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Biomedical subjects

H Metzger

Publications and source records attributed to H Metzger.

At least 127 records · Page 7Linked to original sources
PubMed1983

Use of the airfuge for analysis and preparation of receptors incorporated into liposomes: studies with the receptor for immunoglobulin E.

A Beckman Airfuge has been employed for studying the interaction between lipids and the receptor for immunoglobulin E (IgE). For analytic experiments, samples were applied underneath a discontinuous sucrose gradient. After a 30-min centrifugation in a fixed-angle rotor, liposomes floated toward the top of the gradient whereas unincorporated receptor-IgE complexes remained at the bottom of the tube. Liposomes with incorporated receptors were also efficiently separated in the ACR-90 preparative rotor. These methods of "Airfuge flotation" can provide useful adjuncts to more traditional methods for density-gradient centrifugation especially when rapid analysis of small samples is desired.

Animals↗
PubMed1983

Analysis of the rate-limiting step in a ligand-cell receptor interaction: the immunoglobulin E system.

Theory predicts that the kinetics of simple interactions between a ligand and a receptor bound on the surface of a cell will be affected by the occupancy of receptors on the same cell. In a diffusion-limited reaction the effect will be on the rate of dissociation but not on the rate of association until the cell is virtually saturated with ligand. If the rate of reaction is not diffusion limited, then the opposite holds; i.e., the forward velocities will be proportional to the concentration of vacant receptors, but the reverse reactions will not be. We examined the kinetics of reaction between immunoglobulin E (IgE) and its receptor and clearly demonstrated that the reaction is not diffusion controlled. The substantial (congruent to 30-fold) increase in the forward rate constant observed for the reaction of IgE with solubilized receptors as opposed to cell-bound receptors is therefore not an artifact of calculation. Since the reverse rate constants show little difference, we postulate that the presence of other surface components (rather than conformational differences in the receptor) affects the reaction with the cells. As an aid to the analysis, the theory has been extended so that not only the rate constants but also the entire course of the reaction of ligand with cell receptors can be predicted for diffusion-limited vs. non-diffusion-limited interactions.

Animals↗
PubMed1983

Structure-activity relationships for activation of adenylate cyclase by the diterpene forskolin and its derivatives.

Forskolin (7 beta-acetoxy-8,13-epoxy-1 alpha, 6 beta, 9 alpha-trihydroxylabd-14-en-11-one), a diterpene from the Indian plant Coleus forskohlii, activates cyclic AMP generating systems in a number of mammalian tissues in a rapid and reversible fashion. Derivatives of forskolin have been tested for their ability to stimulate membrane adenylate cyclase from rat brain and rabbit heart, as well as cyclic AMP generation in guinea pig brain vesicular preparations, a model system for intact cells. Derivatives at the 6 beta- and 7 beta-hydroxy functions retain activity, but none have greater activity than that of forskolin. Reduction of the 11-keto function affords an active 11 beta-hydroxy derivative. Reduction of the 14,15-vinyl (alpha) substituent reduces activity, while epoxidation abolishes activity. Derivatization or lack of the 1 alpha- and 9 alpha-hydroxy functions results in a marked reduction in activity, emphasizing the importance of the alpha aspect of the molecule. However, the 1 alpha, 6 beta-di-O-acetyl derivative does retain activity. None of the inactive derivatives, which include the 14,15-epoxy, the 1,9-dideoxy, and the 1,6-diketo derivatives, antagonize the stimulatory effects of forskolin.

Adenylyl Cyclases↗
PubMed1983

[Prenatal diagnosis of congenital cystic neck hygroma. Comparison of sonographic and computed tomography findings].

In a case of prenatal diagnosis of lymphangioma (hygroma) of the neck the results of sonographic and computer tomographic investigations were compared. It was shown that the methods are complimentary and lead to very similar conclusions. The first investigation should be by ultra-sound because of the lack of radiation. The computer tomography is then indicated with specific questions in mind and the radiation exposure can be minimized. In the presented case the combination of sonography and computer tomography permitted a prognosis of poor operability of the lesion. Since the prognosis cannot be determined with complete certainty prior to delivery the obstetrician and neonatalogist need to proceed with the possibility of a treatable anomaly in mind. The neckopsy findings in our case confirmed the inoperability of the lesion.

Adult↗
PubMed1983

Crosslinking of the receptors for immunoglobulin E depolarizes the plasma membrane of rat basophilic leukemia cells.

Aggregation of the receptor for IgE on mast cells, basophils, and a tumor analog, rat basophilic leukemia (RBL) cells, induces a calcium-dependent degranulation of the cells. We have measured the membrane potential (delta psi) of RBL cells during this reaction by using the tetraphenylphosphonium ion (Ph4P+) equilibration technique. We observed a 20-45% reduction in ionophore-sensitive Ph4P+ accumulation. The phenomenon persisted under conditions expected to collapse the mitochondrial membrane potential, consistent with the effect being due to a change in delta psi of the plasma membrane. We estimated that the change reflects a depolarization of 20 mV (from -90 to -70 mV, interior negative). Whereas degranulation fails to occur in the absence of external Ca2+, this was not true of the depolarization, indicating that the latter was not a consequence of secretion. When aggregation of the receptor is induced by reaction of the cell-bound IgE with a multivalent antigen, the secretory reaction can be halted by adding a univalent hapten. In this case, complete repolarization occurs. Equivalent depolarization was observed in the absence of Na+ but was diminished when both Ca2+ and Na+ were absent. Together, the data suggest that aggregation of the receptor opens ion channels and that the latter disappear promptly when the receptors are disaggregated. It is plausible that formation of these channels leads to the entry of Ca2+ and is an early and critical consequence of the aggregation of the receptors, thereby leading to degranulation.

