Development of large numbers of mast cells at sites of idiopathic chronic dermatitis in genetically mast cell-deficient WBB6F1-W/Wv mice

The normal skin and other tissues of adult mast cell-deficient WBB6F1- W/Wv or WCB6F1-Sl/Sld mice contain less than 1.0% the number of mast cells present in the corresponding tissues of the congenic normal (+/+) mice. As a result, genetically mast cell-deficient WBB6F1-W/Wv or WCB6F1-Sl/Sld mice are widely used for studies of mast cell differentiation and function. We found that mast cells developed at sites of idiopathic chronic dermatitis in WBB6F1-W/Wv mice and that the number of mast cells present in the skin of WBB6F1-W/Wv mice was proportional to the severity of the dermatitis (in ear skin, there were 33 +/- 4 mast cells/mm2 of dermis at sites of severe dermatitis v 9 +/- 3 at sites of mild dermatitis, 0.8 +/- 0.3 in skin without dermatitis, and 100 +/- 7 in the normal skin of congenic WBB6F1-+/+ mice; in back skin, the corresponding values were 2.0 +/- 0.6, 1.1 +/- 0.9, 0.025 +/- 0.025, and 26.2 +/- 3.2). The development of mast cells was a local, not systemic, consequence of the dermatitis. Thus, WBB6F1-W/Wv mice with severe dermatitis lacked mast cells in skin not showing signs of dermatitis and also in the peritoneal cavity, stomach, cecum, and tongue. Idiopathic chronic dermatitis was not associated with the local development of mast cells in WCB6F1-Sl/Sld mice, a mutant whose mast cell deficiency is due to a mechanism distinct from that of WBB6F1-W/Wv mice. These findings may have implications for understanding the nature of the mast cell deficiency in WBB6F1-W/Wv and WCB6F1-Sl/Sld mice and for the use of these mutants to analyze mast cell differentiation and function.     

CML patients in blast crisis have breakpoints localized to a specific region of the BCR

Chronic myelogenous leukemia (CML) is associated with the Philadelphia (Ph) chromosome, which results from a reciprocal translocation between chromosomes 9 and 22. This activates the abl oncogene by moving it from chromosome 9 and combining it with sequence located on chromosome 22. The new fusion gene, with chromosome 22 sequence at its 5' end and chromosome 9-abl sequence at its 3' end, generates a new messenger RNA (mRNA) and protein that are implicated in the pathogenesis of CML. The breakpoint near the c-abl locus on chromosome 9 can occur within a large area. In contrast, the breakpoints on chromosome 22 are concentrated within a 6 kilobase (kb) region termed the breakpoint cluster region (bcr). This study was designed to determine whether chronic-phase and blast crisis patients had identifiable differences in the structure of their Ph chromosomes. Restriction mapping of the chromosome 22 translocation breakpoints performed for 26 patients showed that the breakpoints of eight of the nine patients in blast crisis were in the 3' portion of the bcr, whereas the breakpoints in the 17 patients in the chronic phase were clustered in the 5' portion of the bcr. This suggests a strong correlation between a 3' bcr breakpoint and blast crisis in CML.     

Calcium-dependent cysteine protease activity in the sera of patients with thrombotic thrombocytopenic purpura

