Steroidogenesis

Sex steroids appear to be secreted primarily by the ovaries of female frogs (30,31). During breeding season, serum estradiol levels increase, which stimulates vitellogenin production by the liver. The direct effects of estradiol on amphibian ovarian follicle development are still not well understood. Sex steroid production reaches a maximum during ovulation, when gonadotropins secreted from the pituitary stimulate both estrogen and androgen production; however, the exact amounts of the various ovarian steroids made in the ovaries during natural ovulation is not well documented. In contrast, both serum and ovarian steroid levels in female frogs injected with exogenous gona-dotropins have been measured (32).

Injection of hCG promotes a rapid increase in ovarian sex steroid production that peaks after approx 8 h, which roughly coincides with ovulation. Exogenous hCG promotes moderate increases in estradiol production and dramatic increases in testosterone and androstenedione production. Although hCG stimulates ovarian progesterone production as well, its levels in the serum and ovaries relative to testosterone and estradiol remain quite low. This contrasts with gonadotropin-stimulated steroidogen-esis in mammalian ovaries, where progesterone production exceeds that of androgens and estrogens.

Several studies have been directed toward characterizing the steroidogenic pathway in the Xenopus ovary. Together, these studies explain the high androgen, moderate estradiol, and low progesterone levels described. The classical ovarian steroidogenic pathway is depicted in Fig. 2 (33).

After synthesis of the steroid pregnenolone from cholesterol, sex steroid production relies on four important enzymes. First, the cytochrome p450 enzyme CYP17 converts pregnenolone and progesterone to dehydroepiandrosterone (DHEA) and androstenedione, respectively. Second, the steroid dehydrogenase 3PHSD converts A5 steroids (such as pregnenolone and DHEA) to A4 steroids (progesterone and androstenedione, respectively). Third, 17p-hydroxysteroid dehydrogenase (17PHSD) metabolizes androstenedione to testosterone. Finally, the aromatase enzyme CYP19 converts androstenedione and testosterone to estrone and estradiol, respectively.

Several studies have demonstrated the presence of both 3PHSD and 17PHSD activities in Xenopus ovaries, localizing these enzymes primarily to follicle cells (34,35). In contrast, Xenopus CYP17 appears to be exclusively expressed in the oocytes themselves (32,35). Interestingly, pregnenolone is a very poor substrate for 3pHSD-mediated conversion to progesterone but is an excellent substrate for Xenopus CYP17. Thus, pregnenolone is preferentially converted to DHEA, which in turn is a good substrate for 3pHSD-mediated conversion to androstenedione. This selective

Fig. 2. Diagram of Xenopus ovarian steroidogenesis. CYP17 is expressed exclusively in the oocyte, which is represented by an oval shape in the center of the figure. All other steroidogenic enzymes are present in the surrounding follicle cells. Sex steroid production is therefore dependent on the germ cells, or oocytes. Estrogen and estrone enter the circulation to promote vitel-logenesis in the liver. Vitellogenin then returns to the ovary and is taken up by oocytes. Androstenedione and testosterone are produced in large amounts prior to ovulation and may promote oocyte maturation. Although progesterone is also capable of promoting maturation, its production prior to ovulation is quite low; thus, the physiologic role of progesterone in regulating oocyte maturation is uncertain.

Fig. 2. Diagram of Xenopus ovarian steroidogenesis. CYP17 is expressed exclusively in the oocyte, which is represented by an oval shape in the center of the figure. All other steroidogenic enzymes are present in the surrounding follicle cells. Sex steroid production is therefore dependent on the germ cells, or oocytes. Estrogen and estrone enter the circulation to promote vitel-logenesis in the liver. Vitellogenin then returns to the ovary and is taken up by oocytes. Androstenedione and testosterone are produced in large amounts prior to ovulation and may promote oocyte maturation. Although progesterone is also capable of promoting maturation, its production prior to ovulation is quite low; thus, the physiologic role of progesterone in regulating oocyte maturation is uncertain.

metabolism of pregnenolone via the A5 pathway likely explains why little progesterone is produced by the Xenopus ovary. Ovarian CYP19 activity, which seems to be primarily in follicle cells, is also relatively low, even in the presence of gonadotropins, thus explaining the reduced estradiol levels relative to testosterone (34).

