Characterization of sequential immune complexes in infective endocarditis by Western blot analysis

A patient with cutaneous vasculitis during infective endocarditis due to Lactobacillus casei was studied. Immune complexes (IC) were isolated from serum at the time of diagnosis and after 4 wk of therapy. Purification of IC used differential polyethylene glycol precipitation and competitive binding to staphylococcal protein A. In situ radioiodination of IC was performed, followed by SDS-polyacrylamide gel electrophoresis (PAGE). Anti-IC antisera were raised in rabbits by immunization with purified IC. IC were characterized by SDS-PAGE followed by electrophoretic transfer to nitrocellulose, incubation with antiserum and then with 125I protein A, and autoradiography. Although early and late IC differed quantitatively, there were no differentiating immunochemical features. Both IC contained a 60,000 dalton component that did not react with preimmune serum nor with anti-normal human serum. This component reacted with antiserum rendered specific for L. casei by affinity chromatography. The restricted antigen-antibody representation in IC contrasted with a wider panel of antibody activity in patient serum. The Western blot analysis proves to be an ideal method for the characterization of IC because of its sensitivity, dissociative capability, and preservation of immunoreactivity. IC isolated at a time removed from the original antigenic challenge may provide insight into the nature of the inciting antigen.     

Chemotactic macrophage subpopulations defined by macrophage chemotactic factors from delayed hypersensitivity reaction sites

The chemotactic specificity of ia-positive and -negative macrophages was studied by using three macrophage chemotactic factors (MCF), -a, -b, and -c, isolated from delayed hypersensitivity reaction (DHR) skin sites in guinea pigs. Listeria-elicited macrophages migrated toward MCF-a, -b, and -c. The chemotactic responses suggested responsive subpopulations to MCF. The electronic programmable individual cell sorter (EPICS) was used to separate macrophages with anti-la monoclonal antibodies. Ia-positive subpopulations responded to MCF-c, although they did not migrate toward MCF-a and -b. In contrast, Ia-negative subpopulations migrated toward MCF-a and -b, but not toward MCF-c. Furthermore, MCF-c attracted Ia-positive macrophages, whereas MCF-a and -b were Ia-negative in vitro; MCF did not induce Ia-negative macrophages to express surface Ia-antigens in vitro. MCF-c was able to produce massive Ia-positive macrophage accumulations when injected i.p., whereas MCF-a accumulated Ia-negative macrophages. The data suggest that MCF-a and -b, which mediate initial macrophage reactions, attract Ia-negative macrophages, and that MCF-c, which mediates predominant macrophage reactions, attract Ia-positive macrophages in the DHR.     

A Freeze-Fracture Electron Microscope Study of Trichomonas vaginalis Donné and Tritrichomonas foetus (Riedmüller)1

Two strains of Trichomonas vaginalis, JH162A, with low pathogenicity, and Balt 44, with high pathogenicity, as well as one highly pathogenic strain, KV-1, of Tritrichomonas foetus were studied by freeze-fracture electron microscopy. The protoplasmic faces (PFs) of the cell membranes of all three strains of both species had similar numbers of intramembranous particles (IMPs); however, the particles in the external faces (EFs) of these membranes were least abundant in Trichomonas vaginalis strain Balt 44 and most numerous in those of strain JH162A of this species. In Tritrichomonas foetus strain KV-1 the number of IMPs in the EF was close to but somewhat lower than that in the mild strain of the human urogenital trichomonad. In both species, the anterior, but not the recurrent, flagella had rosette-like formations, consisting of ～9 to 12 IMPs on both the PFs and EFs. The numbers and distribution of the rosettes appeared to vary among different flagella and in different areas of individual flagella of a single organism belonging to either species. The freeze-fracture electron micrographs provided a more complete understanding of the fine structure of undulating membranes of Trichomonadinae, as represented by Trichomonas vaginalis, and of Tritrichomonadinae (the Tritrichomonas augusta-type), as exemplified by Tritrichomonas foetus, than was gained from previous transmission and scanning electron microscope studies. Typically three longitudinal rows of IMPs on the PF of the recurrent flagellum of Trichomonas vaginalis were noted in the area of attachment of this flagellum to the undulating membrane. The functional aspects of the various structures and differences between certain organelles revealed in the two trichomonad species by the freeze-fracture method are discussed.     

The critical weed-free period in organically-grown winter wheat

Two experiments were conducted in central southern England between September 1994 and August 1996 to identify the critical weed-free period in organically grown winter wheat (Triticum aestivum, cv. Mercia). In competition with a mixed weed infestation of predominately Alopecurus myosuroides and Tripleurospermum inodorum it was found that wheat yield decreased as the duration of the weed-infested period increased and that the crop needed to be kept free of weeds from sowing in order to completely avoid any yield loss. Also, weeds emerging in the wheat crop (predominately T. inodorum) during the growing season had a significant and detrimental effect on yield. The existence of the critical period, therefore, depends on the imposition of an acceptable yield loss. If a 5% yield loss gives a marginal benefit compared with the cost of weed control, the critical period will begin at 506°C days after sowing (November) and end at 1023°C days after sowing (February). This information could be used by farmers to target mechanical weeding operations to control weeds at a time that will have maximum benefit to the crop.     

