Effects of dehydroepiandrosterone and 16 alpha-hydroxydehydroepiandrosterone on the reduction of glucose to glucitol by the human placenta.

The metabolism of glucose by subcellular preparations of human full term placentae has been investigated. It has been shown that in the presence of NADPH two transformation products can be detected of which one has been identified as glucitol. The effects of dehydroepiandrosterone and 16alpha-hydroxydehydroepiandrosterone on the reduction of glucose to glucitol have also been studied. It has been found that at a concentration of DHA 1.2 X 10(-4)M, the reduction of glucose is strongly inhibited (35-51%), while at a concentration of DHA 5.8 X 10(-6)M this reaction is stimulated by 13 +/- 2.3%. 16alpha-hydroxyepiandrosterone at concentrations ranging from 1.2 X 10(-4)M to 3 X 10(-6)M inhibits the formation of glucitol from 63% to 9%.     

Synthesis of two new haptens of 16alpha-hydroxydehydroepiandrosterone (3beta,16alpha-dihydroxyandrost-5-en-17-one)

Synthetic routes leading to 19E and 7Z O-(carboxymethyl)oximes derived from 16alpha-hydroxydehydroepiandrosterone were developed using two independent methods for introduction of the 16alpha-hydroxy group. Firstly, the oxime moiety was built, and then, either epoxidation of the enol acetate followed by the boron trifluoride mediated rearrangement or alkaline hydrolysis of the corresponding alpha-bromide in aqueous N,N-dimethylformamide were employed. The last step in both methods was removal of the protecting groups, which consisted of acid deprotection of the acetates and gentle alkaline hydrolysis of the methyl ester. Final haptens were designed as components for immunoanalytical kits.     

Purification and properties of 16 alpha-hydroxyprogesterone dehydroxylase from Eubacterium sp. strain 144

Eubacterium sp. strain 144 converts 16 alpha-hydroxyprogesterone to 17-isoprogesterone. The first step of this reaction is catalyzed by 16 alpha-hydroxyprogesterone dehydroxylase (16 alpha-dehydroxylase). This enzyme was purified 40-70-fold and characterized. 16 alpha-Dehydroxylase was found to be active in two molecular weight forms of Mr 181 000 and 326 000. A subunit relative molecular weight of 42 400 was determined by sodium dodecyl sulfate gel electrophoresis of the purified enzyme. Although active with both 16 alpha-hydroxyprogesterone and 16 alpha-hydroxypregnenolone, the affinity of 16 alpha-dehydroxylase for the latter steroid was twice that of the former based on the apparent Km values. Evidence of possible substrate inhibition at high concentrations was seen with 16 alpha-hydroxypregnenolone. 16-Ketoprogesterone was found to be a competitive inhibitor of 16 alpha-dehydroxylase with respect to both steroid substrates. Although generally unaffected by low concentrations of non-ionic detergents, 16 alpha-dehydroxylase activity was stimulated 3-7-fold by sodium dodecyl sulfate and inhibited strongly by cetyltrimethylammonium bromide.     

Plasma corticosteroid patterns in the fetus

In umbilical vein blood samples collected in 137 fetuses between 19 and 31 weeks of gestation, cortisol (F), cortisone (E), 17-hydroxyprogesterone (17-OHP) and 11-deoxycortisol (S) were radioimmunoassayed after column chromatography on Sephadex LH-20 of plasma extracts. While F levels plateaued throughout the period considered those of E displayed an increasing pattern which appeared to be comparable with that of unbound F in pregnant women. The declining pattern of S and more particularly of 17-OHP would suggest an increasing utilization and metabolization of these F precursors by the maturing fetus. E was not correlated with either 17-OHP or S but showed a significant correlation with F. S and 17-OHP were correlated with each other and with F. The significance of these correlations was discussed according to the different origin of these steroids and to their metabolic relationships. The application of this method for the prenatal diagnosis of inborn errors of steroid biogenesis is suggested.     

Diurnal rhythms of cortisol,ACTH, and beta-endorphin levels in neonates and adults.

To determine whether a diurnal rhythm exists in neonates admitted to neonatal intensive care units where there is continuous artificial lighting and periodic nursing and medical care, plasma cortisol, adrenocorticotropin (ACTH), and beta-endorphin concentrations were measured in two groups of infants and in adult human volunteers. As expected, a diurnal rhythm was seen in adults. A diurnal rhythm was also found for cortisol and endorphin levels in neonates (3 to 4 days postnatally) with minimal stress and in infants who were clinically severely stressed. There was not a significant difference between the morning and afternoon concentrations of ACTH in these infants, but the afternoon concentrations were lower than the morning''s, as would be expected. We found that a diurnal rhythm does exist in neonates within the first few days of postnatal life and that the continuous lighting and medical and nursing interventions do not interfere with this rhythm.     