Plasma and serum from patients with thrombotic thrombocytopenic purpura (TTP) can cause activation and aggregation of normal human platelets in vitro. It is possible that this platelet-activating factor contributes to the disease. In this report we describe studies designed to identify the platelet-activating factor in TTP. Platelet activation by sera from 15 patients with TTP was inhibited by leupeptin, iodoacetamide, and antipain but not by phenylmethylsulphonylfluoride, epsilon-aminocaproic acid, soybean trypsin inhibitor, aprotinin, and D-phenylanyl-1-prolyl-1- arginine chloromethyl ketone. These studies suggested that the platelet- activating factor in TTP serum was a cysteine protease. We confirmed that a calcium-dependent cysteine protease (CDP) was present in the sera of each of the 15 patients when we used an assay based on the ability of CDP to proteolyse platelet membrane glycoprotein 1b (GP1b) and hence to abolish the ability of CDP-treated normal platelets to agglutinate in the presence of ristocetin and von Willebrand factor. This proteolytic activity was inhibited by EDTA, leupeptin, antipain, iodoacetamide, and by N-ethyl-maleamide (NEM) but not by the serine protease inhibitors. Activity was detected in 15 of 15 patients with TTP tested before therapy was begun. In contrast, no activity was detected in the serum of any of five of the TTP patients tested in remission or in any of the sera from 36 patients with thrombocytopenia and 423 nonthrombocytopenic controls. To look for in vivo CDP activity in patients with TTP, we studied platelets from two patients with acute TTP (drawn into acid-citrate-dextrose, NEM, and leupeptin). These platelets showed a loss of GP1b from the platelet surface. Both patients were also studied in remission: GP1b on the platelet surface had returned to normal. These studies provide evidence that CDP is present in the sera of patients with TTP, that it is specific to this disease, and that is is active in vivo as well as in vitro. We postulate that a disorder of CDP homeostasis plays a major role in the pathophysiology of TTP.     

A subpopulation of immunoglobulin G in man selectively interacts with the hormone-binding sites of estrogen receptors

Since the binding sites of hormone receptors may be similar to those of antihormone antibodies, we wondered whether the former might not be recognized by the idiotypic network. To test this hypothesis we investigated the interaction of plasma immunoglobulin G (IgG) with the binding sites of estrogen receptors (ER) from uterine or mammary tissue. Using ER isolated from uterine cytosol we found that IgG from normal subjects shifted the position of purified receptor in sucrose gradients and displaced [3H]estradiol (E2) from its receptor-binding sites. Equilibrium studies revealed competitive inhibition by IgG of E2 binding to the ER. IgG isolated by adsorption on a rat uterine cytosol-Blue B matrix gel column also bound to the ER, and this binding was inhibited by an excess of E2. After an 18-h exposure of MCF-7 mammary carcinoma cells in monolayer culture to IgG (2 mg/ml), Scatchard analysis of [3H]E2 binding revealed a reproducible decrease in the available receptor sites from 2.52 +/- 0.56 (+/- SEM) to 0.68 +/- 0.48 fmol/microgram DNA (n = 10). This effect was selective, since enriched anti-ER IgG obtained by adsorption on purified receptor was 20 times more potent than total IgG, whereas IgG identically prepared but not retained by affinity chromatography had no activity. Exposure of the cells to the IgG for 45 min also revealed, as with isolated ER, specific competition of the IgG with E2 for the E2-binding sites; the Kd increased from 10.5 +/- 1.6 to 27.5 +/- 7.2 X 10(-11) M (n = 7). Enriched antireceptor IgG was a 20 times more effective competitor, and the IgG not retained by affinity chromatography had no activity. In conclusion, our observations indicate the presence of ER on the cell surface, interaction of ER with IgG from plasma of normal subjects, and competitive antagonism of these IgG with E2 uptake leading to a decrease in effective ER concentrations.     

Profoundly different prion diseases in knock-in mice carrying single PrP codon substitutions associated with human diseases

In man, mutations in different regions of the prion protein (PrP) are associated with infectious neurodegenerative diseases that have remarkably different clinical signs and neuropathological lesions. To explore the roots of this phenomenon, we created a knock-in mouse model carrying the mutation associated with one of these diseases [Creutzfeldt–Jakob disease (CJD)] that was exactly analogous to a previous knock-in model of a different prion disease [fatal familial insomnia (FFI)]. Together with the WT parent, this created an allelic series of three lines, each expressing the same protein with a single amino acid difference, and with all native regulatory elements intact. The previously described FFI mice develop neuronal loss and intense reactive gliosis in the thalamus, as seen in humans with FFI. In contrast, CJD mice had the hallmark features of CJD, spongiosis and proteinase K-resistant PrP aggregates, initially developing in the hippocampus and cerebellum but absent from the thalamus. A molecular transmission barrier protected the mice from any infectious prion agents that might have been present in our mouse facility and allowed us to conclude that the diseases occurred spontaneously. Importantly, both models created agents that caused a transmissible neurodegenerative disease in WT mice. We conclude that single codon differences in a single gene in an otherwise normal genome can cause remarkably different neurodegenerative diseases and are sufficient to create distinct protein-based infectious elements.     