Impaired beta-cell regeneration in perinatally malnourished rats: a study with STZ.

We investigated the mechanisms implicated in beta-cell mass reduction observed during late fetal and early postnatal malnutrition in the rat. Beta-cell regeneration, including proliferation and neogenesis, was studied after neonatal beta-cell destruction by streptozotocin (STZ). STZ was injected at birth and maternal food restriction was continued until weaning. Beta-cell mass, proliferation, and islet number were quantified by morphometrical measurements on pancreatic sections in STZ-injected normal (C-STZ) and malnourished (R-STZ) rats, with noninjected C and R rats as controls. At day 4, only 20% of the beta cell-mass remained in C-STZ rats. It regenerated to 50% that of noninjected controls, mainly through active neogenesis, as shown by the entire recovery of islet number/cm(2), and also through moderately increased beta-cell proliferation. In contrast, beta-cell mass from R-STZ animals poorly regenerated, despite a dramatic increase of beta-cell proliferation, because islet number/cm(2) recovered insufficiently. In conclusion, perinatal malnutrition impairs neogenesis and the capacity of beta-cell regeneration by neogenesis but preserves beta-cell proliferation, which remains the elective choice to increase beta-cell mass. These results provide an explanation for the impaired capacity of malnourished animals to adapt their beta-cell mass during aging or pregnancy, which aggravate glucose tolerance.     

Energy metabolism and lipid peroxidation of human erythrocytes as a function of increased oxidative stress.

To study the influence of oxidative stress on energy metabolism and lipid peroxidation in erythrocytes, cells were incubated with increasing concentrations (0.5-10 mM) of hydrogen peroxide for 1 h at 37 degrees C and the main substances of energy metabolism (ATP, AMP, GTP and IMP) and one index of lipid peroxidation (malondialdehyde) were determined by HPLC on cell extracts. Using the same incubation conditions, the activity of AMP-deaminase was also determined. Under nonhaemolysing conditions (at up to 4 mM H2O2), oxidative stress produced, starting from 1 mM H2O2, progressive ATP depletion and a net decrease in the intracellular sum of adenine nucleotides (ATP + ADP + AMP), which were not paralleled by AMP formation. Concomitantly, the IMP level increased by up to 20-fold with respect to the value determined in control erythrocytes, when cells were challenged with the highest nonhaemolysing H2O2 concentration (4 mM). Efflux of inosine, hypoxanthine, xanthine and uric acid towards the extracellular medium was observed. The metabolic imbalance of erythrocytes following oxidative stress was due to a dramatic and unexpected activation of AMP-deaminase (a twofold increase of activity with respect to controls) that was already evident at the lowest dose of H2O2 used; this enzymatic activity increased with increasing H2O2 in the medium, and reached its maximum at 4 mM H2O2-treated erythrocytes (10-fold higher activity than controls). Generation of malondialdehyde was strictly related to the dose of H2O2, being detectable at the lowest H2O2 concentration and increasing without appreciable haemolysis up to 4 mM H2O2. Besides demonstrating a close relationship between lipid peroxidation and haemolysis, these data suggest that glycolytic enzymes are moderately affected by oxygen radical action and strongly indicate, in the change of AMP-deaminase activity, a highly sensitive enzymatic site responsible for a profound modification of erythrocyte energy metabolism during oxidative stress.     

Calcium-channel blockers and cognitive function in elderly people: results from the Canadian Study of Health and Aging

BACKGROUND: Concern has been raised about the potential for adverse cognitive effects associated with the use of calcium-channel blockers (CCBs) in older people. This study was undertaken to examine prospectively the association between the use of these and other antihypertensive drugs and cognitive function. METHODS: The authors examined data from the Canadian Study of Health and Aging (CSHA), a population-based, prospective 5-year investigation of the epidemiology of dementia and other health problems in Canadians 65 years of age and older. The risk of cognitive decline, as indicated by a decline in performance on the Modified Mini-Mental State (3MS) examination over the 5-year period, was assessed in relation to the use of antihypertensive and diuretic drugs by 205 subjects with a history of hypertension and no evidence of dementia at baseline. RESULTS: The proportion of subjects whose cognitive performance declined over the study period was significantly higher in the group using CCBs than in the group using other antihypertensive agents (75% v. 59%). The adjusted odds ratio (OR) for a significant decline in cognitive performance (defined as a decrease in 3MS score of 10 points or more) was 2.28 (95% confidence interval [CI] 1.12-4.66) for subjects using CCBs. The adjusted ORs (and 95% CIs) for cognitive decline in subjects using selected antihypertensive agents or diuretics relative to those exposed to beta-blockers were as follows: angiotensin-converting-enzyme inhibitor, OR 1.36 (95% CI 0.41-4.55); diuretic or other antihypertensive drug, OR 1.45 (95% CI 0.51-4.14); dihydropyridine CCB (nifedipine), OR 1.94 (95% CI 0.52-7.27) and non-dihydropyridine CCB (diltiazem or verapamil), OR 3.72 (95% CI 1.22-11.36). INTERPRETATION: Older people taking CCBs were significantly more likely than those using other agents to experience cognitive decline. These findings are consistent with the results of previous cross-sectional research and emphasize the need for further trials to examine the associations between CCB use, blood pressure and cognitive impairment in elderly patients.