Animals↗
PubMed1983

Isolation of cross-linked IgE-receptor complexes from rat macrophages.

Receptors for IgE on macrophages have been characterized by binding assays (1-3), but to date there has been only one report on the isolation of this receptor from macrophages, with use of the cell line U937 (4). In that report the receptor was isolated by using a heavily absorbed polyclonal antibody raised against lymphocytes bearing receptors for IgE (5). Monomeric IgE binds so weakly to macrophages that affinity chromatography of IgE-receptor complexes, such as has been used for isolation of the receptors for IgE on basophils (6) and for IgG on macrophages (7), cannot be readily accomplished. We have used oligomers of IgE to enhance the binding of IgE to macrophages (3), but this alone would not be sufficient because--depending on whether the receptors are multi- or univalent--once the cells are solubilized, multipoint attachment would again be reduced if not abrogated. In this report we describe the use of cross-linking reagents to stabilize further the interaction between IgE and its receptor on peritoneal macrophages. With this approach we have found that the receptor is likely to be composed of two chains whose gross properties are similar to the polypeptides constituting the receptor with high affinity for monomeric IgE on rat basophilic leukemia cells and mast cells.

Animals↗
PubMed1983

The action of forskolin on muscle cells is modified by hormones, calcium ions and calcium antagonists.

Forskolin, a drug isolated from the Indian plant Coleus forskohlii, exerts its actions on cells by directly activating the catalytic subunit of adenylatecyclases. The primary effect on heart muscles is the positive inotropic one, at higher forskolin concentrations, an acceleration of the pacemaker activity can be observed. External Ca2+ is required for this augmentation of contraction, verapamil, prenylamine and tetrodotoxin depress these effects. The action of forskolin and digitalis glycosides are to a certain degree additive. Incubation of rat heart slices with forskolin depresses the membrane bound Na+K+-ATPase activity. In smooth muscle cells, forskolin lowers the contractions, evoked by K+, norepinephrine and by angiotensin II. The results are discussed in respect to a forskolin induced enlargement of the Ca2+-uptake of heart muscle cells and a lowering of the Ca2+-sensitivity of the contractile system of smooth muscle cells.

Adenylyl Cyclases↗
PubMed1982

Phospholipids stabilize the interaction between the alpha and beta subunits of the solubilized receptor for immunoglobulin E.

The cell-surface component (alpha) which binds monomeric immunoglobulin E with high affinity is associated with a second polypeptide (beta) in the plasma membrane. The latter component tends to dissociate during purification of the alpha chain from detergent extracts of cells, even at neutral pHs and physiological ionic strengths. We now report that the interaction of alpha and beta can be stabilized by maintaining an appropriate phospholipid to detergent ratio. Under such conditions, other discrete components reproducibly copurify with the alpha and beta chains. These results suggest that the subunits of this membrane protein--or the interaction of it with other constituents in the cell--may be stabilized in ways not observed with ordinary soluble proteins.

Animals↗
PubMed1982

Phosphorylation of the receptor of immunoglobulin E.

Specific immune precipitation of immunoglobulin E(IgE)-receptor complexes from detergent extracts of 32P-labeled rat basophilic leukemia cells yielded a phosphoprotein of Mr approximately 35,000 on gel electrophoresis in sodium dodecyl sulfate. This phosphoprotein was shown by several criteria to be the beta chain of the receptor for IgE. Phosphorylation occurs at a serine residue (or residues) in a region (beta 2) of the beta chain that is thought to be exposed on the cytoplasmic face of the plasma membrane. Our results suggest that phosphorylation probably takes place after the insertion of the beta chain into the membrane. The IgE-binding alpha chain of the receptor and the IgE associated with it are not phosphorylated. We have so far been unable to detect any changes in the state of phosphorylation of either chain of the receptor or of IgE itself after IgE-mediated triggering of the cells.

Animals↗
PubMed1982

Gel filtration in 6 M guanidine hydrochloride of the alpha-subunit (and its fragments) of the receptor for immunoglobulin E.

The mol. wts of the alpha-chain of the receptor for immunoglobulin E and several of its enzyme-cleaved fragments have been evaluated by gel filtration on Sepharose 6B in 6 M guanidine HCl. The mol. wt of alpha-chains treated with endoglycosidase was 30% less than that of untreated alpha-chains. alpha-Chains digested with papain eluted in a single peak with a mol. wt approximately one-half of that of undigested alpha-chains. The results support the proposal that papain cleaves alpha-chains into two fragments of similar size [Goetze et al. (1981) Biochemistry 20, 6341-6349].

Chromatography, Gel↗