Modulation of CXC chemokine receptor expression and function in human neutrophils during aging in vitro suggests a role in their clearance from circulation.

In mice, differential regulation of CXC chemokine receptor expression in circulating polymorphonuclear neutrophils (PMNs) undergoing senescence results in homing to the bone marrow. However, the role of this compartment and of the chemokine receptor CXCR4 is still under discussion, and only scarce data exist about CXCR4 function in human PMN. In our study, we provide evidence that also in human neutrophils, expression (cell surface and mRNA), chemotactic and signaling functions of the homing-related chemokine receptor CXCR4 are upregulated during aging in vitro, independent of addition of stimulatory cytokines (TNF, IL-1, IL-8, G-CSF).

The Zygosaccharomyces bailii antifungal virus toxin zygocin: cloning and expression in a heterologous fungal host.

Zygocin, a monomeric protein toxin secreted by a virus-infected killer strain of the osmotolerant spoilage yeast Zygosaccharomyces bailii, kills a broad spectrum of human and phytopathogenic yeasts and filamentous fungi by disrupting cytoplasmic membrane function. The toxin is encoded by a double-stranded (ds)RNA killer virus (ZbV-M, for Z. bailii virus M) that stably persists within the yeast cell cytosol.

Transendothelial migration of hematopoietic progenitor cells. Role of chemotactic factors.

There is increasing evidence that hematopoietic stem cell mobilization and homing is regulated not only by adhesion molecules and cytokines, but also by chemotactic factors that support transendothelial migration across the bone marrow sinusoidal endothelium. Many receptors for chemotactic mediators belong to the family of G protein-coupled seven-transmembrane receptors (7-TMR). Signaling via G proteins, particularly Gi proteins, results in a chemotactic response of the cells towards a gradient of the corresponding ligand.

Chemotaxis and transendothelial migration of CD34(+) hematopoietic progenitor cells induced by the inflammatory mediator leukotriene D4 are mediated by the 7-transmembrane receptor CysLT1.

Recent studies suggest that bone marrow (BM)-derived chemotactic mediators such as chemokines play key roles in hematopoietic stem cell trafficking. Lipid mediators, particularly leukotrienes, are involved in leukocyte chemotaxis during inflammation but have also been detected in the normal BM. Therefore, the effects of leukotrienes on hematopoietic progenitor cells were analyzed.

Functional response of leukaemic blasts to stromal cell-derived factor-1 correlates with preferential expression of the chemokine receptor CXCR4 in acute myelomonocytic and lymphoblastic leukaemia.

The chemokine stromal cell-derived factor-1 (SDF-1) that is released by bone marrow (BM) stromal cells and contributes to stem cell homing may also play a role in the trafficking of leukaemic cells. We analysed SDF-1-induced intracellular calcium fluxes in leukaemic blasts from the peripheral blood of patients with newly diagnosed acute myeloid leukaemia (AML) and lymphoblastic leukaemia (B-lineage ALL), determined the effect of BM stromal cell-conditioned medium on in vitro transendothelial migration (TM) and measured expression of the SDF-1 receptor, CXCR4, by flow cytometry. AML FAB M1/2 blasts did not show calcium fluxes and TM was not stimulated.

Expression and secretion of vascular endothelial growth factor-A by cytokine-stimulated hematopoietic progenitor cells. Possible role in the hematopoietic microenvironment.

In the hematopoietic microenvironment, bone marrow endothelial cells may play an important role in trafficking and maintenance of progenitor and stem cells due to adhesive interactions and paracrine secretion of hematopoietic growth factors. However, it is unknown whether progenitors in turn modulate endothelial proliferation and function. We analyzed mRNA expression (Northern blot) and release of vascular endothelial growth factor-A (VEGF-A), which specifically acts on endothelial cells, by cytokine-stimulated peripheral blood-derived CD34+ hematopoietic progenitor cells.

Identification of an alpha-helical epitope region on the PM/Scl-100 autoantigen with structural homology to a region on the heterochromatin p25beta autoantigen using immobilized overlapping synthetic peptides.

The polymyositis-scleroderma overlap syndrome (PM/Scl) autoantigen is a nucleolar multiprotein particle, presumably participating in the maturation of 5.8S rRNAs. The major target antigens of this particle are two polypeptides with apparent molecular masses of 100 and 75 kDa.

Overexpression of the chemokine receptor CXCR4 in B cell chronic lymphocytic leukemia is associated with increased functional response to stromal cell-derived factor-1 (SDF-1).

The chemokine receptor CXCR4 and its ligand stromal cell-derived factor-1 (SDF-1) play an important role in trafficking of normal lymphocytes, monocytes, as well as hematopoietic stem- and progenitor cells. SDF-1 constitutively produced by bone marrow stromal cells acts as a chemoattractant supporting the homing of stem cells and may also contribute to the tropism of malignant cells for the bone marrow. Low-grade lymphoproliferative disorders, particularly B cell chronic lymphocytic leukemia (B-CLL), are characterized by the presence of bone marrow infiltration.

