Structure-function relationships for equine and human aromatases. A comparative study.
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Animals↗
Biomedical subjects
M A Drosdowsky
Publications and source records attributed to M A Drosdowsky.
In vitro effects of Celiptium and MR 14504 on mature rat Leydig cell testosterone production.
Percoll-purified mature rat Leydig cells have been used to evaluate the testicular toxicity of two highly potent intercalating agents (Celiptium and MR 14505). Testosterone secretion in the absence and in the presence of human chorionic gonadotropin (hCG) was measured to assess Leydig cell function. Celiptium and MR 14504 induce time- and dose-related inhibitory effects on the production of testosterone by Leydig cells, both in the presence and in the absence of hCG, whatever the concentration of hCG used. We have observed that MR 14504 is about 5 times more potent as an inhibitor of rat Leydig cell steroidogenesis than Celiptium without inducing any cell toxicity. The present study indicates that the Leydig cell is an additional potential site for the primary toxic effects of these drugs in the adult rat testis.
Animals↗
Postprandial lipoprotein metabolism in normotriglyceridemic non-insulin-dependent diabetic patients: influence of apolipoprotein E polymorphism.
Non-insulin-dependent diabetes mellitus (NIDDM) is associated with postprandial lipoprotein clearance defects that are correlated with the fasting hypertriglyceridemia widely observed in NIDDM patients. The aim of this study was to determine if such postprandial disturbances are found in NIDDM patients strictly normotriglyceridemic in the fasting state, and if the apolipoprotein E (apo E) polymorphism influences postprandial metabolism of intestinally derived lipoproteins. The vitamin A-fat loading test was used in 18 normotriglyceridemic NIDDM patients and seven normotriglyceridemic obese controls, and postprandial triglyceride (TG) and retinyl palmitate (RP) concentrations were evaluated in total plasma, and in the chylomicron (Sf > 1,000) and nonchylomicron (Sf < 1,000) fractions isolated by ultracentrifugation. NIDDM patients exhibited an amplified response of both TG and RP as compared with obese controls in the three fractions. Incremental TG response to the oral fat load was strongly correlated with fasting TG level (r = .80, P < .0001) in the whole study population. Postprandial lipoprotein profiles were distinguished in NIDDM patients according to apo E phenotype: despite normal fasting TG levels in E3/3 (n = 6), E2/3 (n = 6), and E3/4 (n = 6), postprandial RP response was twofold to threefold higher in E2/3 and E3/4 patients than in the common E3/3 phenotype. Contrasting lower postprandial TG increment and lower fasting and postprandial high-density lipoprotein (HDL) and HDL3 cholesterol levels were observed in E3/4 versus E3/3 patients, possibly reflecting modifications in lipid content of the postprandial lipoproteins driven by a differential lipid transfer activity depending on apo E isoform. These data indicate an enhanced postprandial lipemia in normotriglyceridemic NIDDM patients, and demonstrate the influence of apo E polymorphism on their lipoprotein clearance. Postprandial alterations of lipoprotein remnants may thus accelerate atherogenesis even in normotriglyceridemic NIDDM patients.
Adult↗
Equine cytochrome P450 aromatase exhibits an estrogen 2-hydroxylase activity in vitro.
Aromatase (estrogen synthetase) is a steroidogenic enzyme complex which catalyzes the conversion of androgens to estrogens (termed aromatization). This enzyme was purified from adult equine testis to homogeneity by five chromatographic steps. The ability of purified and reconstituted equine aromatase to exhibit an estrogen 2-hydroxylase activity was tested and compared to testosterone aromatization. Enzymatic activities were assessed by tritiated water release from labelled estradiol and testosterone. Kinetic analysis of estradiol 2-hydroxylation showed an apparent K(m) of 23 microM and a V(max) of 18 nmol/min/mg, whereas the values for testosterone aromatization were a K(m) of 15.7 nM and a V(max) of 34.6 pmol/min/mg. A specific antiserum raised against purified testicular equine P450arom and known to inhibit aromatase activity [1] was also found to inhibit the estrogen hydroxylase activity of equine placental microsomes in a dose-dependent manner with an IC50 value of 15 microl serum: 0.5 ml incubate. The estrogen hydroxylase activity was inhibited in a dose-dependent manner by two classes of aromatase inhibitors, i.e. steroidal-- (4-hydroxyandrostenedione and 7alpha-([4-aminophenyl]thio)-androst-4-ene-3, 17-dione)--and non-steroidal--(fadrozole and miconazole). The IC50 values were approximately 300 and 890 nM for 4-hydroxyandrostenedione and 7alpha-([4-aminophenyl]thio)-androst-4-ene-3, 17-dione, and 92 and 285 nM, for fadrozole and miconazole, respectively. Furthermore, 4-hydroxyandrostenedione caused a time-dependent inactivation of estrogen hydroxylase activity. We conclude that equine aromatase is able to use estradiol as a substrate, and converts it to catechol estradiol in vitro, possibly using the active site of aromatization. This is the first demonstration that equine aromatase functions as an estrogen 2-hydroxylase, in addition to transforming androgens into estrogen.
