Iridoids from Scrophularia scorodonia.
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Glycosides↗
Biomedical subjects
R Faure
Publications and source records attributed to R Faure.
Quinovic acid glycosides from Nauclea diderrichii.
From the bark of Nauclea diderrichii (de Wild) Merr. seven saponins were isolated and characterized by 2D-NMR and FABMS. These compounds are quinovic acid glycosides previously reported for the Rubiaceae, six of them are described for the first time in this drug.
Carbohydrate Conformation↗
The myotonin-protein kinase phosphorylates tyrosine residues in normal human skeletal muscle.
As a first approach to study the cellular events involved in myotonic dystrophy, we have produced a polyclonal antibody against a peptide sequence of the predicted gene product. This antibody specifically recognizes a 54 kDa protein in human skeletal muscle. This protein phosphorylates a co-polymer Glu/Tyr but not Myelin Basic Protein. This indicates that the myotonin-protein kinase has a tyrosine kinase activity in human skeletal muscle. This is the first demonstration of the kinase activity of the myotonin-protein kinase.
Amino Acid Sequence↗
Peroxovanadium compounds. A new class of potent phosphotyrosine phosphatase inhibitors which are insulin mimetics.
Twelve peroxovanadium (pV) compounds, each containing an oxo ligand, one or two peroxo anions, and an ancillary ligand in the inner coordination sphere of V, were synthesized, crystallized, and characterized by 51V NMR as > 95% pure. These compounds activated the insulin receptor kinase (IRK) of cultured hepatoma cells, stimulated lipogenesis in adipocytes, and inhibited the in situ dephosphorylation of autophosphorylated IRs and epidermal growth factor receptors of rat liver endosomes. The phosphotyrosine phosphatase inhibitory and IRK activating potencies of these compounds were linearly correlated (r = 0.74; p < 0.003), decayed in parallel in solution, and varied considerably with the ancillary ligands within these compounds. In vivo administration activated rat liver IRK in parallel with its tyrosine phosphorylation. Co-administration of insulin plus pV was markedly synergistic in both respects. pV administration significantly decreased circulating insulin and plasma glucose concentrations; the latter to levels seen after a dose of insulin yielding > or = 50% occupancy of IRs in vivo. Two compounds (mpV(pic) and mpV(2,6-pdc)) displayed relative specificity as phosphotyrosine phosphatase inhibitors by inhibiting IR dephosphorylation to a significantly greater degree than epidermal growth factor receptor dephosphorylation. Thus, pV compounds are the most potent phosphotyrosine phosphatase inhibitors described to date. Their capacity to activate IRK appears to derive from their phosphotyrosine phosphatase inhibitory activity. Their hypoglycemic action is due to a direct tissue effect.
Adipocytes↗
Differential intracellular compartmentalization of phosphotyrosine phosphatases in a glial cell line: TC-PTP versus PTP-1B.
Intracellular levels of protein-phosphotyrosine are regulated by the protein-tyrosine phosphatase (PTP) family. Cellular compartmentalization may play an important role in modulating the function of these enzymes. The recent demonstration that PTP-1B is localized to the endoplasmic reticulum (Frangioni et al: Cell 68:545, 1992) is consistent with this proposition. In this study we have examined the intracellular distribution of TC-PTP in a glial cell line (C6). Using indirect immunofluorescence we have shown that this enzyme is distributed differently from PTP-1B and is mainly concentrated in the perinuclear region of these cells.
Amino Acid Sequence↗
1H-NMR study of suloctidil-A23187 calcium ionophore interaction.
Suloctidil is a molecule with calcium antagonist properties, whose anti-ionophoretic effect has previously been reported. In the presence of A23187 calcium ionophore free acid (A+), the NMR spectra of suloctidil (S +/-) are modified at the level of H-1 protons and to a lesser degree in the CH3-3 and aromatic regions. Experiments with one of the enantiomers of suloctidil and decoupling investigations led us to postulate the existence of diastereoisomers S+/A+, S-/A+ in the suloctidil +/-/A23187 + mixture. Moreover our results allow the hypothesis that suloctidil and calcium compete for the same binding site of the ionophore molecule.
Calcimycin↗
Triterpenoid saponins from Opilia celtidifolia.
Six saponins were isolated from leaves and stems of Opilia celtidifolia and their structures established. These saponins are described for the first time in this plant. One of them is a new saponin: 3-O-[alpha-L-rhamnopyranosyl(1-->3) beta-D-glucuronopyranosyl]-28-O-beta-D-glucopyranosyl-hederagen in.
Africa↗
The dephosphorylation of insulin and epidermal growth factor receptors. Role of endosome-associated phosphotyrosine phosphatase(s).
