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B Holzmann

Publications and source records attributed to B Holzmann.

At least 73 records · Page 4Linked to original sources
PubMed1991

Identification of a 107-kD glycoprotein that mediates adhesion between stromal cells and hematolymphoid cells.

The mechanism of cell complex formation between lymphocytes and stromal cells was investigated. We found that lymphoid lines of both T and B lineages could form cell complexes with stromal cells from the thymus as well as bone marrow but not with macrophages or typical fibroblast lines. Formation of these cell complexes is temperature dependent and requires the presence of Mg2+, active cellular metabolism, and microfilament assembly of cytoskeleton. We raised an antiserum against a thymic stromal cell clone (BATE-2) in rats and found that, after absorption, this serum could effectively block cell complex formation between lymphocytes and stromal cells from both thymus and bone marrow. An efficient blocking was obtained only when the antiserum was added at the initial stage of cell interaction. From the blocking experiments and the SDS-PAGE analysis of immunoprecipitated materials from the stromal cell surface, we identified a unique 107-kD glycoprotein on the stromal cells as a molecule for mediating stromal cell-lymphocyte interaction. This is further supported by the findings that an antiserum raised in hamsters against the excised gel band corresponding to 107 kD, which specifically immunoprecipitated the 107-kD molecule, effectively blocked the lymphocyte-stromal cell interaction. The possible function of this molecule in hematolymphoid development is discussed.

Animals↗
PubMed1991

Cloning and expression of cDNAs for the alpha subunit of the murine lymphocyte-Peyer's patch adhesion molecule.

cDNA clones encoding the alpha chain of the murine lymphocyte-Peyer's patch adhesion molecule (LPAM), which is associated with lymphocyte homing, have been isolated by screening with the human VLA-4 (alpha 4h) probe. Several alpha 4 antigenic determinants were identified on COS-7 cells after transfection. From overlapping clones, approximately 5 kb of contiguous nucleotide sequence have been determined, encoding a protein sequence of 1039 amino acids for the LPAM alpha chain (alpha 4m). LPAM is a member of the integrin family of cell-surface heterodimers, and alpha 4m is the murine homologue of the human alpha 4 h chain. The two proteins have a total sequence similarity of 84%, with an almost perfect conservation (31/32 amino acids) in the cytoplasmic domain. Like alpha 4h, alpha 4m is distinct from other integrin alpha chains because it has neither an I-domain nor a COOH-terminal cleavage site. The positions of the characteristic Cysteine residues are conserved, and a putative protease cleavage site is located near the middle of the protein sequence. The NH2-terminal part of the protein contains seven homologous repeats, and three of them include putative divalent cation-binding sites. These sites are among the most conserved between the alpha 4m sequence and other alpha chains, and may therefore be involved in the binding of integrin alpha and beta chains. An additional cDNA clone was isolated which shares a sequence of perfect homology with the alpha 4m encoding cDNAs, but has a unique 3' poly-A end. This observation correlates with the fact that three discrete murine RNA bands are observed in Northern blot experiments using alpha 4m as a probe, whereas only two human RNA species are described for alpha 4h, indicating a higher complexity for murine than for human sequences.

Amino Acid Sequence↗
PubMed1990

Characterization of a rat monoclonal antibody specific for a determinant encoded by the V beta 7 gene segment. Depletion of V beta 7+ T cells in mice with Mls-1a haplotype.

We have generated a rat mAb, TR310, which recognizes a determinant encoded by the murine V beta 7 gene segment of the TCR. TR310 immunoprecipitates TCR from cell lysates, co-modulates with CD3, and can be used for immunofluorescence staining of T cells. By using this antibody, we found that the average percentage of V beta 7+ peripheral T cells in Mls-1b mice was 3.8%, but only 0.8% in Mls-1a mice. A similar difference was also observed in the mature TCRhi thymocyte subsets, suggesting that V beta 7+ T cells are deleted during intrathymic maturation in Mls-1a mice. TR310 should prove to be a valuable reagent in further studies of the TCR repertoire and the analysis of factors which alter it.

Animals↗
PubMed1989

Identification of a murine Peyer's patch--specific lymphocyte homing receptor as an integrin molecule with an alpha chain homologous to human VLA-4 alpha.

