Stability and reaction characteristics of reverse-phase evaporation vesicles (REVs) as enzyme containers

The ion-dependent enzymes, activities of which are affected by ions, were entrapped in the inside aqueous phase of reverse-phase evaporation vesicles (REVs) to protect the enzyme activities from inhibitory cations. Their activities were still preserved in the presence of the inhibitory cations because the permeabilities of the inhibitory cations through the lipid membranes of REVs were much lower than those of substrates and products. The REVs were relatively stable in hypertonic condition, while they were unstable in hypotonic condition. Changes in osmotic pressure difference largely affected solute permeabilities of REVs. The shrinking and swelling of REVs resulting from osmotic pressure differences led to these changes in stabilities and solute permeabilities. Although the entrapment of the enzyme into REVs showed good protective effects against divalent cations, it was not effective against univalent cations. The measurements of cation permeabilities revealed that the enzyme trapped in the bilayer regions of REVs acts as a selective channel for the univalent cations. The REVs containing the enzyme could be used without any activity losses in a hollow fiber module.     

Use of single heart cells from chick embryos for the Na+ current measurements

To record the fast Na⁺ current, spheroidal heart cells enzymatically-dispersed from 3 18-day-old chick embryos were used for voltage clamping. The peak of currents in response to voltage steps of 200 ms long from holding potentials of -90 -105 mV were measured. The current-voltage curves for the peak inward current showed U-shaped relations; the averaged peak current of about -1400 pA was observed at about -30 mV and the current reversed sign at +40 + 50 mV. Both the peak current and the reversal potential values showed marked [Na]_o- dependence, i.e. reduced by 36% and by 20 mV, respectively, for a halved [Na]_o. Tetrodotoxin (TTX) partially (10^-6 M) or completely (10^-5 M) suppressed the current. The steady-state inactivation of the current (h) was characterized by the half inactivation voltage of around -80 mV and the slope factor of -4 -8 mV. The half activation voltage and the slope factor for the steady-state activation (m) were -55 mV and 4-6 mV, respectively. The electrophysiological and pharmacological properties were similar between young (3-day-old) and old (15-18-day-old) embryonic heart cells, excepting the much smaller current and the slower onset of TTX action in young embryonic hearts.     

Start-up of chemostat: Application of fed-batch culture

A time-optimal strategy for start-up of a chemostat has been established by applying Miele's extremization method based on Green's theorem. The start-up time is always minimized by traveling along the uppermost boundary of the set of admissible domains on the (vx, μ) phase plane. The solutions are batch culture for monotone increasing growth kinetics and exponentially fed-batch culture followed by batch culture for substrate-inhibition kinetics. A time-suboptimal start-up strategy, exponentially fed-batch culture operated the same μ as that of chemostate, is proposed as more feasible than and physiologically preferable to the time-optimal strategies. The complete set of equations of operations necessary for performing each of the optimal and suboptimal start-ups is formulated. Numerical comparisons on the basis of continuous operation giving maximum productivities of cell-mass outputs show that differences in times required for start-up between optimal and suboptimal strategies are small.     

Full-length sequence analysis of subacute sclerosing panencephalitis (SSPE) virus, a mutant of measles virus, isolated from brain tissues of a patient shortly after onset of SSPE

Subacute sclerosing panencephalitis (SSPE) virus, a measles virus (MeV) mutant, was isolated from brain tissues of a patient shortly after the clinical onset, and the entire viral genome was sequenced. The virus, named SSPE-Kobe-1, formed syncytia on B95a and Vero/SLAM cells without producing cell-free infectious virus particles, which is characteristic of SSPE virus. Phylogenetic analysis classified SSPE-Kobe-1 into genotype D3. When compared with an MeV field isolate of the same genotype (Ich-B strain), SSPE-Kobe-1 exhibited mutation rates of 0.8-1.6% at the nucleotide level in each of the proteincoding regions of the viral genome. It is noteworthy that the mutation rate of the M gene (1.2%) of SSPE-Kobe-1 was considerably lower than for other SSPE virus strains reported so far, but that the majority of the mutations (75%) were the uridine-to-cytidine biased hypermutation characteristic of the SSPE virus M gene. At the amino acid level, the viral proteins, such as N, P, C, V, M, F, H and L proteins, had point-mutations on 3, 7, 1, 4, 3, 9, 8 and 14 residues, respectively, compared with the Ich-B strain. In addition, the F and H proteins had mutated C-termini due to single-point mutations near or at the stop codons. Two of the three mutations in the M protein were Leu-to-Pro mutations, which are likely to affect the conformation and, therefore, the function of the protein. Because of the relatively small number of mutations, SSPE-Kobe-1 would be a useful tool to study genetic evolution of SSPE virus.     