The autophosphorylation, from [gamma-32P]ATP, of insulin and epidermal growth factor receptors in rat liver endosomes peaked at 2-5 min and declined thereafter. When autophosphorylation from either [gamma-32P]ATP or unlabeled ATP was stopped after 5 min by adding excess EDTA +/- ATP, the phosphotyrosine (PY) content of each receptor decreased at 37 degrees C with a t 1/2 of 1.6 min. This was equally so whether the PY content of 32P-labeled receptors was analyzed by autoradiography of KOH-treated gels or by Western blotting with PY antibodies of immunoprecipitated receptors. The dephosphorylation reaction was strictly dependent on the presence of sulfhydryl, was unaffected by the addition of rat liver cytosol, and was temperature-dependent. The phosphotyrosine phosphatase(s) (PTPase(s)) appeared to be tightly anchored to the endosomal membrane, since the dephosphorylation reaction was unaffected by sodium carbonate and 0.6 M KCl treatments. However, treatment with Triton X-100 abolished dephosphorylation, implying an intimate association between the PTPase(s) and its substrate in an intact membrane environment. The powerful insulinomimetic agent pervanadate was the most potent inhibitor (50% inhibition at 1 microM). Increasing the dose of injected ligand augmented the rate of insulin and decreased that of EGF receptor dephosphorylation, respectively. Immunoblotting with specific antibodies failed to identify PTPase 1B or T-cell PTPase in ENs, whereas positive signals were seen in plasma membrane. These studies indicate that the phosphorylation state of receptor tyrosine kinases is dynamically regulated, with dephosphorylation, by closely associated PTPase(s), playing an important role.
Adenosine Triphosphate↗
Limits on neutron emission following deuterium absorption into palladium and titanium.
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Journal Article↗
Interaction of sodium molybdate with the thyroid hormone receptor.
The 3,5,3'-triiodothyronine (T3) binding activity of solubilized nuclear proteins from rat liver was decreased when molybdate (10 mM) was present in the incubation medium in the absence of thiol reagents. The equilibrium affinity constant was reduced by 40%. The rate of degradation of T3-receptor complexes at 37 degrees C remained unchanged, but when the extracts were further reincubated in the presence of beta-mercaptoethanol, molybdate had a protective effect after 5 h incubation at 37 degrees C. In contrast, the thyroxine (T4) binding activity was not affected by heating at 37 degrees C or by molybdate. Ion-exchange chromatography confirmed the existence of a molybdate-receptor interaction: the T3-receptor complexes shifted from elution at 0.22 to 0.20 M NaCl with the progressive appearance of a small leader peak, whereas the T4-receptor complexes eluted in a large and split peak (0.22-0.4 M NaCl). The destabilizing effect on T3 binding induced by exogenous dephosphorylation is more efficiently reversed by beta-mercaptoethanol when the extracts were pretreated by molybdate. In controls, the loss of saturable T3 binding activity was recovered by 50% at a 10 mM concentration of beta-mercaptoethanol, but in the presence of molybdate, the loss of T3 binding activity was recovered by 50% at a 5 mM concentration of beta-mercaptoethanol. This molybdate-receptor interaction is similar to that with nuclear receptor models in term of (i) stabilization of hormone binding, (ii) dependency on a thiol, and (iii) reversibility of the destabilizing effect by exogenous dephosphorylation.
Alkaline Phosphatase↗
Hepatic protein phosphotyrosine phosphatase. Dephosphorylation of insulin and epidermal growth factor receptors in normal and alloxan diabetic rats.
Polypeptide hormone signal transmission by receptor tyrosine kinases requires the rapid reversal of tyrosine phosphorylation by protein phosphotyrosine phosphatases (PPTPases). We studied hepatic PPTPases in the rat with emphasis on acute and chronic regulation by insulin. PPTPase activity with artificial substrates ([32P]Tyr-reduced, carboxyamidomethylated, and maleylated lysozyme and [32P]Tyr-poly[glutamic acid:tyrosine] 4:1) was present in distinct membrane, cytoskeletal, and cytosolic fractions. These PPTPase activities were unaffected by alloxan diabetes. Acute administration of insulin to normal animals also did not change PPTPase activity in liver plasma membranes or endosomal membranes. Although alloxan diabetes did not affect PPTPase activity measured with artificial substrates or with epidermal growth factor receptors, a decrease in insulin receptor dephosphorylation was noted. Dephosphorylation of hepatic receptors from normal and diabetic rats by membrane PPTPase from control rats was similar. These results indicate that alloxan diabetes does not lead to a generalized effect on hepatic PPTPase activity, although a substrate-specific decrease in activity with the insulin receptor may occur.
Animals↗
Immunocytochemical localization of the nuclear 3,5,3'-triiodothyronine receptor in the adult rat: liver, kidney, heart, lung and spleen.
