Ultrastructural aspects of cells with 3 beta-HSDH (delta 5-3 betahydroxysteroid dehydrogenase) activity in female ducks. II. Female Barbary ducks (Cairina moschata)]

The cells making up the thin internal theca of the follicle of the Barbary duck and which contain are 3 beta HSDH enzyme show all of the ultrastructural characteristics of steroid cells: a well-developed smooth reticulum, tubular cristae mitochondria, a considerable load in lipid inclusionologous cells of the Peking duck since they are constantly more dense and their cristae are bigger and less numerous.     

Ultrastructure of the granulosa cells of the ovarian follicle of ducks (Barbary duck: Cairina moschata, Pekin duck: Anas platyrhynchos]

In the granulosa cells of the ovarian follicle of the ducks, the Golgi apparatus is developed, the granular endoplasmic reticulum is important. Owing to these characters, these cells are probably oriented towards the elaboration of proteins. The seasonal variations of the aspect of these cells are important: in January all organits are not very developed when in May they are all very developed. For a determined season all cells present the same aspect.     

Human placental delta5-3beta hydroxysteroid dehydrogenase activity (delta5-3beta HSDH): intracellular distribution, kinetic properties, retroinhibition and influence of membrane delipidation.

Delta5-3beta HSDH activity has been assayed either by spectrophotometric method or by use of radioactive substrates. The enzymatic activity is equally distributed between mitochondrial and microsomal fractions verified by electronic microscopy. The specific activity is comparable in both fractions, as well as the optimal pH and the Km for NAD and for the substrates. The delta5-3beta Hut optimal pH, specific activity and sensitivity to the inhibitory action of various steroids are different when C19 and C21 steroids are used as substrates. Estrogens and cyclic AMP have also an inhibitory action on the oxidation of C21 steroids. Treatment of microsomal or mitochondrial membranes with phospholipase A releases fatty acids (mainly arachidonic) and decreases the enzymatic activity. "Adsorbtion" of the fatty acids on bovine serum albumin partially reactivates the delta5-3beta HSDH.     

Ultrastructural aspects of cells with 3 beta-HSDH (delta-5-betahydroxysteroid dehydrogenase) activity in female ducks. I. Female Peking duck (Anas platyrhynchos)]

The cells making up the thick internal theca of the follicle of the Peking duck and which contain an active 3 beta HSDH enzyme have a developed smooth reticulum, tubular crested mitochondria, a considerable load in slightly osmiophilic lipid inclusions; all of these characteristics are those of steroid cells. Various physiological states of these cells juxtapose themselves in different proportions according to the season.     

The genetic variability in the amylase isoenzymes of hybrid muscovy (Cairina moshata) and domestic (Anas platyrhynchos) ducks compared to the initial species]

Genetically conditioned polymorphism of alpha-amylases in musk ducks, domestic ones and their hybrids has been described. Two isoenzymes (AMY-1, AMY-2) are in blood serum of the ducks. One of them is apparently formed in crope gland and other--in pancreas. AMY-2 is monomorphic in both species. AMY-2 is polymorphic with three alleles (Amy-1A, Amy-1B, Amy-1B) in musk ducks and monomorphic in domestic ducks (Amy-1C). The hybrids have also allele obtained by them from domestic ducks.     

Is there any delta 5-3 beta hydroxysteroid dehydrogenase activity in preimplantation embryo of rhesus monkey?

This is the first report on the histochemical assessment of delta 5-3 beta hydroxysteroid dehydrogenase activity in all the preimplantation embryonic stages in the rhesus monkey (Macaca mulatta). An apparent stage dependent increase in enzyme activity was obtained, however, distinctively a high degree of non-specificity in enzyme reaction was noted primarily in morulae and blastocysts. Such marked non-specificity in the histochemical enzyme reaction for delta 5-3 beta hydroxysteroid dehydrogenase activity was not found in mouse blastocysts. High amounts of endogenous steroids present within rhesus embryos, or the participation of non-specific dehydrogenases could account for the observed non-specificity. Furthermore, the present report documents the pattern and degree of association (r = 0.9; P less than 0.01) between developmental stage and gestational age of preimplantation rhesus embryos, and thus provides a normal in situ cell cleavage rate of preimplantation embryo in the rhesus monkey.     

