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Biomedical subjects

K Beiske

Publications and source records attributed to K Beiske.

At least 19 recordsLinked to original sources
PubMed2006

Polyclonal expansion of large granular lymphocytes in common variable immunodeficiency - association with neutropenia.

Common variable immunodeficiency (CVID) is the most frequent symptomatic primary immunodeficiency disease, characterized by low levels of circulating immunoglobulins and recurrent bacterial infections, particularly of the respiratory tract. T cell dysfunction is often present, and lymphoproliferative and autoimmune disorders as well as haematological cytopenias are frequently observed. In this study, we report a polyclonal expansion of large granular lymphocytes (LGL) in a substantial proportion of CVID patients, associated with splenomegaly, increased numbers of CD8(+) T cells, inverted CD4 : CD8 T cell ratios and neutropenia. CVID patients who had both increased numbers of LGL and granulocytopenia had elevated levels of soluble Fas ligand (sFasL). Our observations indicate that CVID may be added to the list of inflammatory diseases associated with increased numbers of LGL. Furthermore, our findings suggest common pathogenic mechanisms of granulocytopenia in CVID and lymphoproliferative disease of granular lymphocytes.

Adult↗
PubMed2001

Plasmacytoid dendritic cells (natural interferon- alpha/beta-producing cells) accumulate in cutaneous lupus erythematosus lesions.

Plasmacytoid dendritic cell (P-DC) precursors in peripheral blood produce large amounts of interferon (IFN)-alpha/beta when triggered by viruses. However, when incubated with interleukin-3 and CD40 ligand, the same precursors differentiate into mature DCs that stimulate naïve CD4(+) T cells to produce Th2 cytokines. We recently reported that P-DCs accumulate in nasal mucosa of experimentally induced allergic rhinitis, supporting a role for this DC subset in Th2-dominated inflammation. Here we examined whether P-DCs accumulate in cutaneous lesions of lupus erythematosus (LE), a disorder associated with increased IFN-alpha/beta production. Our results showed that P-DCs were present in 14 out of 15 tissue specimens of cutaneous LE lesions, but not in normal skin. Importantly, the density of P-DCs in affected skin correlated well (r(s) = 0.79,P < 0.0005) with the high number of cells expressing the IFN-alpha/beta-inducible protein MxA, suggesting that P-DCs produce IFN-alpha/beta locally. Accumulation of P-DCs coincided also with the expression of L-selectin ligand peripheral lymph node addressin on dermal vascular endothelium, adding further support to the notion that these adhesion molecules are important in P-DC extravasation to peripheral tissue sites. Together, our findings suggested that P-DCs are an important source of IFN-alpha/beta in cutaneous LE lesions and may therefore be of pathogenic importance.

Antigens, Surface↗
PubMed2000

A splice variant of human ephrin-A4 encodes a soluble molecule that is secreted by activated human B lymphocytes.

Ephrin-A4 is a ligand for the erythropoietin-producing hepatocellular (Eph) receptor family of tyrosine kinases. We have identified a secreted form of ephrin-A4, denoted ephrin-A4 (s), which is encoded by an alternatively spliced mRNA and is produced by in vivo activated B cells in tonsils. Blood B cells secrete ephrin-A4 (s) upon stimulation via the B-cell antigen receptor. A subpopulation of tonsil cells in the crypts with a dendritic cell phenotype was shown to express EphA2, an Eph receptor tyrosine kinase that was found to be capable of binding an ephrin-A4 immunoglobulin chimeric protein. We conclude that ephrin-A4 (s) may play a role in the interaction between activated B lymphocytes and dendritic cells in human tonsils. (Blood. 2000;95:221-230)

Alternative Splicing↗
PubMed2000

Co-induction of Mad1 and c-Myc in activated normal B lymphocytes.

