that the intakes of foods used in traditional Japanese diet were identified as first principal component for both former and latter periods. As second principal component, food intakes related to the Western diet were identified. During the former period, the reduction rate in ADR from stomach cancer had a significant positive correlation with the fist principal component (p<0.05). On the other hand, during the latter period, a significant negative correlation was obtained between the reduction rate in ADR from stomach cancer and the second principal component. These results indicate that westernization of the Japanese style diet, while preserving its major traditional features, is related to both the reduction and regional differences in deaths rate from stomach cancer in Japan

Quantitative activation-induced manganese-enhanced MRI reveals severity of Parkinson’s disease in mice

We demonstrate that activation-induced manganese-enhanced magnetic resonance imaging with quantitative determination of the longitudinal relaxation time (qAIM-MRI) reveals the severity of Parkinson’s disease (PD) in mice. We first show that manganese ion-accumulation depends on neuronal activity. A highly active region was then observed by qAIM-MRI in the caudate-putamen in PD-model mice that was significantly correlated to the severity of PD, suggesting its involvement in the expression of PD symptoms

Branched-chain amino acids-induced cardiac protection against ischemia/reperfusion injury

Aims: Amino acids, especially branched chain amino acids (BCAAs), have important regulatory roles in protein synthesis. Recently studies revealed that BCAAs protect against ischemia/reperfusion (I/R) injury. We studied the signaling pathway and mitochondrial function affecting a cardiac preconditioning of BCAAs. Main methods: An in vivo model of I/R injury was tested in control, mTOR+/+, and mTOR+/−. Mice were randomly assigned to receive BCAAs, rapamycin, or BCAAs + rapamycin. Furthermore, isolated cardiomyocytes were subjected to simulated ischemia and cell death was quantified. Biochemical and mitochondrial swelling assays were also performed. Key findings: Mice treated with BCAAs had a significant reduction in infarct size as a percentage of the area at risk compared to controls (34.1 ± 3.9% vs. 44.7 ± 2.6%, P = 0.001), whereas mice treated with the mTOR inhibitor rapamycin were not protected by BCAA administration (42.2 ± 6.5%, vs. control, P = 0.015). This protection was not detected in our hetero knockout mice of mTOR. Western blot analysis revealed no change in AKT signaling whereas activation of mTOR was identified. Furthermore, BCAAs prevented swelling which was reversed by the addition of rapamycin. In myocytes undergoing simulated I/R, BCAA treatment significantly preserved cell viability (71.7 ± 2.7% vs. 34.5 ± 1.6%, respectively, p < 0.0001), whereas rapamycin prevented this BCAA-induced cardioprotective effect (43.5 ± 3.4% vs. BCAA, p < 0.0001). Significance: BCAA treatment exhibits a protective effect in myocardial I/R injury and that mTOR plays an important role in this preconditioning effect.This work was supported by JSPS KAKENHI, Japan [grant number 19K09353]

Magnetic Ordering in V-Layers of the Superconducting System of Sr2VFeAsO3

Results of transport, magnetic, thermal, and 75As-NMR measurements are presented for superconducting Sr2VFeAsO3 with an alternating stack of FeAs and perovskite-like block layers. Although apparent anomalies in magnetic and thermal properties have been observed at ~150 K, no anomaly in transport behaviors has been observed at around the same temperature. These results indicate that V ions in the Sr2VO3-block layers have localized magnetic moments and that V-electrons do not contribute to the Fermi surface. The electronic characteristics of Sr2VFeAsO3 are considered to be common to those of other superconducting systems with Fe-pnictogen layers.Comment: 4 pages, 4 figures, To appear in JPSJ 79 (2010) 12371

Suppression of HBV replication by the expression of nickase-and nuclease dead-Cas9

Kurihara, T., Fukuhara, T., Ono, C. et al. Suppression of HBV replication by the expression of nickase- and nuclease dead-Cas9. Sci Rep 7, 6122 (2017). https://doi.org/10.1038/s41598-017-05905-

Farnesylation of Retinal Transducin Underlies Its Translocation during Light Adaptation

SummaryG proteins are posttranslationally modified by isoprenylation: either farnesylation or geranylgeranylation. The γ subunit of retinal transducin (Tα/Tβγ) is selectively farnesylated, and the farnesylation is required for light signaling mediated by transducin in rod cells. However, whether and how this selective isoprenylation regulates cellular functions remain poorly understood. Here we report that knockin mice expressing geranylgeranylated Tγ showed normal rod responses to dim flashes under dark-adapted conditions but exhibited impaired properties in light adaptation. Of note, geranylgeranylation of Tγ suppressed light-induced transition of Tβγ from membrane to cytosol, and also attenuated its light-dependent translocation from the outer segment to the inner region, an event contributing to retinal light adaptation. These results indicate that, while the farnesylation of transducin is interchangeable with the geranylgeranylation in terms of the light signaling, the selective farnesylation is important for visual sensitivity regulation by providing sufficient but not excessive membrane anchoring of Tβγ

Starch biosynthesis in rice endosperm requires the presence of either starch synthase I or IIIa

Starch synthase (SS) I and IIIa are the first and second largest components of total soluble SS activity, respectively, in developing japonica rice (Oryza sativa L.) endosperm. To elucidate the distinct and overlapping functions of these enzymes, double mutants were created by crossing the ss1 null mutant with the ss3a null mutant. In the F2 generation, two opaque seed types were found to have either the ss1ss1/SS3ass3a or the SS1ss1/ss3ass3a genotype. Phenotypic analyses revealed lower SS activity in the endosperm of these lines than in those of the parent mutant lines since these seeds had different copies of SSI and SSIIIa genes in a heterozygous state. The endosperm of the two types of opaque seeds contained the unique starch with modified fine structure, round-shaped starch granules, high amylose content, and specific physicochemical properties. The seed weight was ∼90% of that of the wild type. The amount of granule-bound starch synthase I (GBSSI) and the activity of ADP-glucose pyrophosphorylase (AGPase) were higher than in the wild type and parent mutant lines. The double-recessive homozygous mutant prepared from both ss1 and ss3a null mutants was considered sterile, while the mutant produced by the leaky ss1 mutant×ss3a null mutant cross was fertile. This present study strongly suggests that at least SSI or SSIIIa is required for starch biosynthesis in rice endosperm

Epinephrine unmasks latent mutation carriers with LQT1 form of congenital long-QT syndrome

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