Phylogeography of the temperate grassland plant Tephroseris kirilowii (Asteraceae) inferred from multiplexed inter-simple sequence repeat genotyping by sequencing (MIG-seq) data

Journal of Plant Research - A group of temperate grassland plant species termed the “Mansen elements” occurs in Japan and is widely distributed in the grasslands of continental East...     

Effects of endothelin-1 and conversion of big endothelin-1 in the isolated perfused rabbit lung.

We examined the effects of endothelin-1 (ET-1) on pulmonary hemodynamic and transvascular fluid filtration and the conversion of big endothelin-1 (big ET-1), a precursor of ET-1, in isolated perfused rabbit lungs at constant vascular and airway pressures. Furthermore we examined whether ET-1 contributes to cyclooxygenase metabolism. The perfusate flow decreased significantly after bolus administration of 1 or 0.1 nmol of ET-1. Lung weight did not increase throughout the experimental period. Big ET-1- (1 nmol) induced decrease in the flow was slow in developing, although the maximum response was comparable to that induced by the same dose of ET-1. The concentration of bit ET-1 in the perfusate progressively decreased, while that of ET-1 increased in a time-dependent manner. Phosphoramidon, an inhibitor of metalloproteinase, suppressed the pressor effect of big ET-1 (P less than 0.01) and the increase in the concentration of ET-1 in the perfusate (P less than 0.05). The present findings provide the first evidence suggesting that the potent vasocontractile effect of big ET-1 in pulmonary circulation can be attributed to the production of ET-1 by the conversion from big ET-1 in the vascular bed. ET-1-induced perfusate flow changes were not affected by indomethacin, and the concentration of 6-ketoprostaglandin F1 alpha, a metabolite of prostacyclin, did not increase after ET-1 administration.     

Development of human cerebellar nuclei. Morphometric study

The morphometric development of the human cerebellar nuclei was examined in 9 fetuses (16-40 weeks of gestation; WG), an infant (2 months old) and 2 adults (16 and 63 years old). With the morphological observation of serial sections of the brain containing the cerebellar nuclei, the authors measured sections to get several morphometric parameters: the volume of nuclear column and number, packing density and cell body area of neurons. Each nucleus (dentate, emboliform, globose and fastigial nucleus) was recognized even at 16 WG. Nerve cells containing Nissl bodies were observed in all nuclei after 23 WG. Degenerative changes were detected in some neurons for every nucleus at 21 and 23 WG. Three stages were observed in the developmental course of nuclear volume and neuronal packing density: the primary or undifferentiated stage at 16 WG, the secondary stage with variability at 21-32 WG and the tertiary stage with monotonous increase (nuclear volume) or gradual decrease (neuronal packing density) after 35 WG. No significant correlation between neuronal number and gestational age was noticed for every nucleus. The analysis of cell body area (neuronal size) demonstrated that the dentate neurons developed after the intermediate or fastigial neurons. It is concluded that there is a critical period between slightly before 20 WG and slightly after 30 WG, matched with the secondary stage in the development of the cerebellar nuclei.     

Coevolution of Bacteriophage PP01 and Escherichia coli O157:H7 in Continuous Culture

The interaction between Escherichia coli O157:H7 and its specific bacteriophage PP01 was investigated in chemostat continuous culture. Following the addition of bacteriophage PP01, E. coli O157:H7 cell lysis was observed by over 4 orders of magnitude at a dilution rate of 0.876 h⁻¹ and by 3 orders of magnitude at a lower dilution rate (0.327 h⁻¹). However, the appearance of a series of phage-resistant E. coli isolates, which showed a low efficiency of plating against bacteriophage PP01, led to an increase in the cell concentration in the culture. The colony shape, outer membrane protein expression, and lipopolysaccharide production of each escape mutant were compared. Cessation of major outer membrane protein OmpC production and alteration of lipopolysaccharide composition enabled E. coli O157:H7 to escape PP01 infection. One of the escape mutants of E. coli O157:H7 which formed a mucoid colony (Mu) on Luria-Bertani agar appeared 56 h postincubation at a dilution rate of 0.867 h⁻¹ and persisted until the end of the experiment (~200 h). Mu mutant cells could coexist with bacteriophage PP01 in batch culture. Concentrations of the Mu cells and bacteriophage PP01 increased together. The appearance of mutant phage, which showed a different host range among the O157:H7 escape mutants than wild-type PP01, was also detected in the chemostat culture. Thus, coevolution of phage and E. coli O157:H7 proceeded as a mutual arms race in chemostat continuous culture.     