The exclusivity of steroidogenic enzyme activities to specific cells within the ovary implies that Xenopus sex steroid production depends on both follicle cells and germ cells. This suggests an unusual positive-feedback model by which germ cells, or oocytes, are controlling their own growth and development by regulating synthesis of the steroids (estrogens) that promote vitellogenesis in the liver.

6. Oocyte Maturation and Ovulation

Just prior to ovulation, the microvilli between follicle cells and oocytes retract from the vitelline membrane so that the oocytes start to detach from the membrane (3). At the same time, chromosomal condensation begins, with spindle formation and loss of nuclear membrane definition (germinal vesicle breakdown). These features are part of oocyte maturation, which is defined as the resumption of meiosis beyond prophase I to metaphase II. Oocyte maturation is regulated by both pituitary and ovarian hormones.

In the 1930s, Rugh first demonstrated a role for pituitary factors in triggering amphibian ovulation by inducing the ovulation process with pituitary extracts (36). Shortly afterward, a role for ovarian factors in moderating ovulation was shown to be important through experiments in which ovulation in female frogs was triggered using ovarian tissue that had first been exposed to pituitary extract (37,38).

In the 1960s, progesterone was proposed to be the ovulation-inducing factor produced in the ovaries because submicromolar concentrations of progesterone promoted maturation in vitro (39). Since that time, nearly all of the seminal work studying meio-sis in Xenopus oocytes has used progesterone as the trigger for maturation. However, many other steroids are equally or more capable of promoting oocyte maturation in vitro, including the androgens testosterone and androstenedione (32,39,40). Furthermore, as mentioned, gonadotropins stimulate very little ovarian progesterone production relative to these two androgens in vivo.

These observations suggest that androgens, rather than progesterone, may be the primary physiologic mediators of Xenopus oocytes in vivo. In addition, the CYP17 expressed in isolated oocytes rapidly converts progesterone to androstenedione; thus, in vitro "progesterone-induced maturation" likely involves androgen as well as progesterone actions (32). These observations suggest that, similar to the positive-feedback loop involving oocyte-dependent estrogen synthesis and oocyte growth via vitellogen-esis, oocyte-mediated androgen production might also play a positive role in oocyte development by promoting maturation.

Investigation of the signaling mechanisms regulating steroid-induced oocyte maturation has been a subject of considerable interest for many decades (41). In contrast to most steroid-mediated signals, maturation appears to be transcription independent or nongenomic. Some nongenomic signals triggered by steroids during maturation include changes in intracellular cyclic adenosine monophosphate, promotion of the mitogen-activated protein kinase cascade, and activation of cyclin-dependent kinase 1. Many of these signals appear to involve steroid-induced changes in mRNA translation.

Interestingly, as mentioned, although transcription is very active during late oogenesis, only 20% of mRNA is translated (3). During maturation, a shift in the complement of translated mRNAs occurs, most likely because of changes in mRNA polyadenylation. One example is the MOS protein, a potent regulator of meiosis; its translation is increased by the addition of steroids (10,11,41). Studies suggest that these steroid-induced signals might in part involve classical steroid receptors that are signaling in a nongenomic fashion outside the nucleus. Other studies propose a "release of inhibition" model by which constitutive G protein-mediated signaling might be holding oocytes in meiotic arrest, and steroids might promote maturation by overcoming these inhibitory signals (42,43). More work is needed to confirm the validity of these models; however, studies in mammalian systems have shown that similar mechanisms may be present (44,45), emphasizing the usefulness of Xenopus oocytes as a general model for meiosis as well as for ovulation and oocyte development.

Acknowledgments

We thank Johne Liu for comments regarding this chapter. S. R. H. is funded by the National Institutes of Health (DK59913) and the Welch Foundation (I-1506). M. A. R. is funded in part by the Medical Scientist Training Program at the University of Texas

Southwestern Medical Center.

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  • kirsten
    How dhea works on oocyte?
    2 years ago

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