The fallopian tubes

The fallopian tubes have critical functions following ovulation. The inner linings and secretions facilitate bi-directional transport of oocyte and spermatozoa; create favourable conditions for oocyte maturation, sperm storage, capacitation, fertilization and successive cleavage of the fertilized egg; and aid its transport towards the designated implantation site in the uterus (Leese et al 2001, Shafik et al 2005) (Fig. 29.1).

Figure 29.1 Female reproductive tract.

(Reproduced with permission from Carlson 1994:15; Fig. 1-13.)

Anatomically part of the uterus, inner layers of the fallopian tubes are continuous with those in the cavity. They are composed of an internal mucosa of ciliated and secretory epithelium sensitive to changes in pituitary gonadotrophins and ovarian steroids, across the cycle (Casan et al 2000, Lei et al 1993). Intermediate layers of smooth muscle contain blood, lymph vessels, and steroid-sensitive adrenergic neurons that enable coordinated tubular motility, for the journey of the blastocyst to the uterus (Habayeb et al 2008; Wang et al 2004, 2006).

Around ovulation, smooth muscles display characteristic movements that bring the infundibulum, or distal portion of the tube, into apposition with the ovary containing the dominant follicle, by a change in orientation of muscles surrounding the ovarian fimbriae (Hunter 1988, Zervomanolakis et al 2009). One fimbria – slightly longer than the rest – reaches out to the tubal pole of the ovary and involutes, in synchrony with ovulation, to pick up the COC from the peritoneal cavity, into the enlarged trumpet-shaped infundibulum, lined with a very dense layer of ciliated secretory epithelium.

Oocyte and blastocyst transport within the tube is facilitated by:

Following fertilization, the blastocyst enters the isthmus, which has thick mucosal folds and the greatest concentration of muscle fibres. The diameter increases from 1–2 mm at the uterotubal junction, to more than 1 cm at its distal end, with a lumen ranging from 1 to 100 mm.

The interstitial portion is continuous with the uterine cavity and characterized by a marked increase in ciliated cells, and alterations in the shape of secretory cells. Muscles at the uterotubal junction are formed from four bundles. These hormonally sensitive interlacing spiral fibres allow strong constriction and relaxation of the interstitial portion of the tube. This regulates sperm transport and storage, and movement of the blastocyst towards the uterine cavity (Hunter 1988, Wildt et al 1998).

Cyclical changes

Across the ovarian cycle, the tubular mucosa undergoes cyclical alterations similar to the endometrial lining of the uterus. In the first half of the cycle, secretory and ciliated cells become larger under the influence of oestrogens. Around ovulation, ciliated cells become broader and lower while secretory cells become more distended with fluid. Following ovulation, microscopic holes appear in secretory cell membranes, which coalesce to release secretions accumulated during the first half of the cycle (Hunter 1988).

Higher rates of ciliary movements within the tube ipsilateral to the dominant follicle are regulated by increased blood flow, higher temperature and higher concentrations of ovarian steroids, compared to the contralateral tube (Zervomanola et al 2009). Around ovulation, beating of the dense concentration of cilia in the fimbriated portion is closely synchronized, propelling the COC into the ampulla. During this period, cilia in the ampulla also beat in the direction of the isthmus, suggesting that they further propel the COC towards the site of fertilization, taking the newly fertilized egg and surrounding cells from the ampulla and aiding subsequent movements of the zygote towards endometrial implantation (Hunter 1988).

The tubular environment is highly responsive to changing metabolic needs of the oocyte and blastocyst, through rhythms in secretion of growth factors, antioxidant enzymes and other regulatory molecules, and the supply of chemical and nutritive factors within the fallopian tubes following ovulation (Lapointe et al 2005). The oocyte and blastocyst are exposed to rhythmic secretion of growth regulatory molecules, and precisely timed modulations occur in oxygen tension, glucose concentrations, electrolytes and macromolecules, in different parts of the tube. The composition of tubular fluid complements the low rate of metabolite consumption in the oocyte and newly fertilized egg (Fleming et al 2004, Leese 1995, 2002, Leese et al 2001).

During their journey through the fallopian tube, the oocyte and blastocyst are composed of avascularized cells that undergo maturational changes and a highly regulated series of cleavage divisions, while utilizing exogenous sources of nutrients and growth factors.

