From fertilization to implantation.

The process of early human embryo development involves a complex series of remarkable events. Prior to the union of the human gametes, the oocyte first requires timed completion of meiosis. This vital step does not occur throughout a womans life until the one day event of ovulation and subsequent fertilization. Once the sperm enters the egg, its DNA associated proteins are replaced by oocyte histones. The two pronuclei become enveloped with oocyte derived membranes, which fuse and begin the zygotes mitotic cell cycle. Cellular division follows a predictable 12-18 hour cycle resulting in two to sixteen cell preembryos over the first several days. The sperm centrosome controls the first mitotic divisions until day four when genomic activation occurs within the morula stage. The individual blastomeres are totipotent until the morula begins compaction when cells initiate polarization. The outer cells differentiate towards a placental lineage the trophoectoderm, while the inner blastomeres become the inner cell mass or eventually the fetus. The blastocyst forms approximately 24 hours after the morula stage by the development of an inner fluid filled cavity, the blastocele. The blastocyst must first hatch from the thinning zona pellucida by alternating expansion and contraction. Implantation of the hatched blastocyst requires several steps, including apposition, attachment, penetration and trophoblast invasion. The panel members that follow will discuss each of these peri implantation steps in detail.

Functional proteomics: examining the effects of hypoxia on the cytotrophoblast protein repertoire.

The outcome of human pregnancy depends on the differentiation of cytotrophoblasts, specialized placental cells that physically connect the embryo/fetus to the mother. As cytotrophoblasts differentiate, they acquire tumor-like characteristics that enable them to invade the uterus. In a novel feedback loop, the increasingly higher levels of oxygen they encounter within the uterine wall influence their differentiation into vascular-like cells. Together, the invasive and cell surface properties of cytotrophoblasts enable them to form vascular connections with uterine blood vessels that divert maternal blood flow to the placenta, a critical hurdle in pregnancy. It is therefore important to understand how cytotrophoblasts respond to changes in oxygen tension. Here we used a proteomics approach, two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) combined with mass spectrometry, to characterize the protein repertoire of first trimester human cytotrophoblasts that were maintained under standard tissue culture conditions (20% O(2)). 2-D PAGE showed a unique protein map as compared to placental fibroblasts and human JEG-3 choriocarcinoma cells. Mass spectrometry allowed the identification of 43 spots on the cytotrophoblast map. Enzymes involved in glycolysis and responses to oxidative stress, as well as the 14-3-3 signaling/adapter proteins, were particularly abundant. Hypoxia in vitro (2% O(2)) produced discrete changes in the expression of a subset of proteins in all the aforementioned functional categories. Together, these data offer new information about the early gestation cytotrophoblast protein repertoire and the generalized mechanisms the cells use to respond to changes in oxygen tension at the maternal-fetal interface.

Does human chorionic gonadotropin have human thyrotropic activity in vivo?

Current evidence indicates that thyroid cells are sensitive to human chorionic gonadotropin (hCG) stimulation. In turn, thyroid hormones appear to influence ovarian and endometrial physiology and reproductive function. Our studies addressed the possible effect of endogenous and exogenous hCG on in vivo thyroid function in normal pregnancy and controlled ovarian hyperstimulation, respectively. Circulating concentrations of hCG in pregnant women during gestation were positively correlated with serum free thyroxine (r = 0.43, p = 0.02) and negatively correlated with thyrotropin levels in the same patients (r = 0.42, p = 0.02). By contrast, exogenous administration of hCG to effect follicular maturation in non-pregnant patients undergoing ovarian hyperstimulation resulted in lower circulating hCG concentrations than seen in pregnancy and failed to alter free thyroxine or thyrotropin levels (p > 0.22). Endogenous isoforms of hCG in early pregnancy appear to have thyrotropic activity in vivo. However, the results indicate that, under clinical conditions of controlled ovarian hyperstimulation for assisted reproduction, exogenous hCG does not affect the hypothalamic-pituitary-thyroid axis.

Hyperreactio luteinalis differentiated from severe ovarian hyperstimulation syndrome in a spontaneously conceived pregnancy.

