Joseph C. Gambone
The Menstrual Cycle
Each menstrual cycle represents a complex interaction among the hypothalamus, pituitary gland, ovaries, and endometrium. Cyclic changes in gonadotropins (peptide hormones) and steroid hormones induce functional as well as morphologic changes in the ovary, resulting in follicular maturation, ovulation, and corpus luteum formation. Similar changes at the level of the endometrium allow for successful implantation of the developing embryo or a physiologic shedding of the menstrual endometrium when an early pregnancy does not occur.
The reproductive cycle can be viewed from the perspective of each of the aforementioned organ systems. The cyclic changes within the hypothalamic-pituitary axis, ovary, and endometrium are approached separately in this chapter, but these endocrinologic events occur in concert in a uniquely integrated fashion. In addition, fertilization, implantation, and placentation are discussed.
The pituitary gland lies below the hypothalamus at the base of the brain within a bony cavity (sella turcica) and is separated from the cranial cavity by a condensation of dura mater overlying the sella turcica (diaphragma sellae). The pituitary gland is divided into two major portions (Figure 4-1). The neurohypophysis, which consists of the posterior lobe (pars nervosa), the neural stalk (infundibulum), and the median eminence, is derived from neural tissue and is in direct continuity with the hypothalamus and central nervous system. The adenohypophysis, which consists of the pars distalis (anterior lobe), pars intermedia (intermediate lobe), and pars tuberalis—which surrounds the neural stalk—is derived from ectoderm.
FIGURE 4-1 Hypophyseal-pituitary portal circulatory system.
The arterial blood supply to the median eminence and the neural stalk (pituitary portal system) represents a major avenue of transport for hypothalamic secretions to the anterior pituitary.
The neurohypophysis serves primarily to transport oxytocin and vasopressin (antidiuretic hormone) along neuronal projections from the supraoptic and paraventricular nuclei of the hypothalamus to their release into the circulation.
The anterior pituitary contains different cell types that produce six protein hormones: follicle-stimulating hormone (FSH), luteinizing hormone (LH), thyroid-stimulating hormone (TSH), prolactin, growth hormone (GH), and adrenocorticotropic hormone (ACTH).
The gonadotropins, FSH and LH, are synthesized and stored in cells called gonadotrophs, whereas TSH is produced by thyrotrophs. FSH, LH, and TSH are glycoproteins, consisting of α and β subunits. The α subunits of FSH, LH, and TSH are identical. The same α subunit is also present in human chorionic gonadotropin (hCG). The β subunits are individual for each hormone. The half-life for circulating LH is about 30 minutes, whereas that of FSH is several hours. The difference in half-lives may account, at least in part, for the differential secretion patterns of these two gonadotropins.
Prolactin is secreted by lactotrophs. Unlike the case with other peptide hormones produced by the adenohypophysis, pituitary release of prolactin is under tonic inhibition by the hypothalamus. The half-life for circulating prolactin is about 20 to 30 minutes. In addition to its lactogenic effect, prolactin may directly or indirectly influence hypothalamic, pituitary, and ovarian functions in relation to the ovulatory cycle, particularly in the pathologic state of chronic hyperprolactinemia (see Chapter 32).
GONADOTROPIN SECRETORY PATTERNS
A normal ovulatory cycle can be divided into a follicular and a luteal phase (Figure 4-2). The follicular phase begins with the onset of menses and culminates in the preovulatory surge of LH. The luteal phase begins with the onset of the preovulatory LH surge and ends with the first day of menses.
FIGURE 4-2 Hormone levels during a normal menstrual cycle.
Decreasing levels of estradiol and progesterone from the regressing corpus luteum of the preceding cycle initiate an increase in FSH by a negative feedback mechanism, which stimulates follicular growth and estradiol secretion. A major characteristic of follicular growth and estradiol secretion is explained by the two-gonadotropin (LH and FSH), two-cell (theca cell and granulosa cell) theory of ovarian follicular development. According to this theory, there are separate cellular functions in the ovarian follicle wherein LH stimulates the theca cells to produce androgens (androstenedione and testosterone) and FSH then stimulates the granulosa cells to convert these androgens into estrogens (androstenedione to estrone and testosterone to estradiol), as depicted in Figure 4-3. Initially, at lower levels of estradiol, there is a negative feedback effect on the ready-release form of LH from the pool of gonadotropins in the pituitary gonadotrophs. As estradiol levels rise later in the follicular phase, there is a positive feedback on the release of storage gonadotropins, resulting in the LH surge and ovulation. The latter occurs 36 to 44 hours after the onset of this midcycle LH surge. With pharmacologic doses of progestins contained in contraceptive pills, there is a profound negative feedback effect on gonadotropin-releasing hormone (GnRH) so that none of the gonadotropin pool (ready-release or storage) is released. Hence, ovulation is (generally) blocked (see Chapter 26).
