George Betz

Estriol concentrations in blood during pregnancy.

Abstract Total plasma estriol concentrations and 24 hour total urinary estrogen excretions were measured at frequent intervals during the last half of gestation in a group of selected subjects. The blood analyses frequently gave a more reliable indication of well-being of the infant in utero than the urine analyses and was less inconvenient to the subject. Plasma estriol determination is not substantially more difficult or costly than accurate measurement of the hormone in urine, and the blood analysis therefore should replace the older method.

Impact of histocompatibility antigens on pregnancy outcome

OBJECTIVE The role of histocompatibility antigens in pregnancy outcome is controversial. This controversy may be because the initial studies were of small numbers or the patient groups were not homogeneous. The purpose of this study is to clarify these discrepant results by carrying out histocompatibility antigen typing and mixed lymphocyte culture on couples with idiopathic recurrent spontaneous abortion and by comparing results with those of fertile couples. STUDY DESIGN Sixty couples with at least three spontaneous abortions and 60 normal couples with at least two successful pregnancies were included. Histocompatibility antigen typing and mixed lymphocyte culture were performed by using the standard techniques. The data were analyzed statistically with the Fisher exact test and chi 2. RESULTS Our results failed to show any difference between normal and aborting couples with regard to HLA-A, HLA-B, and HLA-DR distribution or sharing or to mixed lymphocyte culture responsiveness. CONCLUSION Our study, along with other studies, emphasizes that the histocompatibility antigen system does not have an impact on pregnancy outcome.

Reaction mechanism of 20β-hydroxysteroid dehydrogenase determined by equilibrium rate exchange☆

Abstract The reaction mechanism of 20β-hydroxysteroid dehydrogenase (20β-hydroxysteroid:NAD-oxidoreductase; EC 1.1.1.53) has been investigated by measuring the rate of isotopic exchange between substrate-product pairs while varying concentrations of unlabeled reactants. The results show that the reaction is partially compulsory in that the major pathway involves an enzyme:cofactor binary complex. A detectable degree of reaction, however, occurs with steroid forming the binary complex. It was also determined that the conversion of the ternary complexes was not a rate-limiting step in the reaction.

Participation of cytocheome P-450 in the steroid 17α-hydroxylase of testis microsomes

Abstract The effect of metyrapone on the activity of the steroid 17α-hydroxylase from rat testis was evaluated. A competitive pattern of inhibition vas observed after analysis of data using a least mean squares computer analysis. The substrate for the hydroxylase induced a Type I difference spectrum in an active suspension of Triton treated microsomes. The magnitude of this spectral change was dependent on steroid concentration and was diminished by metyrapone. The effect of metyrapone was abolished at infinite steroid concentration. These results confirm the participation of cytochrome P-450 as a reactant in the 17α-hydroxylase reaction.

Steroid 17,20-lyase: the effect of detergents on enzymic activity and microsomal composition.

Abstract The activity of the steroid 17, 20-lyase from rat testis microsomes was determined following exposure of the microsomes to detergents. Only Triton N-101 and X-100 produced enzymically active supernatants. The supernatant from Triton N-101 treatment consisted of submicrosomal particles enriched in cytochrome P450, flavin, and phospholipid; depleted in RNA and NADPH oxidase; and unchanged in the concentration of cytochrome b 5 and non-heme iron. The activities of the NADPH and NADH cytochrome C reductases were also intact. Lubrol produced an inactive supernatant which contained all the components thought to be necessary for microsomal electron transport with the exception of cytochrome b 5 . An assay, specific for cleavage and more expeditious than the chromatographic separation of reactants, is also described.

Actions of danazol in vivo on cytochrome P-450 and steroidogenic enzymes in rat testis and liver microsomal preparations

The mechanism of action of danazol has not been established and the drug may act at multiple loci. Effects suggesting inhibition of pituitary gonadotropin release have been described while some in vitro studies have demonstrated competitive inhibition of steroidogenic enzymes. In addition, destruction of cytochrome P-450 by the acetylenic moiety of danazol is a possible mechanism. Following 14 days of danazol treatment (10 mg/kg/day), the specific content of rat testis microsomal cytochrome P-450 and the serum testosterone were decreased, in spite of no significant change in serum luteinizing hormone. Furthermore, simultaneous administration of human chorionic gonadotropin with danazol still resulted in a decrease in cytochrome. The activities of two testicular microsomal steroid-converting enzymes (17 alpha-hydroxylase and 17,20-lyase) were not altered when based on cytochrome P-450 content but were markedly depressed when based on microsomal protein. In liver, danazol showed a similar but less marked dimunition of the cytochrome. Surprisingly, steroid 17 alpha-hydroxylase activity in liver was significantly increased while other cytochrome P-450 dependent enzyme activities were not altered.

Stimulatory effect of soluble supernatant on hydroxylase activity of rat testis microsomes

Addition of soluble supernatant to testis microsomes results in 42% increase in steroid 17,20-lyase activity and a 65% increase in 17alpha-hydroxylase activity. This stimulatory activity could be partially purified by salt fractionation. The activating factor(s) was not removed by dialysis nor did it appear to be lipid. It was destroyed by trypsin. Differential effects of heat were observed with the hydroxylase and lyase activators. The activation did not affect Km but only increased Vmax. The supernatant could be added to each enzyme to the point of maturation. No binding of steroids by the supernatant could be detected. Corpus luteum and placental supernatant did not stimulate enzymic activity, but supernatant from an adrenal adenoma was active.

Steroid interactions with cytochrome P-450 from testis microsomes.

The cytochrome P-450 of gonadal microsomes is an integral component of the steroid converting enzymes, 17 alpha-hydroxylase and 17,20-lyase. Interaction of the steroid substrates with this cytochrome results in a shift in the Soret band as measured by difference spectroscopy. In these studies it is shown that in contrast to placental microsomal cytochrome P-450 which binds C19 steroids, testis microsomal cytochrome P-450 primarily binds C21 steroids. However, addition of a 17 alpha- methyl, 17 beta-acetate or a 17 beta-benzoate group to testosterone permits interaction. The addition of hydroxyl or methyl groups to other positions does not affect binding. The presence of multiple oxygen functions on C21 steroids, as in cortisol and corticosterone, precludes interaction. At least one oxygen function seems necessary for binding as 5 alpha- and 5 beta-pregnane do not bind whereas 20-deoxypregnenolone (5-pregnen-3 beta-ol) does bind. These findings indicate that factors in addition to hydrophobic interactions dictate the binding of steroid substrates to testis microsomal cytochrome P-450.

Increased Urinary Estriol Excretion without Change in Blood Estriol Concentration Resulting from Bed Rest in the Third Trimester of Pregnancy in the Human

Placing normally pregnant women at bed rest during the third trimester significantly increased the amount of total estrogen excreted into the urine to 122% of the control value, but the concentration of total serum estriol was not significantly altered. Estriol biosynthesis, secretion, conjugation, metabolism, and excretion are regulated by many poorly understood factors, but for purposes of clinical evaluation of the well-being of the infant, bed rest would appear to be another circumstance under which more reliable information can be obtained by blood instead of urine analysis.