Expression of vascular endothelial growth factor (VEGF) and its receptors in human neuroblastoma.

Angiogenic factors may play a role in the biology of neuroblastoma, a well vascularised tumour, which frequently spreads haematogenously. Therefore, we analysed expression of vascular endothelial growth factor (VEGF) in six human neuroblastoma cell lines and five primary neuroblastomas. High VEGF levels (1-3 ng/10(6) cells/day) were found in the supernatant of all cell lines examined (SK-N-LO, SK-N-SH, LS, SH-SY5Y, IMR-32, Kelly).

Regulation of transendothelial migration of hematopoietic progenitor cells.

Transendothelial migration of hematopoietic progenitor cells occurs in the bone marrow during mobilization and homing, and may therefore play a key role in the trafficking of hematopoietic stem cells. We hypothesize that adhesion molecules, chemokines, and paracrine cytokines are involved in this multifactorial process. As suggested in several studies, downregulation of adhesion molecules (e.

Identification of major linear epitopes on the sp100 nuclear PBC autoantigen by the gene-fragment phage-display technology.

Approximately 20-30% of sera from patients suffering from primary biliary cirrhosis contain autoantibodies against a nuclear protein termed sp100. By indirect cytoimmunofluorescence it was shown that the sp100 autoantigen is distributed in up to 20 dot-like structures per nucleus co-localizing with the so-called nuclear bodies. In western blots these sera react with a protein with an apparent molecular mass of 100kDa.

The chemokine receptor CXCR-4 is expressed on CD34+ hematopoietic progenitors and leukemic cells and mediates transendothelial migration induced by stromal cell-derived factor-1.

The chemokine stromal cell-derived factor-1 (SDF-1) and its receptor CXCR-4 (fusin, LESTR) are likely to be involved in the trafficking of hematopoietic progenitor and stem cells, as suggested by the reduced bone marrow hematopoiesis in SDF-1-deficient mice and the chemotactic effect of SDF-1 on CD34+ progenitor cells. Migration of leukemic cells might also depend on the expression of chemokine receptors. Therefore, we analyzed expression of CXCR-4 on mobilized normal CD34+ progenitors and leukemic cells.

Antibodies against a peptide sequence located in the linker region of the HMG-1/2 box domains in sera from patients with juvenile rheumatoid arthritis.

Objective: To extend our work on the mapping of B cell epitopes on nucleosomal high mobility group (HMG) proteins in the sera of patients with juvenile rheumatoid arthritis (JRA).

Methods: Seventy-seven pauciarticular-onset JRA serum samples from antinuclear antibody (ANA)-positive patients and 42 polyarticular-onset JRA patient sera found to react with HMG-2 by immunoblotting were used in this study. To identify B cell epitopes on HMG-2, recombinant HMG-2 protein fragments were used in enzyme-linked immunosorbent assay (ELISA) and in competition ELISA experiments with a set of overlapping synthetic peptides.

Mapping of epitopes recognized by PM/Scl autoantibodies with gene-fragment phage display libraries.

Sera from patients suffering from the polymyositis/scleroderma overlap syndrome (PM/Scl) recognize two antigenically non-related proteins with apparent molecular masses of 100 kDa and 75 kDa respectively. The two proteins are part of a particle termed PM/Scl localized in the granular component of the nucleolus. The predominant immunoreactivity of the PM/Scl sera was shown to be directed against the 100 kDa protein.

A member of the mouse LRR transcript family with homology to the human Sp100 gene.

A previously isolated cDNA sequence with homology to the long-range repeat (LRR) cluster in chromosome 1 of the house mouse, Mus musculus, was identified as derived from a 1.3 kb polyadenylated RNA. This transcript belongs to a family of polyadenylated RNAs which are synthesized from a multicopy gene included in the LRR copies.

Sera from JRA patients contain antibodies against a defined epitope in chromosomal protein HMG-17.

Autoantibodies against the nonhistone nucleosomal protein HMG-17 have been detected in a high percentage of ANA-positive patients with pauciarticular-onset JRA4. Here we report on the epitope mapping of the HMG-17 autoantigen with a set of overlapping and nested synthetic peptides spanning the entire amino acid sequence of the human HMG-17 protein. Competition ELISA experiments defined a proline and lysine rich octapeptide PKPEPKPK as the major epitope recognized by more than 70% of the HMG-17 positive JRA sera.

Antineutrophil cytoplasmic autoantibodies (ANCA) recognizing a recombinant myeloperoxidase subunit.

Sera from 76 patients with diverse vasculitis, systemic lupus erythematosus (SLE) and hydralazine-induced lupus (HiL) were analysed by ELISA for their reactivity with native, reduced or urea-denatured MPO, respectively, as well as with bacterially expressed heavy and light subunits of MPO. All sera (n = 20) recognizing native MPO showed a positive reaction with reduced MPO, while 12 recognized the denatured protein. Most of the linear epitopes are located in the light subunit, since 9 MPO-positive sera recognized significantly the bacterially expressed, denatured light subunit, while only one serum recognized the bacterially expressed heavy subunit purified under denaturing conditions.