FIGURE 4-3 The two-gonadotropin (LH and FSH), two-cell (theca cell on top and granulosa cell below) theory of follicular development. Each cell is theorized to perform separate functions; LH stimulates the production of androgens (androstenedione and testosterone) in the theca cell, and FSH stimulates the aromatization of these androgens to estrogens, estrone, and estradiol in the granulosa cell.
During the luteal phase, both LH and FSH are significantly suppressed through the negative feedback effect of elevated circulating estradiol and progesterone. This inhibition persists until progesterone and estradiol levels decline near the end of the luteal phase as a result of corpus luteal regression, should pregnancy fail to occur. The net effect is a slight rise in serum FSH, which initiates new follicular growth for the next cycle. The duration of the corpus luteum’s functional regression is such that menstruation generally occurs 14 days after the LH surge in the absence of pregnancy.
Five different small peptides or biogenic amines that affect the reproductive cycle have been isolated from the hypothalamus. All exert specific effects on the hormonal secretion of the anterior pituitary gland. They are GnRH, thyrotropin-releasing hormone (TRH), somatotropin release-inhibiting factor (SRIF) or somatostatin, corticotropin-releasing factor (CRF), and prolactin release-inhibiting factor (PIF). Only GnRH and PIF are discussed in this chapter.
GnRH is a decapeptide that is synthesized primarily in the arcuate nucleus. It is responsible for the synthesis and release of both LH and FSH. Because it usually causes the release of more LH than FSH, it is less commonly called LH-releasing hormone (LH-RH) or LH-releasing factor (LRF). Both FSH and LH appear to be present in two different forms within the pituitary gonadotrophs. One is a releasable form and the other a storage form. GnRH reaches the anterior pituitary through the hypophyseal portal vessels and stimulates the synthesis of both FSH and LH, which are stored within gonadotrophs. Subsequently, GnRH activates and transforms these molecules into releasable forms. GnRH can also induce immediate release of both LH and FSH into the circulation. Some recent research that found receptors for GnRH in other tissues including the ovary suggests that GnRH may have a direct effect on ovarian function as well.
GnRH is secreted in a pulsatile fashion throughout the menstrual cycle as depicted in Figure 4-4. The frequency of GnRH release, as assessed indirectly by measurement of LH pulses, varies from about every 90 minutes in the early follicular phase to every 60 to 70 minutes in the immediate preovulatory period. During the luteal phase, pulse frequency decreases while pulse amplitude increases. A considerable variation among individuals has been identified.
FIGURE 4-4 The pulsatile release of GnRH during the normal menstrual cycle.
Intravenous and subcutaneous administration of exogenous pulsatile GnRH has been used to induce ovulation in selected women who are not ovulating as a result of hypothalamic dysfunction. A continuous (nonpulsatile) infusion of GnRH results in a reversible inhibition of gonadotropin secretion through a process of “downregulation” or desensitization of pituitary gonadotrophs. This represents the basic mechanism of action for the GnRH agonists (nonapeptides, containing only nine amino acids) that have been successfully used in the therapy of such ovarian hormone–dependent disorders as endometriosis, leiomyomas, hirsutism, and precocious puberty.
Several mechanisms control the secretion of GnRH. Estradiol appears to enhance hypothalamic release of GnRH and may help induce the midcycle LH surge by increasing GnRH release or by enhancing pituitary responsiveness to the decapeptide. Gonadotropins have an inhibitory effect on GnRH release. Catecholamines may play a major regulatory role as well. Dopamine is synthesized in the arcuate and periventricular nuclei and may have a direct inhibitory effect on GnRH secretion through the tuberoinfundibular tract that projects onto the median eminence. Serotonin also appears to inhibit GnRH pulsatile release, whereas norepinephrine stimulates it. Endogenous opioids suppress release of GnRH from the hypothalamus in a manner that may be partially regulated by ovarian steroids.
