Focal adhesion kinase mediates endothelin-induced cyclin D3 expression in rat cultured astrocytes

Focal adhesion kinase (FAK), a non-receptor type tyrosine kinase, is involved in the G1/S phase cell cycle transition of astrocytes induced by endothelin-1 (ET-1). In this study, the roles of FAK in the expression of cyclin D1 or D3, which are pivotal in G1/S phase transition, were examined in cultured astrocytes. Accompanied with increases in bromodeoxyuridine (BrdU) incorporation, ET-1 (100 nm) increased the numbers of cyclin D1- and D3-positive astrocytes. PD98059 (a MEK inhibitor) and PP-2 (a Src kinase inhibitor) inhibited ET-induced cyclin D1 expression and BrdU incorporation without affecting cyclin D3 expression. In contrast, cytochalasin D, lovastatin (a 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor) and Y-27632 (a rho-kinase inhibitor) prevented both cyclin D3 expression and BrdU incorporation. FAK phosphorylation by ET-1 was inhibited by cytochalasin D, lovastatin and Y-27632, but not by PD98059 or PP-2. Transfection with wild-type FAK increased expression of cyclin D3 in astrocytes, while that of cyclin D1 was not affected. Dominant-negative FAK mutants prevented an ET-induced increase in cyclin D3 expression, but not D1. These results suggest that activation of FAK causes cyclin D3 expression in cultured astrocytes, which would underlie the FAK-mediated astrocytic G1/S phase transition by ET-1.     

Clostridium glycyrrhizinilyticum sp. nov., a glycyrrhizin-hydrolysing bacterium isolated from human faeces

Screening of faecal bacteria for glycyrrhetic acid (GA) production by hydrolysing of glycyrrhizin (GL) resulted in the isolation of two strains, designated ZM35T and ZM38. Strains ZM35T and ZM38 were Gram-positive, obligate anaerobic, non-spore-forming and rod-shaped bacteria. Analysis of the 16S rRNA gene sequences indicated that strains ZM35T and ZM38 belonged to cluster XIVa of the genus Clostridium. The 16S rRNA gene sequences of strains ZM35T and ZM38 were identical. Strain ZM35T exhibited approximately 94% to 95% identity with the validly described species, Clostridium oroticum(94.5%), Eubacterium contortum(93.8%), Ruminococcus gnavus(94.5%) and R. torques(95.1%). In an experiment of DNA-DNA hybridization, it was confirmed that strains ZM35T and ZM38 were the same species. The guanine-plus-cytosine (G+C) content of strain ZM35T is 45.7 mol%. Based on the phylogenetic and phenotypic findings, we propose that strains ZM35T and ZM38 be assigned to a novel species named Clostridium glycyrrhizinilyticum. The type strain is ZM35T (=JCM 13368T=DSM 17593T).     

A signal adaptor SLAM-associated protein regulates spontaneous autoimmunity and Fas-dependent lymphoproliferation in MRL-Faslpr lupus mice

Autoantibody production and lymphadenopathy are common features of systemic autoimmune disease. Targeted or spontaneous mutations in the mouse germline have generated many autoimmune models with these features. Importantly, the models have provided evidence for the gene function in prevention of autoimmunity, suggesting an indispensable role for the gene in normal immune response and homeostasis. We describe here pathological and genetic characterizations of a new mutant strain of mice, the mutation of which spontaneously occurred in the Fas-deficient strain, MRL/Mp.Faslpr (MRL/lpr). MRL/lpr is known to stably exhibit systemic lupus erythematosus-like diseases. However, the mutant mice barely displayed autoimmune phenotypes, though the original defect in Fas expression was unchanged. Linkage analysis using (mutant MRL/lpr x C3H/lpr)F2 mice demonstrated a nucleotide insertion that caused loss of expression of small adaptor protein, signaling lymphocyte activation molecule (SLAM)-associated protein (SAP). SAP is known to be a downstream molecule of SLAM family receptors and to mediate the activation signal for tyrosine kinase Fyn. Recent studies have shown pleiotropic roles of SAP in T, B, and NK cell activations and NKT cell development. The present study will provide evidence for an essential role for SAP in the development of autoimmune diseases, autoantibodies, and lymphadenopathy in MRL/lpr lupus mice.     

Prolactin-releasing peptide is essential to maintain the prolactin level and osmotic balance in freshwater teleost fish

We administered prolactin-releasing peptide (PrRP) or anti-PrRP antiserum to goldfish in fresh water and analyzed their effects on prolactin and osmoregulatory mechanisms. The pituitary mRNA level of prolactin increased by PrRP but decreased by anti-PrRP. The rate of water inflow in the gills decreased by PrRP and increased by anti-PrRP, showing that PrRP restricts branchial water permeability, as also restricted by prolactin. PrRP also expanded the mucous cell layers on the scales, which may restrict efficiently water inflow by the mucous system. Eventually, the plasma osmotic pressure decreased by anti-PrRP. We conclude that PrRP is essential to maintain prolactin levels and osmotic balance in fresh water.     