Lymphocyte homing is controlled by organ-specific interactions of lymphocytes and high endothelial venules (HEV). Adhesion of lymphocytes to Peyer's patch HEV, but not to peripheral node HEV, is inhibited by an antibody recognizing the murine lymphocyte antigen LPAM-1. Lymphoma cell variants were selected on the FACS for differences in LPAM-1 expression: the binding capacity of these variants to Peyer's patch HEV directly correlates with the level of LPAM-1 expression. The anti-LPAM-1 antibody recognizes the alpha subunit of an Mr 160,000/130,000 cell surface alpha beta heterodimer. The association of LPAM-1 alpha and beta chains requires the presence of Ca2+ ions. Proteins of Mr 84,000 and Mr 62,000 present in LPAM-1 immunoprecipitates appear to be products of the proteolytic processing of alpha chains. The structure of LPAM-1 is virtually identical to that of the human integrin receptor VLA-4. The cross-reactivity of a monospecific rabbit antiserum demonstrated the similarity between the human VLA-4 alpha chain and the alpha subunit of LPAM-1.

Animals↗
PubMed1989

Peyer's patch-specific lymphocyte homing receptors consist of a VLA-4-like alpha chain associated with either of two integrin beta chains, one of which is novel.

Lymphocytes home to various lymphoid organs by adhering to and migrating through specialized high endothelial venules (HEV). The murine cell surface heterodimer LPAM-1 is involved in the homing of lymphocytes to mucosal sites (Peyer's patches). LPAM-1 has an alpha subunit (alpha 4m) analogous to the alpha chain of the human integrin molecule VLA-4. Here we show that the LPAM-1 beta subunit (beta p) is immunochemically and biochemically distinct from previously defined integrin beta subunits, suggesting that beta p represents a novel integrin beta subunit. Depending on the cellular source two alternative beta subunits, beta p and integrin beta 1, can be isolated in association with alpha 4m. Therefore, alpha 4m is the common subunit of the unique integrin LPAM-1 (alpha 4m beta p) and of the heterodimer LPAM-2 (alpha 4m beta 1), which is analogous to VLA-4. Antibody-blocking experiments suggest that, in addition to LPAM-1, LPAM-2 is also involved in the organ-specific adhesion of lymphocytes to Peyer's patch HEV.

Animals↗
PubMed1989

De novo expression of intercellular-adhesion molecule 1 in melanoma correlates with increased risk of metastasis.

The 89-kDa cell surface glycoprotein, P3.58, is detectable on advanced human melanomas in situ but not on benign melanocytes or early melanomas. cDNA cloning of P3.58 from melanoma cells was accomplished by screening a lambda zap expression vector library with monoclonal antibodies produced against the denatured antigen. Nucleotide sequencing of the clones revealed that P3.58 is identical to the intercellular-adhesion molecule 1. No qualitative differences in P3.58 mRNA species could be seen between melanoma cells and hematopoietic cells and no differences in gene organization were observed between peripheral blood leukocytes and melanoma cells. Inspection of the deduced amino acid sequence of P3.58 indicated the presence of the consensus sequence characteristic for complement-binding proteins. The acquisition of this cell-adhesion molecule during the process of tumor progression is speculated to contribute to the development of metastasis in melanoma.

Amino Acid Sequence↗
PubMed1989

Integrin molecules involved in lymphocyte homing to Peyer's patches.