The hypothalamus produces PIF, which exerts chronic inhibition of prolactin release from the lactotrophs. A number of pharmacologic agents (e.g., chlorpromazine) that affect dopaminergic mechanisms influence prolactin release. Dopamine itself is secreted by hypothalamic neurons into the hypophyseal portal vessels and inhibits prolactin release directly within the adenohypophysis. Based on these observations, it has been proposed that hypothalamic dopamine may be the major PIF. In addition to the regulation of prolactin release by PIF, the hypothalamus may also produce prolactin-releasing factors (PRFs) that can elicit large and rapid increases in prolactin release under certain conditions, such as breast stimulation during nursing. All PIFs and PRFs have not been clearly characterized biochemically as of 2008. TRH serves to stimulate prolactin release as well. This phenomenon may explain the association between primary hypothyroidism (with secondary TRH elevation) and hyperprolactinemia. The precursor protein for GnRH, called GnRH-associated peptide (GAP), has been identified to be both a potent inhibitor of prolactin secretion and an enhancer of gonadotropin release. These findings suggest that this GnRH-associated peptide may also be a physiologic PIF and could explain the inverse relationship between gonadotropin and prolactin secretions seen in many reproductive states.
During early follicular development, circulating estradiol levels are relatively low. About 1 week before ovulation, levels begin to increase, at first slowly, then rapidly. The conversion of testosterone to estradiol in the granulosa cell of the follicle occurs through an enzymatic process called aromatization and is depicted in Figure 4-3. The levels generally reach a maximum 1 day before the midcycle LH peak. After this peak and before ovulation, there is a marked and precipitous fall. During the luteal phase, estradiol rises to a maximum 5 to 7 days after ovulation and returns to baseline shortly before menstruation. Estrone secretion by the ovary is considerably less than secretion of estradiol but follows a similar pattern. Estrone is largely derived from the conversion of androstenedione through the action of the enzyme aromatase (Figure 4-5).
FIGURE 4-5 Steroidogenic pathways showing aromatization in red. Cmpd B, corticosterone; cmpd S, II-deoxycortisol; DOC, desoxycorticosterone; OH, hydroxylase.
During follicular development, the ovary secretes only very small amounts of progesterone and 17α-hydroxyprogesterone. The bulk of the progesterone comes from the peripheral conversion of adrenal pregnenolone and pregnenolone sulfate. Just before ovulation, the unruptured but luteinizing graafian follicle begins to produce increasing amounts of progesterone. At about this time, a marked increase also occurs in serum 17α-hydroxyprogesterone. The elevation of basal body temperature is temporally related to the central effect of progesterone. As with estradiol, secretion of progestins by the corpus luteum reaches a maximum 5 to 7 days after ovulation and returns to baseline shortly before menstruation. Should pregnancy occur, progesterone levels and therefore basal body temperature remain elevated.
Both the ovary and the adrenal glands secrete small amounts of testosterone, but most of the testosterone is derived from the metabolism of androstenedione, which is also secreted by both the ovary and the adrenal gland. Near midcycle, an increase occurs in plasma androstenedione, which reflects enhanced secretion from the follicle. During the luteal phase, a second rise occurs in androstenedione, which reflects enhanced secretion by the corpus luteum. The adrenal gland also secretes androstenedione in a diurnal pattern similar to that of cortisol. The ovary secretes small amounts of the very potent dihydrotestosterone (DHT), but the bulk of DHT is derived from the conversion of androstenedione and testosterone. The majority of dehydroepiandrosterone (DHEA) and virtually all DHEA sulfate (DHEA-S), which are weak androgens, are secreted by the adrenal glands, although small amounts of DHEA are secreted by the ovary.
Circulating estrogens and androgens are mostly bound to specific sex hormone–binding globulins (SHBG) or to serum albumin. The remaining fraction of sex hormones is unbound (free), and this is the biologically active fraction. It is unclear whether steroids bound to serum proteins (e.g., albumin) are accessible for tissue uptake and utilization. The synthesis of SHBG in the liver is increased by estrogens and thyroid hormones but decreased by testosterone.
Serum prolactin levels do not change strikingly during the normal menstrual cycle. Both the serum level of prolactin and prolactin release in response to TRH are somewhat more elevated during the luteal phase than during the mid-follicular phase of the cycle. This suggests that high amounts of circulating estradiol and progesterone may enhance prolactin release. Prolactin release varies throughout the day, with the highest levels occurring during sleep.
Prolactin may participate in the control of ovarian steroidogenesis. Prolactin concentrations in follicular fluid change markedly during follicular growth. The highest prolactin concentrations are seen in small follicles during the early follicular phase. Prolactin concentrations in the follicular fluid may be inversely related to the production of progesterone. In addition, hyperprolactinemia may alter gonadotropin secretion. Despite these observations, the physiologic role of prolactin during the normal menstrual cycle has not been clearly established.
