Interstitial-tissue localization of high-molecular-weight kininogen in guinea-pig skin

A rabbit antibody against the light-chain of guinea-pig high-molecular-weight (HMW) kininogen, which was specific to HWM kininogen and did not recognize low-molecular-weight kininogen, was prepared. This antibody demonstrated the presence of HMW kininogen antigen at the interstitial-tissue space in the guinea-pig skin by means of immunohistochemistry. The interstitial-tissue HMW kininogen antigen was extracted from the skin. This antigen molecule in the skin extract behaved identically as HWM kininogen of plasma in slab-polyacrylamide gel electrophoresis under the presence of sodium dodecyl sulfate followed by immunoblotting. Therefore, it was concluded that HMW kininogen was present in the interstitial-tissue fluid in the skin. The amount of HMW kininogen in the skin extract was quantified by a sandwich enzyme-linked immunosorbent assay with the anti-light-chain antibody and a goat anti-guinea-pig HMW kininogen antibody. On the assumption that the interstitial-tissue volume is 50 ml/100 g wet skin tissue, the average concentration of HMW kininogen in the interstitial-tissue fluid of the skin was calculated to be 23% of the plasma concentration. On the other hand, the proportion of intravascular HMW kininogen (derived from blood remaining in the vessels of the harvested skin) in relation to the total HMW kininogen in the skin extract was quantified by measuring the radio-labelled HMW kininogen which had been injected intravenously as a tracer of the intravascular HMW kininogen. About 5% of the total HMW kininogen in the skin extract was calculated to be derived from the intravascular blood volume of the skin, indicating that the majority of the HMW kininogen in the skin extract was derived from the extravascular-tissue space.     

Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice

Tissue resident mononuclear phagocytes (Mophs), comprising monocytes, macrophages, and dendritic cells (DCs), play important roles under physiological and pathological conditions. The presence of these cells in the kidney has been known for decades, and studies of renal Mophs (rMophs) are currently underway. Since no unified procedure has been identified to isolate rMophs, results of flow cytometric analysis of rMophs have been inconsistent among studies. We therefore first evaluated a preparative method for rMophs using collagenous digestion. The yield of rMophs greatly increased after the collagenase digestion. In particular, F4/80^high rMophs, which were positive for CD11c, a specific marker of DCs, dramatically increased. In addition, since neutrophils are sometimes mixed among rMophs in the analysis of flow cytometry, we established a gating strategy for eliminating neutrophils. To determine the contribution of rMophs to the development of autoimmune nephritis, we analyzed an experimental model of autoimmune nephritis that was applied to Shp1 conditional knockout mice (Shp1 CKO). This knockout strain is generated by crossing a mouse line carrying floxed Shp1 allele to mice expressing Cre recombinase under the control of the CD11c promoter. Shp1 CKO therefore specifically lack Shp1 in cells expressing CD11c. As a result, Shp1 CKO were susceptible to that experimental glomerulonephritis and F4/80^high rMophs of Shp1 CKO increased dramatically. In conclusion, our preparative methods for collagenase digestion and gating strategy for neutrophils are necessary for the analysis of rMophs, and Shp1 suppresses the development of autoimmune nephritis through the control of rMophs.     

Assessing support for Blaberoidea phylogeny suggests optimal locus quality

Phylogenomics seeks to use next‐generation data to robustly infer an organism's evolutionary history. Yet, the practical caveats of phylogenomics motivate investigation of improved efficiency, particularly when quality of phylogenies are questionable. To achieve improvements, one goal is to maintain or enhance the quality of phylogenetic inference while severely reducing dataset size. We approach this by assessing which kinds of loci in phylogenomic alignments provide the majority of support for a phylogenetic inference of cockroaches in Blaberoidea. We examine locus substitution rate, saturation, evolutionary divergence, rate heterogeneity, stabilizing selection, and a priori information content as traits that may determine optimality. Our controlled experimental design is based on 265 loci for 102 blaberoidean taxa and 22 outgroup species. Loci with high substitution rate, low saturation, low sequence distance, low rate heterogeneity, and strong stabilizing selection derive more support for phylogenetic relationships. We found that some phylogenetic information content estimators may not be meaningful for assessing information content a priori. We use these findings to design concatenated datasets with an optimized subsample of 100 loci. The tree inferred from the optimized subsample alignment was largely identical to that inferred from all 265 loci but with less evidence of long branch attraction, improved statistical support, and potential 4‐6x improvements to computation time. Supported by phylogenetic and morphological evidence, we erect three newly named clades (Anallactinae Evangelista & Wipfler subfam. nov., Orkrasomeria tax. nov. Evangelista, Wipfler, & Béthoux and Hemithyrsocerini Evangelista tribe nov.) and propose other taxonomic modifications. The diagnosis of Pseudophyllodromiidae Grandcolas, 1996 is modified to accommodate Anallactinae and Pseudophyllodromiinae Vickery & Kevan, 1983. The diagnosis of Ectobiidae Brunner von Wattenwyl, 1865 is modified to add novel morphological characters.     

