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PubMed1982

Cytotoxic material released from Staphylococcus epidermidis. II. Fractionation and some effects of the fractions on lymphocytes and hepatocytes.

Cytotoxic substance(s) of about 4 X 10(3) molecular weight, containing 9.5% peptide and 73% carbohydrate was released from Staphylococcus epidermidis in phosphate buffered saline. The material was soluble in ethanol and was heat-resistant. It blocked amino acid uptake and E-rosette formation of human tonsillar and blood lymphocytes. In isolated mouse hepatocytes the toxin inhibited protein synthesis, but only in the presence of calcium ions. The results suggest that eukaryotic cell membranes are damaged by the coccal agent.

Animals↗
PubMed1976

Studies on the structure and function of chick-oviduct chromatin. 1. Fractionation by ECTHAM-cellulose chromatography and physico-chemical characterization.

Chick oviduct chromatin was separated into a ribonucleoprotein fraction and two chromatin fractions (early and late eluting). We utilized a gentle procedure in which moderately hydrated chromatin was subjected to chromatography on a weak ionic-exchange resin (ECTHAM-cellulose) eluted with a combined pH-salt gradient. Chemical analysis of the early (fraction I) and late (fraction II) eluting fractions revealed that their histones were identical and their nonhistone proteins were markedly different. Control experiments showed that these differences were not due to protein rearrangements during chromatin preparation and/or fractionation. The physical properties of fraction I and II differed in certain aspects. The aggregation response of fraction I to increasing concentrations of monovalent cations was five times lower than that of fraction II but the aggregation response to divalent cations was identical. Thermal denaturation assays of DNAs isolated from fractions I and II revealed identical derivative profiles of hyperchromicity vs temperature, thereby indicating similar base composition in the two fractions. Circular dichroism, spectra of the purified DNAs isolated from both fractions showed identical B-type conformations. However, DNA renaturation kinetics analyzed by computer technique indicated that fraction I DNA contained less than half the amount of highly repetitive sequences as compared to either unfractionated chromatin or fraction II. Circular dichroism spectra of fraction I and II chromatins (at room temperature) showed significant differences in a wavelength region were only DNA is optically active (i.e. 255-320 nm). These results indicated that the DNA complexed to proteins in fraction II assumed a more C-type conformation than the DNA in fraction I. The differences in the circular dichroism spectra could not be accounted for by differences in the RNAs or protein chromophores contained in fraction I and fraction II. When the circular dichroism spectra of fraction I and II were recorded at 55 degrees C, the differences between the two fractions were abolished. These results were interpreted to mean that the differences in the DNA conformations found in fractions I and II were due to the differences in their nonhistone proteins. These proteins were effective in maintaining DNA conformation differences only when they were in their native form but not when heated to 55 degree C. Comparison of the sedimentation coefficients of fractions I and II with their calculated molecular weights suggested a more extended structure in fraction I as compared to a more compact structure in fraction II. Only small differences were observed between fraction I and fraction II with respect to either buoyant density analysis in a metrizamide gradient or in the number of phosphate charges accessible to polylysine.

Animals↗
PubMed2006

National survey of radiotherapy fractionation practice in 2003.