The purpose of the present study was to examine the expression of the Myc network proteins c-Myc, Mad1 and Max in normal cells under different growth and differentiation conditions. A dominant view has been that Mad1 as a c-Myc antagonist plays a role in growth inhibition linked to differentiation. Of particular interest to us was therefore to study the regulation of Mad1 in cells undergoing differentiation in the absence of growth cessation. To do so we utilized normal B lymphocytes isolated from peripheral blood. The cells were induced to concomitant proliferation and differentiation by stimulation with a combination of anti-IgM antibodies (anti-mu) and the phorbol ester TPA. Thus, by 72 h of stimulation the percentage of plasmablasts increased from 3 to 17%, and the percentage of lymphocytes decreased from 89 to 27%. The most intriguing observation we made using this cell system was a pronounced coinduction of Mad1 and c-Myc. The levels of c-Myc and Mad1 mRNAs and proteins increased within 3 h of anti-mu stimulation, and the levels were further enhanced by TPA. Furthermore, the expressions of both c-Myc and Mad1 were reduced by forskolin, which also inhibited the anti-mu + TPA driven growth and differentiation of the B lymphocytes. The level of Max remained virtually unchanged. Taken together, our results indicate that a high level of Mad1 in normal human B cells is linked to differentiation and not to growth inhibition. Furthermore, the results demonstrate that Mad1 and c-Myc are not necessarily expressed in a reciprocal manner, which underlines an independent role of Mad1 unrelated to its function as a c-Myc antagonist.

B-Lymphocytes↗
PubMed1999

High-dose therapy supported with immunomagnetic purged autologous bone marrow in high-grade B cell non-Hodgkin's lymphoma.

From August 1987 to March 1995, 25 patients with high-grade B cell non-Hodgkin's lymphoma (NHL) were treated with high-dose therapy (HDT) followed by bone marrow purged with immunomagnetic beads. At the time of transplantation, 20 patients were in sensitive relapse and five in first complete or partial remission. Ten patients had secondary high-grade NHL transformed from low-grade NHL. The HDT consisted of TBI followed by high-dose cyclophosphamide. All patients engrafted, except for two patients with early treatment-related death. Eleven patients relapsed, of whom nine died of lymphoma, and two are alive in new CR. The estimated event-free and overall survivals at 5 years were 40% and 48%, respectively, with a median follow-up of 48 months (range 1-123). Eight of the tumours contained the translocation t(14;18) at the major breakpoint region (MBR) of BCL-2. In these patients the presence of tumour cells in the bone marrow graft before and after purging were assessed by PCR. Four of five patients infused with non-detectable minimal residual disease in their autografts are in complete remission, while two of three patients reinfused with t(14;18) positive cells after purging, experienced a fast and aggressive relapse. As found by others, our data suggest that reinfusion of tumour-free autografts obtained by efficient in vivo purging using chemotherapy before harvesting, and/or by in vitropurging of the stem cell products, influence the patients remission status after HDT.

Adolescent↗
PubMed1998

Increased sensitivity for detection of micrometastases in bone-marrow/peripheral-blood stem-cell products from breast-cancer patients by negative immunomagnetic separation.

Immunocytochemical detection (ICC) of isolated tumor cells in bone marrow (BM) is currently the most established method for monitoring early dissemination in epithelial cancer. However, the low sample size that can practically be analyzed restricts the sensitivity and reliability of the ICC method. To be able to analyze larger samples, a negative immunomagnetic separation (IMS) technique, utilising anti-CD45-conjugated Dynabeads, has been developed. Tumor-cell enrichment by depletion of CD45-expressing mononuclear cells (MNC) is followed by ICC for detection of the cytokeratin (CK)-positive (+) epithelial cells. In this study, bone-marrow samples (n = 165) and peripheral-blood-progenitor-cell (PBPC) apheresis products (n = 22) from breast-cancer patients were analyzed. The negative IMS analysis of 1 to 2 x 10(7) MNC was compared with ICC analysis of 2 x 10(6) unseparated MNC. Negative IMS resulted in 85% mean depletion of MNC. The results showed that 11.7% of the samples were positive by ICC analysis of unseparated MNC, as compared with 23.5% after negative IMS. In samples presenting > 10 CK+ cells, a 4-fold higher number of positive cells was detected by the negative IMS technique. Moreover, there was no evidence for general enrichment of false-positive cells. Altogether our results show that negative IMS is an efficient enrichment method for sensitive detection of CK+ cells in BM/PBPC products from breast-cancer patients. This opens the possibility for further characterization of micrometastatic tumor cells.

Bone Marrow Cells↗
PubMed1998

Immunocytochemical detection of isolated epithelial cells in bone marrow: non-specific staining and contribution by plasma cells directly reactive to alkaline phosphatase.

