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Biomedical subjects

J Garnier

Publications and source records attributed to J Garnier.

At least 91 records · Page 5Linked to original sources
PubMed1981

Changes in Leydig cells and luteinizing hormone receptors in porcine testis during postnatal development.

LH receptors have been characterized in porcine testis (Sus crofa L.) from birth to 220 days of age and have been related to interstitial tissue development (especially Leydig cells). The mean association constant (Ka) of ovine [3H]LH, was 7 +/- 6 X 10(9) M-1, with no apparent age-dependent variation but with some significant individual variations. The concentration of specific LH receptor sites reached a maximum of 9 X 10(-12) M/g testis between days 20-70, decreased to 3 X 10(-12) M/g testis at the onset of puberty (up to 100 days), and remained stable at the same level in the adult. The total number of sites per testis essentially reflected the growth of the testis. Interstitial tissue occupied up to 80% of the volume of the whole testis during the occupied up to 80% of the volume of the whole testis during the first 30 neonatal days. It decreased to about 25% after 120 days. The Leydig cells in this tissue occupied the same proportion (70% of the volume of the whole testis) regardless of the age of the animals. The mean Leydig cell diameter reached a maximum of 20 micrometer at 30 days of neonatal life, decreased to a minimum of 10 micrometer at 90 days, and then increased to a stable value of 15 micrometer after puberty. The number of Leydig cells per unit volume varied with age, with a maximum of 3 X 10(8) cells/ml testis at 90 days, reaching a constant value of 6 X 10(7) cells/ml testis after puberty. These data suggest that each Leydig cell contains 80,000 specific LH-binding sites/cell 30 days after birth and 35,000 in the adult, with no observed periods without receptor sites. The number of receptors per cell is correlated to cell size rather than stage of sexual maturation. (Endocrinology 108: 625, 1981)

Aging↗
PubMed1980

[Basic principles for characterization of hormonal membrane receptors. Application to assay methods (author's transl)].

Basic calculations for membrane receptor characterization, concentration and the affinity constant, are given including usual data linearization procedures. Determination of the specific radioactivity and maximum binding activity of the tracer and development of saturation curves from binding-inhibition experiments together with criteria for correction of non-specific binding, are presented both theoretically and practically in giving examples from the author's study of Luteinizing hormone (Lutropin) receptors in boar testis. The interest for physiologists of the receptor characterization is emphasized by underlying the regulatory aspect in hormone response of the variation of receptor concentration at the target cell surface in the case of spare receptors.

Animals↗
PubMed1980

[Malignant pheochromocytoma of Zuckerkandl's body; clinical data and treatment: one case (author's transl)].

A forty year old man with malignant pheochromocytoma of Zuckerkandl's body is reported. The case history is a severe hypertension with acute episodes, tachycardia and psychiatric disorders. The tumor extra adrenal site was got by abdominal scanning and its malignant type was demonstrated by pathological study and metastasis of lymph nodes occurring 4 months after surgery. The surgical treatment was associated to antimitotic and alpha blocking therapy. Clinical and biological status is still good after a twelve months follow-up.

Adult↗
PubMed1979

Photooxidation of the cytochrome b-559 in the presence of various substituted 2-anilinothiophenes and of some other compounds, in Chlamydomonas reinhardtii.

