Diagnostic value of D-dimer in outpatients with suspected deep venous thrombosis receiving oral anticoagulation.

D-dimer has proved a useful diagnostic tool for the exclusion of deep venous thrombosis (DVT). The objective of this paper was to evaluate the diagnostic performance of a diagnostic algorithm combining clinical probability and D-dimer in outpatients receiving oral anticoagulant treatment (OAT) similar to those regularly applied to nonanticoagulated individuals. We enrolled 70 outpatients on OAT who presented with clinically suspected DVT; a standard diagnostic algorithm including clinical evaluation using the modified Wells score and a quantitative immunoturbidimetric D-dimer assay (STA Liatest D-Di; Diagnostica Stago, Asniéres sur Seine, France) was used. A 3-month follow-up period was applied for those patients in whom DVT was initially excluded. The prevalence of DVT was 18.5% (13/70); four of the diagnoses were made during the 3-month follow-up period. The sensitivity, specificity and negative predictive value of D-dimer were 69.2% (95 confidence interval, 42.4-87.3), 47.4% (95% confidence interval, 35.0-60.1) and 87.1% (95% confidence interval, 71.1-94.9), respectively. In conclusion, D-dimer is of limited value in outpatients on OAT presenting with clinically suspected DVT and should be omitted in such individuals; these patients should always undergo compression venous ultrasound, and repeat ultrasonography within 1 week might be warranted in cases with an initial negative examination.     

Gastro-duodenal digestion products of the major peanut allergen Ara h 1 retain an allergenic potential

BACKGROUND: The process of gastro-duodenal digestion may play a role in determining the allergenic properties of food proteins. The sensitizing and allergenic potential of digestion products of highly degraded allergens, such as the major peanut allergen Ara h 1, is currently under debate. We evaluated the effect of in vitro gastro-duodenal digestion of Ara h 1 on T cell reactivity and basophil histamine release. METHODS: An in vitro model of gastro-duodenal digestion was used to investigate changes in the allergenic properties of Ara h 1 using in vitro assays monitoring T cell reactivity (proliferation, cytokine production) and histamine release of basophils from peanut allergic individuals. The digestion process was monitored using an SDS-PAGE gel. RESULTS: In vitro gastric digestion led to rapid degradation of Ara h 1 into small fragments M(r) L5600. Gastric digestion did not affect the ability of Ara h 1 to stimulate cellular proliferation. Gastro-duodenal digestion significantly reduced its ability to stimulate clonal expansion (P<0,05; Wilxocon's signed rank test). The Th-2 type cytokine polarization of T cells from peanut allergic donors (IFN-gamma/IL-13 ratio and IFN-gamma/IL-4 ratio of CFSE(low) CD4(+) T cells) remained unchanged regardless of the level of digestion. Histamine release of basophils from peanut allergic individuals was induced to the same extent by native Ara h 1 and its digestion products. CONCLUSION: Gastro-duodenal digestion fragments of Ara h 1 retain T cell stimulatory and IgE-binding and cross-linking properties of the intact protein.     

Phenotype traits associated with different alleles at the RPS5 locus in Saccharomyces cerevisiae

Summary The RPS5 gene has been characterised through its ability to reduce invertase production by the SUC5 gene. In this paper we show that RPS5 acts by maintaining low levels of SUC5 mRNA. We also show that RPS5 acts on the SUC1 and SUC4 genes but not on SUC2 and SUC3, which are members of the SUC family. RPS5 also shows a pleiotropic effect on the amount of mitochondrial cytochromes.     

Immunohistochemical detection of VEGF in the bone marrow of patients with acute myeloid leukemia. Correlation between VEGF expression and the FAB category

We studied vascular endothelial growth factor (VEGF) expression in bone marrow sections obtained from 3 healthy donors and 41 patients with acute myeloid leukemia (AML) of various French-American-British (FAB) subtypes by immunohistochemical analysis using an anti-VEGF antibody. In normal bone marrow, the anti-VEGF antibody reacted with myeloid progenitor cells and megakaryocytes but not with erythroid cells or mature granulocytic cells. High levels of VEGF were found in the bone marrow in patients with AML-M1, -M2, -M3, -M4, -M4Eo, and -M5. In these leukemias, the vast majority of myeloblasts (> 90%) expressed VEGF. By contrast, in AML-M0, the percentage of VEGF-positive blasts was lower in most cases (median, 42%), and if at all detectable, these blast cells contained only trace amounts of VEGF. In AML-M3 and -M4Eo, maturing granulocytes failed to express VEGF similar to granulocytes in normal bone marrow. In AML-M6, myeloblasts exhibited VEGF, whereas erythroid cells did not. In AML-M7, blast cells and megakaryocytes were identified as major sources of VEGF. In summary, VEGF expression in the bone marrow is restricted to certain stages of differentiation and maturation of myeloid cells and correlates with the FAB category.     

