Complete amino acid sequence of Proteus mirabilis PR catalase. Occurrence of a methionine sulfone in the close proximity of the active site

The catalase of Proteus mirabilis PR, a peroxide-resistant (PR) mutant of Proteus mirabilis, binds strongly NADPH, which is a unique property among known bacterial catalases. The enzyme subunit consists of 484 amino acid residues for a mass of 55,647 daltons. The complete amino acid sequence was resolved through the combination of protein sequencing, mass spectrometry, and nucleotide sequencing of a PCR fragment. The sequence obtained was compared with that of other known catalases. Amino acids of the active site are all conserved as well as essential residues involved in NADPH binding. Among the amino acids interacting with the heme, a methionine sulfone was found at position 53, in place of a valine in most other catalases. The origin of oxidation of this methionine is unknown, but the presence of this modification could change iron accessibility by large substrates or inhibitors. This posttranslational modification was also demonstrated in the wild-type P. mirabilis catalase.     

Dissociation of airway responsiveness and bronchoalveolar lavage (BAL) cell composition in sensitized guinea-pigs after daily inhalation of ovalbumin

The association between inflammatory cell influx, cell activation status and change of airway responsiveness to acetylcholine (ACh) after daily inhalation of ovalbumin (OA) in sensitized guinea-pigs was investigated. Starting 3 weeks after sensitization (OA at 50 mg/kg s.c.+i.p.) guinea-pigs were exposed daily to 2% OA (10 min; under cover of 0.5 mg/kg mepyramine i.p. 15 min before OA) for 2 weeks. Concentration-response curves (CRCs) for inhaled ACh were performed 24 h after the last OA-challenge and 24 h after another single OA-inhalation 1 week later. CRCs for inhaled ACh were neither affected 24 h after the last OA challenge (daily for two weeks) nor 24 h after another OA-inhalation one week later. In contrast, bronchoalveolar lavage (BAL) from repeatedly OA- sensitized/-challenged guinea-pigs immediately after the last CRC showed a significant increase of total cell count by about tenfold and increases in eosinophils by about 20-fold, neutrophils by 30-fold, macrophages by about fivefold and lymphocytes by about tenfold (P < 0.05, multiple Wilcoxon-test). In contrast, markers of cell activation (EPO, MPO) were significantly decreased (P < 0.05). Methylprednisolone almost completely prevented these changes in increased cell numbers and decreased cell activation (vs OA contr., P < 0.05). The lack of increased airway hyperresponsiveness despite a massive inflammatory cell influx suggests other factors controlling airway responsiveness than inflammation.     

Determination of free acetaldehyde in total blood for investigating the effect of aspartate on metabolism of alcohol in mice

The cerebral cortex of anaesthetised 2- to 12-day-old rats was superfused with artificial cerebrospinal fluid containing 100 mM acetate substituted for chloride to condition the brain for spreading depression (SD). After such superfusion, the earliest SD-like events were found at day 9 and full blown SD at day 10, whereas in the unconditioned brain the first SD occurred between days 12 and 15. Acetate conditioning of the cerebral cortex may be used to unmask neuronal and glial properties that are hidden in early stages of development.     

Taking the V out of genes V environment

Tumor necrosis factor-alpha (TNF-alpha) is thought to be an important mediator in the pathogenesis of neonatal septicemia. To assess the role of TNF-alpha in neonatal sepsis, serum levels of TNF-alpha were measured in a group of neonates with septicemia and compared with the levels of gestational-postnatal, age-matched healthy controls. The relationships between severity of infection, the nature of causative microorganisms, and TNF-alpha levels were also investigated in this prospective study. A total of 49 infants (25 full-term, 24 preterm) with proven sepsis and 40 healthy infants (20 full-term, 20 preterm) were included. Serum TNF-alpha levels were measured using the TNF-alpha immunoradiometric assay. The median level of TNF-alpha was found to be significantly higher in infants suffering from sepsis (154 pg/mL) particularly in those with septic shock (242.5 pg/mL) as compared to healthy controls (61.5 pg/mL) (p < 0.001). No correlation was found between TNF-alpha and postnatal ages, gestational ages or birth weights of the infants. TNF-alpha levels were not different in surviving and terminal neonates. Although serum, TNF-alpha levels were found to be slightly higher in gram-negative septicemia, the difference was not significant. These findings suggest that TNF-alpha plays an important role in the pathophysiology of neonatal septicemia, but its importance as a prognostic factor is not yet clear.     

