Hyperhomocysteinemia and low B vitamin levels are independently associated with venous thromboembolism: results from the EDITH study: a hospital-based case-control study.

BACKGROUND: Moderate hyperhomocysteinemia and B vitamins deficiency are thought to be risk factors for venous thromboembolism (VTE). The causality and independence of those associations are still questioned. METHODS: We measured fasting serum total homocysteine, folates, and vitamin B12 levels as well as 5,10-methylenetetrahydrofolate reductase (MTHFR) C677T genotypes in 467 patients hospitalized with a first well-documented deep vein thrombosis and/or pulmonary embolism not related to a major acquired risk factor and 467 controls matched for gender and age. RESULTS: Mild hyperhomocysteinemia, low serum folates, and vitamin B12 were associated with VTE independently of each other. In multivariate analysis, odds ratios (OR) (95% CI) for VTE associated with mild hyperhomocysteinemia (>15 micromol L(-1)), low serum folates (< or = 4.9 nmol L(-1)), and vitamin B12 (< or = 253 pmol L(-1)) were 1.48 (1.05-2.08), 3.14 (1.35-7.32) and 1.42 (1.03-1.98), respectively. An MTHFRC677T genotype was not significantly associated with VTE; OR (95% CI): 1.13 (0.70-1.81) CONCLUSIONS: The current data provides further knowledge in the complex relationship between hyperhomocysteinemia, low vitamin levels, and VTE.

Androgen regulation of secretions in the sebaceous-like uropygial gland of the male Japanese quail.

The uropygial gland of the male quail is a sebaceous-like, androgen-dependent structure. The waxes secreted by this gland consist of fatty acids esterified by alkane-2,3-diols. In adult male quail the relative concentrations of all fatty acids were not affected by castration and testosterone treatment; but in contrast, the relative concentration of dodecane diol was found to be correlated with the androgen levels. In castrated quail administration of testosterone induced an increase in the dodecane diol percentage which was blocked by protein-synthesis inhibitors. Furthermore, the effects of testosterone were found to be neutralized by the administration of cyproterone acetate. Consequently it may be stated that the relative concentration of dodecane diol is indeed a very good marker for androgenicity in the uropygial gland of the male quail. This experimental model seems well suited for evaluating the stimulators and inhibitors of the activity of the sebaceous gland.

Androgen regulation of the androgen receptor of the quail uropygial gland: application of a [3H]mibolerone exchange assay.

An exchange assay for androgen receptors in the quail uropygial gland using [3H]mibolerone was established. The most efficient exchange conditions were 3 days of incubation at 15 degrees C. Under these conditions, androgen receptors were stable in the presence of sodium molybdate, and the exchange of [3H]mibolerone with endogenous testosterone bound to cytosolic or nuclear androgen receptors was maximal. Less than 5% of [3H]mibolerone-binding sites occurred in the extracted nuclear pellets. Using this exchange technique, it was shown that androgen receptors in the uropygial gland of photostimulated male quail or castrated quail treated with testosterone were activated and that their concentrations in both cytosolic and nuclear fractions were increased. These results confirm the androgen dependency of the quail uropygial gland, and show that it is an organ which can be used as a model for the study of androgen action in sebaceous glands.

Cloning, sequencing and expression of Pseudomonas testosteroni gene encoding 3 alpha-hydroxysteroid dehydrogenase.

We describe the cloning, sequencing and expression of the 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD) gene of Pseudomonas testosteroni. A genomic library of P. testosteroni total DNA constructed from SauIIIA digests ligated to an lambda gt11 vector was probed with a polyclonal antibody raised against purified enzyme. Subclones derived from a recombinant phage containing a 1746 bp insert were sequenced and found to contain an open reading frame of 696 bp that corresponds to a protein of 231 amino acid residues. A search for homologous proteins was performed. No similarity was observed when comparing 3 alpha-HSD with known members of the short-chain dehydrogenase family. However a small proteic fragment (80 amino acids) shows homology with the N-terminal sequence of bacterial L7/L12 ribosomal proteins.

Partial purification of 3 alpha- and 3 beta-hydroxysteroid dehydrogenases from human hyperplastic prostate. Comparison between the two enzymes.