The clinical presentation of hyperreactio luteinalis can mimic ovarian hyperstimulation. Historically, though, the former most often leads to unnecessary surgery whereas the latter is treated supportively. We present a case of a 32-year-old woman who was initially seen with markedly enlarged multicystic ovaries, ascites, and pleural effusions in the tenth week of a spontaneously conceived gestation. Despite a noniatrogenic cause, the patient received supportive management, as would be given with ovarian hyperstimulation syndrome. Making the distinction between hyperreactio luteinalis and ovarian hyperstimulation syndrome has important consequences for diagnosis and management.

Decrease in fibronectin occurs coincident with the increased expression of its integrin receptor alpha5beta1 in stress-deprived ligaments.

Stress deprivation secondary to immobilization leads to atrophic changes in periarticular soft tissues. The changes in ligaments include a disorganization of collagen and cellular ultrastructure with varied biochemical alterations resulting in a functionally weaker tissue. This study tests the hypothesis that alterations in fibronectin (Fn) and the expression of its integrin receptor alpha5beta1 in ligament fibroblasts accompany the extracellular matrix remodeling which occurs in stress-deprived knee ligaments. The left knees of eighteen New Zealand white rabbits were surgically immobilized in acute flexion. Fibroblasts within three nine week and three twelve week stress-deprived anterior cruciate ligaments (ACLs) and medial collateral ligaments (MCLs) demonstrated markedly increased immunostaining for the beta1 and alpha5 integrin subunits, as compared to fibroblasts in the contralateral unoperated control ligaments. The effects of stress deprivation on the concentration of Fn was measured by competitive ELISA on the remaining twelve rabbits. Decreases in Fn of 54.0 percent and 63.7 percent occurred in the ACL after nine and twelve weeks of stress deprivation when compared to contralateral controls. The MCL had less of a decrease, losing 37.7 percent and 41.7 percent at nine and twelve weeks, respectively. These results suggest an important role for the Fn-specific integrin receptor alpha5beta1 in remodeling stress-deprived periarticular ligamentous tissue, and the importance of maintaining normal stresses on periarticular ligaments to prevent the degradation of extracellular matrix components such as Fn.

Operative management of ectopic pregnancy: a cost analysis.

OBJECTIVES: The purpose of this study was to analyze the cost for hospital-based services related to the operative management of ectopic pregnancies and determine the most cost-conscious approach by distinguishing the constituent components. STUDY DESIGN: This is a retrospective comparative review of every ectopic pregnancy that was surgically managed at the Memorial Medical Center of Long Beach. Unit cost estimates that are based on a cost accounting system were derived and compared between different procedures according to resources used among separate services. RESULTS: Hemodynamic instability significantly increases the cost of management by increasing the length of stay and laboratory costs. Among stable patients laparoscopic excision of ectopic pregnancies saves nearly 25% per case (p < 0.001) compared with laparotomy. However, when we compared all intended laparoscopic excisions (i.e., including the 21% of cases in which laparotomy was eventually done), the savings were markedly reduced. In addition, the cost savings was lost if patients undergoing laparotomy were discharged on or before postoperative day 2. Discharging patients after laparotomy on postoperative day 1 is the least costly management for operative treatment of ectopic pregnancy. CONCLUSIONS: Whereas laparoscopy may decrease recuperation time and incisional scarring, operative cost is not a significant reason to choose laparoscopy over laparotomy in a hemodynamically stable patient, especially as postoperative stays decrease.

Quantitative assessment by competitive ELISA of fibronectin (Fn) in tendons and ligaments.

A method utilizing the enzyme-linked immunosorbent assay (ELISA) is presented for quantitating fibronectin in periarticular soft tissues from rabbits. The concentrations of fibronectin were determined in the medial collateral ligament, anterior cruciate ligament, posterior cruciate ligament, and patellar tendon. The anterior cruciate and posterior cruciate ligaments, surrounded by a synovial sheath, had similar amounts of fibronectin that were each over twice as high as that found in either medial collateral ligament or patellar tendon which have no sheath covering.