Phosphorylation and gene expression of p53 are not affected in human cells exposed to 2.1425 GHz band CW or W-CDMA modulated radiation allocated to mobile radio base stations

A large-scale in vitro study focusing on low-level radiofrequency (RF) fields from mobile radio base stations employing the International Mobile Telecommunication 2000 (IMT-2000) cellular system was conducted to test the hypothesis that modulated RF fields induce apoptosis or other cellular stress response that activate p53 or the p53-signaling pathway. First, we evaluated the response of human cells to microwave exposure at a specific absorption rate (SAR) of 80 mW/kg, which corresponds to the limit of the average whole-body SAR for general public exposure defined as a basic restriction by the International Commission on Non-Ionizing Radiation Protection (ICNIRP) guidelines. Second, we investigated whether continuous wave (CW) and wideband code division multiple access (W-CDMA) modulated signal RF fields at 2.1425 GHz induced apoptosis or any signs of stress. Human glioblastoma A172 cells were exposed to W-CDMA radiation at SARs of 80, 250, and 800 mW/kg, and CW radiation at 80 mW/kg for 24 or 48 h. Human IMR-90 fibroblasts from fetal lungs were exposed to both W-CDMA and CW radiation at a SAR of 80 mW/kg for 28 h. Under the RF field exposure conditions described above, no significant differences in the percentage of apoptotic cells were observed between the test groups exposed to RF signals and the sham-exposed negative controls, as evaluated by the Annexin V affinity assay. No significant differences in expression levels of phosphorylated p53 at serine 15 or total p53 were observed between the test groups and the negative controls by the bead-based multiplex assay. Moreover, microarray hybridization and real-time RT-PCR analysis showed no noticeable differences in gene expression of the subsequent downstream targets of p53 signaling involved in apoptosis between the test groups and the negative controls. Our results confirm that exposure to low-level RF signals up to 800 mW/kg does not induce p53-dependent apoptosis, DNA damage, or other stress response in human cells.     

Retroviral DNA integration: reaction pathway and critical intermediates

The key DNA cutting and joining steps of retroviral DNA integration are carried out by the viral integrase protein. Structures of the individual domains of integrase have been determined, but their organization in the active complex with viral DNA is unknown. We show that HIV-1 integrase forms stable synaptic complexes in which a tetramer of integrase is stably associated with a pair of viral DNA ends. The viral DNA is processed within these complexes, which go on to capture the target DNA and integrate the viral DNA ends. The joining of the two viral DNA ends to target DNA occurs sequentially, with a stable intermediate complex in which only one DNA end is joined. The integration product also remains stably associated with integrase and likely requires disassembly before completion of the integration process by cellular enzymes. The results define the series of stable nucleoprotein complexes that mediate retroviral DNA integration.     

Protein tyrosine phosphatase receptor type Z is inactivated by ligand-induced oligomerization

Receptor-type protein tyrosine phosphatases (RPTPs) are considered to transduce extracellular signals across the membrane through changes in their PTP activity, however, our understanding of the regulatory mechanism is still limited. Here, we show that pleiotrophin (PTN), a natural ligand for protein tyrosine phosphatase receptor type Z (Ptprz) (also called PTPzeta/RPTPbeta), inactivates Ptprz through oligomerization and increases the tyrosine phosphorylation of substrates for Ptprz, G protein-coupled receptor kinase-interactor 1 (Git1) and membrane associated guanylate kinase, WW and PDZ domain containing 1 (Magi1). Oligomerization of Ptprz by an artificial dimerizer or polyclonal antibodies against its extracellular region also leads to inactivation, indicating that Ptprz is active in the monomeric form and inactivated by ligand-induced oligomerization.     

An N-glycan structure correlates with pulmonary metastatic ability of cancer cells

N-Glycan structures on the surface of cancer cells have diverse structures and play significant roles in metastatic process. However, little is known about their roles in organ-selective metastasis. Our study revealed that an alpha1,6-fucosylated biantennary N-glycan structure designated A2G2F is characteristic of lungs, with far more abundant expression in normal human and murine lungs than in other organs. In this study, we further examined the role of A2G2F in pulmonary metastasis. We stained metastatic cancers by alpha1,6-fucose-specific Lens culinaris agglutinin lectin and revealed that pulmonary metastatic nodules more abundantly expressed alpha1,6-fucosylated N-glycans than hepatic metastatic nodules from common primary cancers. The most specific alpha1,6-fucosylated N-glycan structure in pulmonary metastatic cancer was identified to be A2G2F. Using a B16 melanoma cell metastasis model, we showed that A2G2F-rich B16 cells formed more pulmonary metastatic nodules than A2G2F-poor cells. Our results suggest that A2G2F plays a critical role in pulmonary metastasis.     

Evaluation of bactericidal effects of low-temperature nitrogen gas plasma towards application to short-time sterilization

To develop a novel low-temperature plasma sterilizer using pure N(2) gas as a plasma source, we evaluated bactericidal ability of a prototype apparatus provided by NGK Insulators. After determination of the sterilizing conditions without the cold spots, the D value of the BI of Geobacillus stearothermophilus endospores on the filter paper was determined as 1.9 min. However, the inactivation efficiency of BI carrying the same endospores on SUS varied to some extent, suggesting that the bactericidal effect might vary by materials of sterilized instruments. Staphylococcus aureus and Escherichia coli were also exposed to the N(2) gas plasma and confirmed to be inactivated within 30 min. Through the evaluation of bactericidal efficiency in a sterilization bag, we concluded that the UV photons in the plasma and the high-voltage pulse to generate the gas plasma were not concerned with the bactericidal effect of the N(2) gas plasma. Bactericidal effect might be exhibited by activated nitrogen atoms or molecular radicals.     

Radiocesium distribution in the tissues of Japanese black beef heifers fed fallout-contaminated roughage due to the fukushima daiichi nuclear power station accident

This study examined the accumulation and tissue distribution of radioactive cesium nuclides in Japanese Black beef heifers raised on roughage contaminated with radioactive fallout due to the accident at the Fukushima Daiichi Nuclear Power Station on March 2011. Radiocesium feeding increased both (134)Cs and (137)Cs levels in all tissues tested. The kidney had the highest level and subcutaneous adipose had the lowest of radioactive cesium in the tissues. Different radioactive cesium levels were not found among parts of the muscles. These results indicate that radiocesium accumulated highly in the kidney and homogenously in the skeletal muscles in the heifers.