3 alpha-Hydroxysteroid dehydrogenase (3 alpha-HSD) activity has been purified to homogeneity, the enzyme is a monomer with a Mw of 32,000 Da. 3 beta-Hydroxysteroid dehydrogenase (3 beta-HSD) activity has been partially purified and has an apparent Mw of 30,000 Da. Both enzymes have the same cofactor requirements, optimal pH. However, 3 beta-HSD appeared to be an integral protein dependent on protein environment for its activity while 3 alpha-HSD activity is a protein more loosely associated to membranes.

Ontogeny of steroid metabolizing enzymes in rat oligodendrocytes.

Rat oligodendroglial cells were isolated from newborn and developing brains and used immediately after, for quantification of steroid metabolizing activities. Oligodendrocytes (Ol) and their progenitor cells were incubated with [(14)C] testosterone, [(14)C] progesterone, [(14)C] pregnenolone or [(14)C] dehydroepiandrosterone (DHEA). Oligodendrocytes and their progenitor cells expressed different steroid metabolizing enzymes. The main activities were 5 alpha reduction of testosterone and progesterone and 3 beta hydroxy steroid dehydrogenase-isomerase which transformed pregnenolone into progesterone and DHEA into Delta 4 androstenedione. 5 alpha reductase activity increased in male and female rats in parallel with testosterone or progesterone. Contrary to this, 3 beta hydroxysteroid dehydrogenase-isomerase activity was found to be high in the young rat and to decrease when testosterone and progesterone plasma concentration increased.

Cloning, DNA sequencing and expression of (3-17)beta hydroxysteroid dehydrogenase from Pseudomonas testosteroni.

We describe the cloning, sequencing and overexpression of the (3-17)beta hydroxysteroid dehydrogenase gene of Pseudomonas testosteroni. A genomic library of Ps. testosteroni total DNA constructed from SauIIIA digests ligated to a lambda gt11 vector was probed with polyclonal antibody raised against purified enzyme. Subclones derived from a recombinant phage containing a 2661-base-pair insert were sequenced and found to contain an open reading frame of 765 base pairs that corresponds to a protein of 254 amino acid residues. A 1492-base-pair fragment was inserted into pBR322 plasmid vector and used to construct a strain of E. coli HB101 that overexpressed the steroid dehydrogenase gene.

Characterization and solubilization of testosterone 17 beta-hydroxysteroid dehydrogenase in human hyperplastic prostate.

17 beta-Hydroxysteroid dehydrogenase is a membrane-bound enzyme in human prostate. Solubilization of this enzyme can only be obtained in the presence of detergents. The optimal solubilization mixture contained 50 mM Tris-HCl buffer pH 9.0, 20% glycerol, 0.1 M KCl and 5 mg/ml of the non-ionic detergent N-octyl glucoside. In these conditions, the soluble fraction contained more than 90% of the enzymatic activity. A 2.5-fold increase of specific activity was obtained during solubilization under optimal conditions.

Purification of testosterone 5 alpha-reductase from human prostate by a four-step chromatographic procedure.

Nuclear membrane bound testosterone 5 alpha-reductase solubilized in active form from human prostatic tissue by 0.5% n-octyl beta-D-glucopyranoside was purified by a four-step chromatographic procedure including DEAE-Trisacryl ion exchange, hydroxylapatite adsorption, testosterone-Sepharose affinity and Sepharose 4B gel filtration. A purification of approximately 30-fold was achieved judging from the increase in the specific enzymatic activity. We have purified the acidic pH-optimum 5 alpha-reductase type 2 isoenzyme. The apparent molecular weight of the purified enzyme was estimated as 42,000 by SDS-PAGE. At the same time we isolated a 38 kDa protein characterized by a real affinity for testosterone and by a possible association to the 5 alpha-reductase enzyme.

Unusually high rates of metabolism of DHT in cytosols of the quail uropygial gland.

The metabolism of DHT in the cytosol of the quail uropygial gland was found to be so high that the steroid was almost completely inactivated within 2 hours of incubation at 0 C. In these conditions, DHT cannot be used for the characterization of androgen receptors. By contrast, R 1881 and mibolerone, which are not metabolized, can be used as alternative ligands. Moreover, the extremely high metabolism of DHT questions the physiologic role of this steroid in the quail uropygial gland.

Levels of collagen degradation products (telopeptides) in the tear film of patients with keratoconus.