This review summarizes experiments designed to analyze lymphocyte receptors mediating recognition of and adhesion to HEV in mucosal lymphoid organs. A monoclonal antibody (R1-2) was selected which inhibits the adhesion of murine lymphocytes to Peyer's patch HEV. Antibody R1-2 recognizes the alpha chain (alpha 4m) of the murine lymphocyte cell-surface alpha beta heterodimer LPAM-1. The association of LPAM-1 alpha and beta chains requires the presence of Ca++ ions. Two proteins of Mr 84,000 and 62,000 which are also precipitated by antibody R1-2 most likely represent fragments of alpha 4m. The cross-reactivity of a monospecific rabbit anti-serum indicated that alpha 4m is analogous to the alpha chain of the human integrin molecule VLA-4. In addition, a cDNA clone encoding the human VLA-4 alpha chain hybridized with RNA from alpha 4m+ but not alpha 4m- cell lines. However, the LPAM-1 beta subunit (beta p) was shown to be immunochemically and biochemically distinct from integrin beta 1, beta 2, and beta 3, indicating that beta p represents a unique integrin beta chain. When the beta subunits associated with alpha 4m on a panel of lymphoma cell lines were analyzed, it was found that, depending on the cellular source, alpha 4m can associate with either of two beta chains: beta p or integrin beta 1. Therefore alpha 4m appears to be the common subunit of the two lymphocyte cell surface heterodimers LPAM-1 (alpha 4m/beta p) and LPAM-2 (alpha 4m/beta 1). LPAM-2 is analogous to the human VLA-4 molecule, whereas LPAM-1 represents a unique integrin heterodimer. Antibody R1-2 inhibited Peyer's patch HEV-adhesion of normal mouse lymphocytes and every lymphoma cell line tested including LPAM-1 and LPAM-2 single-positive cell lines. We also showed that the binding capacity of variants of a clonal lymphoma cell line to Peyer's patch HEV correlates with the level of LPAM-1 expression. It therefore appears that both heterodimers are involved in lymphocyte-Peyer's patch HEV interactions and that the adhesion of lymphocytes to Peyer's patch HEV is generally LPAM-1- or LPAM-2-dependent. We further investigated whether VLA-4, the human analog of LPAM-2, can mediate adhesion of human lymphocytes to HEV in mucosal lymphoid organs.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1989

[Step-wise expression of melanoma-associated antigens in tumor progression].

Selection of monoclonal antibodies for differential reactivity with benign and malignant melanocytic lesions has led to the identification of molecules which may be involved in the development of metastases. Based on the observed alterations in the antigenic profile we propose a scheme representing the tumor progression of melanocytes to metastatic melanoma.

Antigens, Neoplasm↗
PubMed1988

Glycoprotein P3.58, associated with tumor progression in malignant melanoma, is a novel leukocyte activation antigen.

The P3.58 antigen is defined by monoclonal antibodies (MAbs) selected to discriminate between benign and malignant melanocytic cells. Its expression in malignant cells has been shown to correlate with an increased risk of metastasis. A survey of a wide range of tissues revealed that, on normal tissue, expression of P3.58 antigen is restricted to a subset of cells involved in the immune response. The antigen was found not only on certain endothelia, but also on activated macrophages in vivo and in vitro and as well as on activated B lymphocytes. A comparison with known B-lymphocyte and leukocyte activation antigens indicated that P3.58 is a novel leukocyte activation antigen. Biochemical analysis of the P3.58 antigen isolated from cells of different histogenic origin indicated that different molecular forms of the antigen exist, apparently depending on the cell type of origin. P3.58 molecules precipitated from tunicamycin-treated cells were identical in all cell types, suggesting that the variation observed is due to variable N-glycosylation.

Antigens, Neoplasm↗
PubMed1988

Inflammatory cell infiltrates in human melanoma at different stages of tumor progression.

Progression of human melanoma is associated with changes in antigenic phenotypes of tumor cells. To establish whether inflammatory infiltrates in progressing melanoma also change, we studied 146 cutaneous melanomas at different stages of progression. Monoclonal antibodies (MAbs) against lymphocyte and macrophage subpopulations, interleukin-2 receptor (IL-2 R), immune interferon (IFN-gamma), and the IFN-gamma-inducible, progression-associated melanoma antigens HLA-DR and gp89 were applied in situ. During the course of melanoma progression, decreased amounts of peritumoral T cells, IL-2 R-expressing lymphocytes and dermal T6+ dendritic cells were found, while increased numbers of intratumoral T cells, inflammatory (27E10+) and mature (25F9+) macrophages were associated with local progression of primary melanomas. In metastases, most infiltrate components except 25F9+ macrophages were rare. Positive correlations were observed between: (1) dermal T6+ cells and IL-2 R+ lymphocytes, and (2) presence of IFN-gamma in the infiltrate and HLA-DR and gp89 antigens on tumor cells. In all stages, HLA-DR expression on tumor cells was correlated with: (1) a shift towards T8+ lymphocytes in the infiltrates and (2) a loss of IL-2 R expression. Our data suggest mutual influences between melanoma cells and mononuclear cell infiltrates in situ.