Cloning and characterization of the cDNA coding for a polymyositis-scleroderma overlap syndrome-related nucleolar 100-kD protein.

About 50% of patients with the polymyositis-scleroderma overlap syndrome are reported to have autoantibodies to a nucleolar particle termed PM/Scl. The particle consists of several polypeptides of which two proteins of 75 and 100 kD have been identified as the major antigenic components. Here we report on the cDNA cloning and partial epitope mapping of the 100-kD autoantigen from human placenta and HeLa lambda gt11 libraries.

Autoantibodies to the chromosomal protein HMG-17 in juvenile rheumatoid arthritis.

Objective: To determine the antibody profiles in sera from patients with juvenile rheumatoid arthritis (JRA).

Methods: Immunoblotting using nuclear extracts and recombinant high-mobility group (HMG) nonhistone chromosomal proteins.

Results: Antibodies directed against HMG-17 were found in 47% of antinuclear antibody (ANA)-positive patients with pauciarticular-onset JRA and in 16% of ANA-positive patients with polyarticular-onset JRA.

Autoantibodies to nonhistone chromosomal proteins HMG-1 and HMG-2 in sera of patients with juvenile rheumatoid arthritis.

IgG antibodies against the high mobility group (HMG) nonhistone chromosomal proteins HMG-1 and/or HMG-2 were detected in the sera of 49 (39%) of 126 antinuclear antibody (ANA)-positive patients with juvenile rheumatoid arthritis (JRA), by immunoblotting. Clinical diagnosis classified these patients in 2 major groups, 105 with pauciarticular-onset JRA and 21 with polyarticular-onset JRA. Anti-HMG-1 and/or anti-HMG-2 antibodies were found in 8 (25%) of 32 pauciarticular-onset JRA patients with uveitis and in 34 (47%) of 73 patients without uveitis, whereas anti-HMG-1 and/or anti-HMG-2 antibodies were found in 4 (24%) of 17 children with polyarticular-onset JRA without uveitis.

A monoclonal antibody directed against the head region of vimentin.

The production and identification of a monoclonal antibody directed against an epitope in the aminoterminal head region of vimentin is described. Enzyme-linked immunosorbent assay, protein blotting and indirect immunofluorescence were used. The wide range of cross-reactivity within cytoskeletal proteins observed for this antibody gives evidence for a determinant in an evolutionarily conserved region.

Autoimmune sera recognize a 100 kD nuclear protein antigen (sp-100).

Autoimmune sera from patients suffering from undifferentiated connective tissue diseases (UCTD), Sjögren's syndrome (SS), primary biliary cirrhosis (PBC) and other disorders were found to contain antibodies that produce a distinctive nuclear spot pattern with HEp-2 cells in immunofluorescence studies. These spots which vary in size and number, are spread over the whole nucleus with the exception of the nucleoli. This pattern is easily distinguishable from the staining patterns of anti-centromere, anti-RNP, anti-nucleolar and anti-Scl-70 antibodies.

[Nuclear antibodies as serologic markers in progressive systemic scleroderma].

In all, 36 patients with progressive systemic sclerosis (29 women, 7 men) were studied clinically and immunologically; 15 patients had acrosclerosis (type I) and 21, sclerosis extending beyond the wrist (type II). The sera of all patients were evaluated for ANA (HEp-2-cells), Scl-70, centromere and other ENA antibodies. The centromere antigen was characterized by immunoblotting.

Anti-(U1)RNP and anti-Sm autoantibody profiles in patients with systemic rheumatic diseases: differential detection of immunoglobulin G and M by immunoblotting.

Autoimmune sera from patients with systemic lupus erythematosus, scleroderma, or both disorders reactive with the (U1)RNP and Sm antigens were analyzed according to their IgG- and IgM-autoantibody profiles by immunoblotting with HeLa nuclear extracts. Anti-(U1)RNP-specific autoantibodies directed against the 68-kDa polypeptide were found to be predominantly of the IgG type, whereas for the other (U1)RNP-specific protein, 33 kDa, a concomitant occurrence of IgG and IgM class autoantibodies was observed for most patients. In contrast, Sm-specific anti-29/28-kDa autoantibodies were found to be more frequently of the IgM than of the IgG type, while Sm-specific anti-16-kDa antibodies of both classes were present simultaneously in most sera.

Scl 70 autoantibodies from scleroderma patients recognize a 95 kDa protein identified as DNA topoisomerase I.

Sera of patients suffering from the autoimmune disease progressive systemic sclerosis (PSS) are known to contain autoantibodies which have been reported to recognize a 70 kDa antigenic protein, designated the Scl 70 antigen. By immunoblotting of nuclear extracts from HeLa cells with sera from scleroderma patients we observed that the size of the antigen present in such cells depends on the conditions of antigen isolation. When protease inhibitors were included in the extraction buffer, a 95 kDa protein was identified instead of a 70 kDa protein.