Safety and efficacy of low-dose intravenous enoxaparin and GP IIb/IIIa inhibitor therapy during PCI

The use of intravenous enoxaparin, a glyco-protein (GP) IIb/IIIa inhibitor, during percutaneous coronary intervention (PCI) has been shown to be safe and to possibly reduce in-hospital and 30-day major adverse cardiac events(MACE). NICE-4, a recent PCI observational study, evaluated a reduced dose of intravenous (IV) enoxaparin (0.75 mg/kg) with abciximab. However, prior PCI studies evaluating IV enoxaparin have not used percutaneous closure devices. The purpose of this study was to observe the safety and efficacy of a lower dose of IV enoxaparin (0.5 mg/kg) in conjunction with any GP IIb/IIIa inhibitor. The Angio-Seal femoral closure device was also employed as part of the treatment strategy. We administered 0.5 mg/kg IV enoxaparin and a GP IIIb/IIIa inhibitor to 75 eligible PCI patients. None received anticoagulation 24 hours prior to PCI; all received pre-procedural aspirin, post-procedural deployment of the Angio-Seal and clopidogrel therapy, and were discharged home within 36 hours. TIMI minor bleeding was 1.3%; there were no TIMI major bleeding events or major adverse cardiac events during in-hospital stay or at 30-day follow-up. Our small observational study shows that IV enoxaparin is safe and efficacious during PCI when given at a dose 33% lower than previously reported in conjunction with any GP IIb/IIIa inhibitor and Angio-Seal. However, large, randomized PCI trials are needed to confirm the clinical efficacy, safety and cost-effectiveness of lower doses of enoxaparin with GPIIb/IIIa inhibitors and vascular closure devices.     

Influence of Relative Displacement on Surface Roughness in Longitudinal Turning of X37CrMoV5-1 Steel

The shaping process of surface texture is complicated and depends on many factors and phenomena accompanying them. This article presents the author’s test stand for the measurement of relative displacements in a tool–workpiece system during longitudinal turning. The aim of this study was to determine the influence of edge radius on the relative displacement between the tool and workpiece. The cutting process was carried out with inserts with different edge radii for X37CrMoV5-1 steel. As a result of the research, vibration charts of the tool–workpiece system were obtained. In the range of feed 0.03–0.18 mm/rev, the values of the standard deviation of relative displacements in the x-axis were obtained in the range of 0.36–0.78 μm for the insert with an edge radius of r_n = 48.8 μm. As a result of the work, it was determined that for the feed value of 0.12 mm/rev for all inserts, the relative displacements are the smallest. As the final effect, the formula for forecasting the Ra roughness parameter was presented.     

Novel targets for the pharmacotherapy of diarrhoea: A view for the millennium

Abstract Acute diarrhoea continues to carry a high morbidity and mortality worldwide. Intestinal infection is the major cause of acute diarrhoea although the prevalence of individual pathogens varies according to geographic location. In many countries in the industrialized world, reports of intestinal infections continue to increase; these are largely related to waterborne and foodborne outbreaks. Acute diarrhoea may be due to increased intestinal secretion, commonly as a result of infection with enterotoxin-producing organisms (enterotoxigenic Escherichia coli , Vibrio cholerae ) or to decreased intestinal absorption from infection with organisms that damage the intestinal epithelium (enteropathogenic E. coli , Shigella sp., Salmonella sp.). Although oral rehydration therapy has reduced the mortality associated with acute diarrhoea, the diarrhoea attack rate remains unchanged and stool volume often increases during the rehydration process. The search for agents that will directly inhibit intestinal secretory mechanisms and thereby reduce stool volume has been going on for more than 20 years. Research during the past decade has highlighted the importance of neurohumoral mechanisms in the pathogenesis of diarrhoea, notably the role of 5-hydroxtryptamine, substance P, vasoactive intestinal polypeptide and neural reflexes within the enteric nervous system. Cholera toxin, E. coli enterotoxins and Clostridium difficile toxin A are known to invoke these mechanisms in diarrhoea pathogenesis. This new dimension of intestinal pathophysiology has already exposed possible novel targets for anti-secretory therapy, namely, 5-HT receptor antagonists, substance P antagonists and the possibility for potentiating the proabsorptive effects of endogenous enkephalins by use of enkephalinase inhibitors. There now seems to be a real possibility that anti-secretory therapy will become more widely available in the future.     