A monoclonal antibody was used for the localization of the nuclear T3 receptor in different tissues of the adult rat: the liver, kidney, heart, lung, spleen, testis, and pituitary. In the liver, the immunoreactivity was found uniformly distributed in the nuclei of hepatocytes. Sections incubated with a control ascitic fluid or with the same ascitic fluid pre-adsorbed with purified receptor showed no specific staining. In the kidney, the immunoreactivity was higher in the epithelial cell of the proximal convoluted tubes and juxtaglomerular cells. In the heart, only the myocardial cells were stained. In the lung, the immunoreactivity was confined to type II pneumocytes and alveolar macrophages. In the spleen, only a few mature lymphocyte and macrophage cell nuclei were stained. These results show that: 1) the abundance of the nuclear T3 correlates with previous studies using hormone binding techniques; 2) the nuclear T3 receptor is selectively located in certain cell types, which possess a precise local function.
Animals↗
[Flavonoids of 3 cultivars vine leaves, Vitis vinifera L. var. tinctoria (Alicante, Carignan, Grand noir). Value in chemical control].
Three flavonoids were isolated and identified from the leaves of 3 cultivars vine Vitis vinifera L. var. tinctoria (Alicante, Carignan and Grand noir): hyperin, isoquercitrin and quercetin 3-O-beta-D glucuronic acid. TLC analytic control and HPLC determination are proposed in this paper.
Chromatography↗
A monoclonal antibody to the rat nuclear triiodothyronine receptor: production and characterization.
The nuclear T3 receptor (NTR) was affinity-labeled with bromoacetyl-[125I]T3, purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and used to immunize BALB/c mice. Spleen cells from one strongly immunoreactive mouse were fused with Sp2 mouse myeloma cells, and 328 hybridomas were screened by a dot-blot immunoassay using as antigen, a preparation of NTR partially purified by diethylaminoethyl-Sephadex chromatography. Four positive cultures were thus found; three of which were confirmed by comparing Western blotting patterns with the electrophoretic mobility of the affinity-labeled NTR. One of these 3 hybridomas was further subcloned by limiting dilution and gave rise to the 2B3 clone, which produces an immunoglobulin of the immunoglobulin G1 subclass. Several lines of evidence indicated that the 2B3 monoclonal antibody was indeed directed against the NTR. The antibody recognized a protein with the same electrophoretic mobility as the affinity-labeled receptor. Thus, Western blotting revealed a predominant protein with a mol wt of 57,000 and a less abundant 45,000 component on sodium dodecyl sulfate gels, and multiple isoelectric variants of the 57,000 protein, with a predominant form at pI 6.2, were detected on two-dimensional gels. Incubation of the 2B3 antibody with the NTR labeled with [125I]T3 resulted in the formation of an antibody-receptor complex, as indicated by a shift of the radioactivity peak upon gel filtration on Sephacryl S-300. In contrast, control ascitic fluid did not change the elution profile of the labeled NTR. The 2B3 antibody is able to remove the T3-binding activity from rat liver nuclear extracts. Finally, in accordance with previous T3-binding experiments, expected amounts of NTR were found in pituitary, liver, brain, kidney, spleen, and testis with the use of the Western blotting technique and immunohistochemistry on frozen tissue sections. This antibody should prove useful in the characterization and purification of the NTR and also in the study of its distribution in different tissues and cell types.
Animals↗
Effects of adenosine triphosphate and alkaline phosphatase on solubilized 3,5,3'-triiodothyronine-binding activity.
The T3-binding activity of salt-extractable nuclear proteins from rat liver was affected when ATP (2-10 mM; pH 8.0) was added concomitantly with T3 in the incubation medium. Scatchard analysis revealed that the equilibrium association constant was significantly reduced [5 mM ATP, 0.3 +/- 0.1 (+/- SE) 10(10) M-1; control, 1.1 +/- 0.15 X 10(10) M-1], but the maximum binding capacity remained unchanged. Similar values of inhibition were obtained when unbound receptors were preincubated with ATP. ATP achieved its maximal effect after 45 min of incubation at 30 C. Dilution experiments indicated that the effect of ATP was reversible. The inhibiting potency of nucleoside triphosphates at pH 8.0 was in the following order: ATP = CTP greater than GTP, whereas UTP had no effect. Nonhydrolyzable analogs of ATP were also inhibitory, and HPLC fractionation showed an approximately 98% recovery of ATP after incubation with nuclear extract. The adenine ring with at least two phosphates was essential, since ADP was as potent as ATP, whereas AMP had no effect. When the pH of the incubation medium was lowered to 7.3, the T3-binding activity was inhibited by ATP in the 0.1-1 mM range. Magnesium (3 mM) greatly increases the ATP effect at pH 7.3, but not at pH 8. The T3-binding activity was also drastically reduced when calf intestine alkaline phosphatase was added concomitantly in the incubation medium. Eight micrograms per ml enzyme were necessary to inhibit the T3 specific binding by 50% (30 C for 45 min). Scatchard analysis showed that the receptor affinity for T3 was decreased (control, 1.1 +/- 0.02 x 10(10) M-1; alkaline phosphatase, 0.41 +/- 0.03 x 10(10) M-1; n = 6), whereas the maximum binding capacity remained unchanged. Incubations performed with increasing concentrations of beta-mercaphoethanol (2.5, 5, 10, and 25 mM) revealed that the phosphatase inhibitory effect is thiol dependent. The inhibition was maximal at 2.5 mM and progressively decreased at 5 and 10 mM. No inhibition occurred at 25 mM. When a saturating concentration of T3 was employed, the specific binding was decreased at low thiol concentrations. These observations show that the nuclear T3 receptors may be modulated by ATP/ADP and phosphorylation/dephosphorylation processes. It is proposed that in vitro dephosphorylation leads to rapid oxydation of sulfhydryl groups which are essential for optimum T3 binding.
