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J Mollenhauer

Publications and source records attributed to J Mollenhauer.

At least 73 records · Page 4Linked to original sources
PubMed1990

Cellular immune response toward human articular chondrocytes. T cell reactivities against chondrocyte and fibroblast membranes in destructive joint diseases.

Articular cartilage is one of the major targets in destructive joint diseases in humans. We studied cellular immune reactions against cartilage cell-surface membranes, because it has recently been suggested that these represent possible antigenic structures, based upon the observation of autoantibodies with this specificity in certain joint diseases. A striking T cell reactivity toward chondrocyte membranes was found both in blood and synovial tissue from patients with rheumatoid arthritis. This reactivity was strongly dependent on the presence of monocytes and had all the characteristics of an antigen-driven process. Clonal analysis demonstrated high precursor frequencies in peripheral blood T cells that were reactive against chondrocyte membranes. This response to chondrocyte membranes greatly exceeded the T cell stimulation induced by membranes from other sources such as fibroblasts or epithelial cells. In contrast to patients with rheumatoid arthritis, individuals with osteoarthritis showed a strong peripheral blood and synovial fluid T cell response not only to chondrocyte membranes, but also to fibroblast membrane material. However, there was no reactivity to epithelial cell membranes. Normal donors generally did not show significant responses to any membrane preparation. These data indicate that there is a strong T cell reactivity toward chondrocyte membranes in destructive joint disorders, and this may significantly contribute to the pathogenetic processes that occur in these diseases.

Adult↗
PubMed1990

Type-VI collagen in the human iris and ciliary body.

The distribution of type-VI collagen in the human iris and ciliary body was investigated by means of immunohistochemical techniques and compared with that of type-IV collagen, fibronectin and laminin. As has been described for other tissues, type-VI collagen surrounds type-I and -III collagen fibers. The aggregated form of type-VI collagen (the "long-spacing" or "curly" collagen), which has already been described in the trabecular meshwork and sclera, was also observed at the ciliary muscle tips surrounding the anterior elastic tendons of this muscle. In addition, staining for type-VI collagen was seen directly adjacent to the basement membranes of the ciliary muscle cells, the iris muscles, the uveal vascular endothelia and nerves, but not adjacent to the epithelial basement membranes. The staining did not form a discrete line like the immunoreaction for type-IV collagen, but bundles of marked fibrils extended into the surrounding connective tissue. We assume that type-VI collagen similar to type-VII collagen forms part of an anchoring system for these tissues. As type-VII collagen has been described only in connection with epithelial basement membranes, both type-VI and type-VII collagens may represent anchoring fibrils, however for different tissue components.

Aged↗
PubMed1990

Laminin interactions with ductal pancreatic adenocarcinoma cells: identification of laminin- and collagen-binding proteins.

Laminin promotes the modulation of human pancreatic adenocarcinoma cells from a proliferative to a resting phenotype. This process includes restoration of cell polarity, increase of protein biosynthesis, and increase of glycoprotein secretion. The growth correlates with the amount of laminin coated on the culture dish. Adenocarcinoma cells do not synthesize collagen type I, fibronectin or laminin. Prolonged propagation of the cells on laminin substratum enhances the expression of laminin-binding sites on the cell surface. Laminin binds to cell plasma membrane vesicles with a KB of about 2.2 x 10(10). By affinity chromatography of [35S]methionine-labeled, detergent-extracted, cells on immobilized laminin, a Mr 82,000 polypeptide could be enriched. By simultaneous chromatography on immobilized collagen type I, a Mr 34,000 polypeptide was retained. In laminin overlay experiments on transblotted plasma membrane proteins, Mr 82,000 and 70,000 polypeptides were labeled. Affinity chromatography on laminin-Sepharose of tumor cell membranes from cells grown in nude mice tumors retained Mr 100,000, 82,000, 70,000 and 55,000 polypeptides bound to the column. Endoglycosidase F treatment of these proteins reduced the number of higher Mr proteins, leaving a Mr 70,000 polypeptide, together with smaller peptides. Using the enriched binding protein fraction as antigens, monoclonal antibodies (mAbs) were prepared in mice. Two of the mAbs were further analysed; they recognized simultaneously the complete pattern of high and low Mr polypeptides identified to date by the other methods. Antibody 2A1-H7 was capable of inhibiting attachment and spreading of the cells on laminin and collagen I, but not on tissue-culture plastic. These data may indicate a molecular heterogeneity, partly based on diverse glycosylation, responsible for the variations in the molecular weight of the binding proteins. The adaptation processes of the tumor cells during growth on the extracellular matrix may indicate a regulatory function on tumor growth and metastasis in vivo.

