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Biomedical subjects

A Hildebrand

Publications and source records attributed to A Hildebrand.

35 records · Page 2Linked to original sources
PubMed1993

Localization of transforming growth factor beta binding site in betaglycan. Comparison with small extracellular matrix proteoglycans.

The most abundant binding molecule for transforming growth factor beta (TGF-beta) on many cell types is betaglycan, a transmembrane proteoglycan. To localize the binding site for TGF-beta in betaglycan, parts of the betaglycan extracellular portion were expressed as recombinant fusion proteins in bacteria and tested for their ability to compete for the binding of TGF-beta 1 to Hep G2 cells. One fragment encompassing 226 residues near the transmembrane domain (amino acids 543-769) (López-Casillas, F., Cheifetz, S., Doody, J., Andres, J. L., Lane, W. S., and Massagué, J. (1991) Cell 67, 785-795) was active, whereas fusion proteins representing the other parts of the betaglycan ectodomain were inactive. Affinity measurements revealed two classes of binding sites (Kd = 3.9 nM and Kd = 145 nM) for the active fusion protein. The binding of the betaglycan fusion protein to immobilized TGF-beta 1 was inhibited by fusion proteins representing the core proteins of the small interstitial proteoglycans decorin, biglycan and fibromodulin, each also known to bind TGF-beta. The effective concentrations of TGF-beta for binding to these other proteoglycans were similar to those required for binding to betaglycan, indicating similar affinities for the binding of proteoglycans and betaglycan. Affinity cross-linking showed that, at low concentrations, the betaglycan fragment enhanced the binding of TGF-beta to the type II receptor and to endogenous betaglycan but had no effect on the binding to type I receptor. At high concentrations, the TGF-beta binding fragment inhibited the binding of TGF-beta to all these receptors. The fragment enhanced the activity of TGF-beta in mink lung cell bioassay at all active concentrations. The results indicate that betaglycan and the decorin type proteoglycans all bind to the same or closely spaced sites in TGF-beta and compete with one another for the binding. In addition, betaglycan may cooperate with the type II receptor on TGF-beta binding.

Animals↗
PubMed1993

A novel integrin specificity exemplified by binding of the alpha v beta 5 integrin to the basic domain of the HIV Tat protein and vitronectin.

Several studies have addressed the interaction of the HIV Tat protein with the cell surface. Our analysis of the cell attachment-promoting activity of Tat and peptides derived from it revealed that the basic domain of Tat, not the arg-gly-asp (RGD) sequence, is required for cell attachment to Tat. Affinity chromatography with Tat peptides and immunoprecipitation with various anti-integrin antibodies suggest that the vitronectin-binding integrin, alpha v beta 5, is the cell surface protein that binds to the basic domain of Tat. The Tat basic domain contains the sequence RKKRRQRRR. A related sequence, KKQRFRHRNRKG, present in the heparin-binding domain of an alpha v beta 5 ligand, vitronectin, also bound alpha v beta 5 in affinity chromatography and, in combination with an RGD peptide, was an inhibitor of cell attachment to vitronectin. The alpha v beta 5 interaction with these peptides was not solely due to high content of basic amino acids in the ligand sequences; alpha v beta 5 did not bind substantially to peptides consisting entirely of arginine or lysine, whereas a beta 1 integrin did bind to these peptides. The interaction of alpha v beta 5 with Tat is atypical for integrins in that the binding to Tat is divalent cation independent, whereas the binding of the same integrin to an RGD-containing peptide or to vitronectin requires divalent cations. These data define an auxiliary integrin binding specificity for basic amino acid sequences. These basic domain binding sites may function synergistically with the binding sites that recognize RGD or equivalent sequences.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Staphylococcus aureus alpha-toxin. Dual mechanism of binding to target cells.