PURPOSE: Levels of collagen degradation products (telopeptides) in the tear film of patients with keratoconus were measured to study the release of telopeptides in the tears. METHODS: Tear samples were collected from 26 keratoconus patients and 36 age-similar human control subjects. Levels of telopeptides were quantified and compared between the two groups. RESULTS: The telopeptide level in tears from keratoconus patients was 2.5-fold higher than in tears from the control group. The telopeptide concentration was age-dependent in both groups. In tears from young people, telopeptide level was 2-fold higher than in tears from the older people. CONCLUSION: The tear film of patients with keratoconus contains higher levels of telopeptides than those of control subjects. Determination of telopeptide levels in tears could be useful for the follow-up of keratoconus development in patients.

[Effect of testosterone on the levels of nucleic acids and proteins in the uropygial gland of the male quail]. / Influence de la testostérone sur les taux d'acides nucléiques et de protéines dans la glande uropygienne de la caille mâle.

A close correlation was evidenced between the length of a silastic capsule filled with testosterone propionate and the resulting plasma concentrations in testosterone in castrated male Quail. 5 days after the beginning of the treatment, testosterone induced in the spleen glands a conspicuous increase in total RNA, DNA and protein contents.

[Antigen-antibody type precipitation reactions between secretory products of different regions of the oviduct of Pleurodeles waltlii Michah]. / Réactions de précipitation de type "antigène-anticorps" entre produits de sécrétion de différentes régions de l'oviducte de Pleurodeles waltlii Michah

"Antigen-antibody" like reactions between products of secretion of different parts of the Pleurodeles waltlii Michah's oviduct have been observed using the usual immunodiffusion methods. The possible physiological importance of such reactions in egg-jelly formation and in the phenomena associated with the fecondation (particularly capacitation) is discussed.

[Sensitivity to various antibiotics of spiroplasmas isolated from mosquitoes in France]. / Sensibilité à divers antibiotiques de six souches de spiroplasmes isolés de moustiques en France.

Spiroplasmas are helical mycoplasmas that play a significant role in plant diseases. They are also found in arthropods that are likely to bite humans, such as ticks and mosquitoes. These arthropods can act as vectors and therefore may be of epidemiologic significance. Furthermore, mainly on the grounds of morphologic evidence, spiroplasmas have been incriminated in the genesis of human Creutzfeld-Jacob disease. We recovered six strains of Spiroplasma sp. from 1927 female mosquitoes. In vitro susceptibility of each strain to the following antibiotics was studied: tetracycline, oxytetracycline, doxycycline, erythromycin, chloramphenicol, rifampin, kanamycin, gentamicin and pefloxacin. Minimal inhibitory concentrations (MICs) were determined by dilution in liquid SP4 medium using microtiter plates. Plates were incubated for 24 to 48 hours at 30 degrees C. The inoculum contained approximately 5 X 10(5) CFU/ml. Each of the six strains was found to be highly susceptible to tetracycline, oxytetracycline, doxycycline, erythromycin, chloramphenicol and pefloxacin (MICs less than or equal to 0.16 microgram/ml, 0.63 microgram/ml, 0.08 microgram/ml, 0.16 microgram/ml and 0.32 microgram/ml respectively). On the opposite, the strains exhibited resistance to rifampin and variable degrees of susceptibility to kanamycin (12.5 micrograms/ml less than MIC less than 50 micrograms/ml) and gentamicin (3.12 micrograms/ml less than MIC less than 50 micrograms/ml). From our results, spiroplasmas seem to have more or less the same susceptibility to antibiotics as mycoplasmas.

Testosterone regulation of androgen receptor levels in the uropygial gland of quails (Coturnix coturnix): a further proof for the androgen dependency of the uropygial gland.

In castrated quails supplemented with testosterone by a chronically implanted silastic capsule, removal of the implant resulted in the gradual disappearance of the androgen receptors in both cloacal and uropygial glands. However, after a 7-day time span, the receptors reappeared in both glands but did not reach the original level of implanted birds. On the other hand, the photostimulation of intact birds induced an increase of the receptor content of their uropygial and cloacal glands when compared to sexually quiescent birds. These results show that, the concentration of the androgen receptors of the uropygial and cloacal glands of adult male quails is partly controlled by testosterone. By comparison with the mechanism of the action of testosterone in mammal target organs, our results add weight to the androgen dependency of quails uropygial glands (a sebaceous like organ).

Oestrogen secreting Leydig cell tumour and GnRH agonist in-vivo and in-vitro studies.