<Emphasis Type="Italic">Boletus kermesinus</Emphasis>, a new species of <Emphasis Type="Italic">Boletus</Emphasis> section <Emphasis Type="Italic">Luridi</Emphasis> from central Honshu,Japan

Boletus kermesinus, a new species of Boletus section Luridi, is fully described and illustrated based on the materials collected in subalpine coniferous forests of central Honshu, Japan. It has distinctive features of dark-red basidiomata having distinct viscidity in the pileus surface, usually unchanging flesh, discolorous red pores, and an entirely reticulate stipe becoming coarsely lacerate-rimose with age.     

Rnd2 differentially regulates oligodendrocyte myelination at different developmental periods

In the CNS, oligodendrocyte precursor cells differentiate into oligodendrocytes to wrap their plasma membranes around neuronal axons, generating mature neural networks with myelin sheaths according to spatial and temporal patterns. While myelination is known to be one of the most dynamic cell morphological changes, the overall intrinsic and extrinsic molecular cues controlling myelination remain to be fully clarified. Here, we describe the biphasic roles of Rnd2, an atypical branch of the Rho family GTPase, in oligodendrocyte myelination during development and after maturation in mice. Compared with littermate controls, oligodendrocyte-specific Rnd2 knockout mice exhibit decreased myelin thickness at the onset of myelination but increased myelin thickness in the later period. Larger proportions of Rho kinase and its substrate Mbs, the signaling unit that negatively regulates oligodendrocyte myelination, are phosphorylated at the onset of myelination, while their smaller proportions are phosphorylated in the later period. In addition, we confirm the biphasic role of Rnd2 through experiments with oligodendrocyte-specific Rnd2 transgenic mice. We conclude that Rnd2 positively regulates myelination in the early myelinating period and negatively regulates myelination in the later period. This unique modulator thus plays different roles depending on the myelination period.     

Ketone bodies: A double-edged sword for mammalian life span

Accumulating evidence suggests health benefits of ketone bodies, and especially for longevity. However, the precise role of endogenous ketogenesis in mammalian life span, and the safety and efficacy of the long-term exogenous supplementation of ketone bodies remain unclear. In the present study, we show that a deficiency in endogenous ketogenesis, induced by whole-body Hmgcs2 deletion, shortens life span in mice, and that this is prevented by daily ketone body supplementation using a diet containing 1,3-butanediol, a precursor of β-hydroxybutyrate. Furthermore, feeding the 1,3-butanediol-containing diet from early in life increases midlife mortality in normal mice, but in aged mice it extends life span and prevents the high mortality associated with atherosclerosis in ApoE-deficient mice. By contrast, an ad libitum low-carbohydrate ketogenic diet markedly increases mortality. In conclusion, endogenous ketogenesis affects mammalian survival, and ketone body supplementation may represent a double-edged sword with respect to survival, depending on the method of administration and health status.     

Kinetic study on the dimer-tetramer interconversion of phosphorylase b by a stopped-flow X-ray scattering method

The dimer-tetramer interconversion of phosphorylase b induced by the binding of AMP and Mg2+ was monitored using a stopped-flow X-ray scattering method. The rate constants of this second-order reaction have been determined by a nonlinear least-squares method. Burst phases in both radii of gyration and zero-angle intensities were detected at the initial step of the reaction. This suggests that rapid association might take place, followed by a slow association process of which the kinetics were measured in the present study. The radius of gyration of tetrameric phosphorylase b was determined and found to be in excellent agreement with that of phosphorylase a, but different from that of phosphorylase b reported elsewhere (G. Puchwein, O. Kratky, C. F. Golker and E. Helmreich, Biochemistry 9 (1970) 4691). The reason for this inconsistency is discussed.     

Morphology and molecular phylogeny ofTretopileus sphaerophorus, a synnematous hyphomycete with basidiomycetous affinities

Tretopileus sphaerophorus, a synnematous hyphomycete with basidiomycetous affinities was newly isolated from the decaying petiole and peduncle ofCocos nucifera collected in Depok, Indonesia. The species produced first a bulbil as a propagule on the top of a synnema. After the bulbil had fallen, the synnema proliferated about seven times to produce new bulbils, each time making conspicuous nodes at the upper part. By careful morphological observation, clamp connections were confirmed on the hyphae in the specimens and culture. In culture, each hyphal cell with or without a clamp was found to be dikaryotic by DAPI nuclear staining. Germination of the bulbils occurred first from projecting hyphal tips on their upper surface, which have been treated as germ pores. The inner structure of the bulbils, the hyaline mucus of the bulbils, and conidium-like hyphal fragments were also examined. Phylogenetically,T. sphaerophorus was inferred to be related to the Aphyllophorales based on the nuclear encoded small subunit (18S) rDNA using the homology search system (FASTA) and the neighbour-joining method. Part 10 in a series on the taxonomy of synnematous fungi.