Bordetella bronchiseptica dermonecrotizing toxin induces reorganization of actin stress fibers through deamidation of Gln-63 of the GTP-binding protein Rho

Bordetella dermonecrotizing toxin causes assembly of actin stress fibers and focal adhesions in some cultured cells and induces mobility shifts of the small GTP-binding protein Rho on electrophoresis. We attempted to clarify the molecular basis of the toxin action on Rho. Analysis of the amino acid sequence of toxin-treated RhoA revealed the deamidation of Gln-63 to Glu. The substitution of Glu for Gln-63 of RhoA by site-directed mutagenesis caused a mobility shift on electrophoresis, which was indistinguishable from that of the toxin-treated RhoA. Neither mutant RhoA-bearing Glu-63 nor toxin-treated RhoA significantly differed from untreated wild type RhoA in guanosine 5'-[gamma-thio]triphosphate binding activity but both showed a 10-fold reduction in GTP hydrolysis activity relative to untreated RhoA. C3H10T1/2 cells transfected with cDNA of the mutant RhoA bearing Glu-63 showed extensive formation of actin stress fibers similar to the toxin-treated cells. These results indicate that the toxin catalyzes deamidation of Gln-63 of Rho and renders it constitutively active, leading to formation of actin stress fibers.     

Synergistic immunosuppression caused by high-dose methylprednisolone and cardiopulmonary bypass

BACKGROUND: Steroid use during cardiac operations may reduce the risk of postperfusion lung syndrome, but both cardiopulmonary bypass and steroids are immunosuppressive. The synergistic effects of the bypass and steroids on patients' immunologic activities, hemodynamics, and metabolisms during and after heart operations have not been clarified systematically. METHODS: Twenty-four patients undergoing valve replacement were studied in a randomized, double-blind trial. Twelve of these patients (S group) received bolus methylprednisolone, 20 mg/kg body weight, and the remaining 12 patients (C group) received a placebo intravenously before and after bypass. Blood cell count, C-reactive protein, lymphocyte surface markers (CD3, CD4, CD8, CD16, and CD20), phytohemagglutinin response, interleukin-2 production, and natural killer cell activity were examined on admission through day 7. Cardiac output, blood gas, electrolyte, lactate, and serum glucose levels were examined perioperatively. RESULTS: The peak white blood cell count in the S group was higher than that in the C group (analysis of variance: p [group] = 0.0436). The peak C-reactive protein level was higher in the C group than in the S group (p [group] < 0.0001). From the analysis of the surface markers, the steroid increased the natural killer cells before and soon after bypass (p [group] = 0.0117), and later tended to increase the CD4+ T and B cells during the postoperative recovery period. The phytohemagglutinin response in both groups decreased after bypass (p [time] < 0.0001), but the steroid caused exaggerated decreases before (p < 0.01 by Student's t test) and soon after (p < 0.001) bypass in the S group (analysis of variance: p [group] = 0.0127). The interleukin-2 production was suppressed by bypass alone after the bypass in the C group, but was further suppressed by the steroid before and after bypass in the S group (p [group] = 0.0446). The cardiac index, water balance, electrolytes, arterial oxygen tension, and timing of extubation were not different between the groups. In contrast, the glucose (p [group] = 0.0486) and lactate (p [group] = 0.0525) levels were higher in the S group than those in the C group. CONCLUSIONS: T-cell functions are synergistically suppressed by cardiopulmonary bypass and high-dose methylprednisolone in heart operations. The hemodynamic benefits of the steroid are negligible, whereas glucose tolerance is worsened by the steroid during bypass.     

Inhibition of azoxymethane-initiated colon tumor by bovine lactoferrin administration in F344 rats

The influence of bovine lactoferrin (bLF) on colon carcinogenesis was investigated in male F344 rats treated with azoxymethane (AOM). Following three weekly injections of AOM, the animals received 2 or 0.2% bLF for 36 weeks. No effects indicative of toxicity were noted, but significant reduction in both the incidence and number of adenocarcinomas of the large intestine was observed with both doses. Thus, the incidences of adenocarcinomas in the groups receiving 2% and 0.2% bLF were 15% and 25%, respectively, in contrast to the 57.5% control value (P < 0.01 and P < 0.05, respectively). The results indicate that bLF might find application for chemoprevention of colon cancer.     

Two Japanese wildcats, the Tsushima cat and the Iriomote cat, show the same mitochondrial DNA lineage as the leopard cat Felis bengalensis

We previously revealed, based on mitochondrial DNA sequence analysis, that the Iriomote cat is very closely related to the leopard cat Felis bengalensis, which is widespread in Asia [24]. In this study, in order to understand the phylogenetic status of the Tsushima cat which is the other wildcat in Japan, partial sequences (402 bases) of the mitochondrial cytochrome b region were determined and compared with those of the Iriomote cat and other feline species. The phylogenetic tree of the cytochrome b sequences indicated that the Tsushima cat and the Iriomote cat have the same mitochondrial DNA lineage as the leopard cat. One or two transitional substitutions were observed among the two Japanese wildcats and the leopard cat. The divergence time (approximately 100,000 years ago) of the Tsushima cat and the leopard cat, estimated by sequence data, was in concordance with the formation date of the Tsushima Island. These results suggest that genetic drift after geographic isolation has brought fixation of some genetic and morphological characters to the Tsushima cat and the Iriomote cat, while these two Japanese wildcats are still genetically close to the continental leopard cat. Considering morphological differences and molecular phylogeny, it is reasonable for the two Japanese wildcats to be classified as two subspecies of F. bengalensis.     