AIMS: To document UK practice in radiotherapy fractionation. METHODS: All radiotherapy centres in the UK participated in a 1-week audit from 29 September 2003. Fractionation data were collected for all patients starting external beam radiotherapy. This included 2498 patients who were prescribed 32 547 fractions. RESULTS: For the radical treatment of non-skin malignancy (n = 708), the prescribed dose ranged from a single fraction of 8 Gy for total-body irradiation to 75 Gy in 43 fractions for prostate cancer. Postoperative treatment for breast cancer was dominated by three regimens: 40 Gy in 15 fractions; 45 Gy in 20 fractions; and 50 Gy in 25 fractions. Palliative treatment was given in a single fraction to 393 patients (36%) with doses of up to 15 Gy. Three hundred and ninety patients (36%) received four to seven fractions delivering 20-25 Gy. Only 89 patients (8%) received more than 10 fractions with palliative intent but used 29% of such fractions. In the treatment of bone metastases, the most common prescriptions were 8-10 Gy in a single fraction and 20 Gy in five fractions. CONCLUSION: UK radiotherapy practice has become more uniform and moved closer to practice in North America and Europe over the past 15 years. For radical radiotherapy, 54% of prescriptions were for a fraction size of 1.8-2.0 Gy but the distribution was bi-modal and 20% of patients were prescribed fraction sizes of 2.7-3.0 Gy. Evidence-based practice now supports hypo-fractionated palliative treatment favouring single fractions for bone metastases and one or two fractions for many patients with advanced lung cancer. Two fractions are advised for some patients with brain metastasis. If these guidelines had been applied uniformly, then the number of treatments prescribed for palliation could have fallen by 36% from 5197 to 3313. This would have represented a 6% reduction in the overall radiotherapy workload. Not all patients are suitable for such hypo-fractionated treatments, but this is an area in which resource use can be improved. In the postoperative management of breast cancer, a change in practice to use 15 fractions uniformly would reduce overall radiotherapy workload by 4%. By contrast, a change to 25 fractions would increase overall workload by 7%.

Data Collection↗
PubMed1996

Mortality and morbidity after coronary artery bypass surgery related to pre-operative left ventricular ejection fraction: a follow-up study.

OBJECTIVE: To study the pre-operative level of left ventricular ejection fraction that may be indicative of an increased risk of early and late mortality and of recurrent angina pectoris and late non-fatal myocardial infarction. MATERIAL AND METHODS: A total of 934 patients with known left ventricular ejection fraction, 80 women and 854 men, were submitted to coronary artery bypass grafting at the Cardiovascular Unit of Rikshospitalet, Oslo, between August 1982 and December 1986. The closing date was the 1st of January 1993, with a mean follow-up of time of 7.4 years. The patients were divided in to four subgroups according to their level of left ventricular ejection fraction: < or = 40%, 41-60%, 61-80% and > 80%. The left ventricular ejection fraction varied from 13-98%. A chi-square test of linear trend was used to calculate the relative risk between the different subgroups. Cumulative survival was determined using survival curves. RESULTS: Early mortality. Twenty-five patients (2.7%) died within 30 days of operation. Patients with left ventricular ejection fraction < or = 40% had a relative risk of 10.2 (1.9-17.2), for left ventricular ejection fraction 41-60% the relative risk was 0.9 (0.1-8.9) and for left ventricular ejection fraction 61-80% the relative risk was 2.8 (0.6-17.2). Left ventricular ejection fraction > 80% was defined as relative risk = 1. Late mortality. Altogether, 174 patients died in the late phase (18.6%). For patients with left ventricular ejection fraction < or = 40% the relative risk was 3.6 (2.8-10.9), for left ventricular ejection fraction 41-60% the relative risk was 1.8 (1.1-3.6), and for left ventricular ejection fraction 61-80% the relative risk was 1.5 (0.9-2.8). Recurrent angina pectoris. A total of 138 patients developed recurrent angina pectoris during the follow-up period, giving an incidence of 14.8%. Here, for left ventricular ejection fraction < or = 40% the relative risk was 0.5 (0.2-1.3), for left ventricular ejection fraction 41-60% the relative risk was 1.0 (0.5-1.8) and for left ventricular ejection fraction 61-80% the relative risk was 1.2 (0.7-2.0). Late non-fatal myocardial infarction. Altogether, 90 patients (9.6%) experienced non-fatal myocardial infarction in the late phase. For left ventricular ejection fraction < or = 40% the relative risk was 0.6 (1.2-1.8), for left ventricular ejection fraction 41-60% the relative risk was 1.0 (0.5-2.0) and for left ventricular ejection fraction 61-80% the relative risk was 0.7 (0.41-1.3). Cumulative survival. When pooled together, the cumulative survival for patients with left ventricular ejection fraction > 40% was 95.9, 91.9 and 79% after 1, 5 and 10 years, respectively. For the patients with left ventricular ejection fraction < or = 40% cumulative survival was 87.5, 73.1 and 55.2%, respectively. CONCLUSION: When the left ventricular ejection fraction was 40% or lower, there was a substantial increase in the risk of early mortality in patients submitted to coronary artery bypass grafting. As for the risk of late mortality, there was a practically linear increase in risk with falling values of left ventricular ejection fraction. We found no difference in risk of developing recurrent angina pectoris or of late non-fatal myocardial infarction related to values of left ventricular ejection fraction.