Both demonstrate optimal functioning in an alkaline environment, with low concentrations of glucose and oxygen and plentiful supplies of albumin and growth factors (Leese 1995). The nutritive environment of the tubular lumen is regulated by hormonally induced secretions of the tubular mucosa and metabolic activities of the cumulus cell mass which surrounds the oocyte and blastocyst until implantation (Leese et al 2001) (Fig. 29.2).

Figure 29.2 The cumulus–oocyte complex and the corpus luteum following ovulation.

(Reproduced with permission from Blackburn 2007:74.)

Sperm transport

Spermatozoa enter the genital tract in approximately 3–4 mL of seminal fluid which buffers the acidity of vaginal secretions. Over 99% are immediately lost by leakage from the vagina. Those remaining spend variable times in the cervix, showing differential states of motility and rates of transport through the uterus which may increase the chances of fertilization. This reservoir of spermatozoa in the cervix are actively transported in successive waves of peristalsis to a second sperm reservoir site in the isthmus, and finally to the fertilization site, at the isthmic–ampullary junction (Bahat et al 2003, Kunz et al 1998, Wildt et al 1998, Zervomanolakis et al 2009).

Cyclical changes in the vaginal canal around ovulation provide a protected environment for ejaculated sperm. This is mirrored by cyclical changes in cervical and vaginal tissues, including relaxation and widening of the cervix, and an increase in the quantity of cervical mucus, which demonstrates a characteristic ‘stretchiness’ that facilitates sperm transport (Drobnis & Overstreet 1992).

Sperm capacitation

During their active transport through the genital tract, spermatozoa undergo a final series of maturational changes before a small number are ready for fertilization. The first of these is capacitation, caused by interactions between spermatozoa and secretions of the cervix, uterus and fallopian tube during their journey to the site of fertilization. Capacitation is when the composition of the cell membrane undergoes removal of most glycoprotein molecules added during ejaculation. Capitation seems to induce hyperactive motility, providing increased thrusting power and alterations in cell surface properties enabling entry to the zona pellucida surrounding the oocyte cell membrane (Alberts et al 2002, Drobnis & Overstreet 1992). Sperm also acquire:

Fusion of oocyte and spermatozoon

The final set of morphological transformations in spermatozoa are stimulated by binding to the zona pellucida. During this critical process, the acrosome swells and its membranes fuse with the overlying plasma membrane (see Fig. 29.3). Initially, attachment is very loose, involving a number of spermatozoa. Firmer binding follows as an oocyte-binding protein on the sperm head region is recognized by sperm receptors on the zona pellucida. The inner plasma membrane at the apical end of the spermatozoa fuses with the outer membrane of the acrosome and forms a series of membrane-bound vesicles. Proteolytic enzymes released by the acrosome reaction digest sections of the zona pellucida surrounding the sperm head. Subsequent movement through the zona occurs very rapidly, creating immediate access to the oocyte membrane. Only spermatozoa that have undergone this acrosomal exocytosis can fuse with the oocyte. Usually, the spermatozoon that makes first contact with the oocyte proceeds to fertilization. In the process of fusion, the plasma membrane of the sperm head is enveloped by microvilli on the oocyte surface (Johnson 2007, Tosti & Boni 2004).

Figure 29.3 Fertilization and cortical reaction. After capacitation, the plasma membrane of the sperm and the outer membrane of the acrosome fuse and the membranes break down, releasing enzymes that allow the sperm to penetrate the corona radiata. Sperm digest their way through the zona pellucida via enzymes associated with the inner acrosomal membrane. Sperm are engulfed by the oocyte plasma membrane. Cortical granules are released when the sperm cell contacts the membrane. These granules cause other sperm in contact with the membrane to detach.

(Reproduced with permission from Moore & Persaud 2008:30; Fig. 2-14A,B.)

Immediately after oocyte and spermatozoon fusion, the cortical reaction occurs. This is a series of ionic changes occurring in the oocyte cytoplasm, and cortical granules formed following ovulation bind to the oocyte membrane, thus releasing their content into the space between the surface of the oocyte and the surrounding zona pellucida. The vesicles contain enzymes that modify the structure of the plasma membrane blocking the entry of further spermatozoa.