IFN-gamma-enhanced allergen penetration across respiratory epithelium augments allergic inflammation

BACKGROUND: Respiratory allergen contact is the critical event in the elicitation and boosting of allergen-specific immune responses, as well as in the induction of immediate and late inflammatory reactions. OBJECTIVE: We sought to investigate the influence of various factors of allergic inflammation on the integrity and barrier function of respiratory epithelium for allergens. METHODS: We cultured the human bronchial epithelial cell line 16HBE14o- in a transwell culture system as a surrogate of intact respiratory epithelium and used purified iodine 125-labeled recombinant major birch pollen allergen (rBet v 1) to study the extent, kinetics, and factors influencing transepithelial allergen penetration. RESULTS: Culture supernatants from activated allergen-specific T H 1 clones decreased transepithelial resistance. A screening of various factors (histamine, IFN-gamma, IL-1beta, IL-2, IL-3, IL-4, IL-5, IL-8, IL-12, and TNF-alpha) identified IFN-gamma as a potent factor capable of reducing epithelial barrier properties and enhancing transepithelial allergen penetration. Increased submucosal allergen concentrations caused by IFN-gamma-mediated reduction of epithelial barrier function provoked a more than 7-fold augmentation of histamine release from sensitized basophils. CONCLUSION: These results demonstrate that the T H 1 cell-derived cytokine IFN-gamma facilitates allergen penetration through the respiratory epithelium and thereby can aggravate allergic inflammation.     

Recombinant allergens promote expression of CD203c on basophils in sensitized individuals

BACKGROUND: Traditionally, the diagnosis of type I allergies is based on clinical data, skin test results, and laboratory test results with allergen extracts. During the past few years, several attempts have been made to refine diagnostic assays in clinical allergy by introducing recombinant allergens and novel markers of IgE-dependent cell activation. OBJECTIVES: We have identified the ectoenzyme CD203c as a novel basophil antigen that is upregulated on IgE receptor cross-linkage. In this study we applied CD203c and a panel of recombinant allergens to establish a novel basophil test that allows for a reliable quantification of IgE-dependent responses at the effector cell level. METHODS: Patients allergic to birch (Bet v 1, n = 15; Bet v 2, n = 8) and grass (Phl p 1, n = 15; Phl p 2, n = 10; Phl p 5, n = 14) pollen allergens, as well as 10 nonallergic donors, were examined. Basophils were exposed to various concentrations of recombinant allergens for 15 minutes and then examined for expression of CD203c by means of flow cytometry. CD203c upregulation was correlated with the increase in CD63. RESULTS: Exposure to recombinant allergens resulted in a dose-dependent increase in expression of CD203c on peripheral blood basophils in sensitized individuals, whereas no increase was seen in healthy control subjects. The effects of the recombinant allergens on CD203c expression were also time dependent. There was a good correlation between allergen-induced upregulation of CD203c and upregulation of CD63 (R = 0.76). CONCLUSION: Flow cytometric quantitation of CD203c on blood basophils exposed to recombinant allergens is a useful approach to determine the allergic state in sensitized individuals and represents a basis for a sensitive novel allergy test.     

Telomere length analysis on cord blood cells by the flow-FISH method

Telomerase is the enzyme responsible for synthesizing telomeric repeats at the ends of chromosomes to maintain telomere length. Recent studies have suggested that telomere shortening may serve as a surrogate marker of the progression of malignant disorders and seems to be accelerated in allogeneic bone marrow transplant recipients. In this study, the results of the telomere length of nine cord blood mononuclear cell samples are presented. Telomere length was measured by the flow-FISH method, using a peptide nucleic acid probe. The proportion of cord blood cell subsets (CD19/CD34/CD3) was also evaluated. The telomere length of the internal control 1301 cell line was estimated to be 100%. The mean telomere length of cord blood cells was 18.5 +/- 3.9%, compared with the internal control. The progenitor CD34+ cells were detected as 2.6 +/- 0.7% in the lymphoid gate measured. Linear correlation analysis did not find any connection between the cell subsets (CD3+, CD34+, CD19+) and the telomere length. The findings confirm that the telomere flow-FISH method is sufficient for estimation of the telomere length. Assessment of the current procedures of collection, manipulation, and ex vivo expansion of cord blood cells in terms of their effect on telomere shortening might be important.     

Increase of bone marrow lymphocytes in systemic mastocytosis: reactive lymphocytosis or malignant lymphoma? Immunohistochemical and molecular findings on routinely processed bone marrow biopsy specimens

AIMS: To clarify the nature (reactive or neoplastic) of lesional, perifocally aggregated lymphocytes in bone marrow infiltrates of systemic mastocytosis (SM), the histopathology of which can resemble malignant lymphoma with focal bone marrow involvement, particularly low grade malignant B cell lymphoma of lymphoplasmacytic immunocytoma subtype, which frequently exhibits increased mast cell (MC) numbers. METHODS: Thirteen cases of SM and three of lymphoplasmacytic immunocytoma with predominant focal bone marrow infiltration were investigated. Immunostaining of formalin fixed, paraffin wax embedded bone marrow specimens was performed using antibodies against CD2, CD5, CD20, CD23, and CD25; kappa and lambda immunoglobulin light chains; and MC markers chymase, tryptase, and CD117 (KIT). Monoclonal rearrangements of IgH and TCRgamma were studied using seminested polymerase chain reaction (PCR). c-kit point mutation Asp816-Val was detected by PNA mediated PCR clamping and hybridisation probes. RESULTS: The lymphocytic clusters in SM contained nearly equal numbers of mature T and B cells, the latter with no coexpression of aberrant antigens, such as CD5 or CD23. Most MCs in SM cases constantly coexpressed tryptase, CD25, and CD117. No monoclonal rearrangements were seen for IgH or TCRgamma. In contrast, B cells from immunocytomas showed light chain restriction and monoclonal rearrangement for IgH, confirming their neoplastic nature. c-kit point mutation Asp816-Val was found in ten of 13 SM cases, but in none of the three immunocytomas. CONCLUSIONS: Focal accumulations of lymphocytes in the bone marrow of SM are reactive in nature and could be termed lymphocytosis. A diagnosis of SM-AHNMD/immunocytoma should not be made.