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Biomedical subjects

C Klein

Publications and source records attributed to C Klein.

At least 307 records · Page 17Linked to original sources

Morphometric analysis of the angioarchitecture of the synovial membrane in rheumatoid arthritis and osteoarthritis.

Using three different immunohistochemical methods we measured the number of vessels, vessel area and diameter and their form factor in the synovial membrane of 102 patients suffering from different joint disease. The variables were evaluated by means of immunomorphometric analysis. We found UEA (Ulex europeus) immunostaining to be the optimal method for quantification of data characterizing the vasculature of the synovial membrane. Irrespective of causes of the given joint disease, we found increases in the number of vessels, vessel perimeter, vessel area and the product of the number of vessels and vessel area over and against the control group (patients without arthritis). Consequences are discussed regarding local bioavailability of medicaments in the synovial membrane.

Arthritis, Rheumatoid↗

Structure of cyclodextrin glycosyltransferase refined at 2.0 A resolution.

The previously reported structural model of cyclodextrin glycosyltransferase (EC 2.4.1.19) from Bacillus circulans has been improved. For this purpose the known sequence was built into an electron density map established by multiple isomorphous replacement and subsequent solvent-flattening at 2.5 A resolution. The resulting model was refined at 2.0 A resolution using a simulated annealing refinement method. Based on 70,171 independent reflections in the range 7.0 to 2.0 A resolution, a final R-factor of 17.6% was obtained with a model obeying standard geometry within 0.013 A in bond lengths and 2.7 degrees in bond angles. The final model consists of all 684 amino acid residues, two calcium ions and 588 solvent molecules.

Amino Acid Sequence↗

Yeast signal peptidase contains a glycoprotein and the Sec11 gene product.

Partially purified yeast microsomal signal peptidase appears to be a complex of four polypeptides of 13, 18, 20, and 25 kDa. The 18-kDa chain is the product of the Sec11 gene, which is necessary for signal peptidase activity. The 25-kDa subunit is a glycoprotein that binds Con A. Two related methods for purification of the enzyme are presented; the first includes removal of peripheral membrane proteins from microsomes by alkali extraction, solubilization of the enzyme by nonionic detergent and high salt, and four different chromatographic procedures. An alternative method was developed based on lectin-affinity chromatography.

Chromatography↗

Regulation of protein phosphorylation in Dictyostelium discoideum.

We have examined the phosphorylation of the cyclic adenosine 3':5' monophosphate (cAMP) cell surface chemotactic receptor and a 36 kDa membrane-associated protein (p36) in Dictyostelium discoideum. The activity of CAR-kinase, the enzyme responsible for the phosphorylation of the cAMP receptor, was studied in plasma membrane preparations. It was found that, as in intact cells, the receptor was rapidly phosphorylated in membranes incubated with [gamma 32P] adenosine triphosphate (ATP) but only in the presence of cAMP. This phosphorylation was not observed in membranes prepared from cells which did not display significant cAMP binding activity. cAMP could induce receptor phosphorylation at low concentrations, while cyclic guanosine 3':5' monophosphate (cGMP) could elicit receptor phosphorylation only at high concentrations. Neither ConA, Ca2+, or guanine nucleotides had an effect on CAR-kinase. It was also observed that 2-deoxy cAMP but not dibutyryl cAMP induced receptor phosphorylation. The data suggest that the ligand occupied form of the cAMP receptor is required for CAR-kinase activity. Although the receptor is rapidly dephosphorylated in vivo, we were unable to observe its dephosphorylation in vitro. In contrast, p36 was rapidly dephosphorylated. Also, unlike the cAMP receptor, the phosphorylation of p36 was found to be regulated by the addition of guanine nucleotides. Guanosine diphosphate (GDP) enhanced the phosphorylation while guanosine triphosphate (GTP) decreased the radiolabeling of p36 indicating that GTP can compete with ATP for the nucleotide triphosphate binding site of p36 kinase. Thus was verified using radiolabeled GTP as the phosphate donor. Competition experiments with GTP gamma S, ATP, GTP, CTP, and uridine triphosphate (UTP) indicated that the phosphate donor site of p36 kinase is relatively non-specific.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding, Competitive↗

Dual role of GDP in the regulation of the levels of p36 phosphorylation in Dictyostelium discoideum.