Quantitative real-time PCR primer design, cDNA amplification and sequence analysis from 22 genes mainly associated with lipid metabolism in Pekin (Anas platyrhynchos) and Muscovy (Cairina moschata) ducks

Few genomic tools are available in ducks. To produce some new resources, we have designed Pekin (Anas platyrhynchos) and Muscovy (Cairina moschata) duck-specific primers for 22 genes involved mainly in lipid metabolism, and to a lesser extent in carbohydrate metabolism and other functions. Primers were designed according to duck sequences when available and otherwise from the corresponding conserved regions in chicken and human sequences. These primers allowed quantitative RT-PCR amplification of RNA from Pekin and Muscovy ducks. Amplified cDNA products from both species were sequenced and were found to be very similar to chicken sequences (about 94%). This work provides additional genomic resources and polymorphism information for some genes in duck species and represents a first step towards gene expression analyses in Pekin and Muscovy ducks.     

Cyto-immunological study of the anterior pituitary somatotropin cells of male and female adult Peking, Barbary and hybrid (from crossing male Peking X female Barbary) ducks and of male ducklings of the 3 races)]

The anti-human-STH immunserum applied on hypophysis sections of adults males and females Peking (Anas platyrhynchos L.) Barbary (Cairina moschata L.) and Hybrid (from the mating male Peking X female Barbary) ducks, marks orangeophils, oval- or round shaped, PAS negative cells exclusively localized in the caudal lobe of the pars distalis. In the males ducklings of the three races this immunserum reveals numerous and partially exhausted cells. The authors conclude that the acidophils alpha cells observed in the caudal lobe of the duck are somatotrophic cells.     

Separation of [Sp]-3'-O-(5'-dimethoxytritylthymidyl)-5'-O-(3'camphanoyl thymidyl)-O-methyl phosphite.

First isolation of diastereoisomerically enriched (d.p. 95%) [Sp]-3'-O-(5'-dimethoxytrityl tjymidyl)-5'-O-(3'-camphanoyl thymidyl)-O-methyl phosphite (1) allowed to assign, via sulfuration and deprotection, its absolute configuration as Sp. Its reactions with methyl iodide, dimethoxytrityl chloride, or water, respectively, allowed to confirm correctness of assignment of absolute configuration in diastereisomers of TPMeT, and to assign absolute configuration in corresponding diastereoisomers of TPDMTT and TPHT.     

Effect of lithium chloride on spermatogenesis and testicular delta 5-3 beta, 17 beta-hydroxysteroid dehydrogenase activities in the 'antiserum to luteinizing hormone' treated toad (Bufo melanostictus)

Activities of delta 5-3 beta- and 17 beta-hydroxysteroid dehydrogenase (delta 5-3 beta-HSD and 17 beta-HSD), Leydig cell nuclear area (LCNA) and spermatogenesis in the testis were observed after injection of lithium chloride in the 'antiserum to luteinizing hormone (LH)' treated toad. A significant decrease in the activities of steroidogenic enzymes, LCNA and spermatogenesis were noticed after the injections of 'antiserum to LH' to toads. Further decrease in the activities of the above parameters was observed in the lithium chloride--'antiserum to LH' treated toad. It is suggested that lithium chloride may inhibits testicular function without modulating the pituitary activity.     

Binding characteristics of [3H]delta(5)-androstene-3beta,17beta-diol to a nuclear protein in the rabbit vagina

In this study, we investigated the binding characteristics of [3H]Delta(5)-androstene-3beta,17beta-diol to rabbit vaginal cytosolic and nuclear extracts and in freshly excised intact tissue strips. [3H]delta(5)-Androstene-3beta,17beta-diol bound to a protein(s) in the vaginal nuclear extract with high affinity (K(d)=3-5 nM) and limited capacity (50-100 fmol/mg protein). No specific binding was detected in the cytoplasmic extracts. Competitive binding studies showed that binding of [3H]delta(5)-androstene-3beta,17beta-diol was effectively displaced with unlabeled delta(5)-androstene-3beta,17beta-diol but not with dehydroepiandrosterone, testosterone, dihydrotestosterone, triamcinolone acetonide, or progesterone. However, estradiol at high concentrations partially displaced bound [3H]delta(5)-androstene-3beta,17beta-diol. Incubation of freshly excised vaginal tissue strips with [3H]delta(5)-androstene-3beta,17beta-diol in the absence or presence of excess unlabeled delta(5)-androstene-3beta,17beta-diol for 1h at 37 degrees C resulted in specific binding to a soluble macromolecule in the nuclear KCl extracts. In addition, quantitative measurement of estrogen receptor, androgen receptor and delta(5)-androstene-3beta,17beta-diol binding protein was performed by equilibrium ligand binding assays using extracts of distal vaginal tissue from intact animals or ovariectomized animals treated for 2 weeks with vehicle, estradiol, testosterone, or estradiol plus testosterone. These changes in steroid hormone levels resulted in opposing trends between the estrogen receptor and delta(5)-androstene-3beta,17beta-diol binding protein, suggesting that delta(5)-androstene-3beta,17beta-diol binding protein is regulated differently by the hormonal milieu than the estrogen receptor. These data suggest that rabbit vaginal tissue expresses a novel binding protein which specifically binds delta(5)-androstene-3beta,17beta-diol and is distinct from the androgen and estrogen receptors.     