Kinetics and collagenolytic role of eosinophils in chronic gastric ulcer in the rat

 An association between eosinophils and tissue damage has been observed in numerous disorders. However, few reports have addressed the role of infiltrating eosinophils in gastric ulcer healing. The aim of this study was to investigate the kinetics and role of eosinophils infiltrating experimental chronic gastric ulcers in the rat. We developed a monoclonal antibody against human matrix metalloproteinase 1 (MMP1) purified from conditioned culture medium of human skin fibroblasts. Acetic acid-induced gastric ulcers were resected from rats on days 1, 3, 5, 10, 20, 40, and 180 after the days of induction (day 0). Tissue specimens were immunostained with this antibody and examined with an electron microscope. Few eosinophils were observed in the granulation tissue until day 20. By days 40 and 180, MMP1-positive eosinophils had increased in the granulation tissue of open ulcers. Azan staining revealed dispersed collagen fibers around infiltrating eosinophils. In contrast, scars demonstrated few eosinophils in fibrous tissue on days 40 and 180. Eosinophils which express MMP1 infiltrate granulation tissue at the chronic stage of gastric ulceration. The results suggest that eosinophils may play a role in tissue remodeling and deterioration of ulceration. Accepted: 18 March 1997     

Cell migration within the embryonic limb primordium of Drosophila as revealed by a novel fluorescence method to visualize mRNA and protein

 We report a new technique using fluorescent probes to detect a mRNA and a protein simultaneously in the Drosophila embryo. For in situ hybridization, 3-hydroxy-N-2′-biphenyl-2-naphthalenecarboxamide phosphate ester (HNPP)/Fast Red TR was used as a fluorescent substrate for alkaline phosphatase. It was possible to compare protein and mRNA expression on a cell by cell basis with a laser scanning confocal microscope. We applied this technique to analyse the dynamics of Distal-less (Dll) enhancer activity in the thoracic limb primordium in the early Drosophila embryo. We stained embryos bearing the Dll early enhancer (Dll-304) fused to the Escherichia coli lacZ gene. LacZ mRNA was detectable in the ventral region of the limb primordium, and β-galactosidase protein in the dorsal region. In the middle, both mRNA and protein were detectable. These results suggest that the Dll enhancer is activated in the ventral region of the limb primordium and that Dll-positive cells migrate from a ventral position to a dorsal one within a single limb primordium. Received: 7 April 1997 / Accepted: 15 May 1997     

Phylogenetic Position of the Mitochondrion-Lacking Protozoan Trichomonas tenax,Based on Amino Acid Sequences of Elongation Factors 1α and 2

Major parts of amino-acid-coding regions of elongation factor (EF)-1α and EF-2 in Trichomonas tenax were amplified by PCR from total genomic DNA and the products were cloned into a plasmid vector, pGEM-T. The three clones from each of the products of the EF-1α and EF-2 were isolated and sequenced. The insert DNAs of the clones containing EF-1α coding regions were each 1,185 bp long with the same nucleotide sequence and contained 53.1% of G + C nucleotides. Those of the clones containing EF-2 coding regions had two different sequences; one was 2,283 bp long and the other was 2,286 bp long, and their G + C contents were 52.5 and 52.9%, respectively. The copy numbers of the EF-1α and EF-2 gene per chromosome were estimated as four and two, respectively. The deduced amino acid sequences obtained by the conceptual translation were 395 residues from EF-1α and 761 and 762 residues from the EF-2s. The sequences were aligned with the other eukaryotic and archaebacterial EF-1αs and EF-2s, respectively. The phylogenetic position of T. tenax was inferred by the maximum likelihood (ML) method using the EF-1α and EF-2 data sets. The EF-1α analysis suggested that three mitochondrion-lacking protozoa, Glugea plecoglossi, Giardia lamblia, and T. tenax, respectively, diverge in this order in the very early phase of eukaryotic evolution. The EF-2 analysis also supported the divergence of T. tenax to be immediately next to G. lamblia. Received: 15 February 1996 / Accepted: 28 June 1996     

Mutational analysis of human immunodeficiency virus type 1 (HIV-1) accessory genes: requirement of a site in the nef gene for HIV-1 replication in activated CD4+ T cells in vitro and in vivo.

Human immunodeficiency virus type 1 (HIV-1) accessory genes including nef, vif, and vpr are important factors that determine the replication and pathogenesis of HIV-1. The state of activation is also important for the replication of HIV-1. We evaluated the properties of nef-, vif-, and vpr-minus macrophage-tropic HIV-1(JR) CSF in primary CD4+ Th1- or Th2-like cell cultures which had been activated through CD3 molecules in the presence of interleukin-2 (IL-2) and IL-12 (Th1-like culture) or IL-4 (Th2-like culture), respectively. In activated Th1- or Th2-like cultures, replication of nef-minus HIV-1(JR-CSF) was markedly lower than that of wild-type HIV-1. Subsequent analysis by site-directed mutagenesis showed that (i) the presence of an acidic amino acid-rich domain (amino acid residues 72 to 75) in the Nef protein was critical for the enhancement of viral DNA synthesis, resulting in increased virus growth rate, and (ii) prolines that form part of Src homology 3 binding domain were not essential for viral replication. We also confirmed the importance of sites by using an HIV-1-infected animal model, the hu-PBL-SCID mouse system, representing HIV-1 replication and pathogenesis in activated CD4+ T cells in vivo. These results indicate that Nef accelerates viral replication in activated CD4+ T cells.     