Primordial follicles undergo sequential development, differentiation, and maturation until a mature graafian follicle is produced. The follicle then ruptures, releasing the ovum. Subsequent luteinization of the ruptured follicle produces the corpus luteum.
At about 8 to 10 weeks of fetal development, oocytes become progressively surrounded by precursor granulosa cells, which then separate themselves from the underlying stroma by a basal lamina. This oocyte–granulosa cell complex is called a primordial follicle. In response to gonadotropin and ovarian steroids, the follicular cells become cuboidal, and the stromal cells around the follicle become prominent. This process, which takes place in utero (i.e., in the fetal ovary) at between 20 and 24 weeks’ gestation, results in a primary follicle. As granulosa cells proliferate, a clear gelatinous material surrounds the ovum, forming the zona pellucida. This larger unit is called a secondary follicle.
In the adult ovary, a graafian follicle forms as the innermost three or four layers of rapidly multiplying granulosa cells become cuboidal and adherent to the ovum (cumulus oophorus). In addition, a fluid-filled antrum forms among the granulosa cells. As the liquor continues to accumulate, the antrum enlarges, and the centrally located primary oocyte migrates eccentrically to the wall of the follicle. The innermost layer of granulosa cells of the cumulus, which are in close contact with the zona pellucida, become elongated and form the corona radiata. The corona radiata is released with the oocyte at ovulation. Covering the granulosa cells is a thin basement membrane, outside of which connective tissue cells organize themselves into two coats: the theca interna and theca externa.
During each cycle, a cohort of follicles is recruited for development. Among the many developing follicles, only one usually continues differentiation and maturation into a follicle that ovulates.The remaining follicles undergo atresia. On the basis of in vitro measurement of local steroid levels, growing follicles can be classified as either estrogen predominant or androgen predominant. Follicles greater than 10 mm in diameter are usually estrogen predominant, whereas smaller follicles are usually androgen predominant. Mature preovulatory follicles reach mean diameters of about 18 to 25 mm. Furthermore, in estrogen-predominant follicles, antral FSH concentrations continue to rise while serum FSH levels decline beginning at the mid-follicular phase. In smaller, androgen-predominant follicles, antral fluid FSH values decrease while serum FSH levels decline; thus, the intrafollicular steroid milieu appears to play an important role in determining whether a follicle undergoes maturation or atresia. Additional follicles may be “rescued” from atresia by administration of exogenous gonadotropins.
Follicular maturation is dependent on the local development of receptors for FSH and LH. FSH receptors are present on granulosa cells. Under FSH stimulation, granulosa cells proliferate, and the number of FSH receptors per follicle increases proportionately. Thus, the growing primary follicle is increasingly more sensitive to stimulation by FSH; as a result, estradiol levels increase. Estrogens, particularly estradiol, enhance the induction of FSH receptors and act synergistically with FSH to increase LH receptors.
During early stages of folliculogenesis, LH receptors are present only on the theca interna layer. LH stimulation induces steroidogenesis and increases the synthesis of androgens by thecal cells. In nondominant follicles, high local androgen levels may enhance follicular atresia. However, in the follicle destined to reach ovulation, FSH induces aromatase enzyme and its receptor formation within the granulosa cells. As a result, androgens produced in the theca interna of the dominant follicle diffuse into the granulosa cells and are aromatized into estrogens. FSH also enhances the induction of LH receptors on the granulosa cells of the follicle that is destined to ovulate.These are essential for the appropriate response to the LH surge, leading to the final stages of maturation, ovulation, and the luteal phase production of progesterone. Thus, the presence of greater numbers of FSH receptors and granulosa cells and increased induction of aromatase enzyme and its receptors may differentiate between the follicle of the initial cohort that will develop normally and those that will undergo atresia.
Growth factors such as insulin, insulin-like growth factor (IGF), fibroblast growth factor (FGF), and epidermal growth factor (EGF) may also play significant mitogenic roles in folliculogenesis, including enhanced responsiveness to FSH.
The preovulatory LH surge initiates a sequence of structural and biochemical changes that culminate in ovulation. Before ovulation, a general dissolution of the entire follicular wall occurs, particularly the portion that is on the surface of the ovary. Presumably this occurs as a result of the action of proteolytic enzymes. With degeneration of the cells on the surface, a stigma forms, and the follicular basement membrane finally bulges through the stigma. When this ruptures, the oocyte, together with the corona radiata and some cumulus oophora cells, is expelled into the peritoneal cavity, and ovulation takes place.
