Autocrine stimulation of WI38 cell proliferation in the presence of glucocorticoids. Characteristics of the stimulatory factor(s) involved in this response

Chronic exposure to hydrocortisone (HC) or dexamethasone (DEX) results in a 20-40% extension in the proliferative lifespan of WI38 cells. Within a single growth cycle, the addition of HC or DEX at seeding results in saturation densities 20-40% higher than in control cultures. We have recently reported that, within a single growth cycle, the proliferative response of WI38 cells to glucocorticoids is mediated by a stimulatory factor(s) present in medium conditioned by cells in the presence of the hormone. We report here that chronic exposure to medium conditioned in the presence of HC for the first 24 h after seeding (24-h HC-conditioned medium (24-h HC-CM)) results in a 25% extension in the proliferative lifespan of these cultures. The generation of the stimulatory factor(s) present in glucocorticoid-conditioned medium is apparently dependent upon undefined cellular alterations which result from the subcultivation-procedure; confluent or low-density quiescent cultures did not generate media stimulatory to cell growth in the presence of glucocorticoids. This response was not trypsin-dependent, since cultures subcultivated in the absence of proteolytic treatment generated media equally stimulatory to cell growth. A further characterization of this glucocorticoid-induced activity revealed the stimulatory factor(s) was of low MW (dialyzable and recoverable in the less than 10,000 MW fraction following ultrafiltration), heat-stable (95 degrees C), and resistant to treatment with trypsin, chymotrypsin, or protease (S. griseus).     

A comparison of a monoclonal antibody-based sandwich ELISA and immunomagnetic bead selective enrichment for the detection of Escherichia coli O157 from bovine faeces

AIMS: To compare a recently developed monoclonal antibody (MAb) based sandwich ELISA (sELISA) with an immunomagnetic separation (IMS) method for the detection of Escherichia coli O157 in bovine faeces. METHODS AND RESULTS: Faecal samples from 345 cattle were obtained from eight farms in Northern Ireland, in which human disease due to E. coli O157 had occurred. Both assays detected E. coli O157 on five of the farms and the phage-type of the majority of the bovine strains were the same as the corresponding human isolates. Similar numbers of the organism were detected by the two methods, 59 by the sELISA and 53 by the IMS procedure, 39 of the positive samples being common to both. Twenty samples were sELISA positive/IMS negative. CONCLUSIONS: If the IMS is regarded as the gold standard, then the sELISA is less sensitive and less specific, but under the conditions used sELISA positive results were obtained from all positive farms, and the sELISA gave a presumptive positive a day earlier than the IMS method. SIGNIFICANCE AND IMPACT OF THE STUDY: The sELISA has the potential to be used as a rapid method for screening large numbers of samples for E. coli O157, but further work is required to determine its specificity.     

Formation and stability of the complex formed between human antithrombin-III and thrombin

The inhibition of thrombin by antithrombin-III involves formation of a 1:1 covalent complex between protease and inhibitor and concomitant cleavage of the antithrombin-III peptide chain after Arg-385. The resultant fragment remains connected to the complex via a disulfide bond. This complex spontaneously breaks down into a fragment of approximately 55,000 daltons and smaller peptides. Breakdown is prevented by the presence of hydroxylamine or diisopropylflurophosphate, or by denaturation with urea. It occurs even if the purified complex is treated with diisopropylflurophosphate prior to purification, and can be greatly accelerated by the presence of small amounts of active thrombin. The initial sites of proteolytic attack on the complex are after Arg-13 of the thrombin A chain and Arg-68 of the thrombin B chain. These data indicate that active thrombin can be released from the antithrombin-thrombin complex, and that thrombin becomes more susceptible to proteolytic attack when complexed with antithrombin.     

Antibody responses of human lymphocytes in vitro. Specificity and physical properties of plaque-forming cell precursors.

When human tonsillar lymphocytes are cultured with pokeweed mitogen they generate a larger number of antibody-secreting cells. A substantial number of the plaque-forming cells generated have specificity for sheep erythrocytes but very few are specific for either bovine or horse erythrocytes. We have investigated the physical properties of sheep erythrocyte-specific precursors and have found that they have the characteristics of larger lymphocytes and are heterogeneous with respect to density. Our results support the concept that the percursors which can be induced by pokeweed mitogen to develop into plaque-forming cells belong to a pre-activated sub-population of lymphocytes.     

RNA sequence analysis shows that the symbionts in the ciliate Metopus contortus are polymorphs of a single methanogen species

The polymerase chain reaction was used to amplify and partially sequence the 16S ribosomal RNA genes of symbiotic bacteria within the anaerobic ciliate Metopus contortus. In situ probing with fluorescent oligonucleotides showed that the amplified sequences originated from a single species of archaebacterium which is closely related to Methanocorpusculum parvum. The probed symbionts exhibited a variety of shapes and sizes. These data support the hypothesis, first proposed on the basis of electron microscopy, that the symbionts undergo a morphological transformation as part of the symbiotic process.     

