PROGESTERONE AND 20α-DIHYDROPROGESTERONE IN HUMAN MYOMETRIUM DURING PREGNANCY
Authors: Runnebaum Benno , Zander Josef 
Published: October 2009

The aim of the present investigation was to obtain further knowledge about the concentration and distribution of progesterone and 20α-dihydroprogesterone in the human myometrium during various stages of pregnancy.

For the quantitative determination of these steroids in the myometrium a method was used which consisted of the following main steps: (a) addition of radioactive tracers to the plasma or tissue homogenate (b) precipitation of protein using ethanolether (c) extraction with ethyl acetate (d) paper chromatography (e) reduction of progesterone using crystalline 20β-hydroxysteroid dehydrogenase (f) chloroacetylation of the 20β-reduced progesterone and of 20α-dihydroprogesterone (g) quantitative estimation by gas-liquid chromatography using electron capture detection. With this method the overall recovery for progesterone from the uterine tissue was 36.0 ± 7% and 42.6 ± 6% for 20α-dihydroprogesterone. The precision of the entire method varied between 6 and 8% in the quantitative range of 0.1-3.6 μg of progesterone per sample. The specificity of the method was adequate and it was found that the total progesterone content could be extracted from the uterine muscle by the method.

Progesterone was identified and quantitated in pooled extracts of 274.6 g of human myometrium obtained during 74 Caesarian sections in the last month of pregnancy. In these extracts the mean progesterone concentration was 0.16 ± 0.06 μg per g wet tissue. The identification of progesterone was established by the following criteria: (a) ultraviolet absorption (b) behaviour in paper- and thin-layer chromatography (c) retention times in gas-liquid chromatography with argon ionisation and electron capture detectors (d) formation of derivatives after enzymatic reduction with 20β-hydroxysteroid dehydrogenase (e) proof of radiochemical purity of the 14C-labelled derivative of progesterone.

Results of 84 individual progesterone determinations in the myometrium of 54 pregnant uteri showed that the mean progesterone concentrations remain fairly constant during pregnancy. During the 10th-19th week of pregnancy the mean progesterone concentration was 0.111 ± 0.081 μg per g (26 determinations), during the 33rd-38th week 0.126 ± 0.097 μg per g (23 determinations) and during the 39th-42nd week 0.124 ± 0.073 μg per g of myometrium (35 determinations). The location of the placenta was not considered in these calculations.

20α-dihydroprogesterone was identified tentatively in extracts of 27.2 g of myometrium obtained from 7 uteri at the end of pregnancy. The identification of 20α-dihydroprogesterone was based on the following criteria: (a) behaviour of the free and the chloroacetylated substance in paper- and thin-layer chromatography (b) 20α-oxidation of the compound using placental 20α-hydroxysteroid dehydrogenase (c) 20β-reduction of the 20α-oxidized compound (d) formation of the chloroacetate of the 20β-reduced material (e) retention times of the formed 20α- and 20β-dihydroprogesterone chloroacetates in gas-liquid chromatography.

The concentration of 20α-dihydroprogesterone in pooled myometrial extracts of early pregnancy (13th week) was 0.005 μg per g and in pooled extracts of myometrium at term (40th-42nd week) 0.009 μg per g wet tissue.

20β-dihydroprogesterone was identified tentatively in 210 g of tissue from a uterus in the 14th week of gestation. The mean 20β-dihydroprogesterone concentration in this myometrium was 0.00028 μg per g wet tissue.

The identification of 20β-dihydroprogesterone was based on the following criteria: (a) behaviour of the free and the chloroacetylated substance in paper- and thin-layer chromatography (b) 20β-oxidation of the substance with 20β-hydroxysteroid dehydrogenase (c) 20β-reduction of the 20β-oxidized substance using 20β-hydroxysteroid dehydrogenase (d) formation of the chloroacetate of the 20β-reduced material (e) retention times of the formed 20β-dihydroprogesterone chloroacetates in gas-liquid chromatography.

Comparative determinations of progesterone in the peripheral venous blood in the myometrium of 34 patients showed that the ratio of progesterone concentration in the myometrium versus the concentration in plasma decreases significantly during various stages of pregnancy. The progesterone concentration in early pregnancy (13th-19th week) is about 3 times higher in the myometrium than in the peripheral plasma. During late pregnancy (33rd-42nd week) the progesterone concentrations between the myometrium and peripheral plasma are to some extent adjusted.

The concentrations of progesterone in placental and antiplacental sections of the uterus changed during pregnancy. In early pregnancy (10th-16th week) the mean progesterone concentration in placental sections of the uterus was 0.145 ± 0.056 μg per g and in antiplacental sections 0.059 ± 0.019 μg per g of myometrium (11 uteri). In the last month of pregnancy the mean progesterone concentration in the placental parts of the uterus was 0.167 ± 0.035 μg per g and in the antiplacental parts of the uterus 0.121 ± 0.033 μg per g of myometrium (7 uteri). During labour (40th-42nd week) the mean progesterone concentration in the placental sites of the uterus was 0.133 ± 0.068 μg per g and in the antiplacental sites 0.104 ± 0.031 μg per g of myometrium (12 uteri). The statistical analysis of these data showed no significant difference in the myometrial progesterone concentration at the placental site, whereas the progesterone concentration of the myometrium at the antiplacental site increased significantly during pregnancy (P < 0.01).

Quantitative determinations of progesterone and 20α-dihydroprogesterone in the different sections of a whole uterus in the 14th week of gestation showed that the concentration of these two steroids in the myometrium is dependent on the location of the placenta. The sections of this uterus overlying the placenta have a 2-3 times higher progesterone- and 20α-dihydroprogesterone concentration than the myometrium sections located opposite to the placenta. Between the concentrations of progesterone and 20α-dihydroprogesterone an equal relationship is adjusted in the various sections of the uterus during the 14th week of gestation. On an average, the ratio of progesterone to 20α-dihydroprogesterone was 16:1. The statistical evaluation showed that the progesterone concentration in the myometrium during the 14th week of gestation significantly decreases with the distance from the centre of the placenta (P < 0.001).

These investigations show that there is an asymmetrical distribution of progesterone and 20α-dihydroprogesterone in the uterus during early pregnancy.

Determination of the progesterone concentration in the internal and external layer of the uterus during the 14th week of gestation indicated that the internal layer contains more progesterone than the external layer. At the placental site the mean ratio of the progesterone concentration in the internal layer as against the concentration in the external layer of the myometrium was 1.5:1 and at the antiplacental site 1.3:1. This difference was not statistically significant.

With regard to the progesterone concentration in the myometrium our results demonstrate that the uterus changes from an asymmetrical system in early pregnancy towards a more symmetrical system with a rather homogenous distribution of progesterone in the myometrium near term.