Swelling and mechanical properties of polyvinylalcohol hydrogels

Summary The swelling and mechanical properties of Poly(vinylalcohol) hydrogels were examined. It was found that the degree of swelling of PVA hydrogels depends on annealing temperature, but is almost independent of the initial polymer concentration. Mechanical properties of the hydrogels were also influenced by the degree of swelling. A shoulder was observed in double-logarithmic plots of stress vs. strain for the hydrogels, and became clearer as annealing temperature increased. This shoulder was closely related to the breakdown of the microcrystalline domains acting as crosslinks. Also, the shape of stress-strain curves plotted double-logarithmically for the hydrogels changed with the extension rate.     

An investigation of voltage instability problems

The authors investigate the voltage stability problem based on singular perturbation theory. Possible voltage instability patterns are classified into four types (I, II-1, II-2S, and II-2D) according to the mechanism causing voltage instability. Several features of each type of instability are studied as well as their analysis methods. Voltage instability tends to begin with type I and then leads to one of the remaining types. The load flow Jacobian can be an effective index to approximately assess type I and II-2S instabilities, while types II-1 and II-2D require direct nonlinear analyses and eigenvalue analyses, respectively. The validity of the classification proposed has been verified through numerical simulations and theoretical analyses which take into account dynamic characteristics of generating units, loads, and tap-changing transformers     

Enzymatic determination of butane-2,3-diol in wines.

Butanediol is first oxidised to acetoin by butane-2,3-diol dehydrogenase from a strain of Sarcina, in the presence of NAD⁺ and an excess amount of 2,6-dichloro-phenol-indophenol at an alkaline pH. Then the acetoin formed is reduced back to butane-2,3-diol by the same enzyme in an acidic condition, and NADH thereby oxidised is measured spectrophotometrically. Direct measurement of NADH formation in the first reaction, oxidation of butane-2,3-diol, has not yet been successful. Colorimetric measurement of the reduction of 2,6-dichlorophenol-indophenol was also unsuccessful. Ethanol in sample solutions should have been removed by evaporation before the enzymatic oxidation. Presence of acetoin caused no difficulty in the oxidation of butane-2,3-diol, but made it necessary to correct the measured values for the original acetoin contents. The isomers of butane-2,3-diol present in wines have been reported to be D (-) and meso, on both of which the butane-2,3-diol dehydrogenase from Sarcina appears to be able to act. The butane-2,3-diol contents of some Japanese commercial wines were determined by this enzymatic method. Concentrations ranged from 324 to 768 mg/litre.     

Serial changes of plasma plasminogen activator inhibitor activity in acute myocardial infarction: difference between thrombolytic therapy and direct coronary angioplasty

The fibrinolytic system is impaired in patients with acute myocardial infarction (AMI). The primary regulatory element of fibrinolytic activity is plasminogen activator inhibitor (PAI). There are no reports, however, on the serial changes of PAI activity after thrombolysis or coronary angioplasty in patients with AMI undergoing emergency coronary angiography. This study was designed to examine the difference in the change of fibrinolytic activity between patients with AMI who underwent thrombolytic therapy with recombinant tissue-plasminogen activator (rTPA) and those who underwent direct percutaneous coronary angioplasty (PTCA). We measured the serial changes of PAI activity and tissue plasminogen activator (TPA) antigen after rTPA therapy or direct PTCA. Twenty-two patients received emergency coronary angiography and were treated with rTPA intravenously. Twenty patients underwent direct PTCA. Plasma PAI activity levels were increased on admission and further increased within 24 hours in patients treated with rTPA and in those treated with direct PTCA. In the thrombolysis group, there were two peaks in plasma PAI activity levels (IU/ml) at 4 hours (27.0 +/- 2.9) and at 16 hours (25.6 +/- 2.5) after the initiation of rTPA infusion. However, in the direct PTCA group, there was one peak of PAI activity (IU/ml) at 16 hours (23.9 +/- 2.7) after the initiation of direct PTCA. In conclusion, the PAI activity has two peaks in the thrombolysis group and one peak in the direct PTCA group.     

Polymerization of 1-butyne and isopropylacetylene by transition metal catalysts and geometric structure of polymers

Polymerization of 1-butyne and isopropylacetylene was studied using MoCl₅, WCl₆, and two Ziegler catalysts [Fe(acac)₃-Et₃Al, Ti(On-Bu)₄-Et₃Al]. 1-Butyne was polymerized in high yields with WCl₆ and Fe(acac)₃-Et₃Al to give a yellow, air-sensitive polymer. The cis content of poly(1-butyne), evaluated by ¹³C n.m.r., was about 80% irrespective of polymerization conditions. Isopropylacetylene was polymerized well by any of MoCl₅, WCl₆, and Fe(acac)₃-Et₃Al; the polymer formed was a light yellow air-sensitive powder. The cis content of poly(isopropylacetylene) varied from 65% to 90% according to polymerization conditions. Substituent effects on polymerization and polymer structure are discussed.