Positional and species analysis of membrane phospholipids extracted from goldfish adapted to different environmental temperatures.

1. The proportion of ethanolamine phosphoglycerides in microsomal fractions of goldfish intestine increases at low environmental temperatures. The fatty acyl composition also changes, the proportion of C22:6 and C20:4 fatty acids increasing in positions 1 and 2 and position 2 respectively. The proportion of C16:0 and C18:0 fatty acids falls in position 1 and there is an apparent switch of C18:1 and C20:1 fatty acids from position 2 to position 1. 2. The proportion of choline phosphoglycerides does not depend on the previous environmental temperature of the fish. Temperature-dependent changes in fatty acyl composition in positions 1 and 2 take place in a way similar to that described for ethanolamine phosphoglycerides, but in this case C22:6 substitution is confined to position 2. 3. Choline phosphoglycerides have been further separated into 7 different molecular species. The amounts of species 3 to 7 increase and the amount of species 2 decreases at low adaptation temperature. These changes only account for part of total change in fatty acyl composition. The remaining changes occur by chain substitution within species. 4. Present results show temperature adaptation to be highly complex, involving both quantitative and qualitative changes in different phospholipids. The possible physiological significance of these changes are discussed together with the effects these changes might have on cholesterol-phospholipid interactions.

Phosphatidylserine-dependent adhesion of T cells to endothelial cells.

Phosphatidylserine (PS) was exposed at the surface of human umbilical vein endothelial cells (HUVECs) and cultured cell lines by agonists that increase cytosolic Ca(2+), and factors governing the adhesion of T cells to the treated cells were investigated. Thrombin, ionophore A23187 and the Ca(2+)-ATPase inhibitor 2, 5-di-tert-butyl-1,4-benzohydroquinone each induced a PS-dependent adhesion of Jurkat T cells. A23187, which was the most effective agonist in releasing PS-bearing microvesicles, was the least effective in inducing the PS-dependent adhesion of Jurkat cells. Treatment of ECV304 and EA.hy926 cells with EGTA, followed by a return to normal medium, resulted in an influx of Ca(2+) and an increase in adhering Jurkat cells. Oxidised low-density lipoprotein induced a procoagulant response in cultured ECV304 cells and increased the number of adhering Jurkat cells, but adhesion was not inhibited by pretreating ECV304 cells with annexin V. PS was not significantly exposed on untreated Jurkat cells, as determined by flow cytometry with annexin V-FITC. However, after adhesion to thrombin-treated ECV304 cells for 10 min followed by detachment in 1 mM EDTA, there was a marked exposure of PS on the Jurkat cells. Binding of annexin V-FITC to the detached cells was inhibited by pretreating them with unlabelled annexin V. Contact with thrombin-treated ECV304 cells thus induced the exposure of PS on Jurkat cells and, as Jurkat cells were unable to adhere to thrombin-treated ECV304 cells in the presence of EGTA, the adhesion of the two cell types may involve a Ca(2+) bridge between PS on both cell surfaces. The number of T cells from normal, human peripheral blood that adhered to ECV304 cells was not increased by treating the latter with thrombin. However, findings made with several T cell lines were generally, but not completely, consistent with the possibility that adhesion to surface PS on endothelial cells may be a feature of T cells that express both CD4(+) and CD8(+) antigens. Possible implications for PS-dependent adhesion of T cells to endothelial cells in metastasis, and early in atherogenesis, are discussed.

A light-scattering assay for lymphocyte shape and its application to T and B lymphocyte responses to cultured high-walled endothelial cells.

Lymphocyte shape changes are detectable as changes in the forward light scatter (FLS) profile of a Becton-Dickinson FACStar flow cytometer, with polar cells giving lower values of FLS. The FACScan cell analyser does not detect these changes. Freshly isolated lymph node lymphocytes are round, but when stimulated, for example by incubation with cultured high endothelial cells (HEC), a proportion change shape and become polar. The FLS profiles of lymphocytes stained for T and B cell surface markers show that both subsets change shape in response to HEC, but in a consistently different manner. Light microscopy of sorted T and B populations indicates that equal proportions of both cell types change shape, but that T cells are more likely to become highly elongated.

Recovery of Cryptococcus neoformans from sputum using new technics for the isolation of fungi from sputum.

Three sputum-digesting agents, N-acetyl-l-cytseine, dithiothreitol, and pancreatin-trypsin, were shown to be equally effective in allowing for the isolation of Cryptococcus neoformans from sputum samples in quantitative comparisons. By quantitative plating on bird-seed medium it was also shown the centrifugation after digestion concentrated C. neoformans into a platable sediment and, further, that the organisms, when present in concentrations as low as 10 yeasts per ml. of sputum, could be isolated with much higher frequencies than when no digestion-centrifugation procedure was used.

Tinea Pedis caused by Trichophyton violaceum.

This, to authors' knowledge, is the first report in the United States of Tinea pedis caused by Trichophyton violaceum in a recent Southeast Asian immigrant. Because of the recent entry into this country of refugees from an area of the world where T. violaceum is endemic, this anthropophilic dermatophyte may become a significant cause of dermatophytosis in the United States.