HLA-DR Antigens↗
PubMed1988

Homing receptors and metastasis.

As discussed in the preceding sections, there are several indications that the lymphocyte homing receptors involved in the normal process of lymphocyte recirculation are also relevant to the behavior of metastatic cells. Cell fusion experiments indicate that previously nonmetastatic cells can acquire metastatic capacity from fusion with normal lymphocytes. Murine T lymphomas that bear high levels of functional homing receptors can metastasize to peripheral lymphoid organs, whereas those lymphomas lacking homing receptors cannot. Virtually all lymph node metastases of lymphomas contain a high proportion of MEL-14hi cells, even if the primary tumor has been selected to be relatively deficient in these cells. Further investigations of the biology of lymphocyte homing receptors will reveal whether or not there are additional lymphocyte homing receptors and will clarify the role of lymphocyte homing receptors in metastasis. Antibodies against three lymphocyte homing receptors could therefore be useful for diagnosis and treatment of metastatic disease.

Animals↗
PubMed1988

The melanoma progression-associated antigen P3.58 is identical to the intercellular adhesion molecule, ICAM-1.

The 89kd cell surface glycoprotein, P3.58, is expressed on human malignant melanomas in situ where it is associated with an increased risk of metastatic disease. Monoclonal antibodies detecting denatured P3.58 were produced and used to isolate a P3.58 encoding cDNA clone from a human melanoma lambda expression library. Sequencing of the cDNA revealed that the P3.58 antigen is identical to the leukocyte intercellular adhesion molecule 1 (ICAM-1).

Amino Acid Sequence↗
PubMed1987

Tumor progression in human malignant melanoma: five stages defined by their antigenic phenotypes.

The antigenic profile of melanocytic cells in the course of local and systemic tumor progression of human malignant melanoma was investigated by the reactivity of a panel of monoclonal antibodies (MAbs) in frozen sections of histologically defined melanocytic lesions. Specific antigenic phenotypes made it possible to distinguish 5 groups of lesions which could be ranked in relation to each other due to the sequential acquisition or loss of progression markers. On this basis, a scheme of antigenic changes which accompany the stepwise transformation of normal skin melanocytes into highly malignant metastatic melanoma cells is proposed. The steps of tumor progression identified solely by phenotyping with MAbs were in complete concordance with the concept of melanoma progression derived from histological, statistical and clinical analyses. Furthermore, our finding that the expression of gp89 as well as HLA-DR antigens can be induced by interferon-gamma in vitro provides evidence that immune interferon may play a role in the regulation of genes leading to phenotypic changes in progressing melanoma cells.

Animals↗
PubMed1987

Discrimination between benign and malignant cells of melanocytic lineage by two novel antigens, a glycoprotein with a molecular weight of 113,000 and a protein with a molecular weight of 76,000.

The present study describes two novel antigens, a glycoprotein with a molecular weight of 113,000 and a protein with a molecular weight of 76,000, which are associated with the transformed phenotype of melanocytes. The monoclonal antibodies (MoAb) MUC18 and MUC54, raised against human malignant melanoma, were selected for differential reactivity with normal and neoplastic cells of melanocyte lineage. The antigen defined by MoAb MUC18 is a membrane bound monomeric sialylated glycoprotein with an apparent molecular weight of 113,000. In contrast to the broad reactivity with melanomas, isolated nevus nests were stained in only 1 of 55 nevi investigated. No staining of MoAb MUC18 was observed in a large variety of surgically removed normal and tumor tissues except for smooth muscle cells of the blood vessel wall and hair follicles. MoAb MUC54 immunoprecipitated a cytoplasmic monomeric protein with an apparent molecular weight of 76,000. By immunoperoxidase staining, the antigen was demonstrated on a large number of melanomas and in addition on 1 of 36 nevocellular, 3 of 4 Spitz, and 5 of 14 dysplastic nevi. The Mr 76,000 protein was found in a number of epithelial tissues and various types of neoplasms. Both antibodies presented in this study define structural changes in the antigenic profile of melanocytes occurring during carcinogenesis.