Five substituted 2-anilinothiophenes and two substituted carbonylcyanide-phenylhydrazones were comparatively studied with respect to their capacities for inducing photooxidation of the cytochrome b-559 in chloroplast fragments and in whole cells of Chlamydomonas reinhardtii (wild type and P-700-lacking mutant Fl 5). In addition, some other compounds: antimycin A, picric acid, tetraphenylboron and NH4Cl were also tested. Cytochrome b-559 photooxidations were clearly observed in the presence of 2-(3-chloro-4-trifluoromethyl)anilino-3,5-dinitrothiophene (ANT 2p), 2-(3,4,5-trichloro)anilino-3,5-dinitrothiophene (ANT 2s), 2-(4-chloro)anilino-3,5-dinitrothiophene and, with greater amplitudes, in the presence of carbonylcyanide-p-trifluoromethoxyphenylhydrazone and carbonylcyanide-m-chlorophenylhydrazone, both in whole cells and in chloroplast fragments. Picric acid, antimycin A and tetraphenylboron were also able to induce cytochrome b-559 photooxidation in chloroplast fragments, but not in whole cells. In the wild type, the highest photoinduced redox changes were 1.1 (carbonylcyanide-p-trifluoromethoxyphenylhydrazone, carbonylcyanide-m-chlorophenyl-hydrazone) and 0.6 (ANT 2p, ANT 2s) mumol of oxidized cytochrome b-559/1 mmol of chlorophyll, corresponding to 40% and 23% of the redox changes which could be induced chemically. All these cytochrome b-559 photooxidations, the greater part of which was inhibited by 3-(3,4-dichloropheny)-1,1-dimethylurea and occurred in the mutant Fl 5, appeared to be mainly Photosystem II-dependent reactions. But 3-(3,4-dichlorophenyl)-1,1-dimethylurea-insensitive Photosystem I-dependent photooxidations of cytochrome b-559 occurred also in the wild type. On the other hand, 2-(4-dimethylamine)-anilino-3,5-dinitrothiophene, 2-N-methyl-(3-chloro-4-trifluoromethyl)anilino-3,5-dinitrothiophene and NH4Cl did not induce any cytochrome b-559 photooxidation. These results were discussed taking in consideration the nature of the molecular substitutions of the various tested substances and their respective acceleration of the deactivation reactions of the water-splitting enzyme system Y of photosynthesis capacities which had been defined elsewhere by Renger (Renger, G. (1972) Biochim. Biophys. Acta 256, 428-439) for spinach chloroplasts. Like the acceleration of the deactivation reactions of the water-splitting enzyme system Y effect, the capacity for inducing the cytochrome b-559 photooxidation appeared dependent on the acidity of the NH group and on the number of halogenous substituents in the aromatic ring of the molecule. The greatest action towards cytochrome b-559 photooxidation was obtained with the most active acceleration of the deactivation reactions of the water-splitting enzyme system Y agents: carbonylcyanide-p-trifluoromethoxyphenylhydrazone, ANT 2p and ANT 2s.

Aniline Compounds↗
PubMed1979

Zoological origin of gonadotropin subunits and association kinetics.

Mammalian luteinizing hormone (LH) is an association of two dissimilar subunits, alpha and beta. In vitro studies, mainly using difference spectrophotometry, had shown that this phenomenon was slow, especially at low temperatures. If the situation was the same in poikiloterms, it would probably make gonadotropin (GTH) synthesis difficult for these animals in a cold environment. We have found that the formation of a teleost gonadotropin is in fact strikingly more rapid and less thermodependent than formation of mammalian LH. Also, studies with a fish-mammal hybrid molecule have allowed us to estimate the respective influence of the alpha and beta subunits in determining these differences.

Animals↗
PubMed1979

Effects of drugs on pigeon erythrocyte membrane and asymmetric control or adenylate cyclase by the lipid bilayer.

In pigeon erythrocyte membrane, the beta-adrenergic receptor and the enzyme adenylate cyclase can be uncoupled in two different ways depending on the type of drug used. Cationic drugs: chlorpromazine, methochlorpromazine, tetracaine, n-octylamine and a neutral alcohol, octanol, abolished alprenolol receptor binding ability and in the same range of concentration of the drug, sensitized adenylate cyclase to fluoride or Gpp(NH)p stimulation. Anionic drugs: di- and trinitro-phenols, indomethacin and octanoic acid did not affect the total number of beta-adrenergic receptor sites and, with the exception of trinitrophenol, did not change the association constant for alprenolol but they abolished the stimulation of adenylate cyclase by isoproterenol, fluoride or Gpp(NH)p. These modifications of the adenylate cyclase system occurred in a range of drug concentration where cell shape and protection against hemolysis were also affected. As chemical composition varies widely from one drug to another, it is suggested that these effects are largely nonspecific and mediated by the lipid bilayer. They are probably related to a preferential sidedness of action of the drugs in the lipid bilayer, displaying the role of an asymmetric control of the adenylate cyclase system in the membrane by the two halves of this bilayer.

Adenylyl Cyclases↗
PubMed1979

Receptor binding and conformational properties of bovine and ovine prolactins after chemical modification of the two tryptophan residues.