Detection of isolated tumour cells (TCs) in bone marrow (BM) from epithelial cancer patients by immunocytochemical (ICC) analysis seems to predict future relapse, but the reported percentages of positive BMs among patients with localized cancer show large variations and the number of detected TCs is low. This emphasizes the importance of thoroughly testing the methods in use. This study was performed to clarify to what extent positive staining of haematopoietic cells (HCs) interferes with the ICC detection of epithelial cells in BM. BM mononuclear cells (MNCs) from normal donors and stage I-II breast cancer patients were stained with anti-cytokeratin (CK) and isotype control monoclonal antibodies (MAbs) followed by alkaline phosphatase (AP)-based visualization of immunolabelled cells. In the ICC staining of normal donors by the anti-CK MAbs AE1/AE3 or A45-B/B3, rare immunoreactive cells were detected in 7/20 and 8/19 BMs, respectively. Morphological examination recognized all these cells as typical HCs. In the breast cancer patients (n = 257), anti-CK-positive cells were detected in 26.6 per cent, excluding cells with HC morphology. Using the same morphological criteria, isotype control-positive cells were detected in 5.4 per cent of patients. Some of the false-positive events were further analysed and cells with strong reactivity against the AP enzyme alone were detected. Double ICC staining recognized the majority of these AP directly-reactive cells as CD45-negative and human Ig kappa/lambda-positive, in accordance with the phenotype of mature plasma cells. Morphological evaluation and adequate controls are important to ensure the diagnostic specificity of micrometastases in BM. It is recommended that the number of BM MNCs included in negative controls should equal the number of cells in the diagnostic specimens.

Alkaline Phosphatase↗
PubMed1997

CDw78--a determinant on a major histocompatibility complex class II subpopulation that can be induced to associate with the cytoskeleton.

In the present study we demonstrate that CDw78 monoclonal antibody (mAb) recognizes a distinct subpopulation of major histocompatibility complex (MHC) class II molecules. We show that the CDw78 epitope is present on less than 10% of the total number of MHC class II molecules expressed on different cells, is not linked to a single isotype, and exhibits a characteristic expression pattern in tonsils. While mAb against MHC class II (DR, DP and DQ) stained the majority of cells both in the mantle zone and in germinal centers, the CDw78 staining was more heterogeneous with the strongest reactivity and the highest number of positive cells in the mantle zone and in the light centrocyte-rich part of the germinal centers. Antibodies to this MHC class II subpopulation (e.g. FN1) induced association with the cytoskeleton and a subsequent capping in more than 90% of peripheral blood B cells. In contrast, mAb against MHC class II (DR, DP and DQ) did not induce association with the cytoskeleton and only 10-20% of B cells were induced to cap, suggesting that CDw78 defines a population of MHC class II molecules functionally different from the majority of these antigens. Scatchard plot analysis indicates that FN1 mAb is of relatively low affinity (Ka = 1.5 x 10(8) M(-1)) and monovalent Fab fragments fail to bind to the cell surface with measurable affinity. Our data seen in the context of the ability of FN1 to co-stimulate B cells with a suboptimal dose of anti-mu suggest that CDw78 mAb might recognize a functional important subpopulation of MHC class II molecules so far not described. It seems likely that this subpopulation represents dimerized or aggregated MHC class II molecules that can selectively bind this low-affinity mAb.

Antibodies, Monoclonal↗
PubMed1997

Immunomagnetic techniques for the enrichment and detection of isolated breast carcinoma cells in bone marrow and peripheral blood.

Detection of isolated tumor cells (TC) in bone marrow (BM) from patients with breast cancer is usually accomplished by immunocytochemical (ICC) analysis of up to 2 X 10(6) mononuclear cells (MNC). However, this method is cumbersome if large numbers of BM cells (i.e. > 1 X 10(7) cells) are to be analyzed. This emphasizes the need for TC enrichment strategies. This report describes immunomagnetic separation (IMS) techniques for enrichment and detection of viable breast carcinoma cells in BM and peripheral blood (PB). The positive IMS technique was performed by incubation of MNC with 2.8 microns magnetic particles (rat antimouse IgG1 M280-Dynabeads) coated with monoclonal antibody (mAb) against epithelial surface antigens. The rosetted tumor cells were then visualized by ICC staining using alkaline phosphatase-conjugated A45-B/B3 anticytokeratin mAb (Fab). The negative IMS technique was performed by incubation of MNC with anti-CD45-coated M450-Dynabeads (4.5 microns), followed by ICC staining of the nonrosetted cells. When 1000, 100, and 10 breast carcinoma cells were mixed with 1 X 10(7) MNC, an average of 748 (n = 9), 70 (n = 10), and 7.8 TC (n = 8), respectively, were detected with the positive IMS technique. With the negative IMS technique, 648 (n = 8), 57.8 (n = 6), and 7.3 TC (n = 6), respectively, were detected. The analysis of 1 X 10(7) MNC with the IMS techniques was compared with the ICC analysis of 2 X 10(6) unseparated MNC. A mean 3.7-fold (range 1.5-6.4) to 4.2-fold (2.5-8.2) (positive IMS) and 3.1-fold (range 2.0-5.0) to 3.8-fold (2.0-6.0) (negative IMS) higher TC detection frequency was achieved after enrichment by IMS in experiments with 100 and 1000 TC/10(7) MNC. The IMS techniques were used for examination of BM samples from locally advanced breast cancer patients. A 5.3-fold mean increase (range 2.1-13.3) in the number of TC detected was obtained when the use of positive and negative IMS together was compared with the direct ICC analysis of unseparated MNC (n = 11). Enrichment of TC by IMS techniques enables us to examine large numbers of MNC from BM or PB, which can result in the detection and characterization of minimal residual disease with increased sensitivity and specificity.