From zygote to …

The zygote is the newly formed cell containing maternal and paternal chromosomes and measures about 0.15 mm in diameter (FitzGerald & FitzGerald 1994:12). Within 2–3 hours of fertilization, the zygote proceeds with the final phase of meiosis that was halted immediately following fertilization, transmitting one set of chromosomes to the next generation. The remaining set is discarded to a second polar body, on the periphery, which later undergoes apoptosis, along with the first one formed at ovulation (Fig. 29.3). Female chromosomes then divide mitotically, yielding one haploid set within the main body of the cytoplasm.

The cytoplasmic content of the sperm cell membrane combines with that of the oocyte and over the next 2–3 hours the sperm nuclear membrane gradually breaks down. Between 4 and 7 hours after cell fusion, two sets of haploid chromosomes are formed into male and female pronuclei, as each becomes surrounded by distinct membranes, in opposite poles of the cell. During this period, chromosomes synthesize DNA in preparation for the first mitotic division. As chromosomal content increases, the pronuclear membranes break down, bringing together two sets of male and female chromosomes. These events form the diploid complement of a new individual and the cell immediately proceeds with a first mitotic division, passing its genetic material to two daughter cells and forming the two-cell conceptus (Downs 2008) (Fig. 29.4).

Figure 29.4 Various stages of cleavage and formation of the blastocyst.

(Reproduced with permission from Moore & Persaud 2008: 36; Fig. 2-18A–F.)

… morula

Successive rounds of cleavages occur at approximately 12-hour intervals until 8–16 increasingly smaller cells are formed within the zona pellucida and remaining fragments of the cumulus cell mass. When cell cleavage has produced 8–16 cells, the conceptus changes its morphology and undergoes compaction to become a morula (resembling a mulberry) (Fig. 29.4). This process maximizes contiguity between adjacent cell membranes (see website).

Glucose is consumed in increasing quantities and the morula utilizes a large number of growth factors for replication (Leppens-Luisier et al 2001), including epidermal growth factor (EGF), insulin-like growth factors (IGFs), and hypoxia-inducible factors (HIFs) (Leese 1995). All blastomeres show intense staining for gonadotrophin-releasing hormone (GnRH), in possible readiness to stimulate human chorionic gonadotrophin (hCG) release just before implantation (Casan et al 1999, Kikkawa et al 2002). During this phase of formation, the morula remains within the zona pellucida. This smooth outer covering provides an overall structure preventing premature adhesion of the blastomeres to the wall of the fallopian tube and providing an immunological barrier between maternal tissue and the genetically distinct cells of the morula (Johnson 2007).

… to blastocyst

Over 24 hours, the morula continues cell cleavage to 16–32 cells. Between these two points, outer cells commit to trophoblast lineage, while the inner cell mass (ICM) retains the capacity for pluripotency. Expression of critical genes for blastocyst formation commences in the 16-cell morula (Armant 2005). Outer cells begin to pump fluid internally to form a fluid-filled cavity called the blastocoele. Nudged to one side by this fluid, the ICM expresses gap junctions, allowing transfer of ions and small molecules from one cell to the next (Downs 2008). Meanwhile, the outer layer underlying the zona pellucida differentiates into flattened trophectoderm cells that combine with the zona to protect the ICM from destruction by maternal immune cells and signals the endometrium to initiate adhesion (Schultz 1998).

The fertilized egg has now become a blastocyst (Fig. 29.4). The trophoectoderm expresses mRNA for leptin and both outer and inner cells express mRNA, protein and receptors for GnRH, and the cannabinoid receptor (CB₁) (Battista et al 2008, Casan et al 1999).

Composed of 34–64 cells, the blastocyst is programmed to prepare for the embryonic phase of formation, beginning as a free-living organism immersed in uterine secretions actively accumulated by the trophoblasts and utilized as metabolic substrates, while exchange of oxygen and carbon dioxide occurs by diffusion (Burton et al 2002, Leese 1995). Over the next 3 days, paracrine cross-talk between the blastocyst and endometrium coordinates blastocyst activation with differentiation of the endometrium to a receptive state, from 7–9 days following ovulation (Fitzgerald et al 2008, Simon et al 1997).

The corpus luteum

During the fertile cycle, the functional lifespan of the corpus luteum extends from 14 to around 280 days, but the endocrine–paracrine regulation is not well understood, particularly after the first 6 weeks of pregnancy (Handschuh et al 2007, Muyan & Boime 1997, Oon & Johnson 2000).

Research suggests that the close connections between uterine and ovarian arteriovenous systems deliver hCG and other regulatory factors from the endometrium and blastocyst, to the corpus luteum, from the early luteal phase of fertile cycles (see website).

