Relationship between glucose and 20α-hydroxysteroid dehydrogenase in fetal sheep erythrocytes

Abstract

We have examined whether glucose supply to fetal sheep erythrocytes limits the rate of 20 alpha-reduction of progesterone in blood and as such is associated with the progressive loss of 20 alpha-hydroxysteroid dehydrogenase activity which has been observed from 30 days before term. Enzyme activity in erythrocytes depleted of glucose by washing was regained in the presence of at least 0.167 mmol glucose/1. The cofactor NADPH was necessary to support the reaction in lysed cells. Addition of glucose to whole blood diluted 20-fold for assay of 20 alpha-hydroxysteroid dehydrogenase did not increase the rate of reaction. Infusion of dextrose to increase fetal plasma glucose concentrations had no effect on 20 alpha-hydroxysteroid dehydrogenase activity. Over the period from 114 to 137 days of gestation, both dextrose- and saline-infused fetuses showed a decline in enzyme activity from a combined mean of 1.45 +/- 0.21 (S.E.M.) to a mean of 0.78 +/- 0.18 mumol/ml erythrocytes per h. Fetal leucocytes did not contribute significantly to the activity of 20 alpha-hydroxysteroid dehydrogenase in whole blood. The rate of 20 alpha-reduction of progesterone in the blood of eight fetuses with indwelling carotid catheters declined from 2.31 +/- 0.09 mumol/ml erythrocytes per h at 90-95 days of gestation to 0.73 +/- 0.04 mumol/ml per h at 141-145 days. However, a consistent decline was only observed after 116-120 days. The apparent equilibrium position for progesterone reduction to 20 alpha-dihydroprogesterone varied between 83.9 +/- 1.8 and 65.7 +/- 4.2%.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult Sheep Blood Metabolizes Dihydrotestosterone to 5a-Androstane-3 a,17ß-diol and 5a -Androstane-3 ß, 17ß-diol

“…The latter protocol, with an incubation period of 1 h, was used in most experiments of this series. Comparative values for progesterone reduction by 20a-HSD (Nancarrow et al 1984) in the blood of two sheep from this group were 12·0 and 8·9% by 5 hand 13'3 and 6'8% by 24 h.…”

“…D. Nancarrow and P. J. Connell Methods used for collecting and handling blood and for estimating both the rate of 20a-HSD activity (v) in reducing progesterone and the apparent equilibrium position of the reaction (Nancarrow 1983;Nancarrow et al 1984) were followed in these experiments. DHT was used as the substrate and the products were found to result from both 3a-HSD and 3{3-HSD activities.…”

“…Appropriate regions were cut out, placed in glass vials, dampened with ethanol, extracted into 7 ml scintillation cocktail and assessed for radioactivity. Calculations for v and the equilibrium position were made according to Nancarrow et al (1984). Initial experiments were carried out to determine optimal conditions for estimation of v and the equilibrium position.…”

“…A second sample of washed cells was lysed (Nancarrow et al 1984) before glucose or the cofactors NADH and NADPH were tested for their ability to restore the enzyme activity of both washed cells (glucose) and lysed cells (cofactors). Migeon et al (1962) have shown that glucose added to the incubation mixtures resulted in an increase in oestrone transformation to oestradiol in human cells.…”

“…We examined the possibility that glucose was limiting in the assays by using four sheep. Each ewe was maintained for 12 days on a diet of 750 g of a mixture of crushed oats and lucerne hay (60: 40) which was fed daily at 1400 h. Blood samples for plasma glucose estimations (Nancarrow et al 1984) and DHT reduction assays were taken prior to feeding on the 10th, 11th and 12th day after initiation of the regime. The rate of DHT reduction was estimated in the presence of 0, o· 83, 1· 67, 2· 50 and 3· 33 mmol glucose added per litre.…”