Adenosine Triphosphate↗
Ontogenesis of nuclear T3 receptors in primary cultured astrocytes and neurons.
Nuclear T3 receptor (NTR) have been characterized in separated cultures of neurons and astrocytes. Scatchard analysis indicated the presence of a single class of high-affinity sites in both cell lines. The apparent equilibrium association constant ranged from 1.80 +/- 0.41 X 10(10) M-1 to 3.27 +/- 0.74 +/- 10(10) M-1 in neurons and from 1.01 +/- 0.09 to 1.80 +/- 0.73 X 10(10) M-1 in astrocytes depending on the time in culture. In neurons, the maximal binding capacity (MBC) increased from 0.049 +/- 0.008 ng T3/mg DNA to 0.328 +/- 0.052 ng T3/mg DNA between 3 and 12 days of culture. In astrocytes, the changes in MBC were less pronounced ranging from a minimum of 0.095 +/- 0.024 ng T3/mg DNA at the 7th day of culture to a maximum of 0.198 +/- 0.048 ng T3/mg DNA at the 21st day. The relative binding affinity of the receptor for thyroid hormone analogs was in the order TRIAC greater than L-T3 greater than D-T3 greater than L-T4 in both cell lines. These results show that nuclear T3 receptors similar to those found in vivo are present in primary cultures of both astrocytes and neurons.
Animals↗
Production of an antibody against rat liver nuclear T3 receptor.
Rabbits were immunized with rat liver nuclear L-triiodothyronine (T3) receptor purified by preparative sodium dodecyl sulfate-polyacrylamide (SDS-PAGE) gel electrophoresis using bromoacetyl[125I]T3 as an affinity label. SDS-PAGE confirmed the presence of two receptor forms of the apparent molecular weights 57,000 and 45,000. We describe here a specific antibody, raised against the 57,000 receptor type, which reacts with both receptor forms as assessed by electroimmunoblotting and immunoprecipitation in liquid medium.
Animals↗
Regional distribution of nuclear T3 receptors in rat brain and evidence for preferential localization in neurons.
We examined the distribution of nuclear T3 in mature rat brain with the aim of determining specific targets of thyroid hormones within this tissue. Saturation experiments, performed in 9 different structures of the brain and in 4 parts of the cortex, revealed the presence of a single class of binding sites with a mean Ka of 0.53 X 10(10) M-1. The highest concentrations of receptors were found in the amygdala (0.523 +/- 0.025 ng T3/mg DNA, Mean +/- SE) and the hippocampus (0.438 +/- 0.071 ng T3/mg DNA) while the lowest were in the brain stem (0.058 +/- 0.003 ng T3/mg DNA) and the cerebellum (0.079 +/- 0.026 ng T3/ml DNA). The receptor was not uniformally distributed within the cerebral cortex, its concentration being relatively high in the central sections and intermediate in the remaining portions. The cell type distribution of the T3 receptor was studied by separating glial and neuronal nuclei on a discontinuous sucrose gradient. There was no detectable specific T3 binding in the fraction of oligodendrocyte nuclei (approximately 95% pure). Conversely, the neuron-enriched fraction (approximately 60%) showed a significant increase in receptor concentration compared to total nuclei (35-40% neurons): 0.857 +/- 0.196 vs 0.511 +/- 0.095 ng T3/mg DNA (p less than 0.01) in the cortex and 0.425 +/- 0.018 vs 0.234 +/- 0.24 ng T3/mg DNA (p less than 0.01) in the forebrain. The absence of nuclear T3 receptors in oligodendrocytes may have important implications on the mechanism of action of thyroid hormone in myelination.
Animals↗