Ovulation is now known from ultrasonic studies to be a gradual phenomenon, with the collapse of the follicle taking from several minutes to as long as an hour or more. The oocyte adheres to the surface of the ovary, allowing an extended period during which the muscular contractions of the fallopian tube may bring it in contact with the tubal epithelium. Probably both muscular contractions and tubal ciliary movement contribute to the entry of the oocyte into, and the transportation along, the fallopian tube. Ciliary activity may not be essential because some women with immotile cilia also become pregnant.
At birth, primary oocytes are in the prophase of the first meiotic division. They continue in this phase until the next maturation division occurs in conjunction with the midcycle LH surge. A few hours preceding ovulation, the chromatin is resolved into distinct chromosomes, and meiotic division takes place with unequal distribution of the cytoplasm to form a secondary oocyte and the first polar body. Each element contains 23 chromosomes, each in the form of two monads. The second maturation spindle forms immediately, and the oocyte remains at the surface of the ovary. No further development takes place until after ovulation and fertilization have occurred. At that time, and before the union of the male and female pronuclei, another division occurs to reduce the chromosomal component of the egg pronucleus to 23 single chromosomes (22 plus X or Y), each composed of the one monad. The ovum and a second polar body are thus formed. The first polar body may also divide.
LUTEINIZATION AND CORPUS LUTEUM FUNCTION
After ovulation and under the influence of LH, the granulosa cells of the ruptured follicle undergo luteinization. These luteinized granulosa cells, plus the surrounding theca cells, capillaries, and connective tissue, form the corpus luteum, which produces copious amounts of progesterone and some estradiol. The normal functional life span of the corpus luteum is about 9 to 10 days. After this time it regresses, and unless pregnancy occurs, menstruation ensues, and the corpus luteum is gradually replaced by an avascular scar called a corpus albicans. The events occurring in the ovary during a complete cycle are shown in Figure 4-6.
FIGURE 4-6 Schematic representation of the sequence of events occurring in the ovary during a complete follicular cycle.
(Adapted from Yen SC, Jaffe R [eds]: Reproductive Endocrinology. Philadelphia, WB Saunders, 1978.)
Histophysiology of the Endometrium
The endometrium is uniquely responsive to the circulating progestins, androgens, and estrogens. It is this responsiveness that gives rise to menstruation and makes implantation and pregnancy possible.
Functionally, the endometrium is divided into two zones: (1) the outer portion, or functionalis, that undergoes cyclic changes in morphology and function during the menstrual cycle and is sloughed off at menstruation; and (2) the inner portion, or basalis, that remains relatively unchanged during each menstrual cycle and, after menstruation, provides stem cells for the renewal of the functionalis. Basal arteries are regular blood vessels found in the basalis, whereas spiral arteries are specially coiled blood vessels seen in the functionalis.
The cyclic changes in histophysiology of the endometrium can be divided into three stages: the menstrual phase, the proliferative or estrogenic phase, and the secretory or progestational phase.
Because it is the only portion of the cycle that is visible externally, the first day of menstruation is taken as day 1 of the menstrual cycle. The first 4 to 5 days of the cycle are defined as the menstrual phase. During this phase, there is disruption and disintegration of the endometrial glands and stroma, leukocyte infiltration, and red blood cell extravasation. In addition to this sloughing of the functionalis, there is a compression of the basalis due to the loss of ground substances. Despite these degenerative changes, early evidence of renewed tissue growth is usually present at this time within the basalis of the endometrium.
The proliferative phase is characterized by endometrial proliferation or growth secondary to estrogenic stimulation. Because the bases of the endometrial glands lie deep within the basalis, these epithelial cells are not destroyed during menstruation.
During this phase of the cycle, the large increase in estrogen secretion causes marked cellular proliferation of the epithelial lining, the endometrial glands, and the connective tissue of the stroma(Figure 4-7). Numerous mitoses are present in these tissues, and there is an increase in the length of the spiral arteries, which traverse almost the entire thickness of the endometrium. By the end of the proliferative phase, cellular proliferation and endometrial growth have reached a maximum, the spiral arteries are elongated and convoluted, and the endometrial glands are straight, with narrow lumens containing some glycogen.
FIGURE 4-7 Early proliferative phase endometrium. Note the regular, tubular glands lined by pseudostratified columnar cells.
Following ovulation, progesterone secretion by the corpus luteum stimulates the glandular cells to secrete glycogen, mucus, and other substances. The glands become tortuous and the lumens are dilated and filled with these substances. The stroma becomes edematous. Mitoses are rare. The spiral arteries continue to extend into the superficial layer of the endometrium and become convoluted (Figure 4-8).