The two tryptophan residues in ovine and bovine prolactins were modified by reaction with 2-nitrophenylsulfenyl chloride in 75% formic acid. These derivatives exhibited an important loss of receptor affinity (less than 1% of the native hormones) to a rabbit mammary gland preparation. To a lesser degree, they also lost their binding affinity to specific guinea pig antibodies as detected by radioimmunoassay. The chemical modifications induced a change in the folding of the polypeptide chain, which in itself could be partly or totally responsible for the loss of biological or binding activities. This conformational change has been analyzed by circular dichroism and by prediction of secondary structures from the amino acid sequence using the method of Garnier et al. (Garnier, J., Osguthorpe, D.J. and Robson, B. (1978) J. Mol. Biol. 120, 97--120). The comparison of predicted prolactin and somatotropin structures revealed almost identical alpha-helix, turns and coil regions with an overall content of 67% alpha-helix, 5% beta-sheet, 17% turn and 11% of aperiodic structures. These values were close to those obtained from circular dichroism. The conformational change of the chemically modified hormones as compared to native folding, can be described as a partial loss of alpha-helical structure and an increase in beta-sheet content.

Amino Acid Sequence↗
PubMed1978

[Speed of in vitro reassociation of gonadotropin subunits in a teleost fish, the carp (Cyprinus carpio L.)].

Purified gonadotropin subunits of a teleost Fish, the Carp, reassociated in vitro, according to a second order rate reaction. The rate constant was temperature dependent. Compared to ovine LH, the Q10 was about three times less and the rate constant much greater (160 times for instance at 20 degrees C). The possible physiological significance of these differences is underlined.

Animals↗
PubMed1976

In vitro activation of glycoprotein hormones. Hybridization of subunits from thyrotropin, lutropin and human choriogonadotropin.

In vitro assembly of thyrotropin alpha and beta subunits led to an increase in content of alpha helix and beta sheet very similar to that found for gonadotropins. This association-dependent active folding involved the burying of three tyrosine residues tentatively assigned to Tyr alpha 41, Tyr beta 37 and Tyr beta 59 and common to all studied glycoprotein hormones. In vitro hybridizations between alpha and beta subunits of various hormones (thyrotropin, lutropin and choriogonadotropin) from different species (ovine, bovine and human) triggered the same molecular events as assembly of homologous subunits: the burying of three tyrosine residues and the increase of periodic structure of the folding. These changes are slow, time-dependent processes. Rates and yields of hybrid formation measured by sedimentation analysis and difference spectroscopy of tyrosines are identical, within experimental error, with the rates and yields measured by the recovery of the biological activity either the stimulation of chick thyroids for thyrotropin-beta hybrids or binding to porcine testis receptors for gonadotropin-beta hybrids. Whatever the origin of the alpha subunit, the thyrotropin-beta hybrids were not able to bind to testis receptors although active on chick thyroids. Rates and yields of hybrid formation essentially depended on the origin of the beta subunit. All the hybrids could be dissociated at acid pH with rates similar to those of native hormone. The extension to thyrotropin and various hybrids of the structural features of the in vitro assembly already recognized for gonadotropins strengthens the hypothesis that one deals with a basic activation process which also occurs in vivo after the synthesis of the subunits.

Animals↗
PubMed1976

Binding of gonadotropin-releasing hormone (LH-RH) to the pituitary plasma membranes and the problem of adenylate cyclase stimulation.

Specific binding studies of tritium-labeled LH-RH to sheep anterior pituitary membranes at 37 degrees C showed a maximum of binding capacity of 110 +/- 20 mol/mg protein with an association rate constant of 2 +/- 1 X 10(5) M-1 S-1 and a dissociation rate constant of 0.7 +/- 0.4 X 10(-2) S-1. The Scatchard plot data showed a single type of binding site with Kass = 1.1 +/- 0.4 X 10(8) M-1 in good agreement with the kinetic studies. Various doses of LH-RH in the presence or absence of Ca2+, were unable to stimulate adenylate cyclase either in the rat anterior pituitary homogenates, or in the purified sheep anterior pituitary plasma membranes. To explain these results, it may be argued that the proportion of gonadotrophs in the pituitary gland is too small to show a significant increase in the LH-RH-induced adenylate cyclase activity. Another possibility is the disconnection of the hormonal receptor from the site of activation of adenylate cyclase during the preparation of plasma membranes. Finally, cAMP may not be involved in LH release by LH-RH.