Staphylococcal alpha-toxin was radiolabeled to high specific radioactivity (1,500-3,000 Ci/mmol) under retention of its hemolytic activity. Binding studies with susceptible rabbit erythrocytes and highly resistant human erythrocytes revealed that binding of alpha-toxin to target cells can occur via two different mechanisms. Binding of alpha-toxin to rabbit erythrocytes initially involves specific binding sites and occurs at low concentrations, with half-maximal binding at 1-2 nM. In contrast, toxin binding to human erythrocytes is absorptive and nonspecific, in this case, significant binding as well as hemolysis occur only at alpha-toxin concentrations exceeding 1 microM. Autoradiographic analyses of membrane-associated alpha-toxin from either cell species proved that hemolysis was inevitably associated with the formation of toxin hexamers. Our data indicate that the high susceptibility of certain target cells toward alpha-toxin is caused by the presence of specific binding sites. However, membrane damage of both susceptible and nonsusceptible target cells occurs via a common mechanism involving toxin oligomerization and pore formation.

Animals↗
PubMed1989

A novel beta-endorphin binding protein. Complement S protein (= vitronectin) exhibits specific non-opioid binding sites for beta-endorphin upon interaction with heparin or surfaces.

Human beta-endorphin (1-31) (beta H-endorphin) was found to specifically interact with purified complement S protein from human plasma. As found by chemical cross-linking beta H-endorphin bound to both, the 65- and 75-kDa molecular mass forms of S protein. The interaction of S protein with heparin as well as the adsorption of S protein to surfaces led to an almost 10-fold increase of specific binding which was due to the exposure of further beta H-endorphin-binding sites. The interaction of beta H-endorphin with S protein bore characteristics of a ligand-receptor interaction, such as time dependence, reversibility, high affinity, saturability, and structural specificity and was mediated through the non-opioid COOH terminus of the beta H-endorphin molecule. beta H-Endorphin binding to S protein was observed at physiological pH or cation concentrations, indicating that the interaction may well occur in vivo. Our results provide conclusive evidence that interactions of S protein with very different effectors led to similar conformational changes which uniformly resulted in exposure of a highly specific beta H-endorphin binding domain on S protein. With S protein as major beta H-endorphin-binding protein in the periphery, the molecular basis of a widespread system of humoral target sites of the neuroendocrine effector appears to be established. In view of S protein involvement in processes of inflammation and wound repair and beta-endorphin effects on immunocompetent cells, the demonstrated S protein-beta H-endorphin interaction appears to be of considerable functional significance.

Binding Sites↗
PubMed1989

Identification of the beta-endorphin-binding subunit of the SC5b-9 complement complex: S protein exhibits specific beta-endorphin-binding sites upon complex formation with complement proteins.

Beta-Endorphin has been reported to specifically interact with SC5b-9 complement complexes via non-opioid binding sites. Covalent cross-linking of [125I]beta H-endorphin to SC5b-9 and analysis of the cross-linking products by gel electrophoresis and subsequent autoradiography revealed a single specifically labelled species which was identical with the S protein subunit of the complement complex. In contrast to SC5b-9, no cross-linking of labelled beta-endorphin to subunits of C5b-9(m) could be observed, indicating that beta-endorphin binding to SC5b-9 was mediated exclusively via S protein. Beta-Endorphin binding to SC5b-9 was compared with binding to purified S protein. Whereas beta-endorphin binding to purified S protein was only modest, complex formation of S protein with complement proteins led to a strong increase in beta-endorphin-binding site concentration, compatible with the exposure of primarily cryptic beta-endorphin-binding sites on S protein.

Binding, Competitive↗
PubMed1989

Hydrophobic labelling of membrane-embedded proteins with lipophilic reagents. Incorporation of [125I]INA and [125I]TID into B lymphocyte membrane immunoglobulins.

Hydrophobic labelling is frequently used in the study of membrane-inserted domains of intrinsic proteins. However, the published procedures, fail to incorporate sufficient radioactivity into membrane immunoglobulins of B lymphocytes to permit investigation of their subunit structures and associations with other proteins. In order to increase the specific radioactivity of [125I]iodonaphthylazide ([125I]INA), an improved method for the synthesis of the reagent was developed. In addition, the optimal conditions for labelling B lymphocytes with [125I]INA and the commercially available reagent 3-(trifluoromethyl)-3-(3'-[125I]iodophenyl)diazirine ([125I]TID) were elaborated. Under these optimized conditions, Ig molecules labelled with [125I]INA and [125I]TID were isolated and analysed in detail by SDS-PAGE. The usefulness of the two reagents for the investigation of lipid-embedded domains of membrane proteins is discussed.