Cytopathology of opportunistic infection in bronchoalveolar lavage.

Bronchoalveolar lavage is an important tool for the cytologic and microbiologic examination of the lung. Silver- or Papanicolaou-stained slides from 604 lavage specimens from 344 patients were evaluated for the presence of fungal, parasitic, and viral organisms. Yeast, pseudohyphae, or hyphae occurred in 155 specimens (25.7%). Candida was the most frequent opportunistic fungus in immunosuppressed hosts. Patients with clinically significant Candida infection had many budding yeasts and pseudohyphae on cytologic preparations of their lavage fluid. Aspergillus, which occurred in five patients, was readily identified by cytologic examination and by fungal culture. Pneumocystis carinii was found in 14 patients (4%). Cytopathologic characteristics of Herpes simplex or cytomegalovirus were present in four patients (1.2%). In 414 specimens no pathogens were identified by either microbial culture or cytologic examination; usually, these patients had pulmonary hemorrhage, recurrent neoplasia, or therapy-induced lung disease to account for their abnormal chest x-rays. Cytologic and microbiologic examination of lavage fluid readily identified opportunistic pathogens. The ease and low morbidity of bronchoalveolar lavage support its use in the assessment of pulmonary infiltrates in immunocompromised hosts.

Protection of mice against the lethal effect of an intraperitoneal infection with Haemophilus (Actinobacillus) pleuropneumoniae after vaccination with capsular proteins.

Haemophilus (Actinobacillus) pleuropneumoniae Serotypes 5 and 7 capsular antigens (CA-1) were precipitated from culture supernatants with N-cetyl-N,N,N,-trimethylammonium bromide (Cetavlon). CA-1 contained a carbohydrate to protein ratio of 2:1 for Serotype 5 and 3:1 for Serotype 7. Glucosamine and uronic acid were detected in CA-1 from both serotypes suggesting that the capsule contained hyaluronic acid. All mice immunized intraperitoneally with CA-1 vaccine were protected from death when challenged with 10X the LD50 of the homologous but not the heterologous serotype. Oil adjuvants and the use of young (6 h) cultures were necessary for CA-1 vaccines to be protective. Deproteinization of CA-1 with chloroform and butanol followed by pronase treatment resulted in failure to protect mice from death. The protective capsular protein antigen in CA-1 vaccine may not originate from the outer membrane (OM) since repeated washing of the OM to elute the capsular protein antigen rendered the OM vaccine completely nonprotective for mice. Vaccines prepared from cell-wall lipopolysaccharide also were nonprotective for mice. Passive immunization of mice with anti-CA-1 antibody produced in rabbits to Serotype 5 was highly protective (P less than 0.01) for mice when challenged with 10X the LD50 of the homologous serotype.

Survival of ram spermatozoa at high dilution: protective effect of simple constituents of culture media as compared with seminal plasma.

During incubation of ram spermatozoa at 1 x 10(7) cells mL-1 or less in a simple HEPES-buffered saline medium, high levels of cell death were detected using propidium iodide as a probe of viability (membrane integrity): some 70% of the cells died during 3 h incubation at 37 degrees C. Because the conditions of incubation were similar to those encountered during manipulations for in vitro fertilization, this phenomenon was investigated further. If ram spermatozoa were diluted in an equivalent sucrose-based medium, or if the saline medium was supplemented with 10% seminal plasma, survival was greatly improved (only 5-15% died during a 3-h incubation at 37 degrees C); the protective effect of seminal plasma resided in a 5-10 kDa fraction. Sperm death in the basal saline medium was strongly dependent on cell concentration below 5 x 10(7) spermatozoa mL-1 whereas little effect of concentration was seen in the sucrose medium or in the presence of seminal plasma. The presence of Ca2+ (2 mM), EGTA (1 mM) or mercaptoethanol (1 mM) enhanced sperm survival in saline medium, but no effect was gained by replacing NaCl with KCl, and neither BSA nor fetal calf serum were beneficial. However, when a combination of pyruvate (1 mM), lactate (21.7 mM), Mg2+ (0.4 mM), phosphate (0.3 mM) and Ca2+ (2 mM) was included in the saline medium (to render it similar to Tyrode's medium), cell survival was greatly improved (12% died during the 3-h incubation).(ABSTRACT TRUNCATED AT 250 WORDS)

Randomized trial of artificial surfactant (ALEC) given at birth to babies from 23 to 34 weeks gestation.

Artificial surfactant (ALEC) composed of dipalmitoylphosphatidylcholine and unsaturated phosphatidylglycerol in a ratio of 7:3 (w/w) and a dose of 50-100 mg was suspended in 1 ml of cold saline and used at birth as a prophylaxis against the respiratory distress syndrome and its complications in a two centre randomized prospective trial involving 341 babies from 23 to 34 weeks gestation regardless of their antenatal problems. The surfactant had little effect in babies above 29 weeks gestation and was most beneficial in babies under 30 weeks gestation (67 controls and 69 surfactant treated babies). In this subgroup the artificial surfactant significantly reduced the inspired oxygen and peak ventilator pressure requirements during the first 96 h, the incidence of intraventricular haemorrhages from 40% to 19% (P less than 0.01), the overall mortality from 36% to 17% (P less than 0.02), the mortality due to RDS from 31% to 9% (P less than 0.01), the need for more than 28 days oxygen from 37% to 21% (P = 0.05) and the use of pancuronium in ventilated babies from 52% to 27% (P less than 0.01). There were no apparent side effects. This protein free, artificial surfactant should be a useful addition to the therapy of babies under 30 weeks gestation to reduce the severity of their RDS and the incidence of serious complications.