Aged↗
PubMed1995

Heterogeneity in the fractionation sensitivities of human tumor cell lines: studies in a three-dimensional model system.

PURPOSE: Current concepts to optimize the therapeutic gain of radiotherapy by hyperfractionation assume that human tumors are less sensitive to fractionation than late reacting normal tissues. The aim of this study was to investigate the extent of the intercell line heterogeneity of fractionation sensitivity of a wide variety of human tumor cell lines in a three-dimensional model system under fully oxic conditions using schedules with one to eight fractions. Biological characteristics of the tumors that correlate with fractionation sensitivity should be identified. METHODS AND MATERIALS: A total of 21 cell lines from human tumors maintained as multicellular spheroids consisting of 1000-1500 cells were given fractionated irradiation within a total treatment time of maximally 50 h. Complete dose-spheroid control curves were determined for each fractionation scheme. The spheroid control data were adequately described by the linear quadratic model assuming Poisson statistics. In addition, the induction of a G2 block by a fractionated test dose of seven 3 Gy fractions given at 6-h intervals was determined in spheroid cells using flow cytometry of propidium bromide stained cell nuclei. RESULTS: The fractionation sensitivities of human tumor cells in multicellular spheroids could be characterized by alpha/beta values, ranging from 2.8-37 Gy in dependence on the cell line. The log normally distributed alpha/beta values were positively correlated with the percentage increase in G2/M phase after the fractionated test dose compared to the controls (r = 0.72, p < 0.01), and were associated with the degree of tumor differentiation (p = 0.01, ANOVA F-test). No significant correlation between the log (alpha/beta) values and the surviving fractions at 2 Gy (SF2) or the total doses with 2 Gy per fraction necessary to control 50% of the spheroids (SCD50) was observed. Despite the intercell line variability of the alpha/beta values, the SCD50 values of the different cell lines, given with one and eight fractions or one fraction and 2 Gy per fraction, were closely associated (Spearman rank correlation coefficients: r = 0.89 or r = 0.90, p < 0.0001). CONCLUSION: Human tumor cell lines showed a marked heterogeneity in the fractionation sensitivity when irradiated as multicellular spheroids and assayed in situ using the spheroid control end point. Therefore, the therapeutic gain of altered fractionation also depends on those biological characteristics of each individual tumor that affects its fractionation sensitivity. Parameters that correlate with fractionation sensitivity of the tumor lines in the spheroid system were identified as grade of tumor differentiation and percentage increase in G2/M cells at the end of an eight-fraction schedule.

Humans↗
PubMed2001

Stable hydrogen and carbon isotope fractionation during microbial toluene degradation: mechanistic and environmental aspects.