Bioconversion of 16 alpha-hydroxyprogesterone to 16 alpha-hydroxylated corticosteroids by newborn rat adrenal cells in primary culture

Using newborn rat adrenal cells in primary culture, 16 alpha-hydroxyprogesterone was bioconverted into numerous 16 alpha-hydroxylated steroids. The method of analysis of these steroids comprised the association of column and thin-layer chromatography to gas chromatography-mass spectrometry in order to obtain the mass spectra of pure compounds. The identified compounds resulted principally from the enzymatic reactions of 21-hydroxylation 11 beta-hydroxylation and reduction of the 20-oxo and 3-oxo-4-ene groups. Minor metabolites resulted from 18-hydroxylation and 6 beta-hydroxylation of the substrate. The metabolism of 16 alpha-hydroxyprogesterone is similar to that of progesterone in the same cell-culture system; however, there are two exceptions. The 21-hydroxylation of 16 alpha-hydroxyprogesterone occurs at a rate similar to that of its 11 beta-hydroxylation, whereas the 21-hydroxylation of progesterone is faster than its 11 beta-hydroxylation. The ratio of 11 beta- to 18-hydroxylation of 16 alpha-hydroxyprogesterone is about 3, whereas the ratio of 11 beta- to 18-hydroxylation of progesterone, 20 alpha-dihydroxyprogesterone and DOC is between 1./ and 2. It is most likely the rate of 18-hydroxylation which is decreased by the hydroxyl group at C-16. The use of adrenal cell cultures is a practical, simple method for the preparation of a variety of 16 alpha-hydroxylated steroids from a single substrate. Its adaptation to the production of important amounts of 16 alpha-hydroxylated corticosteroids will permit the study of their biological activity.     

Plasma fenfluramine levels,weight loss,and side effects.

Fifty women with refractory obesity received fenfluramine for 20 weeks. Every two weeks details of weight change, drug dose, degree of anorexia, and any side effects were recorded and plasma was obtained for fenfluramine and norfenfluramine measurements. Of the 41 patients available for final analysis 26 achieved a maximum plateau dose of 160 mg/day. Plasma fenfluramine concentrations did not correlate with the degree of anorexia or with the incidence of side effects other than the severity of dream disturbance. There was a highly significant relation between weight loss and plasma fenfluramine and norfenfluramine concentrations and also between weight loss and the presence of sustained anorexia. Women who achieved mean plateau concentrations over 200 ng/ml lost a mean 8.8 kg while those with concentrations less than 100 ng/ml lost a mean of only 2.1 kg. When fenfluramine is prescribed in refractory obesity the dose should be increased stepwise until either satisfactory weight loss is achieved or troublesome side effects appear.     

Delayed thymocyte maturation in the trisomy 16 mouse fetus

Mouse fetuses with trisomy 16, an animal model for human trisomy 21 (Down syndrome), have severe defects in several hematopoietic stem cell populations and a marked reduction in thymocyte number. To determine whether there are other defects in the development of the trisomic thymus, the ontogeny of the cell surface antigenic determinants, Thy-1, Ly-1, CD3, CD4, CD8, and TCR v beta, was investigated. The trisomy 16 thymocytes were able to express all of determinants either during fetal life (days 14 to 19 of gestation) or in cultures of intact thymus lobes. However, in all instances (except for Thy-1, which already had a high proportion of expressing thymocytes by day 14), there was a delay in the time at which the determinants were first expressed, as manifested by reduced numbers of positively staining cells. Furthermore, there was also a delay in the rate at which the positively staining cells attained maximal Ag densities. Overall, there was an approximate 2 day lag in development of the fetal trisomic thymocytes. This lag permitted the identification of a large population of CD4-8+ cells prior to the appearance of CD4+8+ thymocytes. These findings are consistent with the identification of CD4-8+ as an intermediate stage between CD4-8- and CD4+8+ in fetal thymocyte ontogeny.     

Elevated plasma beta-endorphin levels in pregnant women and their neonates.

Using radioimmunoassay technique β-endorphin levels were measured in the plasma of women undergoing labour and partirition and in the plasma of their neonates. The level of immunoreactive β-endorphin in the plasma of women undergoing labour was found to be significantly elevated (mean values: 38–135 fmoles/ml) above the levels found in non-pregnant women (mean values: 5–10 fmoles/ml). After birth, the level of β-endorphin-like immunoreactivity in maternal venous plasma was significantly higher than that in the umbilical vein and artery plasma of the new-borns, but there was no arterio-venous difference in the neonatal plasma. Since the antiserum used displayed the same avidity for human β-endorphin and β-lipotropin chromatographic separation of the immunoreactive components was performed by gelfiltration. Both peptides were found in the plasma of non-pregnant women, in maternal plasma and in the plasma of the neonates. In addition, high amounts of both peptides were found in the fetal pituitary gland showing that the fetus can probably produce its own peptides.     

Plasma corticotrophin levels in addison-schilder's disease.

Raised plasma immunoreactive corticotrophin (ACTH) levels were found in five boys with the sex-linked disorder progressive leucodystrophy associated with adrenal insufficiency (Addison-Schilder''s disease) and in a symptom-free brother of one of them. Similar ACTH concentrations were found using two antisera, one against the N-terminal part of the ACTH molecule and the other against the C-terminal part. In one patient the circulating ACTH had normal biological activity as measured using the cytochemical ACTH bioassay. Immunoreactive beta/-melanocyte-stimulating hormone was also determined in one patient and found to be raised.