Bone Marrow↗
PubMed1996

[High-dose therapy of cancer with CD34 positive cells as stem cell support].

In this article we report our initial clinical experiences in connection with immunomagnetic isolated CD34-positive cells from peripheral blood progenitor cells. Six patients, five with breast cancer and one with non-Hodgkin's lymphoma, were mobilized by chemotherapy and G-CSF (5ug/kg). CD34-positive cells were isolated by means of immunomagnetic beads (Dynalbeads) and Isolex 300 Cell Separator (Baxter, USA). Mean purity of isolated CD34-positive cells was 97% (94.7-99.7) and mean yield was 54% (35-68). Three patients were treated with high dose therapy followed by reinfusion of CD34-positive cells as stem cell support. Recovery of neutrophils (> 0.5 x 10(9) leucocytes/liter) occurred at day 8, 11 and 13 and of platelets (> 20 x 10(9) platelets/litre) at day 9,14 and 32. It is concluded that immunomagnetic isolated CD34-positive cells give high purity and yield. Although use of CD34-positive cells reduces the content of contaminating tumour cells in the graft, breast cancer cells were still detectable in two out of five CD34-positive cell products.

Antigens, CD34↗
PubMed1996

[Human T-cell lymphotrophic virus type I and T-cell lymphoma].

Human T-cell lymphotropic virus type I is an oncogenic retrovirus, endemic in Southwestern Japan, the Caribbean, some parts of Africa and Central and South America. The virus is etiologically associated with adult T-cell leukemia/lymphoma and a myelopathy called tropical spastic paraparesis or HTLV-I associated myelopathy. Transmission of the virus is almost identical to that of HIV. The latency period before onset of clinical symptoms can last from a few years (tropical spastic paraparesis) up to several decades (adult T-cell leukemia/lymphoma). Four different clinicopathological subtypes of the T-cell neoplasia are known, and in this article we describe two patients with the subtype lymphoma.

Adult↗
PubMed1996

Antigen-presenting function of human peritoneum mesothelial cells isolated from a pancreatic carcinoma patient after mutant Ras peptide vaccination.

Mesothelial cells obtained from ascites fluid of a Ras-peptide vaccinated pancreatic adenocarcinoma patient were cultured in vitro. Fresh isolated cells expressed HLA class II molecules, which were lost upon culture, but could be up-regulated after coculture with human recombinant interferon-gamma. The antigen-presenting capacity of these mesothelial cells was tested with allogeneic peripheral blood mononuclear cells (PBMC) in a mixed lymphocyte mesothelial cell culture and by stimulating autologous PBMC with purified protein derivative of Mycobacterium tuberculosis. Cloned T cells from the same patient allowed us to test the ability of mesothelial cells to present a mutant Ras-derived peptide to specific T cells in a DLA-DR-restricted manner. Mesothelial cells effectively stimulated allogeneic resting T lymphocytes to proliferate and presented soluble protein antigen or a mutant Ras-derived peptide to specific T cells, indicating that they display processing and presenting capabilities. Since mesothelial cells are in close anatomical relationship with intraabdominal malignancies, they may contribute to stimulation of specific T cells by endocytosing tumour-specific antigens and presenting them to T lymphocytes. This could be a possible mechanism in which mesothelial cells participate in maintaining local specific immunity in patients already primed.

Adenocarcinoma↗
PubMed1996

Purging of tumor cells from leukapheresis products: experimental and clinical aspects.