Primary features of hydrogen and carbon isotope fractionation during toluene degradation were studied to evaluate if analysis of isotope signatures can be used as a tool to monitor biodegradation in contaminated aquifers. D/H hydrogen isotope fractionation during microbial degradation of toluene was measured by gas chromatography. Per-deuterated toluene-d(8) and nonlabeled toluene were supplied in equal amounts as growth substrates, and kinetic isotope fractionation was calculated from the shift of the molar ratios of toluene-d(8) and nondeuterated toluene. The D/H isotope fractionation varied slightly for sulfate-reducing strain TRM1 (slope of curve [b] = -1.219), Desulfobacterium cetonicum (b = -1.196), Thauera aromatica (b = -0.816), and Geobacter metallireducens (b = -1.004) and was greater for the aerobic bacterium Pseudomonas putida mt-2 (b = -2.667). The D/H isotope fractionation was 3 orders of magnitude greater than the (13)C/(12)C carbon isotope fractionation reported previously. Hydrogen isotope fractionation with nonlabeled toluene was 1.7 and 6 times less than isotope fractionation with per-deuterated toluene-d(8) and nonlabeled toluene for sulfate-reducing strain TRM1 (b = -0.728) and D. cetonicum (b = -0.198), respectively. Carbon and hydrogen isotope fractionation during toluene degradation by D. cetonicum remained constant over a growth temperature range of 15 to 37 degrees C but varied slightly during degradation by P. putida mt-2, which showed maximum hydrogen isotope fractionation at 20 degrees C (b = -4.086) and minimum fractionation at 35 degrees C (b = -2.138). D/H isotope fractionation was observed only if the deuterium label was located at the methyl group of the toluene molecule which is the site of the initial enzymatic attack on the substrate by the bacterial strains investigated in this study. Use of ring-labeled toluene-d(5) in combination with nondeuterated toluene did not lead to significant D/H isotope fractionation. The activity of the first enzyme in the anaerobic toluene degradation pathway, benzylsuccinate synthase, was measured in cell extracts of D. cetonicum with an initial activity of 3.63 mU (mg of protein)(-1). The D/H isotope fractionation (b = -1.580) was 30% greater than that in growth experiments with D. cetonicum. Mass spectroscopic analysis of the product benzylsuccinate showed that H atoms abstracted from the toluene molecules by the enzyme were retained in the same molecules after the product was released. Our findings revealed that the use of deuterium-labeled toluene was appropriate for studying basic features of D/H isotope fractionation. Similar D/H fractionation factors for toluene degradation by anaerobic bacteria, the lack of significant temperature dependence, and the strong fractionation suggest that analysis of D/H fractionation can be used as a sensitive tool to assess degradation activities. Identification of the first enzyme reaction in the pathway as the major fractionating step provides a basis for linking observed isotope fractionation to biochemical reactions.

Anaerobiosis↗
PubMed1969

Fractionation and characterization of the plasma and mesosome membrane of Listeria monocytogenes.

Protoplasts of Listeria monocytogenes strain 42 were fractionated after control lysis on a Ficoll (a polysucrose) density gradient. Visually, five zones could be recognized in the gradient. The first one was composed of amorphous cytoplasmic solutes (fraction 1a) and a mixture of particles (fraction 1b). These were: (i) light particles that were lipase-sensitive and composed of six subunits and (ii) heavy particles, sensitive to ribonuclease and devoid of fine structure. The second zone consisted of tubules and vesicles still harboring cytoplasmic components (fraction 2), whereas the third zone contained only empty vesicles and protoplast ghosts (fraction 3). The material congregating into the fourth zone was morphologically identical to that of the third (fraction 3a). The fifth and heaviest zone contained a mixture of (i) particles without any substructure and (ii) partly lysed protoplasts (fraction 4). Fractions 1b and 4 were the richest in nucleic acids (ribonucleic acid, 11.4 and 9.4%, respectively; deoxyribonucleic acid, 5.1 and 4.8%, respectively), whereas fraction 1b had the highest protein contents (74.6%). Phospholipids were mainly found in fractions 2 and 3. Except for fraction 1, all materials contained significant amounts of protein-bound phosphorus. The main concentrations of four enzymes were: glucose-6-phosphate dehydrogenase (fraction 1a); adenosine triphosphatase and reduced nicotinamide adenine diphosphate oxidase (fraction 3); nitro blue tetrazolium chloride reductase (fraction 2). Fractionation of strain 42 after addition of (32)P during the mid-log phase of growth revealed that the radio-activity was mainly detected in fraction 1b, when growth in the presence of the marker was allowed for 10 min, and in fraction 2, when growth was allowed for 90 min. The vesicles of fraction 2, often tubular, are probably of mesosomal origin, whereas those of fraction 3, which are always spherical, represent, most likely, the bulk of the cell plasma membrane. Our data showed slight chemical differences between these two fractions, but the differences in enzymatic activities and lipid-phosphorus incorporation during long pulse experiments were most dramatic.

Adenosine Triphosphatases↗
PubMed1969

Studies on protein-polysaccharides from pig laryngeal cartilage. Heterogeneity, fractionation and characterization.

