Interleukin-18 inhibits hepatitis B virus replication in the livers of transgenic mice

Interleukin-18 (IL-18) produced by activated antigen-presenting cells stimulates natural killer (NK) cells, natural killer T (NKT) cells, and T cells to secrete gamma interferon (IFN-gamma). In this study, injection of a single 10- micro g dose of recombinant murine IL-18 rapidly, reversibly, and noncytopathically inhibited hepatitis B virus (HBV) replication in the livers of HBV transgenic mice. Furthermore, HBV replication was inhibited by as little as 1 micro g of IL-18 injected repetitively, and also by a single 0.1- micro g dose of IL-18 injected together with 1 ng of IL-12, neither of which inhibited HBV replication individually, demonstrating synergy between these cytokines in this system. The antiviral effect of IL-18 was mediated by its ability to activate resident intrahepatic NK cells and NKT cells to produce IFN-gamma and by its ability to induce IFN-alpha/beta production in the liver. These results suggest that IL-18 has the potential to contribute to the control of HBV replication during self-limited infection and that it may have therapeutic value for the treatment of patients with chronic hepatitis.     

GPR40 gene expression in human pancreas and insulinoma

To assess gene expression of a membrane-bound G-protein-coupled fatty acid receptor, GPR40, in the human pancreas and islet cell tumors obtained at surgery were analyzed. The mRNA level of the GPR40 gene in isolated pancreatic islets was approximately 20-fold higher than that in the pancreas, and the level was comparable to or rather higher than that of the sulfonylurea receptor 1 gene, which is known to be expressed abundantly in human pancreatic beta cells. A large amount of GPR40 mRNA was detected in tissue extracts from two cases of insulinoma, whereas the expression was undetectable in glucagonoma or gastrinoma. The present study demonstrates that GPR40 mRNA is expressed predominantly in pancreatic islets in humans and that GPR40 mRNA is expressed solely in human insulinoma among islet cell tumors. These results indicate that GPR40 is probably expressed in pancreatic beta cells in the human pancreas.     

Antiangiogenic activity of 3,4-seco-cycloartane triterpenes from Thai Gardenia spp. and their semi-synthetic analogs

Twelve naturally occurring 3,4-seco-cycloartane triterpenes (1-12) isolated from Gardenia sootepensis and Gardenia obtusifolia, and eight semi-synthetic derivatives (13-20) were evaluated for their antiangiogenic activity on a rat aortic sprouting assay, an ex vivo model of angiogenesis. Among these compounds, sootepin B (1) displayed the most potent activity in terms of the inhibition of microvessel sprouting from rat aortic rings in a dose-dependent manner with IC(50) value of 4.46 μM. Its angiogenic effect was found to occur via suppression of endothelial cell proliferation and tubular formation, and was likely mediated by regulation (inhibition) of the Erk1/2 signaling pathway.     

Immunization with a Recombinant Vaccinia Virus That Encodes Nonstructural Proteins of the Hepatitis C Virus Suppresses Viral Protein Levels in Mouse Liver

Chronic hepatitis C, which is caused by infection with the hepatitis C virus (HCV), is a global health problem. Using a mouse model of hepatitis C, we examined the therapeutic effects of a recombinant vaccinia virus (rVV) that encodes an HCV protein. We generated immunocompetent mice that each expressed multiple HCV proteins via a Cre/loxP switching system and established several distinct attenuated rVV strains. The HCV core protein was expressed consistently in the liver after polyinosinic acid–polycytidylic acid injection, and these mice showed chronic hepatitis C-related pathological findings (hepatocyte abnormalities, accumulation of glycogen, steatosis), liver fibrosis, and hepatocellular carcinoma. Immunization with one rVV strain (rVV-N25), which encoded nonstructural HCV proteins, suppressed serum inflammatory cytokine levels and alleviated the symptoms of pathological chronic hepatitis C within 7 days after injection. Furthermore, HCV protein levels in liver tissue also decreased in a CD4 and CD8 T-cell-dependent manner. Consistent with these results, we showed that rVV-N25 immunization induced a robust CD8 T-cell immune response that was specific to the HCV nonstructural protein 2. We also demonstrated that the onset of chronic hepatitis in CN2-29^(+/−)/MxCre^(+/−) mice was mainly attributable to inflammatory cytokines, (tumor necrosis factor) TNF-α and (interleukin) IL-6. Thus, our generated mice model should be useful for further investigation of the immunological processes associated with persistent expression of HCV proteins because these mice had not developed immune tolerance to the HCV antigen. In addition, we propose that rVV-N25 could be developed as an effective therapeutic vaccine.     