Characterization of a Pasteurella multocida bacteriophage.

The Pasteurella multocida bacteriophage B939a produces turbid plaques, which suggests infection with a temperate phage. Attempts to induce production of lytic phage by UV irradiation were unsuccessful, however, Bateriophage B932a is highly stable at 4 C and relatively stable at room temperature. It tolerates moist heat at 60 C, but is inactivated at 70 C in 10 minutes. Morphologically, it is a tailed phage with a hexagonal head.

Variation in the hemolytic activity of Leptospira interrogans serovar canicola.

Leptospira interrogans serovar canicola strain moulton was grown to a high cell density in a protein-free medium with hemin. The hemolysin produced in this culture system showed a greatly expanded spectrum of hemolysis as compared to previous reports of leptospiral hemolysin produced in a more traditional culture system containing serum.

Hemolysin production by Leptospira interrogans serovar canicola in a protein-free medium with hemin.

Leptospira interrogans serovar canicola strain moulton was grown to a high cell density in a protein-free medium. When hemin was added to this medium, hemolysin was produced. Hemolysin was not detected when other porphyrins or cytochrome C were substituted for hemin in the medium.

Cerebellocerebral diaschisis is the likely mechanism of postsurgical posterior fossa syndrome in pediatric patients with midline cerebellar tumors.

BACKGROUND AND PURPOSE: PFS occurs in approximately 25% of pediatric patients receiving surgery for midline posterior fossa tumors. Increasing evidence suggests that PFS represents a complex supratentorial cortical dysfunction related to surgery-induced disruption of critical cerebellocerebral connections. The purpose of this study was to determine whether a consistent surgical damage pattern may be identified in patients with PFS by early postoperative anatomic imaging analysis of the pECP and to test whether DSC can detect corresponding changes in cerebral cortical perfusion to indicate a secondary, remote functional disturbance, which could suggest a diaschisis-like pathomechanism. MATERIALS AND METHODS: Eleven patients with postoperative PFS were evaluated retrospectively and were paired with age- and sex-matched control subjects in whom PFS did not develop. MR imaging work-up included DSC within 3 to 4 weeks after surgery as well as early postoperative anatomic imaging to evaluate components of the pECP. RESULTS: DSC showed significant decreases in CBF within frontal regions (P < .05) and a trend to global cerebral cortical hypoperfusion in patients with PFS. Logistic regression analysis suggested a strong (potentially predictive) relationship between bilateral damage to pECP and the development of PFS (P = .04). CONCLUSIONS: Our data suggest that the primary cause of PFS is the bilateral surgical damage to the pECP. This leads to a trans-synaptic cerebral cortical dysfunction (a form of bilateral crossed cerebellocerebral diaschisis), which manifests with DSC-detectable global, but dominantly frontal, cortical hypoperfusion in patients with patients with PFS compared with age- and sex-matched control subjects.

Onychomycosis caused by Trichosporon beigelii in the United States.

We present the first case report of onychomycosis in the United States caused by Trichosporon beigelii in a normal, healthy man. The fungus, which was demonstrated histologically in the nails, was isolated on three different occasions with multiple cultures obtained each time.

Enhancement of ketoconazole penetration across the blood-brain barrier of mice by dimethyl sulfoxide.

Mice were treated with ketoconazole with and without dimethyl sulfoxide. Concentrations of ketoconazole at 3 and 5 h after treatment were significantly higher in serum (P less than 0.05) and brain tissue homogenate (P less than 0.01) of mice treated with dimethyl sulfoxide than in those of mice not treated with dimethyl sulfoxide.

Comparative study of the effect of thiabendazole and fungizone on Histoplasma capsulatum in macrophages.

The antifungal properties of Fungizone (amphotericin B intravenous solution) and thiabendazole on Histoplasma capsulatum within guinea pig macrophages were compared using the staining method and a newly developed plating method to determine the viability of intracellular H. capsulatum. The two methods were compared to determine the effectiveness of Fungizone and thiabendazole on H. capsulatum within macrophages. Fungizone was fungicidal for intracellular H. capsulatum, killing 99.9% of the yeasts at a concentration of 0.5 microgram/ml. There was some indication that non-viable intracellular yeasts were stained which could result in misinterpretation of the effectiveness of Fungizone using the staining method unless the yeasts are very closely examined for staining abnormalities. There was a good correlation between the two methods to demonstrate suppression of the multiplication of intracellular H. capsulatum in macrophages treated with 50 microgram/ml of thiabendazole. Thiabendazole was lethal for some intracellular H. capsulatum.