Approximation equation for oxygen uptake kinetics in decrement-load exercise starting from low exercise intensity

The purpose of the present study was to determine the degree of fitting an approximation equation for oxygen uptake (Vo(2)) in decrement-load exercise (DLE). Work rate was started from 120 watts and was decreased by a rate of 15 watts per min. The initial work rate of DLE corresponded to 72+/-10% of the work rate at anaerobic threshold determined in incremental-load exercise (ILE). Vo(2) in DLE increased rapidly, reached a peak, and decreased linearly until the end of the exercise. Vo(2) in DLE was higher than that in ILE at the same work rate except in the early periods in ILE and DLE. This difference ranged from 300 to 400 ml/min. This difference is a result of repayment of oxygen debt in DLE and from the oxygen deficit induced by the delay of response of Vo(2) in ILE. As work rate in DLE can be obtained by the difference between work rates in constant-load exercise (CLE) and ILE, we postulated that the approximation equation for Vo(2) kinetics in DLE could be expressed by a combination of approximation equations in CLE and in ILE. When time delay was taken into consideration in this equation, the fitting of data obtained by using the equation was better than that of data obtained by using the equation without a parameter of time delay. The degree of fitting ranged from 94 to 98% (r(2)). Thus, it seems that Vo(2) including oxygen debt in DLE can be approximated by the equation used in this study.     

Photoswitching of peroxidase activity by position-specific incorporation of a photoisomerizable non-natural amino acid into horseradish peroxidase.

Horseradish peroxidase mutants containing L-p-phenylazophenylalanine (azoAla) at various positions were synthesized by using an Escherichia coli in vitro translation system. Among the 15 mutants examined, four mutants containing a single azoAla unit at the 6th, 68th, 142nd, and 179th positions, respectively, retained the peroxidase activity. The activity of the Phe68azoAla mutant was higher when the azobenzene group was in the cis form than in the trans form. On the contrary, the activity of the Phe179azoAla mutant disappeared when the azobenzene group was photoisomerized to the cis form, but recovered in the trans form. In the latter mutant, therefore, an on/off photoswitching of the peroxidase activity was attained.     

Analysis of gene expression profile in p130(Cas)-deficient fibroblasts

p130(Cas) (Cas) is a docking protein that becomes tyrosine phosphorylated in v-Src- or v-Crk-transformed cells and in integrin-stimulated cells. Cas -/- fibroblasts show defects in stress fiber formation, cell spreading, cell migration, and transformation by activated Src. To further characterize the role of Cas in signaling, we compared the expression profile in Cas -/- fibroblasts with that in Cas-re-expressing fibroblasts using the microarray methods. In Cas -/- fibroblasts, the expression of heme oxygenase 1 and caveolin-1 was reduced, but the expression of procollagen 1 alpha 1, procollagen 3 alpha 1, procollagen 11 alpha 1, elastin, periostin, TSC-36, and MARCKS was enhanced. The domains in Cas necessary for the change varied among these genes. Activated Src reduced the expression of most of these genes both in Cas -/- and in Cas +/+ fibroblasts. These results suggest the existence of signaling pathways that emanate from Cas to gene expression.     

The biomechanics of amnion rupture: an X-ray diffraction study

Pre-term birth is the leading cause of perinatal and neonatal mortality, 40% of which are attributed to the pre-term premature rupture of amnion. Rupture of amnion is thought to be associated with a corresponding decrease in the extracellular collagen content and/or increase in collagenase activity. However, there is very little information concerning the detailed organisation of fibrillar collagen in amnion and how this might influence rupture. Here we identify a loss of lattice like arrangement in collagen organisation from areas near to the rupture site, and present a 9% increase in fibril spacing and a 50% decrease in fibrillar organisation using quantitative measurements gained by transmission electron microscopy and the novel application of synchrotron X-ray diffraction. These data provide an accurate insight into the biomechanical process of amnion rupture and highlight X-ray diffraction as a new and powerful tool in our understanding of this process.     

Solution structure of the SWIRM domain of human histone demethylase LSD1

SWIRM is an evolutionarily conserved domain involved in several chromatin-modifying complexes. Recently, the LSD1 protein, which bears a SWIRM domain, was found to be a demethylase for Lys4-methylated histone H3. Here, we report a solution structure of the SWIRM domain of human LSD1. It forms a compact fold composed of 6 alpha helices, in which a 20 amino acid long helix (alpha4) is surrounded by 5 other short helices. The SWIRM domain structure could be divided into the N-terminal part (alpha1-alpha3) and the C-terminal part (alpha4-alpha6), which are connected to each other by a salt bridge. While the N-terminal part forms a SWIRM-specific structure, the C-terminal part adopts a helix-turn-helix (HTH)-related fold. We discuss a model in which the SWIRM domain acts as an anchor site for a histone tail.     