Purification and properties of the 5 alpha-dihydrotestosterone 3 alpha(beta)-hydroxysteroid dehydrogenase from human prostatic cytosol.

5 alpha-Dihydrotestosterone 3 alpha(beta)-hydroxysteroid dehydrogenase [3 alpha(beta)-HSDH] [EC 1.1.1.50/EC 1.1.1.51] which catalyses the conversion of 5 alpha-dihydrotestosterone (5 alpha-DHT) to both 5 alpha-androstane-3 alpha,17 beta-diol and 5 alpha-androstane-3 beta,17 beta-diol was purified to an apparent homogeneous state using cytosol of three human hyperplastic prostates by a 4-step purification procedure. After each purification step 3 alpha-HSDH activity was coincident with 3 beta-HSDH activity. On average, specific 3 alpha-HSDH activity was enriched 856-fold, specific 3 beta-HSDH activity 749-fold compared to human prostatic cytosol using anion exchange, hydrophobic interaction, gel filtration and affinity chromatography. Examination of the purified enzyme by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS) revealed a single protein band with silver staining. The molecular weight of the enzyme was estimated as 33 kDa by SDS-polyacrylamide gel electrophoresis and as 28 kDa by Sephacryl S-200 gel filtration indicating that the native 3 alpha(beta)-HSDH is a monomer. In the presence of the preferred co-factor, NADPH, the purified enzyme had a mean apparent Km for 5 alpha-DHT of 3.9 microM and a Vmax of 93.3 nmol (mg protein)-1 h-1 with regard to 3 alpha-HSDH activity, and a Km of 6.3 microM and a Vmax of 20.6 nmol (mg protein)-1 h-1 with regard to 3 beta-HSDH activity.     

Role of thiol group(s) in the enzymatic activity of the membrane-bound delta5-3beta-hydroxysteroid dehydrogenase from bovine adrenal cortex microsomes.

The enzyme mediated ATP- and, to a lesser extent, CoASH-dependent synthesis of the spirodilactone of 4-(2'-carboxyphenyl)-4,4-dihydroxybutyrate from 2-succinylbenzoate has been demonstrated in membrane-free extracts of Micrococcus luteus and Escherichia coli. The suggestion is made that the spirodilactone is the product of an aberrant reaction involving a compound that is normally an intermediate in the conversion of 2-succinylbenzoate to 1,4-dihydroxy-2-naphthoate.     

Role of reduced glutathione in the delta(5)-3-kitosteroid isomerase reaction of liver.

Partially purified rat liver Δ⁵-3-ketosteroid isomerase (EC 5.3.3.1) is profoundly and specifically activated by reduced glutathione (GSH). This stimulating effect shows normal saturating kinetics, and both K_m and V_max are pH-dependent. The binding of GSH is independent of the concentration of Δ⁵-androstene-3,17-dione, whereas the K_m for Δ⁵-androstene-3,17-dione is markedly reduced by saturating levels of GSH. The same catalytic site appears to isomerize both Δ⁵-androstene-3,17-dione and Δ⁵-pregnene-3,20-dione. Several steroidal inhibitors compete with Δ⁵-androstene-3,17-dione, whereas $\text{S}$-methyl-glutathione competes with GSH. This activation of Δ⁵-3-ketosteroid isomerase is also observed in the livers of other species (calf, guinea pig, human), and represents a hitherto unrecognized function of reduced glutathione.     

beta]-Glucan Synthesis in the Cotton Fiber (III. Identification of UDP-Glucose-Binding Subunits of [beta]-Glucan Synthases by Photoaffinity Labeling with [[beta]-32P]5[prime]-N3-UDP-Glucose

Using differential product entrapment and photolabeling under specifying conditions, we identifIed a 37-kD polypeptide as the best candidate among the UDP-glucose-binding polypeptides for the catalytic subunit of cotton (Gossypium hirsutum) cellulose synthase. This polypeptide is enriched by entrapment under conditions favoring [beta]-1,4-glucan synthesis, and it is magnesium dependent and sensitive to unlabeled UDP-glucose. A 52-kD polypeptide was identified as the most likely candidate for the catalytic subunit of [beta]-1,3-glucan synthase because this polypeptide is the most abundant protein in the entrapment fraction obtained under conditions favoring [beta]-1,3-glucan synthesis, is coincident with [beta]-1,3-glucan synthase activity, and is calcium dependent. The possible involvement of other polypeptides in the synthesis of [beta]-1,3-glucan is discussed.