Preoperative levels of natriuretic peptides and the incidence of postoperative atrial fibrillation after noncardiac surgery: a prospective cohort study

BACKGROUND:Postoperative atrial fibrillation (POAF) is associated with clinically significant short- and long-term complications after noncardiac surgery. Our aim was to describe the incidence of clinically important POAF after noncardiac surgery and establish the prognostic value of N-terminal pro–brain-type natriuretic peptide (NT-proBNP) in this context.METHODS:The Vascular events In noncardiac Surgery patIents cOhort evaluatioN (VISION) Study was a prospective cohort study involving patients aged 45 years and older who had inpatient noncardiac surgery that was performed between August 2007 and November 2013. We determined 30-day incidence of clinically important POAF (i.e., resulting in angina, congestive heart failure, symptomatic hypotension or requiring treatment) using logistic regression models to analyze the association between preoperative NT-proBNP and POAF.RESULTS:In 37 664 patients with no history of atrial fibrillation, we found that the incidence of POAF was 1.0% (95% confidence interval [CI] 0.9%–1.1%; 369 events); 3.2% (95% CI 2.3%–4.4%) in patients undergoing major thoracic surgery, 1.3% (95% CI 1.2%–1.5%) in patients undergoing major nonthoracic surgery and 0.2% (95% CI 0.1%–0.3%) in patients undergoing low-risk surgery. In a subgroup of 9789 patients with preoperative NT-proBNP measurements, the biomarker improved the prediction of POAF risk over conventional prognostic factors (likelihood ratio test p < 0.001; fraction of new information from NT-proBNP was 16%). Compared with a reference NT-proBNP measurement set at 100 ng/L, adjusted odds ratios for the occurrence of POAF were 1.31 (95% CI 1.15–1.49) at 200 ng/L, 2.07 (95% CI 1.27–3.36) at 1500 ng/L and 2.39 (95% CI 1.26–4.51) at 3000 ng/L.INTERPRETATION:We determined that the incidence of clinically important POAF after noncardiac surgery was 1.0%. We also found that preoperative NT-proBNP levels were associated with POAF independent of established prognostic factors. Trial registration: ClinicalTrials.gov, no. {"type":"clinical-trial","attrs":{"text":"NCT00512109","term_id":"NCT00512109"}}NCT00512109

Postoperative atrial fibrillation (POAF) is the most common arrhythmia that occurs after noncardiac surgery¹ and is associated with prolonged hospital stays and an increased risk of stroke and death,^2–8 both of which raise societal costs.⁹ The reported incidence of POAF ranges from 3.7% to 21.0% in patients undergoing thoracic surgery^3,10 and from less than 0.5% to 10.0% in patients undergoing noncardiac, nonthoracic surgery (Appendix 1, Supplementary Table 1, available at www.cmaj.ca/lookup/doi/10.1503/cmaj.200840/tab-related-content).^6,11Brain-type natriuretic peptide (BNP) is a neurohormone released by cardiomyocytes in response to volume and pressure overload.¹² This biomarker has an established role in the diagnosis and management of heart failure and in predicting cardiovascular morbidity and mortality.¹³ Although natriuretic peptides have been shown to predict new-onset atrial fibrillation (AF) in the nonsurgical setting reliably,^14–18 their value in predicting POAF after noncardiac surgery is unclear.^19–21The Vascular events In non-cardiac Surgery patIents cOhort evaluatioN (VISION) Study was a prospective international cohort study involving a representative sample of adults who underwent noncardiac surgery that required an overnight hospital stay. One of the predefined objectives in VISION was to determine the incidence of new-onset, clinically important POAF. We aimed to describe the overall and surgery-specific incidence of POAF after noncardiac surgery and to investigate the association between preoperative N-terminal pro–brain-type natriuretic peptide (NT-proBNP) and POAF. We hypothesized that measurement of NT-proBNP would improve POAF risk prediction beyond conventional prognostic factors.     

A Preliminary Study of Human Calcium Metabolism Using Long-Term Whole Body Tracer Measurements

Long-term bone calcium metabolism was measured using a whole body counter apparatus and the radioisotopes calcium-47 and strontium-85. Strontium was given with calcium in a single intravenous injection in order to estimate long-term retention of calcium. Calcium-47, with a short radio-active half-life, was measured directly for 20 days, and then indirectly from strontium retention for periods up to 200 days.The standard parameters of calcium transport were measured and, as well, a new parameter, calcium accretion to the fixed bone pool. These studies were carried out on 13 subjects, six of whom were selected “normals” and seven were patients with osteoporosis. Calcium accretion to the fixed bone pool, the new index of bone formation, varied from 100 to 210 mg. of calcium daily in normal subjects and from 70 to 340 mg./day in patients with osteoporosis. These values are one-third of those for the accretion to the total bone pool, the previously reported bone formation rate.