Androstenedione↗
Determination of metallothioneins by HPLC: application to zinc metabolism.
Metallothioneins (MT) are well known Zn and Cu tissue proteins. Their level is mainly regulated by Zn. Therefore, they have been suggested as an index of Zn status. The determination of MT has been performed by HPLC (fitted with UV-detector) following a preparation phase (heating, ultracentrifugation and filtration). This method allows direct quantification of MT in tissues and shows a good correlation between MT concentrations and peak heights (r2 = 0.995; a = 774.73; b = -148.38) from 0 to 10 micrograms/100 microL. MT determination by RP-HPLC method allows to assess the tissue zinc status as displayed by the correlation between hepatic zinc and metallothionein concentrations (r2 = 0.82; p < 0.001). Variations of MT in response to Zn intakes are observed within 24 hours and for a physiological range of dietary Zn intakes.
Chromatography, High Pressure Liquid↗
In vitro effects of various albumins on Leydig cell testosterone production in immature rat.
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Albumins↗
Germ cell and Sertoli cell interactions in human testis: evidence for stimulatory and inhibitory effects.
In human Sertoli cell preparations obtained from healthy men (mean age 31.8 +/- 6.8 years; n = 6), we have measured the productions of lactate, 17 beta-oestradiol, transferrin and inhibin between day 4 and day 5 after plating, either in the presence or absence (hypotonic treatment of plated cells on day 2) of germ cells. The results, expressed per 10(6) of cells plated/24 h, showed that lactate production was unchanged, whether or not germ cells were present. However, if we calculated the lactate production per mg protein/24 h, the lactate output was decreased (30-60%) in the presence of germ cells. Whatever the mode of expression, Sertoli cell 17 beta-oestradiol synthesis was diminished 1.5-fold in the presence of germ cells. Conversely, the transferrin output was increased 3.2-fold in non-treated Sertoli cell preparations when compared to the hypotonic-treated plates. A similar observation was recorded for the in-vitro production of inhibin by Sertoli cells, which was enhanced 1.4-fold when germ cells were present. These results, together with a likely potentializing role of germ cells on follicle stimulating hormone control of Sertoli cell function, strongly suggest that germ cells exert both stimulatory and inhibitory effects in regulating human Sertoli cell function through either direct contact and/or via secreted factors.
Adult↗
[Physiological variations of LpA1, HDL2 and principal lipid markers in athletes].
An original approach of the influence of physical activities on lipid metabolism is presented in this work: the authors studied the physiological variations of the lipoparticle LpA1, high-density lipoprotein subfraction HDL2 and the most important lipid markers in serum of a presumably healthy population of 55 high-level sportsmen. They were 18-45 years old and practised endurance sports, whether individual (cycling, long-distance running), or collective activities (soccer, basketball). The authors observed an important increase of LpA1 (+20%, p < 0.001) and C-HDL2 (+23.3%, p < 0.01) after an intense physical activity (CK = 430 +/- 312 U/l); they also found a good correlation between LpA1 and HDL2 (r = 0.85, p < 0.0001) and between LpA1 and CK (r = 0.62, p < 0.001).
Adolescent↗
[Sertoli cells. Functional aspects compared in rats, pigs and man].
