Pseudoaneurysm of the aortic arch associated with cervical spondylodiscitis.

STUDY DESIGN: A case report. OBJECTIVES: To report and discuss a case of pseudoaneurysm of the aortic arch presenting as hemoptysis following a cervical spondylodiscitis. The pseudoaneurysm was remote and any direct extension of the abscess was not observed from the cervical lesion. SETTING: Hamamatsu Medical Center. CASE REPORT: A 73-year-old male being treated with antibiotics for a cervical spodylodiscitis deteriorated tetraplegia. Following a posterior decompression of the cervical spine and subsequent neurological recovery, hemoptysis occurred and a pseudoaneurysm of the aortic arch was identified. Emergent vascular graftings combined with dèbridement of the pseudoaneurysm and the infected cervical intervertebral disc were performed. The patient recovered gradually and the cervical spondylodiscitis disappeared. CONCLUSIONS: The septicemia originating from the remote cervical spondylodiscitis was thought to contribute to this pseudoaneurysm. Attention should be paid to the systemic septicemia as well as the focal spinal infection. As for cervical spondylodiscitis, posterior decompression without drainage cannot be recommended as the initial treatment.

Mechanisms of nitroso compound-induced inhibition of superoxide generation in neutrophils: fluorescence quenching of perylene by nitroso-compounds in the membrane fractions of neutrophils.

To investigate the mechanism of nitroso compound-induced inhibition of the respiratory burst in neutrophils, we studied fluorescence quenching of perylene by nitroso-compounds in the membrane fractions of neutrophils at 17, 27, and 37 degrees C and the reagent-induced inhibition of superoxide generation at 28 and 37 degrees C. With increasing temperature, the quenching of perylene fluorescence and inhibition of superoxide generation by nitrosobenzene (NB) were both diminished, while those by 2-nitrosotoluene (NT) were both enhanced. The temperature dependence of the inhibition constants and the quenching constants indicates that the binding of NB is exothermic (deltaH= -27 kJ/mol for inhibition and deltaH= -29 kJ/mol for quenching) and essentially enthalpy-driven. On the other hand, that of NT is endothermic (deltaH= +16 kJ/mol for inhibition and quenching) and essentially entropy-driven. Quenching studies of perylene fluorescence in synthetic vesicles made of endogenous polar lipids of neutrophils showed that the enthalpy changes of NB- and NT-binding with perylene in lipids were similar to each other. Moreover, their values were in good agreement with that of NT, but not of NB, in the membrane fractions, an assembly of proteins and lipids, of neutrophils. These results suggest that NB inhibits the activity by binding to proteins in the membrane, whereas inhibition by NT occurs through hydrophobic interaction with lipids and/or proteins.

Effect of aromatic nitroso-compounds on superoxide-generating activity in neutrophils.

Aromatic nitroso-compounds such as nitrosobenzene inhibited the respiratory burst of intact neutrophils induced by various stimulants, including phorbol 12-myristate 13-acetate and a chemotactic peptide. The compounds also inhibited NADPH-dependent oxygen consumption by cell-free preparations of neutrophils. This indicates that nitroso-compounds act directly on the NADPH-oxidase system. The inhibitory effects induced by several nitroso-compounds, 2-nitrosotoluene, nitrosobenzene, 4-nitrosophenol, and 1-nitrosopyrrolidine, were examined and their inhibition constants, the concentrations causing 50% reduction of oxygen consumption, were found to be 0.043, 0.173, 0.672, and 32.1 mM, respectively. These values correlated well with the hydrophobicity of the compounds: a more hydrophobic compound was a more potent inhibitor against NADPH oxidase, suggesting that the oxidase has a hydrophobic site(s) for interaction with the inhibitors.

Superoxide production by cytochrome b558 purified from neutrophils in a reconstituted system with an exogenous reductase.

Cytochrome b558, which is considered to be an essential component of the phagocytic superoxide (O2-)-generating system, was highly purified from porcine neutrophils. The isolated cytochrome was resolved into two polypeptides with molecular masses of 60-90 and 19 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. For enzymatic reduction of purified cytochrome b558, we utilized hepatic NADPH-cytochrome P450 reductase purified from rat liver microsomes. More than 80% of the cytochrome was reduced by incubation with the reductase and NADPH under the anaerobic condition, and was quickly reoxidized by the air. As indicated by measurement of oxygen consumption, the purified cytochrome catalytically reduced oxygen at a rate equal to approximately 30% of the activity of the phorbol myristate acetate-activated cells on the basis of cytochrome b558 content. Electron paramagnetic resonance study with a spin trapping agent 5, 5-dimethyl-1-pyrroline-1-oxide demonstrated that O2- is the exclusive primary product in the reduction of oxygen by the cytochrome. This gives direct evidence that cytochrome b558 functions as the terminal oxidizing enzyme in the O2- -generating system of neutrophils. This also establishes a new functional class of heme proteins that catalyzes one-electron reduction of molecular oxygen.

Effect of a light-induced pH gradient on purple-to-blue and purple-to-red transitions of bacteriorhodopsin.