Differential regulation of hyaluronan-induced IL-8 and IP-10 in airway epithelial cells

Airway epithelium is emerging as a regulator of local inflammation and immune responses. However, the cellular and molecular mechanisms responsible for the immune modulation by these cells have yet to be fully elucidated. At the cellular level, the hallmarks of airway inflammation are mucus gland hypertrophy with excess mucus production, accumulation of inflammatory mediators, inflammation in the airway walls and lumen, and breakdown and turnover of the extracellular matrix. We demonstrate that fragments of the extracellular matrix component hyaluronan induce inflammatory chemokine production in primary airway epithelial cells grown at an air-liquid interface. Furthermore, hyaluronan fragments use two distinct molecular pathways to induce IL-8 and IFN-gamma-inducible protein 10 (IP-10) chemokine expression in airway epithelial cells. Hyaluronan-induced IL-8 requires the MAP kinase pathway, whereas hyaluronan-induced IP-10 utilizes the NF-kappaB pathway. The induction is specific to low-molecular-weight hyaluronan fragments as other glycosaminoglycans do not induce IL-8 and IP-10 in airway epithelial cells. We hypothesize that not only is the extracellular matrix a target of destruction in airway inflammation but it plays a critical role in perpetuating inflammation through the induction of cytokines, chemokines, and modulatory enzymes in epithelial cells. Furthermore, hyaluronan, by inducing IL-8 and IP-10 by distinct pathways, provides a unique target for differential regulation of key inflammatory chemokines.     

Characterization of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> isolated from chronically infected children with cystic fibrosis in India

Background  

Pseudomonas aeruginosa is the leading cause of morbidity and mortality in patients with cystic fibrosis (CF). With chronicity of infection, the organism resides as a biofilm, shows multi-drug resistance, diversifies its colony morphology and becomes auxotrophic. The patients have been found to be colonized with multiple genotypes. The present work was carried out to characterize P. aeruginosa isolated from children with cystic fibrosis using phenotypic and genotypic methods.     

Evolutionary history of Chaetognatha inferred from molecular and morphological data: a case study for body plan simplification

Background

Chaetognatha are a phylum of marine carnivorous animals which includes more than 130 extant species. The internal systematics of this group have been intensively debated since it was discovered in the 18^th century. While they can be traced back to the earlier Cambrian, they are an extraordinarily homogeneous phylum at the morphological level - a fascinating characteristic that puzzled many a scientist who has tried to clarify their taxonomy. Recent studies which have attempted to reconstruct a phylogeny using molecular data have relied on single gene analyses and a somewhat restricted taxon sampling. Here, we present the first large scale phylogenetic study of Chaetognatha based on a combined analysis of nearly the complete ribosomal RNA (rRNA) genes. We use this analysis to infer the evolution of some morphological characters. This work includes 36 extant species, mainly obtained from Tara Oceans Expedition 2009/2012, that represent 16 genera and 6 of the 9 extant families.

Results

Cladistic and phenetic analysis of morphological characters, geometric morphometrics and molecular small subunit (SSU rRNA) and large subunit (LSU rRNA) ribosomal genes phylogenies provided new insights into the relationships and the evolutionary history of Chaetognatha. We propose the following clade structure for the phylum: (((Sagittidae, Krohnittidae), Spadellidae), (Eukrohniidae, Heterokrohniidae)), with the Pterosagittidae included in the Sagittidae. The clade (Sagittidae, Krohnittidae) constitutes the monophyletic order of Aphragmophora. Molecular analyses showed that the Phragmophora are paraphyletic. The Ctenodontina/Flabellodontina and Syngonata/Chorismogonata hypotheses are invalidated on the basis of both morphological and molecular data. This new phylogeny also includes resurrected and modified genera within Sagittidae.

Conclusions

The distribution of some morphological characters traditionally used in systematics and for species diagnosis suggests that the diversity in Chaetognatha was produced through a process of mosaic evolution. Moreover, chaetognaths have mostly evolved by simplification of their body plan and their history shows numerous convergent events of losses and reversions. The main morphological novelty observed is the acquisition of a second pair of lateral fins in Sagittidae, which represents an adaptation to the holoplanktonic niche.

     

Increased activity and expression of histone deacetylase 1 in relation to tumor necrosis factor-alpha in synovial tissue of rheumatoid arthritis

Introduction  

The purpose of this study was to investigate the profile of histone deacetylase (HDAC) expression in the synovial tissue of rheumatoid arthritis (RA) compared with that of normal control and osteoarthritis (OA), and to examine whether there is a link between HDAC activity and synovial inflammation.     

Correlation of CrystaL Growth with the Staining of Axons by the Golgi Procedure

The surface of the specimens subjected to a modified Golgi technique (formalin fixed material; specimens in the following solution for 8-10 days at 27 C: 3% K₂Cr₂O₇, 100 ml, with the addition of 2.5-10 ml of 10% formalin and 6-25 gm of sucrose; then in 0.75% AgNO₃ for at least 2 days at 27 C) is sometimes covered with a fur of filamentous crystals and sometimes with a powdery precipitate of laminar crystals. In a series of experiments in which about 500 blocks of tissue were treated with variations of the staining procedure, good axonal stain was positively correlated with the appearance of filamentous crystals. These filaments have a thickness of 1-4 μ and grow at a rate of 160-330 μ/hr, reaching a length of 2-7 mm.