In the mammalian testis, the Sertoli cell plays a key role in the development and maintenance of spermatogenesis. Indeed, within the seminiferous tubule, the structure of the Sertoli cells and the specialized junctions between them and the neighbouring germ cells, contribute to create the sophisticated microenvironment and to bring all the nutriments require for a full development of germ cells. In this review, we have compared the main Sertoli cell functions in the rat and the pig to the available data concerning the human. We have included our recent results obtained from testicular tissues of 15 young men (mean age: 29 years) from which we have prepared the Sertoli cells. In addition to the lactate and estradiol-17 beta syntheses, the human Sertoli cell produces several proteins, namely transferrin, ferritin and inhibin under the control of germ cells.
Adult↗
Germ cell-Sertoli cell interactions and production of testosterone by purified Leydig cells from mature rat.
The addition of seminiferous tubule (ST) culture medium (STM) prepared from testes of either busulfan-treated (Bus) or cryptorchid (Cryp) or genetically sterile (hd) rats, to Percoll purified Leydig cells leads to a further increase of LH-stimulated testosterone (T) output (26, 43 and 14%, respectively). Taking into account that the Sertoli cell number per cm of ST is 2.6, 1.8 and 1.4-fold greater in Bus, Cryp and hd rats than in controls, the above STM effects on T output, expressed per 10(6) Sertoli cells are in fact lower (63, 44 and 43%, respectively) that those of control STM. Similar results have been obtained for the STM transferrin levels which are decreased, 74, 67 and 45%, respectively in Bus, Cryp and hd animals. So, it is likely that the Sertoli cell secretion of both the paracrine factor involved on Leydig cell T production and the transferrin is influenced mainly by spermatids and to a lesser extent by spermatocytes of mature rat testis.
Animals↗
Age-related changes in the function of the pituitary-gonadal axis in a sterile male rat mutant (hd/hd).
Testicular growth is depressed in the genetically sterile male rat (hd/hd) relative to its LE phenotype littermates (by 50% and 73% at 27 and 90 days of age, respectively). Within the hd/hd testis, both the tubular and seminiferous tubule tissues are affected by the mutation. In addition, there is significantly less germ cell production from the primary spermatocyte stage of spermatogenesis onwards and the total number of Sertoli cells observed is less. In the intertubular tissue, the total volume and the total number of Leydig cells per testis is significantly less, but the mean volume of an average Leydig cell is not modified. The serum gonadotropin levels are higher in the hd/hd rat, whereas from 40 days of age onwards the level of testosterone is lower. The FSH and LH binding affinity constants are unchanged by the mutation; however, the total number of FSH binding sites per 10(6) Sertoli cells is lower while that of LH per 10(6) Leydig cells is greater. Indeed, it is likely that the lesser concentration of serum testosterone in the hd/hd rat is a result of a smaller number of Leydig cells since their individual function is not modified. The testicular androgen binding protein (ABP) content and the ABP output towards the epididymis are lower as a consequence of both a lesser number and an altered function of the Sertoli cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Aging↗
[Value of the study of total alkaline phosphatases and bone isoenzyme in a population of subjects with osteoporosis].
The authors have compared the values of total and bone serum alkaline phosphatases in an osteoporotic population and a control group. The total activity of alkaline phosphatases was determined by a kinetic method at 30 degrees C and alkaline phosphatase isoenzymes were separated by agarose gel electrophoresis in presence or absence of wheat germ lectin. The authors have confirmed reported data concerning the physiological variations of alkaline phosphatases in the control group; they have therefore divided each group in several sub-populations according to age and sex in order to obtain accurate comparisons. Mean values of total and bone alkaline phosphatases were greater in the osteoporotic population than in the control group regardless of age or sex. Nevertheless, significant differences were obtained only with 50 to 75 year-old subjects (men or women) for total alkaline phosphatases and with 50 to 75 year-old women for bone alkaline phosphatase.
Adult↗
Differential diagnosis of an hyperamylasemia in an acute abdominal syndrome.
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Abdomen, Acute↗
Combined diagnostic value of biochemical markers in acute pancreatitis.