1. Protein-polysaccharides from pig laryngeal cartilage extracted by two procedures described in the preceding paper (Tsiganos & Muir, 1969) were shown to consist of macromolecules of various sizes as assessed by gel filtration in 4% and 6% agarose. 2. A larger proportion of the smaller molecules was present in the preparation obtained by brief extraction in iso-osmotic sodium acetate (procedure I) than in that obtained by more prolonged extraction in 10% (w/v) calcium chloride (procedure II). 3. Two fractions were separated by gel filtration in 6% agarose and by electrophoresis in compressed glass fibre. These fractions differed in chemical composition and in antigenic determinants. The gel-retarded fraction R and that of higher electrophoretic mobility possessed the same single antigen, whereas the gel-excluded fraction E and the slower electrophoretic fraction contained all the antigens of the starting material including that of fraction R. 4. Five N-terminal amino acid residues were identified in preparation I and fraction E, only two of which were present in fraction R. 5. The relative proportions of gel-excluded and gel-retarded fractions did not change when solutions of high ionic strength, urea or guanidine hydrochloride were used for elution. 6. The differences in chemical and amino acid composition between fractions R and E showed that the latter was not a simple aggregate of the former. Fraction E contained more basic and aromatic amino acids, and some methionine and cystine; the last two were absent from fraction R. Hydroxyproline was not detected in either fraction. 7. The number of glycosidic linkages in both fractions was estimated by alkaline beta-elimination. Appreciable amounts of threonine as well as serine were destroyed in both fractions. An average chain length for chondroitin sulphate was calculated from the galactosamine content of both fractions and the amounts of hydroxy amino acid destroyed. Average chain lengths were also calculated from the xylose and galactosamine content of each fraction. Each independent method gave a value of approximately 28 disaccharide units for the chain length in both fractions and hence their difference in size could not be explained by differences in the length of carbohydrate chains. 8. All fractions contained glucosamine, which was attributed to keratan sulphate. Content of both protein and keratan sulphate increased with the size of the macromolecules. 9. It is suggested, from these results, that chondroitin sulphate-protein complexes normally exist as a heterogeneous population of macromolecules in cartilage, and that keratan sulphate is involved in the formation of larger molecules.

Amino Acids↗
PubMed2006

Outcome of heart failure with preserved ejection fraction in a population-based study.

BACKGROUND: The importance of heart failure with preserved ejection fraction is increasingly recognized. We conducted a study to evaluate the epidemiologic features and outcomes of patients with heart failure with preserved ejection fraction and to compare the findings with those from patients who had heart failure with reduced ejection fraction. METHODS: From April 1, 1999, through March 31, 2001, we studied 2802 patients admitted to 103 hospitals in the province of Ontario, Canada, with a discharge diagnosis of heart failure whose ejection fraction had also been assessed. The patients were categorized in three groups: those with an ejection fraction of less than 40 percent (heart failure with reduced ejection fraction), those with an ejection fraction of 40 to 50 percent (heart failure with borderline ejection fraction), and those with an ejection fraction of more than 50 percent (heart failure with preserved ejection fraction). Two groups were studied in detail: those with an ejection fraction of less than 40 percent and those with an ejection fraction of more than 50 percent. The main outcome measures were death within one year and readmission to the hospital for heart failure. RESULTS: Thirty-one percent of the patients had an ejection fraction of more than 50 percent. Patients with heart failure with preserved ejection fraction were more likely to be older and female and to have a history of hypertension and atrial fibrillation. The presenting history and clinical examination findings were similar for the two groups. The unadjusted mortality rates for patients with an ejection fraction of more than 50 percent were not significantly different from those for patients with an ejection fraction of less than 40 percent at 30 days (5 percent vs. 7 percent, P=0.08) and at 1 year (22 percent vs. 26 percent, P=0.07); the adjusted one-year mortality rates were also not significantly different in the two groups (hazard ratio, 1.13; 95 percent confidence interval, 0.94 to 1.36; P=0.18). The rates of readmission for heart failure and of in-hospital complications did not differ between the two groups. CONCLUSIONS: Among patients presenting with new-onset heart failure, a substantial proportion had an ejection fraction of more than 50 percent. The survival of patients with heart failure with preserved ejection fraction was similar to that of patients with reduced ejection fraction.

Aged↗