Isolation of basidiomycetous anamorphic yeast-like fungus <Emphasis Type="Italic">Meira argovae</Emphasis> found on Japanese bamboo

A basidiomycetous anamorphic yeast-like fungus, isolated from new bamboo shoots collected in Japan, was assigned to Meira argovae by comparison of conidial morphology, physiological characteristics, rDNA sequences, and DNA-DNA relatedness with the ex-type strains of Meira species. This is the first record of the finding of M. argovae from other than mite cadavers and in regions other than Israel. Phylogenetic analysis based on the D1-D2 domain demonstrated that Meira species and teleomorphic Dicellomyces species, which include a bamboo leaf parasite, D. gloeosporus, formed sister clades.     

Solar-simulated light-exposed benzo[a]pyrene induces phosphorylation of histone H2AX

Polycyclic aromatic hydrocarbons (PAHs), wide-spread mutagenic and carcinogenic environmental pollutants, are consistently exposed to sunlight in the environment. The exposure causes structural change, resulting in the generation of a variety of photomodified products having different bioactivities compared with the parent compounds. In this study, we found that benzo[a]pyrene (BaP) exposed to solar-simulated light (SSL)-induced phosphorylation of histone H2AX (gamma-H2AX), which was recently identified as an early event after the induction of DNA double strand breaks (DSBs). Although BaP itself did not produce gamma-H2AX, SSL-exposed BaP significantly generated gamma-H2AX depending on the period of exposure. Furthermore, we revealed that reactive oxygen species produced by the SSL-exposed BaP mainly contributed to the generation of gamma-H2AX. The appearance of gamma-H2AX means the induction of the most serious form of DNA damage, DSBs, suggesting the potential risk of carcinogenesis.     

Species separation in <Emphasis Type="Italic">Curvularia</Emphasis> “geniculata” group inferred from <Emphasis Type="Italic">Brn1</Emphasis> gene sequences

 Brn1, a reductase gene involved in the melanin biosynthetic pathway, was adopted for species delimitation among members in the “geniculata” group of Curvularia species and proved to be useful for this purpose. Phylogenetic trees of these fungal members were constructed from nucleotide sequences of this region. The so-called geniculata group of Curvularia was separated into several clusters. The conidial morphology of the members in each cluster is closely similar but clearly different among discrete clusters. The phylogenetic groups almost concurred with the morphological grouping. Thus, the synonymous treatment of Curvularia affinis, C. fallax, and C. senegalensis to C. geniculata in a previous study was supported. The isolates with warping hilum conidia were clearly different from C. geniculata and separated into two clusters. C. geniculata ATCC 6671 made an independent cluster situated near these clusters. The protuberant hilum species were located separately in the phylogenetic trees. For sound taxonomic treatment of these isolates, we should accumulate more information and retain our species determination for them. Received: September 26, 2002 / Accepted: March 12, 2003     

An efficient system to establish multiple embryonic stem cell lines carrying an inducible expression unit

The growing use of mouse embryonic stem (ES) cells in research emphasizes their importance in studies of molecular mechanisms that maintain pluripotency and direct cellular differentiation. Although systems for regulatable transgene expression are essential for fine analysis of cellular processes at the molecular level, a strategy for the establishment of multiple ES cell lines carrying any of these systems has not yet been described. Here, we report our development of the ROSA-TET system, an effective system for the establishment of multiple ES cell lines carrying a tetracycline (Tc)-regulatable transgene at the Gt (ROSA)26asSor (ROSA26) locus. This system contains a knock-in step of a construct carrying both loxP and its mutant sequences into the ROSA26 locus, followed by a subsequent exchange step that introduces a cDNA to be Tc-regulated to the locus using the recombinase-mediated cassette exchange reaction. Both steps are demonstrated to give desired clones with high efficiency, suggesting that this system can be introduced readily into any ES cell lines, leading to the simultaneous establishment of multiple cell lines carrying different Tc-regulated cDNAs. We believe that use of this system will strongly accelerate molecular biological research using ES cells.     

Noise-triggered regime shifts in a simple aquatic model

We examined the effects of random fluctuations and diffusion on regime shifts in bistable aquatic ecosystems, using a simple two-component reaction–diffusion model with nutrients and phytoplankton. The corresponding mean-field model generates two stable attractors: one is a nutrient-dominated (ND) state or a clear-water state, and the other is a phytoplankton-dominated (PD) state or a turbid-water state. According to our simulations of the reaction–diffusion model without fluctuations, either of two stable states can be dominant corresponding to the given parameters and the configuration of two states, meaning that the long-term coexistence of these two states is difficult. The dominant state expands gradually eliminating the other non-dominant one, which results in the homogeneous distribution of the dominant state in the whole lake. Further simulations including the effects of temporally changing fluctuations show that the regime shifts driven by external noise proceed even within a bistable region, which complicates the detection of hysteresis in real aquatic ecosystems.