Adenylyl Cyclases↗
PubMed1976

Synthetic peptides for chymosin and pepsin assays: pH effect and pepsin independent-determination in mixtures.

Peptide I [H-Phe-Gly-His-Phe(NO2)-Phe-Ala-Phe-OMe] hydrolyzed by chymosin with kcat=.3+/-.3 s-1 and KM=7+/-3 mM (pH 4.7) inhibited competitively peptide II [H-Leu-Ser-Phe(NO2)-Nle-Ala-Leu-OMe] hydrolysis by chymosin with KI=.23 +/- .12 mM at pH 4.7. In reference conditions (.4 mM peptide, .01 M acetate buffer pH 4.7), the specific activities of porcine pepsin and chymosin on peptide I were 470 +/- 70 nM S-1 and .8 nM S-1 per mg of enzyme. This difference in specific activity for peptide I allowed development of a chymosin-independent pepsin assay for mixtures of these enzymes. In addition, peptide II with a specific activity of 2400 +/- 300 nM S-1 and 154 +/- 20 nM S-1 per mg of porcine pepsin and chymosin provides an alternative to measurement of milk clotting for measurement of chymosin- and pepsin-like activities in commercial rennets. Hydrolysis products of peptide II by chymosin exhibited one ionized group of apparent pK of 3.5 +/- .2 and a molar absorption coefficient change of 1000 +/- 100 at pH 4.7 and at 310 nm. From measurements of the kinetic constants, kcat and KM, from pH 2.5 to 7 with peptide II, chymosin activity depends on the protonation of one group of apparent pK 5.3 +/- .2 in the free enzyme. Rennet powder proved to be fairly stable after a 17-month storage at 4 C. Within the same period, a crystalline chymosin solution kept at --18 C lost 30 to 50% of its activity.

Animals↗
PubMed1975

[Cytochrome b-559 photooxidation in the presence of carbonyl cyanide p-trifluorometh-oxyphenylhydrazone and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone or p-benzoquinone in three non-photosynthetic mutants of Chlamydomonas reinhardti (author's transl)].

Studies of absorbance related to the cytochrome b-559 photooxidation induced by FCCP, with and without addition of 3-p-chlorophenyl-1, 1-dimethylurea (CMU), DBMIB or p-benzoquinone, in whole cells and in chloroplast fragments of Chlamydomonas reinhardti, were carried out. In addition to the wild type, three strains of non-photosynthetic mutants were used: Fl 5, which lacks P 700; Fl 9 and Fl 15, which are deficient in bound cytochrome c-553 and in cytochrome b-563. In the presence of FCCP, whole cells and chloroplast fragments of the four strains showed a System II-dependent photooxidation of cytochrome b-559. This photooxidation was inhibited by CMU but it occurred again in presence of FCCP, CMU and DBMIB. In chloroplast fragments, cytochrome b-559 photooxidation was also inhibited by an excess of FCCP; it was recovered, likewise, by addition of DBMIB. In whole cells, the highest measured redox changes were: 1 mu mol oxidized cytochrome b-559 per 1 mmol chlorophyll, corresponding approximately to about one seventh (wild type, Fl5) or one fifth (Fl 9, Fl 15) of the total amount of this cytochrome. Another kind of cytochrome b-559 photooxidation, CMU-insensitive, also occurred in the mutants Fl 9 and Fl 15 and in the wild type, but not in the mutant Fl 5. This latter kind of photooxidation was observed with chloroplast fragments in the presence of FCCP and CMU and also with whole cells in the presence of FCCP, CMU and p-benzoquinone. These reactions can be attributed to the Photosystem I; they do not require the intervention of the cytochrome c-553. A high-potential form of cytochrome b-559, hydroquinone-reducible, was involved in these two kinds of photooxidation. In addition, a lower potential form, reducible only by ascorbate, appeared to be able to interfere also. An interpretation is attempted, taking into consideration the various effects of FCCP and DBMIB, at different concentrations, on photosynthetic electron transport.

Chlamydomonas↗