Suppression of chronic antigen-induced arthritis in rats by a monoclonal antibody against the T cell receptor alpha beta.

We have investigated a role for T cells in chronic antigen-induced arthritis in rats employing a monoclonal antibody (R73 mAb) against the T cell receptor alpha beta. Treatment with R73 mAb from the time of intra-articular antigenic challenge blocked completely the induction of chronic, but not acute ovalbumin-induced arthritis in sensitized rats. Histologically, treatment-controlled arthritic rats exhibited marked hyperplasia of synovial membrane with pronounced infiltration of inflammatory cells including alpha beta + T cells in the chronic phase of arthritis. In contrast, R73 mAb-treated rats had almost normal joint histology. Treatment with R73 mAb after onset of arthritis was also effective in suppressing the progression of chronic antigen-induced joint inflammation. The preventive and suppressive effects of the mAb on chronic antigen-induced arthritis were associated with marked depletion of alpha beta + T cells in peripheral blood. The DTH but not the humoral response to ovalbumin in sensitized rats was suppressed significantly by R73 mAb. Thus, alpha beta + T cells appear to have a central role in both induction and progression of chronic antigen-induced arthritis.     

Binding of both Ca2+ and mastoparan to calmodulin induces a large change in the tertiary structure

The technique of small-angle X-ray scattering has been employed to examine the solution conformation of calmodulin and its complexes with Ca2+ alone, and with both Ca2+ and mastoparan. The radius of gyration decreased by 3.1 +/- 0.3 A upon binding of both 4 mol Ca2+/mol of protein and 1 mol mastoparan/mol of protein to form the ternary complex. A smaller increase was found for the separate binding of 4 mol Ca2+/mol of protein in the absence of mastoparan (0.6 +/- 0.3 A). The analyses of pair distance distribution function showed that the maximal pair distance in calmodulin complex with both Ca2+ and mastoparan decreased by 20-30% in comparison with calmodulin or its complex with Ca2+, and a shoulder near 40 A, which characterizes the dumbbell-shaped molecule of calmodulin, disappeared. These results indicate that the two globular domains of the calmodulin complex with Ca2+ and mastoparan come close together by 8.0-9.5 A on average, if the size and the overall shape of the globular domains are the same in Ca2+-calmodulin-mastoparan complex as in calmodulin or Ca2+-calmodulin complex.     

Calcium-induced shape change of calmodulin with mastoparan studied by solution X-ray scattering

Solution x-ray scattering using synchrotron radiation as an x-ray source was used to analyze the Ca2+-dependent shape change of pig brain calmodulin in detail. The radius of gyration of calmodulin at 10 mg/ml was increased by 0.9 A. The increase was nearly completed when 2.5 mol of Ca2+/mol of calmodulin was added, whereas the radius of gyration of calmodulin with mastoparan decreased by about 3 A with an increasing Ca2+ concentration up to 4 mol of Ca2+/mol of calmodulin. At a moderate angle of region, both scattering profiles from calmodulin with or without Ca2+ displayed clear humps at s = 0.03 A-1 which are characteristic of a dumbbell structure. However, in the presence of mastoparan, the hump in the scattering profile became obscure and later disappeared with the third and fourth Ca2+ binding to calmodulin. These findings are attributable to the Ca2+-induced shape change of calmodulin with mastoparan from a dumbbell structure to a non-dumbbell structure in which the distance between the two lobes of calmodulin become closer by a bend in the central helix.     

Botryococcus braunii carbon/nitrogen metabolism as affected by ammonia addition

Carbon metabolism in photosynthesizing and respiring cells of Botryococcus braunii was radically changed by the presence of 1 mM NH₄Cl in the medium, when the so-called resting state previously had been subjected to a nitrogen-deficient medium. Ammonia addition to the algae photosynthesizing with ¹⁴C-labelled HCO ₃ ^- almost completely inhibited the synthesis of ¹⁴C-labelled botryococcenes and other hexane-extractable compounds, and also inhibited the formation of insoluble compounds; however, it resulted in a large increase in the synthesis of alanine, glutamine, other amino acids, and especially of 5-aminolevulinic acid. Total CO₂ fixation decreased about 60% and O₂ evolution decreased more than 50%.CO₂ fixation in the dark with ammonia present led to labelled products derived from phosphoenolpyruvate carboxylation, such as glutamine, glutamate, and malate. Respiratory uptake of O₂ increased by about 70%.The inhibition of terpenoid synthesis and increased synthesis of C₅ amino acids by Botryococcus upon ammonia addition indicates 1) a diversion of acetyl coenzyme A from synthetic pathways leading to terpenoids and 2) increased operation of pathways leading to the synthesis of amino acids, especially 5-aminolevulinic acid, a precursor to chlorophyll biosynthesis.This work was supported in part by the Office of Energy Research, Office of Basic Energy Sciences, Biological Energy Research Division of the U.S. Department of Energy under Contract No. DE-AC03-76SF00098, in part by a grant from SOHIO, and, in part, by a grant from the Japan-U.S. Cooperative Science Program (The Japan Society for the Promotion of Science, National Science Foundation, Division of International Programs)     

Protective role of urinary trypsin inhibitor in acute lung injury induced by lipopolysaccharide