FIGURE 4-8 Late secretory phase endometrium. Note the tortuous, saw-toothed appearance of the endometrial glands with secretions in the lumens. The stroma is edematous and necrotic during this stage, leading to sloughing of the endometrium at the time of menstruation.
The marked changes that occur in endometrial histology during the secretory phase permit relatively precise timing (dating) of secretory endometrium.
If pregnancy does not occur by day 23, the corpusluteum begins to regress, secretion of progesterone and estradiol declines, and the endometrium undergoes involution. About 1 day before the onset of menstruation, marked constriction of the spiral arterioles takes place, causing ischemia of the endometrium followed by leukocyte infiltration and red blood cell extravasation. It is thought that these events occur secondary to prostaglandin production by the endometrium. The resulting necrosis causes menstruation or sloughing of the endometrium. Thus, menstruation, which clinically marks the beginning of the menstrual cycle, is actually the terminal event of a physiologic process that enables the uterus to be prepared to receive another conceptus.
Spermatogenesis, Sperm Capacitation, and Fertilization
Fertilization, or conception, is the union of male and female pronuclear elements. Conception normally takes place in the fallopian tube, after which the fertilized ovum continues to the uterus, where implantation occurs and development of the conceptus continues.
Spermatogenesis requires about 74 days. Together with transportation, a total of about 3 months elapses before sperm are ejaculated. The sperm achieve motility during their passage through the epididymis, but sperm capacitation, which renders them capable of fertilization in vivo, does not occur until they are removed from the seminal plasma after ejaculation. Interestingly, sperm aspirated from the epididymis and testis can be used to achieve fertilization in vitro employing intracytoplasmic injection techniques directly into the ooplasm.
Estrogen levels are high at the time of ovulation, resulting in an increased quantity, decreased viscosity, and favorable electrolyte content of the cervical mucus. These are the ideal characteristics for sperm penetration. The average ejaculate contains 2 to 5 mL of semen; 40 to 300 million sperm may be deposited in the vagina, 50% to 90% of which are morphologically normal. Fewer than 200 sperm achieve proximity to the egg. Only one sperm fertilizes a single egg released at ovulation.
The major loss of sperm occurs in the vagina following coitus, with expulsion of the semen from the introitus playing an important role. In addition, digestion of sperm by vaginal enzymes, destruction by vaginal acidity, phagocytosis of sperm along the reproductive tract, and further loss from passage through the fallopian tube into the peritoneal cavity all diminish the number of sperm capable of achieving fertilization.
Those sperm that do migrate from the alkaline environment of the semen to the alkaline environment of the cervical mucus exuding from the cervical os are directed along channels of lower-viscosity mucus into the cervical crypts where they are stored for later ascent. Two waves of passage to the tubes may occur. Uterine contractions, probably facilitated by prostaglandin in the seminal plasma, propel sperm to the tubes within 5 minutes. Some evidence indicates that these sperm may not be as capable of fertilization as those that arrive later largely under their own power. Sperm may be found within the peritoneal cavity for long periods, but it is not known whether they are capable of fertilization. Ova are usually fertilized within 12 hours of ovulation.
Capacitation is the physiologic change that sperm must undergo in the female reproductive tract before fertilization. Human sperm can also undergo capacitation after a short incubation in defined culture media without residence in the female reproductive tract, which allows for in vitro fertilization (see Chapter 34).
The acrosome reaction is one of the principal components of capacitation. The acrosome, a modified lysosome, lies over the sperm head as a kind of “chemical drill-bit” designed to enable the sperm to burrow its way into the oocyte (Figure 4-9). The overlying plasma membrane becomes unstable and eventually breaks down, releasing hyaluronidase, a neuraminidase, and corona-dispersing enzyme. Acrosin, bound to the remaining inner acrosomal membrane, may play a role in the final penetration of the zona pellucida. The latter contains species-specific receptors for the plasma membrane. After traversing the zona, the postacrosomal region of the sperm head fuses with the oocyte membrane, and the sperm nucleus is incorporated into the ooplasm. This process triggers release of the contents of the cortical granules that lie at the periphery of the oocyte. This cortical reaction results in changes in the oocyte membrane and zona pellucida that prevent the entrance of further sperm into the oocyte.
FIGURE 4-9 The sperm head.