Animals↗
PubMed1989

Characterization of albumin-binding properties of Peptostreptococcus magnus.

A Peptostreptococcus magnus strain demonstrated binding activity for albumin preparations from humans, mice, and dogs, but not for rabbit or bovine albumin. The albumin binding site appeared to be heat stable and of protein nature. Treatment of P. magnus cells with trypsin under specified conditions enhanced this albumin binding. Electron micrographs and kinetic analyses revealed that this enhancement was the result of the removal of some cell wall associated proteins leading to a higher binding affinity without significant changes in binding site numbers. The albumin-binding proteins could be readily solubilized and purified by affinity chromatography. Upon gel electrophoresis the molecular mass of the albumin-binding proteins was estimated as 130 kilodaltons.

Albumins↗
PubMed1988

Characterization and identification of heparin-induced nonopioid-binding sites for beta-endorphin in human plasma.

We have characterized the specific binding of human beta-endorphin (1-31) to novel binding sites which are formed in human plasma or serum in the presence of heparin. The formation of the binding sites is temperature-dependent and does not occur in the presence of other anticoagulants, such as sodium-EDTA, sodium-oxalate, or sodium-citrate. The specific binding of 125I-beta H-endorphin to heparin-induced binding sites in human plasma is saturable and reversible. It is not inhibited by morphine or naloxone or by various opioid peptides which share their NH2-terminal opioid-active sequence with beta H-endorphin. In contrast, binding is inhibited by the COOH-terminal beta H-endorphin fragment Gly-Glu indicating that binding is to nonopioid sites. Electroimmunoprecipitation techniques revealed that these binding sites are identical with S protein/vitronectin or derivatives thereof. S protein is a plasma alpha 1-glycoprotein involved in attachment and spreading of cells and also in blood coagulation and complement activation. It is possible that the interaction of beta-endorphin with S protein is of physiological significance.

Binding Sites↗
PubMed1988

Binding activity of Streptococcus canis for albumin and other plasma proteins.

All 24 cultures of Streptococcus canis examined bound 125I-labelled human albumin, IgG and fibrinogen; but neither IgA nor haptoglobin. Binding of human albumin was time-dependent, saturable and reversible by the addition of unlabelled albumin. The binding of 125I-labelled human albumin could be inhibited completely by unlabelled albumin preparations from humans, mice and dogs, and partly by bovine albumin. In contrast, binding of 125I-labelled human albumin was not inhibited by unlabelled rabbit albumin, human IgG or human fibrinogen. Data from competition experiments of two S. canis cultures with high 125I-labelled albumin-binding activities yielded KD values of 10 and 15 nmol l-1, respectively. The estimated number of binding sites per bacterial cell ranged from 30,000 to 57,000. The binding factor for albumin could be isolated from S. canis by boiling the bacteria at pH 2, and it was purified by affinity chromatography on human albumin-Sepharose. The isolated albumin-binding proteins had a molecular mass of approximately 51 kDa and inhibited binding of 125I-labelled albumin to S. canis. They formed complexes with human albumin that altered its electrophoretic mobility.

Albumins↗
PubMed1985

Clinical trial of common treatments for low back pain in family practice.

The results of a multicentered randomised clinical trial are reported of bed rest and of a physiotherapy and education programme for patients who presented in family practice with an acute episode of low back pain. No beneficial effect of either treatment was observed on several clinical outcome measures, including straight leg raising, lumbar flexion, activities of daily living, and pain. In fact the results favoured early mobilisation over bed rest and suggested that the physiotherapy and education programme was doing more harm than good. Moreover, additional analyses, which focused on clinically interesting patient subgroups, discovered no subset of patients who benefited from either of the treatments under study. Having failed to identify any clinically important benefits, or other explanations for these negative results, we can only conclude that family doctors have little reason to prescribe either bed rest or isometric exercises to patients who suffer from low back pain.