We examined the dephosphorylation of p36, a protein of D. discoideum that has previously been shown to be phosphorylated in a GDP-dependent manner (Anschutz et al., 1989). Specific dephosphorylation of p36 was found to occur in cell preparations but the activity responsible was strongly dependent upon the concentration of proteins in those extracts. When preparations were diluted, this activity was no longer detectable and the radiolabeled phosphate incorporated into p36 was stable. In contrast, p36 phosphorylation was seemingly unaffected by this treatment. Under the conditions where endogenous dephosphorylating activity was not detectable, the addition of GDP to the reaction resulted in substantial dephosphorylation of p36. The stimulation of this dephosphorylation process occurred at concentrations of GDP that were distinct from those that led to an increased p36 phosphorylation due to the previously reported stimulation of p36 protein kinase activity. Characterization of the dephosphorylation of p36 indicates that the same enzyme is responsible for the endogenous and GDP-stimulated activities. Additionally, these activities are identical when assayed with p36 that had been phosphorylated with ATP or GTP. In contrast to p36 kinase activity, the dephosphorylation of p36 did not display any developmental changes with respect to its regulatory features.

Animals↗

Tiapride as treatment for certain patients with idiopathic torsion dystonia.

Five female patients with idiopathic torsion dystonia (ITD) responded to treatment with tiapride, a selective D-2 dopamine antagonist. Previous treatments with various drugs, including anticholinergics, were ineffective. It is suggested that the previously reported genetic subgroups of ITD respond to different drug regimens. The autosomal dominant group responded to anticholinergics, while the autosomal recessive group responded best to a selective D-2 dopamine antagonist.

Adult↗

Relief of right ventricular angina and increased exercise capacity with long-term oxygen therapy.

Long-term low-flow oxygen therapy can lead to improved exercise capacity and improved hemodynamics in selected patients with pulmonary hypertension. We report a patient who presented with severe exercise limitation and anginal chest pain that appeared to result from pulmonary hypertension and predominantly right ventricular ischemia. Acute oxygen therapy led to relief of pain but no change in exercise capacity or of pulmonary hypertension. After eight months of oxygen therapy, the patient's pulmonary hypertension was unchanged, but right ventricular hypertrophy and marked increases in exercise cardiac output and exercise capacity developed. Thus, oxygen can relieve right ventricular angina and facilitate the development of compensatory hypertrophy.

Angina Pectoris↗

[Serological diagnosis of viral hepatitis].

The differentiation and classification of different types of viral hepatitis was essentially improved in the laboratory diagnosis by the direct determination of the virus in serum, viral antigens or their antibodies. Especially for the hepatitis B it is possible to recognize the infectivity by the serum marker HBeAg, the prognosis by seroconversion (HBeAg/HBeAb), the epidemiology, and vaccination prophylaxis, respectively. Without doubt, immunological marker have an increasing significance for prophylaxis, diagnosis, and therapy of viral hepatitis.

Diagnosis, Differential↗

[Histomorphometrical study of physiological tooth movements in humans].

Quantitative analysis of remodeling activity was done using microradiographs of horizontal ground sections of human alveolar bone obtained from autopsies. The results demonstrated no significant age- or sex-dependent variations. The local distribution of remodeling activity in younger individuals pointed to a tipping mesial movement of the teeth. In older individuals this distribution showed a more random pattern.

Adult↗

Localization of functional domains of the cAMP chemotactic receptor of Dictyostelium discoideum.

The topography and functional domains of the cAMP chemotactic receptor of Dictyostelium discoideum were investigated by protease sensitivity to chymotrypsin. Proteolytic digestion of intact cells produced a 23-kDa fragment of the receptor that retained the photoaffinity label used to identify the receptor. Additionally, this fragment contained the sites phosphorylated by CAR-kinase, the enzyme that phosphorylates the ligand-occupied form of the receptor. The fragment was also found to be phosphorylated in response to cAMP stimulation of cells. Proteolytic digestion of either intact cells or membrane preparations did not appreciably alter the binding properties of the receptor, indicating that the domains which determine the cAMP binding pocket are likely to be transmembrane regions of the protein. Additionally, the sensitivity of down-regulated receptors to chymotrypsin digestion suggests that the initial loss of cAMP binding activity upon incubation of cells with high concentrations of ligand does not require receptor internalization.

