The long-term objective of this research proposal is to explore the molecular basis of the regulation of embryonic implantation by calcitonin (CT), a peptide hormone that regulates calcium homeostasis. The expression of CT is induced in the glandular epithelium of rat uterus in the preimplantation phase of gestation and is switched off once implantation is completed. CT expression in human endometrium is restricted to the mid- secretory phase (days 19-24) of the menstrual cycle, with closely overlaps with the putative window of implantation. These findings suggest that CT may function as an important regulatory signal in the uterus during implantation.
The specific aims of this proposal are: 1. To determine the functional event(s) regulated by CT during implantation. Administration of antisense oligodeoxynucleotides (ODNs), targeted against CT mRNA, into the preimplantation phase uterus results in marked suppression of the steady-state level of uterine CT mRNA. This intervention is also accompanied by a severe reduction in the number of implanted embryos. These results suggest that the impairment of implantation could be a direct phenotypic consequence of the blockade of CT gene expression by the antisense ODN. The effect of antisense ODN-induced CT deficiency on (a) uterine receptivity and (b) the ability of the embryo to implant will be examined by embryo transfer experiments. 2. To elucidate the signal transduction pathway(s) of CT in transformed human endometrial cell line Ishikawa and in primary cultures of human endometrial epithelial cells. CT acts on target cells through specific cell surface receptors. The expression of the CT receptor is also markedly elevated in the preimplantation endometrial epithelium. The second messenger pathways that are activated by CT via its receptor will be investigated in Ishikawa and primary cultures of endometrial epithelial cells. 3. To identify the genes that mediate the cellular actions of CT. CT stimulates the expression of c-fos mRNA while it inhibits the expression of osteopontin mRNA in human endometrial cells. To further understand how CT influences the embryo-uterine interactions, additional genes whose expression in the target cells is modulated in response to this hormone will be identified by subtractive cloning and their spatio-temporal expression in human endometrium during the menstrual cycle will be determine. The proposed study will provide valuable insights into the molecular mechanisms underlying the chain of events that link the transient expression of CT in the uterine glands to the control of embryo-endometrial interactions during implantation.
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