The development of newborn C 57 BL/KsJ-dbm mice produced from eggs fertilized in vitro

The purpose of this study was to examine the development of newly born C57BL/KsJ-dbm mice produced from eggs fertilized in vitro. The embryos derived from fertilization in vitro (which was performed by using db/db eggs and adrenalectomized db/db (Adrex) spermatozoa,) were transferred to the oviduct of MRL/MpJ pseudopregnant recipients 30 hr after insemination. 376 of these embryos yielded 65 young. Weight gain and urine glucose, plasma glucose and insulin levels were measured in these young as well as in Adrex males. The young produced by fertilization in vitro showed hyperglycemia, hyperinsulinemia and obesity. The physiological abnormalities in these young were similar to those in db/db young produced by natural mating between heterozygote (db/+) males and females. Adrex males did not show hyperglycemia but did show hyperinsulinemia. These results indicate that in vitro fertilization and embryo transfer is an effective means of producing fetuses or newborns with an overt genotype in genetically diabetic obese (db) mice.     

Emergence of gp120 V3 Variants Confers Neutralization Resistance in an R5 Simian-Human Immunodeficiency Virus-Infected Macaque Elite Neutralizer That Targets the N332 Glycan of the Human Immunodeficiency Virus Type 1 Envelope Glycoprotein

Neutralization-resistant simian-human immunodeficiency virus AD8 (SHIV_AD8) variants that emerged in an infected macaque elite neutralizer targeting the human immunodeficiency virus type 1 (HIV-1) gp120 N332 glycan acquired substitutions of critical amino acids in the V3 region rather than losing the N332 glycosylation site. One of these resistant variants, carrying the full complement of gp120 V3 changes, was also resistant to the potent anti-HIV-1 monoclonal neutralizing antibodies PGT121 and 10-1074, both of which are also dependent on the presence of the gp120 N332 glycan.     

Differential type I IFN-inducing abilities of wild-type versus vaccine strains of measles virus

Laboratory adapted and vaccine strains of measles virus (MV) induced type I IFN in infected cells. The wild-type strains in contrast induced it to a far lesser extent. We have investigated the mechanism for this differential type I IFN induction in monocyte-derived dendritic cells infected with representative MV strains. Laboratory adapted strains Nagahata and Edmonston infected monocyte-derived dendritic cells and activated IRF-3 followed by IFN-beta production, while wild-type MS failed to activate IRF-3. The viral IRF-3 activation is induced within 2 h, an early response occurring before protein synthesis. Receptor usage of CD46 or CD150 and nucleocapsid (N) protein variations barely affected the strain-to-strain difference in IFN-inducing abilities. Strikingly, most of the IFN-inducing strains possessed defective interference (DI) RNAs of varying sizes. In addition, an artificially produced DI RNA consisting of stem (the leader and trailer of MV) and loop (the GFP sequence) exhibited potential IFN-inducing ability. In this case, however, cytoplasmic introduction was needed for DI RNA to induce type I IFN in target cells. By gene-silencing analysis, DI RNA activated the RIG-I/MDA5-mitochondria antiviral signaling pathway, but not the TLR3-TICAM-1 pathway. DI RNA-containing strains induced IFN-beta mRNA within 2 h while the same recombinant strains with no DI RNA required >12 h postinfection to attain similar levels of IFN-beta mRNA. Thus, the stem-loop structure, rather than full genome replication or specific internal sequences of the MV genome, is required for an early phase of type I IFN induction by MV in host cells.     

Radical scavenging activity of spring mountain herbs in the Shikoku mountain area and identification of antiradical constituents by simple HPLC detection and LC-MS methods

The functionality of spring mountain herbs, which were collected in the Kajigamori mountain area of Shikoku area in Japan, was investigated in the course of our studies for utilizing local plant resources. The radical scavenging activity of the extracts from seventeen herbs was measured. Among these herbs, two extracts from Polystichym ovato-paleaceum (Japanese name: Tsuyanashiinode) and Sambucus racemosa subsp. sieboldiana (Japanese name: Niwatoko) showed potent DPPH radical scavenging activity. The material evidence for the potent activity of the extracts was studied by a combination of our developed method for detecting antiradical compounds, LC-MS/MS, and enzymatic hydrolysis.     

Development of mouse models for analysis of human virus infections

Viruses usually exhibit strict species‐specificity as a result of co‐evolution with the host. Thus, in mouse models, a great barrier exists for analysis of infections with human‐tropic viruses. Mouse models are unlikely to faithfully reproduce the human immune response to viruses or viral compounds and it is difficult to evaluate human therapeutic efficacy with antiviral reagents in mouse models. Humans and mice essentially have different immune systems, which makes it difficult to extrapolate mouse results to humans. In addition, apart from immunological reasons, viruses causing human diseases do not always infect mice because of species tropism. One way to determine tropism would be a virus receptor that is expressed on affected cells. The development of gene‐disrupted mice and Tg mice, which express human receptor genes, enables us to analyze several viral infections in mice. Mice are, indeed, susceptible to human viruses when artificially infected in receptor‐supplemented mice. Although the mouse cells less efficiently permit viral replication than do human cells, the models for analysis of human viruses have been established in vivo as well as in vitro, and explain viral pathogenesis in the mouse systems. In most systems, however, nucleic acid sensors and type I interferon suppress viral propagation to block the appearance of infectious manifestation. We herein review recent insight into in vivo antiviral responses induced in mouse infection models for typical human viruses.     

Adaptive evolution of the uncoupling protein 1 gene contributed to the acquisition of novel nonshivering thermogenesis in ancestral eutherian mammals

Homeotherms possess various physiological mechanisms to maintain their body temperature, thus allowing them to adapt to various environments. Under cold conditions, most eutherian mammals upregulate heat production in brown adipose tissue (BAT), and uncoupling protein (UCP) 1 is an essential factor in BAT thermogenesis. The evolutionary origin of UCP1 was believed to have been a specific event occurring in eutherian lineages. Recently, however, the UCP1 ortholog was found in fishes, which uncovers a more ancient origin of this gene than previously believed. Here we investigate the evolutionary process of UCP1 by comparative genomic approach. We found that UCP1 evolved rapidly by positive Darwinian selection in the common ancestor of eutherians, although this gene arose in the ancestral vertebrate, since the orthologous genes were shared among most of the vertebrate species. Adaptive evolution occurred after the divergence between eutherians and marsupials, which is consistent with the fact that BAT has been found only in eutherians. Our findings indicate that positive Darwinian selection acted on UCP1 contributed to the acquisition of an efficient mechanism for body temperature regulation in primitive eutherians. Phylogenetic reconstruction of UCP1 with two paralogs (UCP2 and UCP3) among vertebrate species revealed that the gene duplication events which produced these three genes occurred in the common ancestor of vertebrates much earlier than the emergence of eutherians. Thus, our data demonstrate that novel gene function can evolve without de novo gene duplication event.