Does endothelin-1 participate in the exercise-induced changes of blood flow distribution of muscles in humans?

Maeda, Seiji, Takashi Miyauchi, Michiko Sakane, MakotoSaito, Shinichi Maki, Katsutoshi Goto, and Mitsuo Matsuda. Does endothelin-1 participate in the exercise-induced changes of blood flowdistribution of muscles in humans? J. Appl.Physiol. 82(4): 1107-1111, 1997.Endothelin-1(ET-1) is an endothelium-derived potent vasoconstrictor peptide thatpotentiates contractions to norepinephrine in human vessels. Wepreviously reported that the circulating plasma concentration of ET-1is significantly increased after exercise (S. Maeda, T. Miyauchi, K. Goto, and M. Matsuda. J. Appl.Physiol. 77: 1399-1402, 1994). Tostudy the roles of ET-1 during and after exercise, we investigatedwhether endurance exercise affects the production of ET-1 in thecirculation of working muscles and nonworking muscles. Male athletesperformed one-leg cycle ergometer exercise of 30-min duration atintensity of 110% of their individual ventilatory threshold. Plasmaconcentrations of ET-1 in both sides of femoral veins (veins in theworking leg and nonworking leg) and in the femoral artery (artery inthe nonworking leg) were measured before and afterexercise. The plasma ET-1 concentration in the femoralvein in the nonworking leg was significantly increased after exercise,whereas that in femoral vein in the working leg was not changed. Thearteriovenous difference in ET-1 concentration was significantlyincreased after exercise in the circulation of the nonworking leg butnot of the working leg, which suggests that the production of ET-1 wasincreased in the circulation of the nonworking leg by exercise. Thepresent study also demonstrated that the plasma norepinephrineconcentrations were elevated by exercise in the femoral veins of boththe working and nonworking legs, suggesting that the sympathetic nerveactivity was augmented in both legs during exercise. Therefore, thepresent study demonstrates the possibility that the increase inproduction of ET-1 in nonworking muscles may cause vasoconstriction andhence decrease blood flow in nonworking muscles through its directvasoconstrictive action or through an indirect effect of ET-1 toenhance vasoconstrictions to norepinephrine and that these responsesmay be helpful in increasing blood flow in workingmuscles. We propose that endogenous ET-1 contributes tothe exercise-induced redistribution of blood flow in muscles.

     

Alcohol dehydrogenase (ADH) isozymes in the AdhN/AdhN strain ofPeromyscus maniculatus (ADH− deermouse) and a possible role of class III ADH in alcohol metabolism

Although the Adh^N/Adh^N strain ofPeromyscus maniculatus (so-called ADH^? deermouse) has been previously considered to be deficient in ADH, we found ADH isozymes of Classes II and III but not Class I in the liver of this strain. On the other hand, the Adh^F/Adh^F strain (so-called ADH⁺ deermouse), which has liver ADH activity, had Class I and III but not Class II ADH in the liver. In the stomach, Class III and IV ADHs were detected in both deermouse strains, as well as in the ddY mouse, which has the normal mammalian ADH system with four classes of ADH. These ADH isozymes were identified as electrophoretic phenotypes on the basis of their substrate specificity, pyrazole sensitivity, and immunoreactivity. Liver ADH activity of the ADH^? strain was barely detectable in a conventional ADH assay using 15 mM ethanol as substrate; however, it increased markedly with high concentrations of ethanol (up to 3M) or hexenol (7 mM). Furthermore, in a hydrophobic reaction medium containing 1.0M t-butanol, liver ADH activity of this strain at low concentrations of ethanol (<100 mM) greatly increased (about sevenfold), to more than 50% that of ADH⁺ deermouse. These results were attributable to the presence of Class III ADH and the absence of Class I ADH in the liver of ADH^? deermouse. It was also found that even the ADH⁺ strain has low liver ADH activity (<40% that of the ddY mouse) with 15 mM ethanol as substrate, probably due to low activity in Class I ADH. Consequently, liver ADH activity of this strain was lower than its stomach ADH activity, in contrast with the ddY mouse, whose ADH activity was much higher in the liver than in the stomach, as well as other mammals. Thus, the ADH systems in both ADH^? and ADH⁺ deermouse were different not only from each other but also from that in the ddY mouse; the ADH^? strain was deficient in only Class I ADH, and the ADH⁺ strain was deficient in Class II ADH and down-regulated in Class I ADH activity. Therefore, Class III ADH, which was found in both strains and activated allosterically, may participate in alcohol metabolism in deermouse, especially in the ADH^? strain.