Læknaneminn - 01.10.1996, Page 103

Rannsóknarverkefni 4. árs læknanema, útdrættir
un tvíhólfa gangráða síðastliðin ár er um 30% af heildinni og
virðist ekki fara vaxandi. Aberandi er hvernig hlutfall sjúks
sínus og III°-AV-block hefur breyst hlutfallslega með tíman-
um.
Hormonal regulation of progesterone
synthesis in cultured luteal cells from
midpregnant mice.
Guðmundur Orn Guðmundsson1.
Brandt Newcomer2, and Frank Talamantes2.
'LHI, 2Department of Biology, Sinsheimer Laboratories,
University of California, Santa Cruz, California.
Introduction: In mice, the corpus luteum is necessary
throughout gestation for progesterone (P4) production and
therefore the maintenance of pregnancy. The corpus luteum
in rodents is under the control of a luteotropic hormonal
complex. Hormones from the pituitary are the most import-
ant part of this complex during the first 10 days of pregnancy.
Thereafter, placental hormones take over the role of the
pituitary hormones. Prolactin (PRL) or prolactin-like hor-
mone (lactogen) is one component of the luteotropic hor-
monal complex. In the mouse, three lactogens are secreted
during pregnancy, each one of them dominating in the cir-
culation at a different time. Pituitary prolactin is the domin-
ant lactogen during the first 8 to 9 days of pregnancy. Ther-
eafter, lactogens from the placenta replace the pituitary
prolactin. At the time when the placenta is taking over the
role of the pituitary in reguiating the function of the corpus
luteum, a large peak of androgens appears in the circulation.
It is not known whether the androgens are important for the
maintenance of the corpus luteum or if they interact with the
placental lactogens in stimulating P4. The androgens may
serve only as a substrate for estrogen production. We were
interested to investigate a) the interaction between androgens
and lactogen in regulating P4 production; b) if the androgens
are only important as substrates for the estrogen production;
and c) how androgens, estrogens, and progestins exert their
effects on the P4 production in the mouse corpus luteum.
Materials and methods: Timed pregnant (vaginal
plug=day 0), 8-10 weeks old Swiss-Webster mice were used
for the studies. The luteal cell culture was prepared by remov-
ing the ovaries from the animals at day 10 and 14 of the
pregnancy. The corpora lutea were excised from the ovaries
and the cells dissociated in collagenase and Dnase. The cell
suspension was filtered through 150pm Nitex and the luteai
cells were then separated from the blood cells by centri-
fugation through 44% Percoll cushion. The luteal cells were
plated onto 96-well plates previously coated with fibronectin
(105 cell/well) and cultured for 3 days. The culture medium
(DME/F12+ NS and 22R-OHC) containing the different
hormonal treatments and steroid inhibitors was changed
daily. Medium was assayed for progesterone by a specific
radioimmunoassay (RIA).
Results and discussion: Mouse (m)PL-I significantly incre-
ased the P4 production in the cultured luteal cells at a con-
centration of 1 ng/ml. The increase was dose dependent
reaching maximum Ievel at 1000 ng/ml. Androstenedione
(AD) also caused a dose dependent increase in P4 secretion
when administered alone to the luteal cell culture. This incr-
ease was significant only at 5*10'6M concentration of AD.
When mPL-I and AD were administered together, the two
hormones acted synergistically to stimulate the release of P4.
To test whether the androgens serve only as substrates for
estrogen productions, two sets of experiments were carried
out. Firstly, the effect of dihydrotestosterone (DHT) on P4
production was tested. This is a nonaromatizable androgen
and, therefore, should not be capable of stimulating P4 rele-
ase if a conversion of the androgens to estrogen is needed.
Secondly, 17fi-estradiol (E2) was used with and without
mPL-I instead of the androgens to test the effect on P4 secre-
tion. The results from these experiments indicated that the
androgens were acting directly to stimulate the P4 product-
ion, as DHT was as effective in enhancing P4 release as AD,
both alone and when administered with mPL-I,. E2, on the
other hand, had no stimulatory effect on the release of P4
regardless of whether it was administered alone or in comb-
ination with mPL-I. The androgens may be affecting various
pathways to enhance P4 secredon such as stimulating
enzymes that regulate P4 production or inhibit enzymes in-
volved in P4 metabolism.
Hydroxyflutamide, an androgen receptor inhibitor, had no
effect on the basal secretion of P4 and slightly increased the
androgen-stimulated release of P4. On the other hand, ICI
182, 780, an estrogen receptor inhibitor, significantly inhi-
bited P4 secretion, while RU486, a progesterone receptor in-
hibitor, significantly stimulated P4 release from the cultured
luteal cells. The effect of the ICI 182,780 and RU486 were
reversible when the steroid receptor inhibitors were admini-
sterd with E2 and the progestin 17°°-hydroxyprogesterone
hexanoate (17^-HPH), respectively. These results show that
the stimulatory effect of androgens on P4 production is not
mediated through the nuclear androgen receptor. The effect
of the ICI 182, 780 show that although estrogens are not in-
volved in the stimulatory effect of lactogens and androgens
on P4 production, they are important for the basal release of
P4, and these effect are mediated through the nuclear estro-
gen receptor. P4 also influences its own secretion, probably
through negative feedback mechanism, as progesterone
receptor inhibitor enhanced P4 secretion, and these effect
appear to be exerted through the progesterone nuclear recept-
or.
LÆKNANEMINN
93
2. tbl. 1996, 49. árg.