The process of capacitation may be inhibited by a factor in the semen, thus preserving maximal release of enzyme to allow effective penetration of the corona and zona pellucida surrounding the oocyte. The cellular investments of the oocyte may further activate the sperm, thus facilitating penetration to the oocyte membrane. The corona is not required for normal fertilization to occur because its removal has no effect on the rate or quality of fertilization in vitro. The major function of these surrounding granulosa cells and their intercellular matrix may be to serve as a sticky mass that causes adherence to the ovarian surface and to the mucosa of the tubal epithelium.
Following penetration of the oocyte, the sperm nucleus decondenses to form the male pronucleus, which approaches and finally fuses with the female pronucleus at syngamy to form the zygote. Fertilization restores the diploid number of chromosomes and determines the sex of the zygote. In couples with infertility resulting from severe sperm abnormalities, fertilization and subsequent pregnancy can be successfully achieved after the injection of a single sperm, with or without its tail, into the cytoplasm of the oocyte (see Chapter 34).
Cleavage, Morula, Blastocyst
Following fertilization, cleavage occurs. This consists of a rapid succession of mitotic divisions that produce a mulberry-like mass known as a morula. Fluid is secreted by the outer cells of the morula, and a single fluid-filled cavity develops, known as the blastocyst cavity. An inner-cell mass can be defined, attached eccentrically to the outer layer of flattened cells; the latter becomes the trophoblast. The embryo at this stage of development is called a blastocyst, and the zona pellucida disappears at about this time. A blastocyst cell can be removed and tested for genetic imperfections without harming further development of the conceptus.
The fertilized ovum reaches the endometrial cavity about 3 days after ovulation.
Hormones influence egg transport. Estrogen causes “locking” of the egg in the tube, and progesterone reverses this action. Prostaglandins have diverse effects. Prostaglandin E relaxes the tubal isthmus, whereas prostaglandin F stimulates tubal motility. It is unknown whether abnormalities of egg transportation play a role in infertility, but in animal studies, acceleration of ovum transportation causes a failure of implantation. Additional cytokines may be released by the tubal epithelium and embryo to enhance embryo transportation and development and to signal the impending implantation to the endometrium.
Initial embryonic development primarily occurs in the ampullary portion of the fallopian tube with subsequent rapid transit through the isthmus. This process takes about 3 days. On reaching the uterine cavity, the embryo undergoes further development for 2 to 3 days before implanting. The zona is shed, and the blastocyst adheres to the endometrium, a process that is probably dependent on the changes in the surface characteristics of the embryo, such as electrical charge and glycoprotein content. A variety of proteolytic enzymes may play a role in separating the endometrial cells and digesting the intercellular matrix.
Initially, the wall of the blastocyst facing the uterine lumen consists of a single layer of flattened cells. The thicker opposite wall has two zones: the trophoblast and the inner cell mass (embryonic disk). The latter differentiates at 7.5 days into a thick plate of primitive “dorsal” ectoderm and an underlying layer of “ventral” endoderm. A group of small cells appears between the embryonic disk and trophoblast. A space develops within them, which becomes the amniotic cavity.
Under the influence of progesterone, decidual changes occur in the endometrium of the pregnant uterus. The endometrial stromal cells enlarge and form polygonal or round decidual cells. The nuclei become round and vesicular, and the cytoplasm becomes clear, slightly basophilic, and surrounded by a translucent membrane. During pregnancy, the decidua thickens to a depth of 5 to 10 mm. The decidua basalis is the decidual layer directly beneath the site of implantation. Integrins, a class of proteins involved in cell-to-cell adherence, peak within the endometrium at the time of implantation and may play a significant role. Additional growth factors act in a synergistic fashion to enhance the implantation process. The decidua capsularis is the layer overlying the developing ovum and separating it from the rest of the uterine cavity. The decidua vera (parietalis) is the remaining lining of the uterine cavity (Figure 4-10). The space between the decidua capsularis and decidua vera is obliterated by the 4th month with fusion of the capsularis and vera.
FIGURE 4-10 Early stage of implantation.
The decidua basalis enters into the formation of the basal plate of the placenta. The spongy zone of the decidua basalis consists mainly of arteries and dilated veins. The decidua basalis is invaded extensively by trophoblastic giant cells, which first appear as early as the time of implantation. Minute levels of hCG appear in the maternal serum at this time. Nitabuch’s layer is a zone of fibrinoid degeneration where the trophoblast meets the decidua. When the decidua is defective, as in placenta accreta, Nitabuch’s layer is absent.
When the free blastocyst contacts the endometrium after 4 to 6 days, the syncytiotrophoblast, a syncytium of cells, differentiates from the cytotrophoblast. At about 9 days, lacunae, irregular fluid-filled spaces, appear within the thickened trophoblastic syncytium. This is soon followed by the appearance of maternal blood within the lacunae as maternal tissue is destroyed and the walls of the mother’s capillaries are eroded.