Adenocarcinoma↗
PubMed1990

[New approaches in laboratory diagnosis of osteoarthritis].

The article elucidates the new laboratory tests for chronic joint diseases, with special attention to quantitative and qualitative serum markers for metabolic and immunologic processes. The assays include the quantification of keratan sulphate, hyaluronan and beta-D-xylosyltransferase in serum and pyridinolin in serum and urine as markers for cartilage catabolism, as well as the detection of autoantibodies to cartilage matrix and cell components and cartilage-directed T-cell reactions in the patients. Taken together, these assays represent models for future laboratory diagnostic tools for the characterization of arthritic diseases. So far, however, it remains unclear whether they will prove to be adequate assays for the practicing physician.

Autoantibodies↗
PubMed1989

Evidence for the direct interaction of chicken gizzard 5'-nucleotidase with laminin and fibronectin.

The ectoenzyme 5'-nucleotidase purified from chicken gizzard is shown to specifically interact with laminin and fibronectin, components of the extracellular matrix, by a number of different techniques: (i) cosedimentation with laminin by sucrose gradient centrifugation; (ii) affinity adsorption to both laminin- and fibronectin-Sepharose 4-B; (iii) specific binding to both laminin and fibronectin dotted onto cellulose filters; and (iv) monoclonal antibodies against 5'-nucleotidase are shown to interfere with the interaction of 5'-nucleotidase with laminin and fibronectin. For all the techniques employed, the interactions were found to be specific, since 5'-nucleotidase did not bind to unrelated proteins such as bovine serum albumin or to monomeric actin. The interaction of purified chicken gizzard 5'-nucleotidase could be demonstrated for the hydrophobic enzyme solubilized in detergent and after its reconstitution into artificial phospholipid vesicles. The affinity adsorption experiments indicate that reconstituted enzyme binds more strongly to both laminin and fibronectin. The 5'-nucleotidase employed in this study is anchored to the plasma membrane by a glycan-phosphatidylinositol linker. After treatment with phosphatidylinositol-specific phospholipase C, the enzyme is transformed into a hydrophilic form, for which interactions with laminin and fibronectin could also be demonstrated by the dot-blot technique. Thus controlled cleavage of the phosphatidylinositol linker of 5'-nucleotidase could enable cells to rapidly alter their adhesiveness to certain components of the extracellular matrix.

5'-Nucleotidase↗
PubMed1989

Immunomicroscopical study of type VI collagen in the trabecular meshwork of normal and glaucomatous eyes.

Cross-strained fiber bundles called long-spacing collagen or curly collagen occur in normal eyes in the trabecular meshwork. It can be seen in the basement membrane of the trabecular lamellae, in the sheath of the elastic-like fibers and underneath the inner wall of Schlemm's canal, where it forms part of the so called plaque material. The amount of this long-spacing collagen increases with age and is significantly more pronounced in glaucomatous eyes. Using immunohistochemical and immuno-electronmicroscopic methods, we have been able to show that type VI collagen is present in the aggregates called long-spacing collagen.

Age Factors↗
PubMed1989

Selective growth inhibition of ductal pancreatic adenocarcinoma cells by the lysosomotropic agent chloroquine.

Pancreatic adenocarcinoma cells show characteristics of macrophages as phagocytosis and exocytosis. These phagocytic activities can be inhibited in macrophages by antirheumatic drugs, e.g. gold compounds and antimalarials. We have now tested the activity of auranofin and chloroquine to inhibit growth of a pancreatic tumor cell line (PaTu II) and of human foreskin fibroblasts (HFF) as a control. We found that auranofin (greater than or equal to 2 microM) inhibits both PaTu II cell and HFF growth. In contrast, chloroquine (4-18 microM) selectively interferes with the growth of PaTu II cells. This specific vulnerability of PaTu II cells was confirmed in cocultures of tumor cells and fibroblasts.

Auranofin↗
PubMed1989

Laminin and other basal lamina proteins with neurite promoting activity in medium conditioned by C6 glioma cells.