Diverging associations of an intended early invasive strategy compared with actual revascularization, and outcome in patients with non-ST-segment elevation acute coronary syndrome: the problem of treatment selection bias

AIMS: In several observational studies, revascularization is associated with substantial reduction in mortality in patients with non-ST-segment elevation acute coronary syndrome (nSTE-ACS). This has strengthened the belief that routine early angiography would lead to a reduction in mortality. We investigated the association between actual in-hospital revascularization and long-term outcome in patients with nSTE-ACS included in the ICTUS trial. METHODS AND RESULTS: The study population of the present analysis consists of ICTUS participants who were discharged alive after initial hospitalization. The ICTUS trial was a randomized, controlled trial in which 1200 patients were randomized to an early invasive or selective invasive strategy. The endpoints were death from hospital discharge until 4 year follow-up and death or spontaneous myocardial infarction (MI) until 3 years. Among 1189 patients discharged alive, 691 (58%) underwent revascularization during initial hospitalization. In multivariable Cox regression analyses, in-hospital revascularization was independently associated with a reduction in 4 year mortality and 3 year event rate of death or spontaneous MI: hazard ratio (HR) 0.59 [95% confidence interval (CI) 0.37-0.96] and 0.46 (95% CI 0.31-0.68). However, when intention-to-treat analysis was performed, no differences in cumulative event rates were observed between the early invasive and selective invasive strategies: HR 1.10 (95% CI 0.70-1.74) for death and 1.27 (95% CI 0.88-1.85) for death or spontaneous MI. CONCLUSION: The ICTUS trial did not show that an early invasive strategy resulted in a better outcome than a selective invasive strategy in patients with nSTE-ACS. However, similar to retrospective analyses from observational studies, actual revascularization was associated with lower mortality and fewer MI. Whether an early invasive strategy leads to a better outcome than a selective invasive strategy cannot be inferred from the observation that revascularized patients have a better prognosis in non-randomized studies.     

Long-Term Outcome and Quality of Life After Continent Ileostomy

Introduction This study was designed to evaluate long-term outcomes for patients undergoing Kock continent ileostomy, identify factors associated with adverse outcomes, and compare changes in quality of life after removal of the reservoir. Methods The records of all patients (n = 330) undergoing continent ileostomy at the Cleveland Clinic Foundation between 1974 and 2001 were reviewed. Patient-related, intraoperative, and postoperative factors were evaluated as predictor variables of long-term pouch survival. Quality of life was evaluated using the continent ileostomy surgery follow-up questionnaire and the Cleveland Global Quality of Life scale (n = 216). These were compared between patients with continent ileostomy (n = 181) and patients who underwent removal of the continent ileostomy and conversion to an end stoma (n = 35). Results The median patient follow-up was 11 (range, 1–27) years. The median revision-free pouch interval was 14 (95 percent confidence interval, 11–17) months. The 10-year and 20-year pouch survival was 87 and77 percent, respectively. Patients had an average of 3.7(range, 1–28) complications and 2.9 (range, 1–27) pouch revisions during follow-up. On multivariate analysis, Crohn's disease (hazard ratio = 4.5), female gender (hazard ratio = 2.4), fistula development (hazard ratio = 3), and body mass index (hazard ratio = 2.4 per 5 unit increase) were independent predictors of pouch failure. Quality of life measurements for patients with a continent ileostomy were higher on all scales in comparison with patients who had the Kock reservoir and then reverted to a Brooke ileostomy. Conclusions Despite the associated morbidity with continent ileostomy surgery, long-term results and quality of life were encouraging. Continent ileostomy may be offered as an attractive long-term option to select patients whose only alternative is an end ileostomy. Poster presentation at The Tripartite Colorectal Meeting, Dublin, Ireland, July 5 to 7, 2005.