OBJECTIVE: The purposes of our study concerning two patients with oestrogen secreting Leydig cell tumour were to determine whether endogenous LH levels are involved in testicular tumour steroidogenesis and whether aromatase activity of oestrogen secreting Leydig cell tumours is directly or indirectly dependent on LH levels. MEASUREMENTS: E2 and T were evaluated after hCG injection (5000 IU) during 96 hours. Bio and immuno LH, T, E2, were determined at the basal state and after administration of D-Trp-6-GnRH agonist (3.75 mg) every 3 weeks. The abnormal testis was removed after the third injection and testicular venous blood was collected during the operation. Testicular tumour was incubated with 4-14C-T. RESULTS: Oestradiol (E2) response to hCG injection (5000 IU) was prolonged and exaggerated while that of testosterone (T) was similar to that of the controls. The aromatase index (E2/T) remained elevated even 96 hours after hCG. Intramuscular injection of the GnRH agonist, D-Trp-6-GnRH (3.75 mg) resulted in a reduction of immunoreactive and bioactive LH. T was decreased to about 10% of baseline levels and E2 fell from 240 to 36 pmol/l. In the blood of the spermatic veins collected in the course of surgery, E2 levels were found to be lower in comparison with the controls. E2 was found to be twofold higher in the spermatic vein draining the tumoral side than in that of the contralateral testis. Incubation of the testicular tumours with 4-14C-T, displayed a reduced aromatase activity (conversion of T to E2: 0.3 and 0.1% in patients 1 and 2 respectively). CONCLUSIONS: The kinetics of E2 response to hCG administration would suggest a modification of the regulation of the aromatase activity in this type of oestrogen secreting tumour. A certain endogenous LH level may be necessary to supply a sufficient quantity of T substrate, and to maintain aromatase activity of such Leydig cell tumours secreting oestrogens. These tumours seem to be responsive to endogenous LH levels.

[Evidence and characterization of an androgen receptor in the uropygial gland of the adult male quail]. / Mise en évidence et caractérisation du récepteur des androgènes dans la glande uropygienne de la Caille mâle adulte.

5 alpha-DHT is bound with a high affinity (Kd = 0.8 nM) and a limited capacity (80 fmol/mg proteins) in the cytosol of the preen gland of quails, an analogue of sebaceous glands. The complex can be precipitated by protamin-sulfate and is sedimented in the 8 S region after ultra-centrifugation in a sucrose gradient. The occurrence of this binding that exhibits the main properties of an androgen receptor, supports the use of the preen gland of quails as a tool for the study of the mechanism of action of androgens in birds and as a model for the study of the androgen control of sebaceous-like glands.

Testosterone stimulation of DNA-dependent DNA polymerase activities in the cloacal and uropygial glands of the male quail (Coturnix coturnix japonica).

DNA polymerase alpha and beta activities were studied and compared in the cloacal gland and the uropygial gland (sebaceous-like gland) taken from castrated quails after testosterone administration. In the cloacal gland, testosterone induced an important increase in DNA polymerase alpha while DNA polymerase beta was not found to be altered. In the uropygial gland, testosterone induced a cyclic variation of DNA polymerase alpha activity which may relate to the holocrine function of this gland.

Absence of 5 alpha-dihydrotestosterone action in the sebaceous-like uropygial gland of the male Japanese quail.

The uropygial gland of the quail, a sebaceous-like gland, has been proven to be androgen-dependent. Waxes secreted by this gland consist of fatty acids esterified by alkane-2,3-diols [12]. In castrated quails, the relative concentration of dodecane diol was enhanced after testosterone treatment; but 5 alpha-DHT could not evoke any increase in the relative concentration of dodecane diol. It is not possible from our present results to know if this lack of gland response to DHT administration is related to a high level of DHT metabolism in the gland cells or to a decreased affinity of the androgen receptor for DHT. However, because of the high similarity existing between uropygial gland of birds and mammalian sebaceous glands, these results give rise to the question of the true role of DHT in mammalian sebaceous glands.

Androgen control of the secretion in the sebaceous-like preen gland.

Uropygial gland of quails was previously proved to have androgen receptors. The waxes secreted by this gland were shown to consist mainly of linear saturated fatty acids esterified by linear saturated diols. In adult male quails, the relative concentration of the C12 diol was found to be correlated to the androgen levels of the birds. Furthermore, in castrated quails implanted with testosterone propionate capsules, 6 days were required for the C12 diol to reach the level found in sexually active birds. These testosterone effects related to the C12 diol were nullified by cyproterone acetate administration. Consequently the C12 diol is a reliable specific marker for androgenicity in the quails' preen gland, a new and accurate model for the study of androgen action in sebaceous glands.