Neurite promoting activities (NPFs) are essential factors in neuronal differentiation. Some of them are associated with proteins of the extracellular matrix (ECM). C6 cells, a rat glioma cell line, release NPF activities into the cell culture medium. We used antibodies against ECM-proteins for enrichment and partial characterization of these activities. Results show that, (1) C6 cells express and release laminin; (2) the C6-laminin consists of 260 kD chains only and is therefore different from typical basal lamina laminin (220 and 440 kD chains), but comparable to other laminins of glial origin (chains in the 200 kD range only); (3) C6-laminin partially purified by affinity chromatography shows NPF-activity; (4) laminin concentration in C6 cell-conditioned medium is not sufficient to account for the total neurite promoting activity of the medium, and (5) in addition to laminin C6 cells express and release fibronectin and possibly type IV collagen.

Animals↗
PubMed1988

The structure of anchorin CII, a collagen binding protein isolated from chondrocyte membrane.

cDNA clones for anchorin CII (Mr = 34,000), a collagen-binding protein, were isolated from a lambda gt 11 cDNA library prepared from chick cartilage mRNA. Several overlapping clones were characterized which gave rise to an open reading frame coding for 329 residues and a 3'-untranslated segment of 500 base pairs. The clones were identified as coding for anchorin by hybrid select translation analysis and by comparing the deduced amino acid sequence with the sequence of 10 tryptic peptides of the protein. A hydrophobic domain of 25 residues interrupted with 3 polar residues was identified with the carboxyl-terminal portion. There was no evidence for an aminoterminal signal peptide. Northern analysis revealed that the 5' probe hybridizes to a single 1.7-kilobases (kb) mRNA species, whereas the 3' probe hybridizes to two mRNA species of 1.7 kb and 5 kb, which are present in many cells including chondrocytes, crop cells, and fibroblasts. The level of anchorin mRNA in chick embryo fibroblasts was increased by infection with Rous sarcoma virus.

Amino Acid Sequence↗
PubMed1988

A defective cell surface collagen-binding protein in dermatosparactic sheep fibroblasts.

Fibroblasts from dermatosparactic sheep fail to contract collagen gels and show a reduced attachment to collagenous substrates. By comparing collagen-binding membrane proteins of normal (+/+), homozygote (-/-), and heterozygote (+/-) fibroblasts, we present evidence that the interaction of normal fibroblasts with native type I collagen involves a protein of apparent Mr = 34,000 which is absent from dermatosparactic fibroblasts and seems to be related to anchorin CII. This conclusion was reached from the following experiments: (a) On a blot of membrane proteins from normal fibroblasts radioactively labeled type I collagen bound predominantly to a protein band of 34 kD; dermatosparactic membranes revealed only a small amount of binding to a component with a molecular mass of 47 kD. (b) After separation of normal fibroblast membrane proteins on type I collagen-Sepharose, a collagen-binding component of 34 kD was found which was absent from the corresponding fraction of dermatosparactic membranes. (c) Antibodies to anchorin CII stained the surface of normal (+/+), but not of dermatosparactic (-/-) fibroblasts and labeled a 34-kD component after immunoblotting of normal fibroblast membrane proteins. (d) After metabolic labeling of fibroblasts with [35S]methionine and immunoprecipitation with anti-anchorin CII, 40- and 34-kD components were precipitated from extracts of normal fibroblasts, while the latter component was absent from affected cells. Similar differences were found after immunoblotting of membranes from whole normal or affected skin. These data indicate that dermatosparaxis of sheep involves a molecular defect of a collagen-binding protein. Therefore this disease represents a model to study the complex interaction of cells with the extracellular matrix on a molecular level.

Animals↗
PubMed1988

Serum antibodies against chondrocyte cell surface proteins in osteoarthritis and rheumatoid arthritis.

Serum autoantibodies to cartilage cell surface proteins have been detected in the blood from patients with osteoarthritis (OA) and rheumatoid arthritis (RA). As tested by ELISA using plasma membrane vesicles as a substrate, patients with RA exhibited significantly elevated antibody titers, whereas titers were somewhat lower in OA, and only marginal in controls. The antibodies did not crossreact with plasma membranes from fibroblasts or epithelial cells, but showed interspecies crossreactivity towards chicken and rat cartilage cell surface proteins. A group of at least 9 distinct immunoreactive polypeptides could be characterized by Western blot analysis. These peptides had molecular weights of Mr28, 30, 38, 42, 66, 76, 78, 116, and 155 kD. The patients each expressed an individual pattern of reactive antibodies against these proteins. Autoantibody reactivity to cell surface associated collagen type II was also observed.