Leprosy patients with lepromatous disease recognize cross-reactive T cell epitopes in the Mycobacterium leprae 10-kD antigen

BACKGROUND: Cough associated with gastroesophageal reflux (GER) may originate in extrathoracic airway receptors made hypersensitive by acid-induced mucosal injury. OBJECTIVE: We investigated the role of laryngeal disease and dysfunction in the pathogenesis of GER-associated cough in nonasthmatic patients. METHODS: Seven patients with GER-associated cough were compared with 7 patients with GER but no cough. The patients underwent fiberoptic endoscopy for assessment of laryngitis and esophagitis (expressed by scores); esophageal manometry; 24-hour pH monitoring; lung function tests; and histamine inhalation challenge with assessment of bronchial threshold (concentration provoking 10% fall in FEV1 [PC10]), extrathoracic airway threshold (concentration provoking 25% fall in the maximal midinspiratory flow [PC25MIF50]), and cough threshold (concentration provoking 5 or more coughs PCcough). The patients were reevaluated after 3 months of medical treatment for GER. RESULTS: Patients with cough, compared with those without cough, had significantly higher laryngitis scores (P = .002), lower esophageal sphincter pressures, longer time with pH below 4 (P = .003), greater number of episodes of reflux longer than 5 minutes (P = .016), longer esophageal clearance time (P = .048), and significantly lower PC25MIF50 (P = .005) and PCcough (P = .008) values. Laryngitis score was significantly inversely related to either PCcough (P < .001) or PC25MIF50 (P <.01) but not to PC10. Laryngitis score, PC25MIF50, and PCcough were all closely related to GER severity. After GER treatment, laryngitis, PC25MIF50, and PCcough were all significantly improved. CONCLUSIONS: These findings suggest that GER-associated cough is strongly associated with laryngeal disease and dysfunction consequent to acid reflux injury in nonasthmatic patients.     

Experimental infection of domestic sheep with culture-derived Leishmania donovani promastigotes

Domestic sheep were intradermally inoculated with culture-derived stationary phase Leishmania donovani promastigotes. Sampling of site of inoculation, liver and spleen for 244 days showed that this parasite can stay alive in the skin for up to 28 days post-inoculation. Apart from pyrexia that was evident in all the animals for 42 days, no other symptoms of kala-azar were seen. No parasites were recovered from the visceral organs throughout the sampling period, suggesting that sheep are not susceptible to infection with L. donovani. It is therefore unlikely that sheep can be synanthropic reservoirs for this parasite.     

Insulin activates the hepatitis B virus X gene through the activating protein-1 binding site in HepG2 cells

OBJECTIVES: To study the turnaround time (TAT) for rendering diagnoses on routine biopsy specimens, to examine pathology practice variables that influence TAT, and to assess the level of surgeons' satisfaction with biopsy TAT. DESIGN: Over a 3-month period, voluntary participants in the College of American Pathologists Q-Probes laboratory quality improvement program prospectively collected TAT data on up to 20 biopsy specimens performed on elective surgical cases, completed questionnaires profiling their institution's practice characteristics, and had surgeons complete questionnaires indicating their satisfaction with biopsy report TAT. SETTING AND PARTICIPANTS: One hundred fifty-seven private and public small hospitals located in 43 American states (n = 153), Canada (n = 1), and Australia (n = 3). MAIN OUTCOME MEASURES: The routine surgical biopsy report TATs for 2 testing intervals, each commencing when surgeons acquired the biopsy specimens. One interval concluded when pathologists signed off the biopsy diagnoses, and the other concluded when surgeons received the hard-copy reports. RESULTS: Pathologists signed off 85.9% of 5384 biopsy diagnoses by the second working day, and surgeons received 88.3% of the hard-copy reports by the fourth working day. In 90% of hospitals participating in this study, pathologists signed off half their biopsy diagnoses between the second and third postcollection days, and 90% of surgeons received half their final hard-copy reports by the fourth postcollection day. Institutional practice variables associated with fewer sign-off and/or hard-copy receipt TATs exceeding the institutional 90th percentile performance benchmarks included yearly surgical caseloads greater than 2000 cases per full-time equivalent pathologist, provision of pathology support services on site, and accreditation of the hospital by the Joint Commission on Accreditation of Healthcare Organizations and of the laboratory by the College of American Pathologists. Most (96.4%) surgeons indicated that they were satisfied with hard-copy TATs and that they believed most (98.1%) of the hard-copy TATs had no effect on the lengths of their patients' hospital stays. CONCLUSIONS: Pathologists are capable of signing off most routine biopsy diagnoses within 2 working days and delivering the final hard-copy reports to surgeons within 4 working days (both intervals measured from the time that surgeons collect biopsy specimens). Most surgeons report they are satisfied with this level of performance.