As the blastocyst burrows deeper into the endometrium, the trophoblastic strands branch to form the solid, primitive villi traversing the lacunae. The villi, which are first distinguished about the 12th day after fertilization, are the essential structures of the definitive placenta. Located originally over the entire surface of the ovum, the villi later disappear except over the most deeply implanted portion, the future placental site.
Embryonic mesenchyme first appears as isolated cells within the cavity of the blastocyst. When the cavity is completely lined with mesoderm, it is termed the extraembryonic celom. Its membrane, the chorion, is composed of trophoblast and mesenchyme. When the solid trophoblast is invaded by a mesenchymal core, presumably derived from cytotrophoblast, secondary villi are formed.
Maternal venous sinuses are tapped about 15 days after fertilization. By the 17th day, both fetal and maternal blood vessels are functional, and a placental circulation is established. The fetal circulation is completed when the blood vessels of the embryo are connected with chorionic blood vessels that are formed from cytotrophoblast. Proliferation of cellular trophoblasts at the tips of the villi produces cytotrophoblastic columns that progressively extend through the peripheral syncytium. Cytotrophoblastic extensions from columns of adjacent villi join together to form the cytotrophoblastic shell, which attaches the villi to the decidua. By the 19th day of development, the cytotrophoblastic shell is thick. Villi contain a central core of chorionic mesoderm, where blood vessels are developing, and an external covering of syncytiotrophoblasts or syncytium.
By 3 weeks, the relationship of the chorion to the decidua is evident. The greater part of the chorion, denuded of villi, is designated the chorion laeve (smooth chorion). Until near the end of the 3rd month, the chorion laeve remains separated from the amnion by the extraembryonic celomic cavity. Thereafter, amnion and chorion are in intimate contact. The villi adjacent to the decidua basalis enlarge and branch (chorion frondosum) and progressively assume the form of the fully developed human placenta (Figure 4-11). By 16 to 20 weeks, the chorion laeve contacts and fuses with the decidua vera, thus obliterating most of the uterine cavity.
FIGURE 4-11 Relationship of the chorion to the placenta.
Throughout normal pregnancy, the amniotic fluid compartment allows the fetus room for growth, movement, and development. Without amniotic fluid, the uterus would contract and compress the fetus. In cases of leakage of amniotic fluid early in the first trimester, the fetus may develop structural abnormalities including facial distortion, limb reduction, and abdominal wall defects secondary to uterine compression.
Toward mid-pregnancy (20 weeks), the amniotic fluid becomes increasingly important for fetal pulmonary development. The latter requires a fluid-filled respiratory tract and the ability of the fetus to “breathe” in utero, moving amniotic fluid into and out of the lungs. The absence of adequate amniotic fluid during mid-pregnancy is associated with pulmonary hypoplasia at birth, which is often incompatible with life.
The amniotic fluid also has a protective role for the fetus. It contains antibacterial activity and acts to inhibit the growth of potentially pathogenic bacteria. During labor and delivery, the amniotic fluid continues to serve as a protective medium for the fetus, aiding dilation of the cervix. The premature infant, with its fragile head, may benefit most from delivery with the amniotic membranes intact (en caul). In addition, the amniotic fluid may serve as a means of communication for the fetus. Fetal maturity and readiness for delivery may be signaled to the maternal uterus through fetal urinary hormones excreted into the amniotic fluid.
Adashi E. The ovarian cycle. In Yen S.S.C., Jaffe R.B., editors: Reproductive Endocrinology, 4th ed., Philadelphia: WB Saunders, 1997.
Olive D.L., Palter S.F. Reproductive physiology. Berek and Novak’s Gynecology, 14th ed.. Philadelphia: Lippincott Williams & Wilkins; 2007.
Speroff L., Glass R.H., Kase N.G. Clinical Gynecologic Endocrinology and Fertility, 6th ed. Baltimore: Williams & Wilkins; 1999.
Strauss J., Gurpide E. The endometrium: Regulation and dysfunction. In Yen S.S.C., Jaffe R.B., editors: Reproductive Endocrinology, 4th ed., Philadelphia: WB Saunders, 1997.
Yen S.S.C. The human menstrual cycle: Neuroendocrine regulation. In Yen S.S.C., Jaffe R.B., editors: Reproductive Endocrinology, 4th ed., Philadelphia: WB Saunders, 1997.