Chemotactic Factors↗

Determination of the active metabolite of molsidomine in human plasma by reversed-phase high-performance liquid chromatography.

A reversed-phase high-performance liquid chromatographic method, with ultraviolet detection, is proposed for the plasma determination of SIN-1, the active metabolite of molsidomine, which involves propoxycarbonyl derivatization. The internal standard is the ethoxycarbonyl derivative of SIN-1 (i.e. molsidomine). Derivatization and extraction are each performed in one step (2 min) with 70% yield. The nature of a by-product is discussed. The method provides rapid elution (less than 15 min), linearity over the range 0.4-200 ng/ml, day-to-day precision between 2.5 and 11.3% and a limit of determination of 0.5 ng/ml. This method is also suitable for the simultaneous determination of molsidomine and SIN-1. In this case the internal standard is an ethoxycarbonyl derivative of a piperazino-3-sydnonimine, a SIN-1 analogue.

Antihypertensive Agents↗

cAMP stimulation of Dictyostelium discoideum destabilizes the mRNA for 117 antigen.

Transcription of the 117 gene and changes in its mRNA levels in Dictyostelium discoideum were studied by mRNA hybridization with a cDNA probe. In wild type cells (Ax-2), the expression is developmentally regulated during cell aggregation, while in the aggregateless mutant, Agip 45, 117 mRNA is not detectable during cell starvation. Low concentrations of cAMP, given in the form of extracellular pulses to induce the development of starved Agip 45 cells to aggregation competence, are able to induce the appearance of 117 mRNA. The induction seems to be via the cell surface cAMP receptor and by a mechanism which does not involve changes in intracellular cAMP. Interestingly, high concentrations of cAMP, which down-regulate the cell surface cAMP receptor, elicit a rapid decrease in the level of 117 mRNA in aggregation-competent cells. Nuclear run-off and pulse-chase experiments show that the high concentrations of cAMP selectively destabilize the mRNA for 117 antigen. This destabilization requires both de novo mRNA synthesis and protein synthesis since the addition of inhibitors of these processes eliminates the effects of cAMP on 117 mRNA. The data suggest that a cAMP-induced protein(s) may be involved in the destabilization of selective mRNAs.

Adenylyl Cyclase Inhibitors↗

Characterization of a GDP-sensitive phosphorylation in plasma membranes of D. discoideum.

In a previous study, we reported the GDP-dependent phosphorylation of a 36 kD membrane protein, p36, in D. discoideum membranes prepared from starved (aggregation competent) cells (Anschutz et al., 1989). Here we show that p36 can be phosphorylated when membranes are supplied either ATP or GTP as the phosphate donor, but that a greater level of p36 phosphorylation is achieved with GTP. The rate of phosphorylation of p36, using either nucleotide triphosphate, is enhanced by GDP. This reflects a decrease in the apparent Km of the enzyme for the particular nucleotide triphosphate. p36 can also be phosphorylated in membranes prepared from vegetative cells. However, the ability of GDP to stimulate p36 phosphorylation is not observed in vegetative cell membranes. Competition experiments indicate that there are also developmental differences in the nucleotide triphosphate site(s) available to phosphorylate p36.

Adenosine Triphosphate↗

Properties of CAR-kinase: the enzyme that phosphorylates the cAMP chemotactic receptor of D. discoideum.

The cell surface cAMP chemotactic receptor of D. discoideum can be phosphorylated in partially purified plasma membrane preparations in a ligand-dependent manner. CAR-kinase, the enzyme responsible for receptor phosphorylation, was shown to be an integral membrane protein. It could utilize either ATP or GTP to phosphorylate the receptor, although ATP was much more efficient. The apparent affinity constant for ATP was approximately 20-25 microM. Maximum CAR-kinase activity was observed between pH 6.5 and pH7, and required the presence of Mg2+. Neither Mn2+ nor Ca2+ could substitute for that divalent cation. The enzyme was found to be sensitive to the ionic strength and temperature of the incubation reaction. Dephosphorylation of the receptor was not observed in the membrane preparations, indicating that the enhanced level of receptor phosphorylation that occurred upon ligand binding was not an indirect reflection of receptor dephosphorylation and subsequent incorporation of radiolabeled phosphate.

