Hiroshi Wakimoto

Regulation of milk protein synthesis by progesterone in cultured mouse mammary gland

The effect of progesterone on the synthesis of milk proteins, casein and alpha-lactalbumin was investigated by culturing mammary explants from mid-pregnant mice in serum-free medium. The addition of progesterone at concentrations above 10 ng/ml inhibited both the casein and alpha-lactalbumin accumulation that were induced by the synergistic actions of insulin, prolactin and cortisol. The maximal inhibition was attained at a progesterone concentration of 100 ng/ml. The maximal level of inhibition of the alpha-lactalbumin accumulation was about 90% in the presence of insulin and prolactin or insulin, prolactin and 0.01 microgram/ml of cortisol. The inhibition of the casein accumulation by progesterone was about 80% in the presence of insulin and prolactin, and about 40% in the presence of insulin, prolactin and 1 microgram/ml of cortisol, indicating that cortisol partially antagonized the action of progesterone on the casein synthesis. When the inhibitory effect of progesterone on the accumulation of both alpha-lactalbumin and casein was examined in cultured mammary tissues from virgin, early pregnant, mid-pregnant and late pregnant mice, the degree of inhibition was markedly reduced in tissue from late pregnant mice. This indicates that the susceptibility of mammary gland to the inhibitory action of progesterone varies with the developmental stage of the tissue.

Specific antibodies to porcine zona pellucida detected by quantitative radioimmunoassay in both fertile and infertile women

The specific radioimmunoassay system was developed for the titration of the antibodies to porcine zona pellucida (ZP) in human sera by using 125I-labeled purified porcine ZP as antigen, which is known to have cross-reactivity with human ZP. The antibodies in human sera were detected in 3 of 11 (27%) women with unexplained infertility, in 16 of 48 (33%) amenorrheic patients, in 4 of 12 (33%) fertile women, and in 3 of 10 (30%) men. Moreover, antibody titers in infertile women were no higher than those in fertile women and in men. These results seem to suggest that the antibodies in human sera that cross-react with porcine ZP may not be an important factor in causing infertility in women.

Second pregnancy with spontaneous ovulation following clomiphene- or gonadotropin-induced pregnancy.

For investigation of the rates of spontaneous pregnancy following termination of pregnancy induced by treatment with clomiphene or human menopausal gonadotropin-human chorionic gonadotropin (hMG-hCG) and of spontaneous abortion in the second pregnancy, 119 women (58 with an anovulatory cycle and 141 with amenorrhea) who desired another pregnancy were studied. The rate of spontaneous pregnancy following pregnancy induced by clomiphene was 46.6%, which was significantly (p less than 0.001) higher than that following pregnancy induced by hMG (14.9%). The mean (+/- SEM) period between the termination of the first induced-ovulation pregnancy and the second spontaneous pregnancy was 16.6 +/- 1.4 months. The spontaneous abortion rates in the second pregnancies after pregnancies induced by clomiphene and hMG were 7.5% and 9.8%, respectively, which were significantly (p less than 0.001, p less than 0.05) lower than those in the first pregnancy (25.7%, 33.0%). There was no difference between the spontaneous abortion rates following a first pregnancy which terminated in abortion and those following a pregnancy which ended in birth. These data suggest that a first pregnancy in anovulatory women may restore the ovulatory cycle and have a beneficial effect on the rates of spontaneous abortion in the second pregnancy.

Roles of antiestrogen binding sites in human endometrial cancer cells

Estrogen-noncompatible antiestrogen binding sites (AEBS) as well as estrogen receptors (ER), and the growth-inhibitory effect of tamoxifen were investigated in two human endometrial cancer cell lines, IK-90 and HEC-IA cells. IK-90 cells contained specific AEBS, but no ER was found in these cells. Scatchard plot analysis of AEBS in 12,000 g supernatant from IK-90 cells showed a high affinity binding site for tamoxifen (Kd:5.6 +/- 1.0 nM) with the maximum binding site of 457 +/- 47 fmol/mg protein. However, no measurable ER or AEBS was found in HEC-IA cells. The effect of tamoxifen on the growth of cells was found to be identical in both cell lines; the addition of 10 microM tamoxifen to culture medium was cytocidal whereas tamoxifen at lower concentrations (1 nM-1 microM) did not significantly affect the growth of both IK-90 and HEC-IA cells. These results demonstrate for the first time the presence of AEBS in human endometrial cancer cells. The present results also suggest that AEBS does not play a fundamental role in mediating the growth-inhibitory effect of tamoxifen in endometrial cancer cells.