Peripheral blood progenitor cell autografts are being used increasingly in conjunction with high-dose therapy of cancer patients, in the belief that these products have a low probability of containing tumor cells. However, recent findings demonstrate that tumor cell involvement is frequent in leukapheresis products. Although the clinical value of purging has not been clinically established by prospective randomized trials, several studies indicate that contaminating tumor cells in autografts contribute to relapse of the disease in the recipients. We describe our experimental and clinical experience in purging tumor cells from leukapheresis products. Based on our work with purging of lymphoma cells from bone marrow by the use of anti-B cell and anti-T cell antibodies and immunobeads, a purging procedure to deplete leukapheresis products of lymphoma cells has been developed. Moreover, we present data showing that breast cancer cells can be efficiently removed from leukapheresis products using antibreast cancer antibodies, either in combination with immunobeads or as immunotoxins. Our experience with enrichment of CD34 cells employing immunobeads in leukaphresis products from patients with breast cancer and lymphomas shows high purity and yield of CD34 cells. In spite of this, contaminating tumor cells can be observed, strongly suggesting that a combination of CD34 cell enrichment and a purging procedure might be warranted.

Antibodies, Monoclonal↗
PubMed1995

TGF-beta 1 and cyclic AMP promote apoptosis in resting human B lymphocytes.

TGF-beta and agents that elevate intracellular cAMP levels are potent inhibitors of B cell activation in vitro and have been shown to arrest stimulated B cells in the G1 phase of the cell cycle. We tested the effects of TGF-beta 1 and the cAMP-inducing agent, forskolin, on the viability of resting B cells from human peripheral blood, and found that both agents caused a significant, dose-dependent increase in cell death relative to spontaneous death in medium alone, as measured by vital dye staining with propidium iodide. Apoptosis was shown to be the overall mode of death by demonstrating DNA fragmentation using DNA nick end labeling and by verifying the characteristic morphologic changes. In contrast with TGF-beta 1 and forskolin, various B cell activation stimuli generally inhibited spontaneous apoptosis of resting cells. The most potent effects were observed with IL-4 and the phorbol ester, O-tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C. IL-4 also partly inhibited TGF-beta 1 and forskolin-induced apoptosis. In contrast, TPA completely reversed cell death in forskolin-treated cultures, but had no effect on TGF-beta 1-induced apoptosis, indicating that TGF-beta 1 and forskolin promote apoptosis by different mechanisms. The relative protein expression of bcl-2, a proto-oncogene that inhibits apoptosis, was unaltered by the apoptotic as well as the survival stimuli tested, suggesting that apoptosis was regulated by a bcl-2-independent mechanism. We conclude that apoptosis is a regulated phenomenon in resting human B cells. Furthermore, TGF-beta and cAMP may inhibit B cell responses not only by blocking cell cycle progression in activated cells, but also by inducing apoptosis in resting cells.

Antigens, CD↗
PubMed1992

Cell cycle-dependent regulation of CDw75 (beta-galactoside alpha-2,6-sialyltransferase) on human B lymphocytes.

Within the hematopoietic system, CDw75 is primarily expressed on cells of the B cell lineage. Cloning and sequencing of the gene has shown CDw75 to be a beta-galactoside alpha-2,6-sialyltransferase. This enzyme plays an important role in the intracellular terminal glycosylation pathways in various cell types. In this article, we demonstrate that COS cells transfected with the CDw75 cDNA clone displayed sialyltransferase activity, in contrast to mock-transfected cells. We also found that activated B cells displayed an increased enzyme activity compared to resting cells, in accordance with the staining data. Moreover, CDw75 expression was found to be up-regulated approximately 7-9-fold from early G1 to the G2/M phases of the cell cycle in peripheral blood leukocyte B cells. This was shown by staining of in vitro activated B cells with the anti-CDw75 monoclonal antibody HH2, using cell fractions corresponding to different stages of the cell cycle. Using a combination of Hoechst 33258 and propidium iodide after bromodeoxyuridine incorporation, it is possible to distinguish between different phases of the first and second cell cycle. By combining this with HH2 immunofluorescence staining, using a multistation multiparameter flow cytometry program, we confirmed the cell cycle-dependent expression of CDw75. Immunocytochemical stainings of cytospin specimens of elutriated B cells showed that the antigen was up-regulated in late G1 before the appearance of the nuclear activation antigen Ki67. Finally, we showed that activated B cells secreted soluble CDw75 into the medium, as demonstrated by a specific blocking of HH2 staining of B cells using suboptimal concentrations of HH2. In accordance with this, we observed small, but detectable levels of soluble sialyltransferase activity in supernatants of activated B cells.

Antibodies, Monoclonal↗