Antibodies, Monoclonal↗
PubMed1987

Thyroglobulin, the major and obligatory exportable protein of thyroid follicle cells, carries the lysosomal recognition marker mannose-6-phosphate.

Thyroglobulin (TG), the major exportable protein of thyroid follicle cells, is conveyed to lysosomes on a complex secretion, storage and recapture pathway by as yet unknown transport mechanisms. This report establishes that the dimeric porcine TG-molecule carries an average of six phosphate residues. Endoglycosidase digestion showed that two phosphate residues are bound to the high-mannose carbohydrate side chains (CHO), while two others are linked to the complex CHO. These four residues are also sensitive to alkaline phosphatase treatment, indicating their terminal linkage. Immunoprecipitation analyses showed that TG obtained from microsomal fractions is already phosphorylated. Most important, an enzymatic assay applied to hydrolysates of TG established that the two phosphate residues at the high mannose CHO are present as mannose-6-phosphate (M-6-P). Alkaline phosphatase treatment of biosynthetically radiophosphorylated CHO followed by hydrolysis and t.l.c. indicated that M-6-P is present at least in part in phosphomonoester linkage. Furthermore, porcine TG binds specifically to the M-6-P receptor of Chinese hamster ovary cells. It is concluded that the M-6-P residues of TG are exposed and able to operate as a ligand for the M-6-P receptor. It is unknown why the lysosomal recognition-marker M-6-P does not convey TG directly on an intracellular route to lysosomes. We propose that for the secretion of newly synthesized TG into the follicle lumen an additional export signal dominating over the M-6-P recognition-marker is required.

Animals↗
PubMed1987

Micrometastatic cancer cells in bone marrow: in vitro detection with anti-cytokeratin and in vivo labeling with anti-17-1A monoclonal antibodies.

The detection of early micrometastasis or disseminated single tumor cells poses a problem for conventional diagnosis procedures. Using a panel of monoclonal antibodies against cytokeratin and the 17-1A epithelial antigen we identified immunocytochemically tumor cells in bone marrow of patients with breast cancer (n = 155) and colorectal cancer (n = 57) at the time of surgery of the primary tumor. Monoclonal antibody CK2, recognizing the human cytokeratin component 18 in simple epithelia, appeared to be the most suitable reagent because of its negative reaction with bone marrow samples of the noncarcinoma patients (n = 75). Its specificity was further demonstrated in a double-marker staining procedure using an anti-leukocyte common antigen monoclonal antibody (T200) as counterstain. A comparative analysis showed that immunocytology was clearly superior to conventional cytology (n = 212) and histology (n = 39). In 9.5-20.5% of patients without distant metastasis, tumor cells could be detected in bone marrow. We found a significant correlation between tumor cells in bone marrow and conventional risk factors, such as distant metastasis or lymph node involvement. In a first approach toward immunotherapy we demonstrated in 3 patients that infused monoclonal antibody 17-1A can label single tumor cells in bone marrow in vivo. We then used this approach to follow up 7 patients undergoing 17-1A therapy in an adjuvant clinical trial.

Antibodies, Monoclonal↗
PubMed1985

In situ analysis of antigens on malignant and benign cells of the melanocyte lineage. Differential expression of two surface molecules, gp75 and p89.

Monoclonal antibodies (mAb) were selected for differential binding to sections of freshly frozen biopsy material of human malignant melanomas and their precursor lesions, the melanocytic nevi. Both melanomas and normal nevi expressed human Ia-like antigens, transferrin receptor and the transferrin-related molecule p97. In contrast, only 1 nevus of 21 tested expressed both glycoprotein gp75, defined by mAb 15.75, and protein p89, defined by mAb P3.58, whereas 12 of 15 melanomas tested expressed both antigens. mAb P3.58 reacted with one additional melanoma and one nevus. The expression of these two molecules therefore appears to be correlated with the appearance of the malignant phenotype of melanocytes.

Animals↗