Interstitial implantation in humans and mice is highly dependent on the ability of trophoblast cells to modify their phenotype, which is that of a transporting epithelium, and invade the tissues of the uterine wall. Trophoblast cell invasion begins as the apical surface of the blastocyst adheres to the basement membrane underlying uterine luminal epithelial cells. Acquisition of adhesive activity in the apical plasma membrane domain of trophoblast cells is marked by translocation of the integrin, alpha5beta1, to that surface after culturing blastocysts for 72 h. However, strong binding to the extracellular matrix protein, fibronectin, requires contact with extracellular fibronectin, initiating a signal transduction cascade that is mediated by the ligation of alpha5beta1 and alpha5beta3 integrins, and is dependent on protein trafficking. Further delineation of the mechanism responsible for ligand-mediated up-regulation of trophoblast fibronectin binding activity is complicated by the vast number of proteins known to interact with integrins and regulate their adhesive activity. A strategy to simplify this investigation is provided by evidence that genes critical for trophoblast adhesion are expressed exclusively during the cavitation stage, approximately 72 h before adhesion-competence is attained. It is hypothesized that genes transcribed during that critical period of development will include proteins that mediate the up-regulation of fibronectin binding activity on the apical surface of the trophoblast. The differential display method will be used to identify genes first transcribed during cavitation. Genes differentially expressed in triplicate RNA preparations from embryos obtained before and during cavitation (10 h and 24 h after morulae are collected) will be isolated, cloned and sequenced for comparison with known sequences in gene data bases. Highest priority for further investigation will be given to known genes that could potentially interact with integrins. Based on prior studies, it is expected that proteins most likely involved in trophoblast adhesion will be those associated with intracellular calcium signaling, tyrosine phosphorylation, or protein trafficking after exposure of intact blastocysts to extracellular matrix. Using probes generated in these experiments and reagents such as antibodies or inhibitors already available for proteins corresponding to previously identified genes, the investigators will pursue evidence in support of a role for the selected molecules in regulating trophoblast adhesion. It is believed that these studies could reveal important mechanisms underlying the developmental regulation of cellular processes that mediate blastocyst implantation.
| Martínez-Hernández, M G; Baiza-Gutman, L A; Castillo-Trápala, A et al. (2011) Regulation of proteinases during mouse peri-implantation development: urokinase-type plasminogen activator expression and cross talk with matrix metalloproteinase 9. Reproduction 141:227-39 |