Eighty-three patients suffering from upper abdominal pain were studied to evaluate the contribution of commonly used biochemical markers in the diagnosis of acute pancreatitis. On admission to hospital, serum amylase, lipase, total bilirubin, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and gamma-glutamyl transferase activities were measured. By stepwise logistic discrimination, only two determinations appeared to be of clinical value: lipase and alkaline phosphatase activities. A classification rule was established including these two measurements and its diagnostic performance evaluated by a jackknifed method amounted .83%. ROC curves were used to assess sensitivity and specificity. Our study clearly shows that serum lipase measurements should be preferred to amylase measurements, and that our two-test classification rule provides an efficient aid in clinical decision-making.
Acute Disease↗
Partial purification of androgen binding protein from bull epididymis.
An androgen binding protein (ABP), with an equilibrium dissociation constant of 4.2 nM and a molecular weight of about 100 kDa, has been purified from bull epididymal extracts using a four-step procedure. These preliminary results underline the main difficulties encountered in the purification of this protein present at a very low concentration (i.e. 50-fold less than in rat or rabbit epididymides). Ammonium sulfate precipitation is not a suitable step due to the formation, in presence of salt, of insoluble material leading to a loss of ABP. Lipids, particularly phospholipids, might be implicated in this phenomenon. Several steps, including anion exchange in batch followed by concentration, affinity chromatography and HPLC gel filtration allowed us to obtain a 7667-fold purified protein with a 9% yield.
Androgen-Binding Protein↗
Evidence for LH-inhibiting activity in ovine peripheral and testicular blood.
Intracellular cyclic AMP and testosterone productions by purified mature rat Leydig cells were stimulated by oLH (25 micrograms/l) 18- and 12-fold, respectively, after a 5-h incubation period. The replacement of the incubation medium by charcoal-treated testicular venous plasma (40%, v/v) from adult rams in the breeding season induced an inhibition of cyclic AMP and testosterone productions (82 and 66%, respectively, of oLH-stimulated values). Testicular arterial plasma is less effective than testicular venous plasma, even when it originates from non-breeding season rams; in that case, testicular venous and arterial plasma strongly inhibit testosterone productions (84 and 67%, respectively of oLH-stimulated values), which probably indicates that the inhibitory activity is higher in the non-breeding season. The addition of peripheral plasma leads to a testosterone production equal to 35 and 65% of the oLH-stimulated values, respectively, for ram blood collected in non-breeding and breeding seasons. The same concentration of ovine testicular lymph or rete testis fluid is without significant effect on cyclic AMP production; however, testosterone is slightly decreased by lymph but enhanced by rete testis fluid. Increasing amounts of venous or arterial testicular blood induce a dose-related decrease of the specific binding of labelled hCG in both rat and ram testicular membranes. This inhibiting factor is present in peripheral and testicular blood of either control or hypophysectomized or castrated rams, is a protein in nature, heat-sensitive, and has an apparent molecular weight higher than 10,000 daltons.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
[Demonstration of LH inhibitor(s) in sheep and human sera].
In mature rat Leydig cells, the testosterone output (24 ng/10(6) Leydig cells/4hrs.) is increased 10 fold by LH; the addition of serum from either control or castrated or hypophysectomized rams inhibits (60%) the LH-stimulated testosterone production. Similarly, the incubation of immature rat Leydig cells with sera from hypophysectomized patients leads to a diminution (70 and 30% respectively) of both basal (0.98 ng) and LH stimulated (3.44 ng) testosterone biosynthesis. These data suggest the existence of an LH inhibitor (or inhibitors) in blood from ram and human; in addition, this substance is not only of testicular origin and is not an LH-related molecule.
Animals↗
In vitro effects of growth factors on rat germ cell RNA synthesis and their modulation by Sertoli cell-secreted proteins.
In vitro rat germ cell RNA synthesis is influenced by growth factors. Basic fibroblast growth factor (0.1 to 100 ng/ml) increases [3H]uridine incorporation in round spermatids (RS) but not in pachytene spermatocytes (PS); this effect is potentiated by insulin (10 micrograms/ml) and blocked in the presence of Sertoli cell-secreted proteins (SCSP). Somatomedin C (0.1 to 100 ng/ml) exhibits a similar effect when used alone without an influence by SCSP. Transforming growth factor beta (0.1 to 10 ng/ml) acts on both cell types, but SCSP amplify this effect only in PS. These data suggest that growth factors synthesized in situ may play a role in the germ cell development and that their effects are modulated by SCSP.
Animals↗