Effect of clomiphene citrate administration during the early luteal phase on the luteal function and pregnancy rate of women

Abstract To study the effect of clomiphene citrate (clomiphene) administration during the early luteal phase of the menstrual cycle on the luteal function and the pregnancy rate in women, 75 infertile women who ovulated but did not conceive after clomiphene treatment during the early follicular phase and 6 normal cycling women were chosen. Clomiphene was administered orally to 35 of the 75 infertile women at a dose of 50 mg per day for 5 days from the second day of the rise in the basal body temperature (BBT) as well as during the follicular phase, while 40 control patients received clomiphene only during the follicular phase. In the test patients, the rate of pregnancy (25.7%) was significantly (p

Lack of metabolism of progesterone to 5α-products in dog and guinea pig testes compared with immature rat testes

Abstract Testicular homogenates from immature and adult dogs and guinea pigs and those from immature rats were incubated with [ 3 H]-progesterone or [ 14 C]-3β-hydroxy-5-pregnen-20-one in the presence of cofactors. After incubation, radioactive products were separated and identified by column and paper chromatography with derivative formation and recrystallization to constant specific activity. All testes from dogs and guinea pigs of different ages converted significant amounts (up to 80%) of 3β-hydroxy-5-pregnen-20-one and progesterone to 17-hydroxy-4-pregnene-3,20-dione, 4-androstene-3, 17-dione and testosterone, but these testes converted very small amounts (less than 1% in dogs and less than 5% in guinea pigs) of these precursors to 5α-reduced metabolites of 4-ene-3-oxosteroids. However, in immature rat testes under the same incubation conditions, conversion of progesterone to 5α-reduced steroids reached 70%. In contrast to rats and mice, immature testes of dogs and guinea pigs do not form large amounts of 5α-reduced C 19 -steroids.

Effect of Clomiphene Citrate Administration during the Early Luteal Phase on the Luteal Function and Pregnancy Rate of Women

To study the effect of clomiphene citrate (clomiphene) administration during the early luteal phase of the menstrual cycle on the luteal function and the pregnancy rate in women, 75 infertile women who ovulated but did not conceive after clomiphene treatment during the early follicular phase and 6 normal cycling women were chosen. Clomiphene was administered orally to 35 of the 75 infertile women at a dose of 50 mg per day for 5 days from the second day of the rise in the basal body temperature (BBT) as well as during the follicular phase, while 40 control patients received clomiphene only during the follicular phase. In the test patients, the rate of pregnancy (25.7%) was significantly (p less than 0.05) higher than that of control patients (10.0%). On the 7th of the rise of BBT, the mean serum progesterone levels of the test patients and normal cycling women treated with clomiphene were significantly (p less than 0.05) higher than those of the control patients. However, the levels of serum estradiol, LH and FSH, the gonadotropin pulsatilities, and the pituitary responses to LH-RH in the test women were not significantly different from those of the control. These data suggest that, when administered during the early luteal phase, clomiphene may act directly on the ovary, enhancing the secretion of progesterone from the corpus luteum, and thereby increasing the rate of pregnancy in infertile women with clomiphene-induced ovulation.

Increase in collagen production with loss of androgen responsiveness in cultured androgen-responsive Shionogi carcinoma 115 cells

The collagen production of androgen-responsive and -unresponsive Shionogi carcinoma 115 cells was investigated by culturing them in a medium with or without testosterone. Androgen-unresponsive cells were obtained by culturing a cloned androgen-responsive cell in a testosterone-free medium for 12 weeks. The collagen production of androgen-responsive cells slightly increased in the absence of testosterone, whereas testosterone did not affect the collagen production of androgen-unresponsive cells. Androgen-unresponsive cells produced 3-4 times more collagen than androgen-responsive cells. The major collagen produced by both androgen-responsive and - unresponsive cells migrated to the same position in sodium dodecylsulfate:polyacylamide gel electrophoresis. The present results indicate that the collagen production of androgen-responsive Shionogi carcinoma 115 cells increases with the loss of androgen responsiveness in culture.