Bacteriorhodopsin-containing vesicles that were able to alkalize the extravesicular medium by greater than 1.5 pH units under illumination, i.e., inside-out vesicles, were reconstituted by reverse-phase evaporation with Halobacterium halobium polar lipids or exogenous phospholipids. Acid titration of a dark-adapted sample was accompanied by a color change from purple to blue (pKa = 2.5-4.5 in 0.15 M K2SO4), and alkali titration resulted in the formation of a red species absorbing maximally at 480 nm (pKa = 7 to greater than 9), the pKa values and the extents of these color changes being dependent on the nature of lipid. When a vesicle suspension at neutral or weakly acidic pH was irradiated by continuous light so that a large pH gradient was generated across the membrane, either a purple-to-blue or a purple-to-red transition took place. The light-induced purple-to-red transition was significant in an unbuffered vesicle suspension and correlated with the pH change in the extravesicular medium. The result suggests that the purple-to-red transition is driven from the extravesicular side, i.e., from the C-terminal membrane surface. In the presence of buffer molecules outside, the dominant color change induced in the light was the purple-to-blue transition, which seemed to be due to a large decrease in the intravesicular pH. But an apparently inconsistent result was obtained when the extravesicular medium was acidified by a HCl pulse, which was accompanied by a rapid color change to blue. We arrived at the following explanation: The two bR isomers, one containing all-trans-retinal and the other 13-cis-retinal, respond differently to pH changes in the extravesicular and the intravesicular medium. In this relation, full light adaptation was not achieved when the light-induced purple-to-blue transition was significant; i.e., only the 13-cis isomer is likely to respond to a pH change at the N-terminal membrane surface.

Effect of partial delipidation of purple membrane on the photodynamics of bacteriorhodopsin.

The effect of lipid-protein interaction on the photodynamics of bacteriorhodopsin (bR) was investigated by using partially delipidated purple membrane (pm). When pm was incubated with a mild detergent, Tween 20, the two major lipid components of pm, phospholipids and glycolipids, were released in different ways: the amount of phospholipids released was proportional to the logarithm of the incubation time; the release of glycolipids became noticeable after the release of approximately 2 phospholipids/bR, but soon leveled off at approximately 50% of the initial content. It was found that the thermal decay of the photocycle intermediate N560 was inhibited by the removal of less than 2 phospholipids per bR. This inhibition was partly explained by an increase in the local pH near the membrane surface. More significant changes in the bR photoreactions were observed when greater than 2 phospholipids/bR were removed: (1) the extent of light adaptation became much smaller, and this reduction correlated with the release of glycolipids; (2) N560 became difficult to detect; (3) the M412 intermediate, which is characterized by a pH-insensitive lifetime, was replaced by a long-lived M-like photoproduct with a pH-sensitive lifetime. The heavy delipidation apparently altered the mechanism by which the deprotonated Schiff base receives a proton. An important conformational change in the protein moiety is suggested to take place during the M412 state, this conformational change being inhibited in the rigid lipid environment.

Bacteriorhodopsin photoreaction: identification of a long-lived intermediate N (P,R350) at high pH and its M-like photoproduct.

An alkaline suspension of light-adapted purple membrane exposed to continuous light showed a large absorption depletion at 580 nm and a small increase around 350 nm. We attribute this absorption change to an efficient photoconversion of bR570 into a photoproduct N (P,R350), which has a major absorption maximum between 550 and 560 nm but has lower absorbance than bR570. N was barely detectable at low pH, low ionic strength, and physiological temperature. However, when the thermal relaxation of N to bR570 was inhibited by increasing pH, increasing ionic strength, and decreasing temperature, its relaxation time could be as long as 10 s at room temperature. N is also photoactive; when it is present in significant concentrations, e.g., accumulated by background light, the flash-induced absorption changes of purple membrane suspensions were affected. Double-excitation experiments showed an M-like photoproduct of N,NM, with an absorption maximum near 410 nm and a much longer lifetime than M412. It may be in equilibrium with an L-like precursor NL. We suggest that N occurs after M412 in the photoreaction cycle and that its photoproduct NM decays into bR570. Thus, at high pH and high light intensity, the overall photoreaction of bR may be approximated by the two-photon cycle bR570----M412----N----(NL----NM)----bR570, whereas at neutral pH and low light intensity it can be described by the one-photon cycle bR570----M412----N----O640----bR570.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteriorhodopsin is a powerful light-driven proton pump.

The activity of bacteriorhodopsin was investigated with Halobacterium halobium cell envelopes, which lack cytoplasmic constituents. It was found that the physiological concentration of magnesium ion greatly enhanced the light-induced pH change; under optimal conditions, the pH change of the external medium was as large as 3.5 pH units, even though the volume fraction of the envelope vesicles was as low as 0.01. This pH change is about three times larger than the largest change reported thus far. This same effect was observed with transition metal ions, but not with other alkaline divalent cations. That is, divalent cations that formed hydroxides below pH 10 were effective in enhancing the light-induced pH change. This result suggests that some divalent cations acted as buffers against a large increase in the internal pH, and that the internal pH was an important factor in determining the activity of bacteriorhodopsin. It was also shown that a high level of the proton-pump activity was maintained in a wide range of external pHs, at least between 4.5 and 9.4.

Preference of oxygenation between alpha and beta subunits of haemoglobin. Results of multidimensional spectroscopic observation.

The distribution of oxygen between the subunits of haemoglobin was studied spectrophotometrically. The difficulty in discriminating the spectral changes upon oxygen binding to the alpha or beta subunit can be surmounted by means of multidimensional spectroscopic observations and a correlation analysis of the data. M-type abnormal haemoglobins are used as a control against normal haemoglobin because only one type of its subunits can bind oxygen. A multidimensional spectroscopic measuring system, which has been developed in our laboratory, makes it possible to carry out simultaneous and continuous acquisition of a set of spectroscopic data at several wavelengths on one sample solution during the course of increasing or decreasing the partial pressure of oxygen. The data-storing function of a magnetic disk memory provides enough precision for a rigorous investigation of the correlation of oxygen equilibrium curves measured at several wavelengths. No chemical modification to enhance the spectral difference between subunits is necessary. In conclusion, by detecting slight differences between the oxygenation-sensitive bands of alpha and beta subunits, the beta subunits are found to have a higher affinity for oxygen than the alpha subunits.