Urinary trypsin inhibitor (UTI), a serine protease inhibitor, has been widely used as a drug for patients with acute inflammatory disorders such as disseminated intravascular coagulation, shock, and pancreatitis. However, direct contribution of UTI to inflammatory diseases has not been established. The present study analyzed acute inflammatory lung injury induced by lipopolysaccharide (LPS) in UTI-deficient (-/-) mice and corresponding wild-type (WT) mice. UTI (-/-) and WT mice were treated intratracheally with vehicle or LPS (125 mug/kg). The cellular profile of bronchoalveolar lavage fluid, lung water content, histology, and expression of proinflammatory molecules in the lung were evaluated. After LPS challenge, both genotypes of mice revealed neutrophilic lung inflammation and pulmonary edema. UTI (-/-) mice, however, showed more prominent infiltration of inflammatory cells and edema than WT mice. After LPS challenge in both genotypes of mice, the lung levels of mRNA and/or protein expression of interleukin-1beta, macrophage inflammatory protein-1alpha, macrophage chemoattractant protein-1, keratinocyte chemoattractant, and intercellular adhesion molecule-1 (ICAM-1) were elevated in both groups, but to a greater extent in UTI (-/-) mice than in WT mice. These results suggest that UTI protects against acute lung injury induced by bacterial endotoxin, at least partly, through the inhibition of the enhanced local expression of proinflammatory cytokines, chemokines, and ICAM-1.     

Local changes in blood flow within the early and midcycle corpus luteum after prostaglandin F(2 alpha) injection in the cow

One of the postulated main luteolytic actions of prostaglandin (PG) F(2 alpha) is to decrease ovarian blood flow. However, before Day 5 of the normal cycle, the corpus luteum (CL) is refractory to the luteolytic action of PGF(2 alpha). Therefore, we aimed to determine in detail the real-time changes in intraluteal blood flow after PGF(2 alpha) injection at the early and middle stages of the estrous cycle in the cow. Normally cycling cows at Day 4 (early CL, n = 5) or Days 10--12 (mid CL, n = 5) of the estrous cycle (estrus = Day 0) were examined by transrectal color and pulsed Doppler ultrasonography to determine the blood flow area, the time-averaged maximum velocity (TAMXV), and the volume of the CL after an i.m. injection of a PGF(2 alpha) analogue. Ultrasonographic examinations were carried out just before PG injection (0 h) and then at 0.5, 1, 2, 4, 8, 12, 24, and 48 h after the injection. Blood samples were collected at each of these times for progesterone (P) determination. The ratio of the colored area to a sectional plane at the maximum diameter of the CL was used as a quantitative index of the changes in blood flow within the luteal tissue. Blood flow within the midcycle CL initially increased (P < 0.05) at 0.5-2 h, decreased at 4 h to the same levels observed at 0 h, and then further decreased to a lower level from 8 h (P < 0.05) to 48 h (P < 0.001). Plasma P concentrations decreased (P < 0.05) from 4.7 +/- 0.5 ng/ml (0 h) to 0.6 +/- 0.2 ng/ml (24 h). The TAMXV and CL volume decreased at 8 h (P < 0.05) and further decreased (P < 0.001) from 12 to 24 h after PG injection, indicating structural luteolysis. These changes were not detected in the early CL, in which luteolysis did not occur. In the early CL, the blood flow gradually increased in parallel with the CL volume, plasma P concentration, and TAMXV from Day 4 to Day 6. The present results indicate that PGF(2 alpha) induces an acute blood flow increase followed by a decrease in the midcycle CL but not in the early CL. This transitory increase may trigger the luteolytic cascade. The lack of intraluteal vascular response to PG injection in the early CL appears to be directly correlated with the ability to be resistant to PG.     

A basic study on universal design of auditory signals in automobiles

In this paper, the impression of various kinds of auditory signals currently used in automobiles and a comprehensive evaluation were measured by a semantic differential method. The desirable acoustic characteristic was examined for each type of auditory signal. Sharp sounds with dominant high-frequency components were not suitable for auditory signals in automobiles. This trend is expedient for the aged whose auditory sensitivity in the high frequency region is lower. When intermittent sounds were used, a longer OFF time was suitable. Generally, "dull (not sharp)" and "calm" sounds were appropriate for auditory signals. Furthermore, the comparison between the frequency spectrum of interior noise in automobiles and that of suitable sounds for various auditory signals indicates that the suitable sounds are not easily masked. The suitable auditory signals for various purposes is a good solution from the viewpoint of universal design.     

Deletion analysis of the superoxide dismutase (sodM) promoter from Aspergillus oryzae

The manganese superoxide dismutase gene (sodM) is very highly expressed in Aspergillus oryzae. To elucidate the basis for this high-level expression, deletion analysis of the promoter was undertaken using β-glucuronidase (GUS) as a reporter. Deletion of a 63-bp sequence from −200 to −138 in the 1,038-bp sodM promoter caused a drastic decrease in GUS activity. In addition, an electrophoretic gel mobility shift assay (EMSA) implicated a 30-bp element from −209 to −178 containing cis-element(s) in the high-level expression. The results of fine structure deletion analysis of this region were consistent with the EMSA results. To confirm these findings, we constructed enhanced sodM promoters by incorporating tandem repeats of this region, which resulted in an approximate twofold increase in expression relative to the native sodM promoter.