Multipotent cells from the human third molar: feasibility of cell-based therapy for liver disease

Abstract Adult stem cells have been reported to exist in various tissues. The isolation of high-quality human stem cells that can be used for regeneration of fatal deseases from accessible resources is an important advance in stem cell research. In the present study, we identified a novel stem cell, which we named tooth germ progenitor cells (TGPCs), from discarded third molar, commonly called as wisdom teeth. We demonstrated the characterization and distinctiveness of the TGPCs, and found that TGPCs showed high proliferation activity and capability to differentiate in vitro into cells of three germ layers including osteoblasts, neural cells, and hepatocytes. TGPCs were examined by the transplantation into a carbon tetrachloride (CCl₄)-treated liver injured rat to determine whether this novel cell source might be useful for cell-based therapy to treat liver diseases. The successful engraftment of the TGPCs was demonstrated by PKH26 fluorescence in the recipient's rat as to liver at 4 weeks after transplantation. The TGPCs prevented the progression of liver fibrosis in the liver of CCl₄-treated rats and contributed to the restoration of liver function, as assessed by the measurement of hepatic serum markers aspartate aminotransferase and alanine aminotransferase. Furthermore, the liver functions, observed by the levels of serum bilirubin and albumin, appeared to be improved following transplantation of TGPCs. These findings suggest that multipotent TGPCs are one of the candidates for cell-based therapy to treat liver diseases and offer unprecedented opportunities for developing therapies in treating tissue repair and regeneration.     

Tamiflu-resistant but HA-mediated cell-to-cell transmission through apical membranes of cell-associated influenza viruses

The infection of viruses to a neighboring cell is considered to be beneficial in terms of evasion from host anti-virus defense systems. There are two pathways for viral infection to "right next door": one is the virus transmission through cell-cell fusion by forming syncytium without production of progeny virions, and the other is mediated by virions without virus diffusion, generally designated cell-to-cell transmission. Influenza viruses are believed to be transmitted as cell-free virus from infected cells to uninfected cells. Here, we demonstrated that influenza virus can utilize cell-to-cell transmission pathway through apical membranes, by handover of virions on the surface of an infected cell to adjacent host cells. Live cell imaging techniques showed that a recombinant influenza virus, in which the neuraminidase gene was replaced with the green fluorescence protein gene, spreads from an infected cell to adjacent cells forming infected cell clusters. This type of virus spreading requires HA activation by protease treatment. The cell-to-cell transmission was also blocked by amantadine, which inhibits the acidification of endosomes required for uncoating of influenza virus particles in endosomes, indicating that functional hemagglutinin and endosome acidification by M2 ion channel were essential for the cell-to-cell influenza virus transmission. Furthermore, in the cell-to-cell transmission of influenza virus, progeny virions could remain associated with the surface of infected cell even after budding, for the progeny virions to be passed on to adjacent uninfected cells. The evidence that cell-to-cell transmission occurs in influenza virus lead to the caution that local infection proceeds even when treated with neuraminidase inhibitors.     

Analytical method for ubiquinone-9 and ubiquinone-10 in rat tissues by liquid chromatography/turbo ion spray tandem mass spectrometry with 1-alkylamine as an additive to the mobile phase

We investigated the application of 1-alkylamines, as additives to the mobile phase, to a quantification method for ubiquinone-9 (CoQ9) and ubiquinone-10 (CoQ10) in rat thigh muscle and heart using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In the optimization of the analytical method, we found that 1-alkylamines mixed with CoQ9 and CoQ10 in the turbo ion sprayed solution formed the 1-alkylammonium adduct molecules of these compounds during the ionization process and that the intensity of the adduct ions was considerably higher than that of the protonated molecules ([M+H]+) of these compounds. Furthermore, we investigated a variety of 1-alkylamines in the mobile phase for LC-MS/MS analysis to select the most appropriate 1-alkylamine for higher sensitivities of CoQ9 and CoQ10. After these examinations, we found that methylamine was the most suitable additive for the mobile phase, allowing a 12.5-fold gain in signal intensity in the full ion mass spectrum compared with that without methylamine. The internal standard (IS) used was ubiquinone-11 (CoQ11) for each analyte. The analytes and IS were extracted with methanol from the tissue homogenates at neutral pH and were injected into an LC-MS/MS with a turbo ion spray interface. The calibration curves for CoQ9 (5-500 microg/g in thigh muscle and 50-10,000 microg/g in heart) and CoQ10 (1-500 microg/g in thigh muscle and 10-10,000 microg/g in heart) showed good linearity. The method was precise; the relative standard deviations of the method for rat thigh muscle were not more than 13.5 and 9.0% for CoQ9 and CoQ10, respectively, and those for rat heart were not more than 6.7 and 5.4% for CoQ9 and CoQ10, respectively. The accuracies of the method for both rat thigh muscle and heart were good, with the deviations between the nominal concentration and calculated concentration of CoQ9 and CoQ10 typically being within 12.3 and 4.3%, respectively. This method provided reliable concentration levels for CoQ9 and CoQ10 in rat thigh muscle and heart.