Rheb (Ras Homologue Enriched in Brain)-dependent Mammalian Target of Rapamycin Complex 1 (mTORC1) Activation Becomes Indispensable for Cardiac Hypertrophic Growth after Early Postnatal Period

Cardiomyocytes proliferate during fetal life but lose their ability to proliferate soon after birth and further increases in cardiac mass are achieved through an increase in cell size or hypertrophy. Mammalian target of rapamycin complex 1 (mTORC1) is critical for cell growth and proliferation. Rheb (Ras homologue enriched in brain) is one of the most important upstream regulators of mTORC1. Here, we attempted to clarify the role of Rheb in the heart using cardiac-specific Rheb-deficient mice (Rheb^−/−). Rheb^−/− mice died from postnatal day 8 to 10. The heart-to-body weight ratio, an index of cardiomyocyte hypertrophy, in Rheb^−/− was lower than that in the control (Rheb^+/+) at postnatal day 8. The cell surface area of cardiomyocytes isolated from the mouse hearts increased from postnatal days 5 to 8 in Rheb^+/+ mice but not in Rheb^−/− mice. Ultrastructural analysis indicated that sarcomere maturation was impaired in Rheb^−/− hearts during the neonatal period. Rheb^−/− hearts exhibited no difference in the phosphorylation level of S6 or 4E-BP1, downstream of mTORC1 at postnatal day 3 but showed attenuation at postnatal day 5 or 8 compared with the control. Polysome analysis revealed that the mRNA translation activity decreased in Rheb^−/− hearts at postnatal day 8. Furthermore, ablation of eukaryotic initiation factor 4E-binding protein 1 in Rheb^−/− mice improved mRNA translation, cardiac hypertrophic growth, sarcomere maturation, and survival. Thus, Rheb-dependent mTORC1 activation becomes essential for cardiomyocyte hypertrophic growth after early postnatal period.     

A Novel Bivalent Vaccine Based on a PB2-Knockout Influenza Virus Protects Mice from Pandemic H1N1 and Highly Pathogenic H5N1 Virus Challenges

Vaccination is an effective means to protect against influenza virus. Although inactivated and live-attenuated vaccines are currently available, each vaccine has disadvantages (e.g., immunogenicity and safety issues). To overcome these problems, we previously developed a replication-incompetent PB2-knockout (PB2-KO) influenza virus that replicates only in PB2 protein-expressing cells. Here, we generated two PB2-KO viruses whose PB2-coding regions were replaced with the HA genes of either A/California/04/2009 (H1N1pdm09) or A/Vietnam/1203/2004 (H5N1). The resultant viruses comparably, or in some cases more efficiently, induced virus-specific antibodies in the serum, nasal wash, and bronchoalveolar lavage fluid of mice relative to a conventional formalin-inactivated vaccine. Furthermore, mice immunized with these PB2-KO viruses were protected from lethal challenges with not only the backbone virus strain but also strains from which their foreign HAs originated, indicating that PB2-KO viruses with antigenically different HAs could serve as bivalent influenza vaccines.     

Adipose‐derived mesenchymal stem cells and regenerative medicine

Adipose tissue‐derived mesenchymal stem cells (ADSCs) are multipotent and can differentiate into various cell types, including osteocytes, adipocytes, neural cells, vascular endothelial cells, cardiomyocytes, pancreatic β‐cells, and hepatocytes. Compared with the extraction of other stem cells such as bone marrow‐derived mesenchymal stem cells (BMSCs), that of ADSCs requires minimally invasive techniques. In the field of regenerative medicine, the use of autologous cells is preferable to embryonic stem cells or induced pluripotent stem cells. Therefore, ADSCs are a useful resource for drug screening and regenerative medicine. Here we present the methods and mechanisms underlying the induction of multilineage cells from ADSCs.     

Impacts of invasive crayfish (Pacifastacus leniusculus) on endangered freshwater pearl mussels (Margaritifera laevis and M. togakushiensis) in Japan

The invasive alien crayfish Pacifastacus leniusculus is considered harmful to freshwater pearl mussels Margaritifera laevis and M. togakushiensis. It also often colonises mussel habitats in Japan. In order to test the negative effects of alien crayfish on mussels, we evaluated the predation impact of signal crayfish on freshwater pearl mussels in vitro. We tested the relationship between the survival/injury rates of mussels and crayfish predation with respect to different sizes of mussels (four classes based on shell length: 10, 30, 50 and 70 mm). Crayfish selectively fed on the flesh of the 10-mm size class mussels after breaking their shells. The shell margins of mussels in all size classes were injured by crayfish. Results also showed that crayfish particularly injured the 50-mm size class of mussels. This observation could be attributed to this mussel size being the most suitable shell size (29.56–37.73 mm in carapace length) that the crayfish can effectively hold. This study shows that the presence of invasive crayfish reduces freshwater pearl mussel populations by damaging the shell margins and/or killing the mussels. This negative impact of invasive crayfish not only decreases the mussel population but could also limit mussel recruitment, growth and reproduction.     