Animals↗

Cell adhesion in transformed D. discoideum cells: expression of gp80 and its biochemical characterization.

Dictyostelium discoideum amoebae were transformed with an expression vector for the gp80, a protein believed to mediate EDTA-resistant cell adhesion in developmental cells. Vegetative cells, that do not normally contain gp80, expressed the protein and this expression was correlated with the formation of cell-cell adhesions. These contacts exhibited minimal EDTA-resistance. Biochemical analyses of the protein synthesized by vegetative cells suggested that it is identical to that produced by aggregation-competent cells, including the presence of a glycolipid anchor. Additional experiments indicated that the anchor was insensitive to hydrolysis by exogenous (glycosly)phosphatidylinositol-specific phospholipase C [G)PI-PLCs) but was sensitive to the endogenous anchor degrading enzyme. This enzyme, initially described in aggregating cells (da Silva and Klein, Exp. Cell Res., in press) was found to be present also in vegetative amoebae.

Blotting, Northern↗

Primary and diagenetic controls of isotopic compositions of iron-formation carbonates.

Mineralogic, chemical, and isotopic compositions have been determined for 97 carbonate microbands in five core segments from the early Proterozoic (2.5 Ga) Dales Gorge Member of the Brockman Iron Formation, Hamersley Basin, Western Australia. Samples were obtained both from banded iron-formation (BIF) macrobands 9-12 at Paraburdoo, on the southern margin of the basin, and from BIF macroband 13 at Wittenoom, 130 km NW. At Paraburdoo, oxygen-isotopic compositions of coexisting chert and magnetite microbands were measured and indicated final equilibration temperatures ranging from 60-160 degrees C. This range is consistent with observed mineral assemblages and indicates a considerable temperature gradient across the basin (cf. T approximately 300 degrees C at Wittenoom; BECKER and CLAYTON, 1976). Carbon-isotopic compositions of carbonates are near -7% vs. PDB at Paraburdoo and -10.5% at Wittenoom, but the greater isotopic depletion at Wittenoom appears related to primary or diagenetic processes, not metamorphism. Contents of organic carbon are consistently low. Isotopic depletion is roughly correlated with iron abundance and, together with petrographic observations and chemical balances, is consistent with the model of BIF deposition introduced by BEUKES et al. (1990): primary siderite (delta approximately -5%) precipitated from an anoxic water column depleted in 13C; additional depletion of 13C is associated with coprecipitation of iron oxides and organic carbon. Oxygen-isotopic abundances of microbanded carbonates are similar to those under- and overlying massive marine carbonates, ranging from 17.6 to 21.0% vs. SMOW (-9.6 to -12.9% vs. PDB). Millimeter-scale variations in abundances of 13C and 18O are associated with diagenetic replacement of primary siderite by secondary ankerite and/or magnetite. It is shown that these isotopic variations cannot result from mineral-dependent fractionations, metamorphism, or the influence of large volumes of water in an open system.

Carbon↗

The effect of age on the sensitivity of the alpha 1-adrenoceptor to phenylephrine and prazosin.

Certain beta-adrenoceptor-mediated functions seem to diminish with age; however, information on alpha-adrenoceptor-mediated function is sparse and often conflicting but overall suggests little age-related change. To assess an age-related alteration in the responsiveness to an alpha 1-adrenergic agonist and to estimate changes in the apparent affinity of the alpha 1-adrenoceptor for the antagonist prazosin, we infused phenylephrine into 12 healthy elderly subjects and 12 healthy young subjects before and after an oral dose of prazosin, and we compared the shift in the dose-response curves for the two groups. With this protocol we were unable to detect any age-related decline in sensitivity of the alpha-adrenoceptor to either agonist or antagonist. However, oral prazosin resulted in higher plasma concentrations and a consistently greater hypotensive effect in the elderly subjects than in the young subjects. We concluded that there was no difference in alpha 1-adrenoceptor sensitivity in the elderly persons, but that the kinetics of prazosin may be altered and that the response of the blood pressure to prazosin was increased because of the kinetic changes and possibly the physiologic changes associated with aging.

Adult↗