Adult↗
PubMed1987

Distribution of extracellular matrix proteins in pancreatic ductal adenocarcinoma and its influence on tumor cell proliferation in vitro.

Affinity-purified antibodies to major components of the extracellular matrix (fibronectin and collagen type I) and basal lamina (laminin) were used in indirect immunofluorescence studies on frozen sections of 12 pancreatic ductal adenocarcinoma of the human and on sections of normal and inflamed pancreatic tissue of the same patients. Laminin-specific immunoreactivity was distributed in close correlation to the grade of differentiation of the tumor tissue. Intact basement membranes, also with some structural irregularities were found only in the highest grade of differentiation where tumor cells grew as tubular structures. With progressive dedifferentiation basal laminae were either absent or the laminin-positive material was focally distributed without spatial association with tumor cells. In all cases of pancreatic tumors a remarkable increase in interstitial connective tissue was observed, as demonstrated by antibodies specific for human collagen type I and for human serum fibronectin. Tumor extracts contained high amounts of collagen type I and V but no significant amount of collagen type III as visualized by analytical SDS gel electrophoresis. A similar distribution of collagen types was observed in lymph node and liver metastases, and in tumors xenografted into nude mice. Since previously a close correlation between grading and growth kinetics of primary tumors had been observed, in vitro experiments were performed analyzing the effect of purified extracellular matrix proteins on tumor cell proliferation. In vitro cultivation of two established cell lines of pancreatic carcinoma on collagen type I or on laminin resulted in an arrest of proliferation on laminin substrates, while normal proliferation comparable to growth on regular culture dishes was found using collagen type I and fibronectin as substrates. Fine structural studies demonstrated a higher degree of cell differentiation in the presence of laminin, as compared to collagen type I or fibronectin.

Adenocarcinoma↗
PubMed1987

Monocyte-related functions expressed in cell lines established from human pancreatic adenocarcinoma. I. Comparative analysis of endocytotic activity, lysosomal enzyme secretion, and superoxide anion production.

Fine structural analyses of human pancreatic carcinoma have demonstrated their composition of one major cell type, a mucin secreting cell. In lower grades of differentiation the tumor cells change polarity and reduce mucin production and secretion. Instead numerous small vesicles in the cytoplasm and a high frequency of coated pits and coated vesicles beneath the plasma membrane suggest endocytotic activity. One monoclonal antibody (BW 227/19) produced against pancreatic tumor cells of low differentiation, besides binding to primary pancreatic carcinoma in cytofluorometric studies, also reacted with 80% blood monocytes and with macrophages in a variety of tissues, indicating common antigenic determinants on both cell types. Therefore, monocyte-related functions--endocytosis, lysosomal enzyme secretion, and superoxide anion production--were measured in 5 cell lines of pancreatic carcinomas (two lines established from adenocarcinoma of the colon, two lines from small lung cell carcinomas, and one line from squamous cell carcinoma of the lung). These functions were measured under basal conditions and after exposure to zymosan or immune complexes and were compared to isolated normal monocytes of the human. Endocytotic activity, as measured by the uptake of radiolabeled colloidal gold in 4 out of 5 pancreatic tumor cell lines, showed 50% of the basal activity of monocytes, while other tumor cells had only 20% of this activity. Only pancreatic tumor cells demonstrated increased endocytotic, secretory, and chemiluminescence activity to a similar extent as human monocytes after exposure to zymosan or immune complexes. Fine structural studies using cationized ferritin demonstrated the regular endocytotic pathway with uptake via coated pits and vesicles into endosomes and their transfer to a membrane compartment associated with the Golgi complex. The findings indicate a striking similarity between pancreatic tumor cells in vitro and human monocytes, which could be of importance in the understanding of the biology of these ill-defined tumor cells.

Adenocarcinoma↗
PubMed1986

The extracellular matrix proteins laminin and fibronectin modify the AMPase activity of 5'-nucleotidase from chicken gizzard smooth muscle.

Laminin and fibronectin, but not collagen, affect the AMPase activity of the purified transmembrane protein 5'-nucleotidase. Laminin stimulates whereas fibronectin inhibits the AMPase activity of this ectoenzyme. The AMPase-modulating effects by these components of the extracellular matrix require a preincubation period of several hours when detergent-solubilized 5'-nucleotidase is employed, they can, however, instantaneously be elicited with liposome-incorporated 5'-nucleotidase.

5'-Nucleotidase↗