Relaxin

Following the LH/FSH surges, large luteal cells (LLCs) and small luteal cells (SLCs) develop the capacity to secrete peptide and steroid hormones, in equal amounts. Relaxin, a peptide hormone of the insulin-like growth factor family, is a major hormone of the corpus luteum of pregnancy. Beginning during the luteal phase, under the stimulatory influence of LH/hCG, relaxin secretion peaks at 10 weeks’ gestation, decreases by around 20% and is present in maternal plasma, at stable concentrations, for the remainder of pregnancy (Bell et al 1987). Ovarian relaxin:

Hormonal signals

Within a week of fertilization, intact hCG (see website) appears in the maternal circulation and rapidly increases in the first 4 weeks after implantation, reaching peak levels of more than 100,000 mIU/mL around 9–10 weeks’ gestation, before falling rapidly to levels under 50,000 mIU/mL from around 18–20 weeks, until the end of pregnancy (Chard et al 1995, Kosaka et al 2002). In contrast, α-hCG levels increase progressively until term (Nagy et al 1994). At present, the exact mechanisms regulating intact hCG secretion are not well understood. Placental GnRH and leptin regulate secretion for the first 8 weeks of pregnancy (Islami et al 2003). Thereafter, hCG secretion as well as uterine and placental expression of LH/hCG receptors seem to be maintained by an autoregulatory mechanism (Cameo et al 2004, Kikkawa et al 2002). As pregnancy progresses, rising levels of fetal adrenal steroids seem to inhibit placental hCG (Tsakiri et al 2002).

hCG appears to have a fundamental role in regulating the establishment, development and maintenance of human pregnancy and the onset of labour (Ambrus & Rao 1994, Handschuh et al 2007, Ticconi et al 2007). Before attachment, trophectoderm cells of the blastocyst secrete hCG (Lopata et al 1997). Following attachment, hCG is released from mitotically active mononuclear cytotrophoblast cells, multinuclear syncytiotrophoblasts and extravillous cytotrophoblasts that differentiate from the trophoblast following implantation (Handschuh et al 2007, Muyan & Boime 1997). This early pregnancy rise in hCG secretion by the syncytiotrophoblast, extraplacental chorion and endometrial lining of the uterus and fallopian tubes constitutes the first neuroendocrine signal that induces maternal recognition of the fertile cycle (Handschuh et al 2007).

Pre-decidualization

During the secretory phase, stromal cells release a growing number of new matrix proteins together with surface expression of their receptors, including laminin, fibronectin, collagen and integrins, while the earlier cross-linking collagen fibrils are degraded. This process of matrix remodelling creates a thicker, looser and more soluble structure for trophoblast infiltration (see website).

Blastocyst–endometrial communication

When fertilization occurs, the designated attachment site undergoes extensive synchronized changes transforming it from a usual state of active rejection of blastocyst attachment, to a brief state of receptivity, forming a ‘window of implantation’ commencing 7 days after ovulation (Fitzgerald et al 2008, Hustin 1992, Hustin & Franchimont 1992, Lessey 2000). Some changes are regulated by paracrine cross-talk with the free floating blastocyst, while others are activated by attachment.

As the blastocyst hatches from the zona pellucida, around 6 days after fertilization, trophoblast cells surrounding the ICM make initial contact with the endometrium, by close apposition of trophoblast plasma membranes with the apical membranes of surface epithelial cells, followed by rapid proliferation and formation of junctional complexes with the surface epithelium. Apical membranes of epithelial cells display a variety of progesterone- and oestrogen-induced changes that facilitate cell recognition and interaction with the trophectoderm. These include a progressive shortening of the microvilli, creating a flatter surface, reduced thickness of the normally dense coating of glycoproteins, and inhibition of gap junctions, facilitating adhesion (Fride 2008, Hustin & Franchimont 1992, Johnson 2007, Lessey 2000).

(For more information, see website.)

Adhesion and attachment

Around the designated site of implantation, the luminal epithelium expresses calcitonin and the blastocyst-dependent heparin-binding epidermal growth factor (HB-EGF). As the zona pellucida is shed, direct communication between cell adhesion molecules and their receptors on trophoblast and surface epithelium begins the dynamically balanced processes of attachment, implantation and trophoblast replication (Fitzgerald et al 2008) (see website).