Back Pain↗
PubMed1984

[The lysozyme liberation test--a method for the demonstration of stimulation of human neutrophil granulocytes and monocytes].

A method for proof of induced secretion of lysozyme as a marker of lysosomal enzymes is described. Secretory cells are neutrophil granulocytes and monocytes. The stimulation of the cells is affected by soluble inductors. We used phytohemagglutinin (PHA), Concanavalin A (Con A), purified protein derivates (PPD) and lipopolysaccharide (LPS). Myelin basic protein has a direct stabilizing action on the membrane of lysosomes and induces an increased secretory reaction of cells after irritation. That is why it is recommended to add this substance as a supplement to the culture medium of the test. The optimal incubation parameters for the lysozyme release assay are ascertained. The method is recommended to apply in the fundamental investigation, for testing of stimulating or antiphlogistic agents and in the clinical functional diagnosis.

Adolescent↗
PubMedDate not supplied

Combined cytomorphologic and immunophenotypic analysis in the diagnostic workup of lymphomatous effusions.

OBJECTIVE: To analyze the results of cytomorphology and immunophenotyping in 54 patients with lymphomatous effusions. STUDY DESIGN: We report the results of cytomorphology and immunophenotyping in 54 patients with lymphomatous effusions. Twenty-three of the 54 had a previous diagnosis of NHL. In the remaining 31 patients, lymphomatous involvement was clinically suspected. RESULTS: Thirty-three lymphomatous effusions were positive for involvement by NHL. Twenty-one of these 33 patients (64%) had a previous diagnosis of NHL. Of the remaining 12 patients with newly diagnosed NHL, 11 had high grade lymphoma, and one had follicular center lymphoma. Twenty effusions were considered to be reactive; only two of these patients had NHL. One effusion revealed involvement by a previously unknown carcinoma. We observed seven false negative results if only one of both methods was considered. A high grade NHL was not diagnosed by immunophenotyping in one case, and six cases of low grade NHL could not be detected by cytomorphology. The combined strategy of cytomorphology and immunophenotyping had a sensitivity of 100% and specificity of 100% in our study, confirmed by follow-up studies. CONCLUSION: Both methods have shown difficulties in the examination of lymphomatous effusions. Cytomorphology has problems distinguishing reactive effusions from low grade NHL. The detection of high grade NHL by immunophenotyping is difficult. However, both methods together offer the advantage of dual staining ability and are most helpful in distinguishing clonal lymphomatous from reactive effusions.

Ascitic Fluid↗
PubMedDate not supplied

Fine needle aspiration cytology in extramedullary plasmacytoma.

OBJECTIVE: Extramedullary plasmacytoma is a rare plasma cell neoplasm. It can occur as the sole manifestation of plasma cell neoplasm, as a metastasis from another extramedullary plasmacytoma, as a solitary plasmacytoma of the bone or as a consequence of multiple myeloma. These plasma cell tumors can occur anywhere and have to be differentiated from other neoplasms, infectious processes and chloroma. STUDY DESIGN: We report the findings of fine needle aspiration cytology (FNAC) in 18 patients with extramedullary plasmacytoma. In six patients extramedullary plasmacytoma was the initial presentation of plasma cell neoplasm. In the remaining 12 patients the tumors occurred under or after treatment of plasma cell disease. RESULTS: Eleven lesions were located in the skin, seven in the lymph nodes, one in the liver and another in the spleen. Two patients with known diagnoses of plasma cell disease were thought, before FNAC, to have an infection, and two had a histologic diagnosis of non-Hodgkin's lymphoma. In 13 of 18 patients, cytologic smears showed anaplastic plasma cells. CONCLUSION: FNAC is a front-line investigative procedure in diagnosing extramedullary plasmacytoma.

Adult↗