Identification of 2,3-disubstituted pyridines as potent,orally active PDE4 inhibitors

A series of 2,3-disubstituted pyridines were synthesized and evaluated for their PDE4 inhibitory activity. We successfully modified undesirable cyano group of initial lead compound 2 to 4-pyridyl group with improvement of in vitro efficacy and optimized the position of nitrogen atoms in pyridine moiety and alkylene linker. The most potent compound showed significant efficacy in animal models of asthma and inflammation.     

Excitotoxicity-induced immediate surge in hippocampal prostanoid production has latent effects that promote chronic progressive neuronal death

Excitotoxicity is involved in neurodegenerative conditions. We investigated the pathological significance of a surge in prostaglandin production immediately after kainic acid (KA) administration [initial phase], followed by a sustained moderate elevation in prostaglandin level [late phase] in the hippocampus of juvenile rats. Numerous pyknotic hippocampal neurons were observed 72 h after KA treatment; this number remained elevated on days 10 and 30. Gross hippocampal atrophy was observed on days 10 and 30. Pre-treatment with indomethacin ameliorated neuronal death on days 10 and 30, and prevented hippocampal atrophy on day 30. Microglial response was moderated by the indomethacin pre-treatment. Blockade of only late-phase prostaglandin production by post-treatment with indomethacin ameliorated neuronal death on day 30. These findings suggest a role for initial-phase prostaglandin production in chronic progressive neuronal death, which is exacerbated by late-phase prostaglandin production. Blockade of prostaglandin production has therapeutic implications in preventing long-term neurological sequelae following excitotoxic brain damage.     

Phytuberol,from Japanese Domestic Tobacco,Nicotiana tabacum cv. Suifu

Carcinogenesis is believed to be induced through the oxidative damage of DNA, and antioxidants are expected to suppress it. So, the polyphenolic antioxidants in daily foods were investigated to see whether they protect against genetic damage by active oxygen. In the evaluation, we used a bioassay and a chemical determination, a Salmonella mutagenicity test for mutation by a N-hydroxyl radical from one of the dietary carcinogens 3-amino-1-methyl-5H-pyrido[4,3-b]indole and the formation of 8-hydroxyl (8-OHdG) from 2′-deoxyguanosine (2′-dG) in a Fenton OH-radical generating system. Thirty-one antioxidants including flavonoids were compared in terms of radical-trapping activity with bacterial DNA and 2′-dG. Antioxidants inhibited the mutation but the IC₅₀ values were in the mM order. Against 8-OHdG formation, only α-tocopherol had a suppressive effect with an IC₅₀ of 1.5 μM. Thus, except α-tocopherol, the dietary antioxidants did not scavenge the biological radicals faster than bacterial DNA and intact 2′-dG, indicating that they failed to prevent oxidative gene damage and probably carcinogenesis.     

Isolation of New Tobacco Constituents, 8,9-Dihydro-8,9-dihydroxymegastigmatrienone,from Japanese Domestic Suifu Tobacco

The tetradecapeptide of a renin substrate, DRVYIHPFHLLVYS, was used as a substrate for assaying several fungal aspartic and acidic proteinases in the acidic pH range. Aspartic and acidic proteinases froll) Phycomycetes, Mucor and Rhizopus, and Deuteromycotina, Aspergillus and Penicillium, cleaved the tetradecapeptide at its tyrosyl⁴-isoleucyl⁵ (Y⁴-I⁵),histidyI⁶-proly⁷ (H⁶_P⁷) and leucyl¹¹-valyl¹² (L¹¹-V¹²) bonds in the acidic pH range, while acidic proteinases type B and type A-I from Scytalidium lignicolumn, and those from Cladosporium and Basidiomycetes, Pycnoporus sanguineus, and the yeast, Rhodotorula glutinis; showed slightly different specificities towards the tetradecapeptide. Pepsin primarily cleaved the valy³-tyrosyl⁴ (V³-Y⁴) and leucyl¹⁰-leucyl¹¹ (L¹⁰-L¹¹) bonds. All of the aspartic and acidic proteinases of fungal origin tested in the present study have different specificities from that of pepsin.