FUNCTION OF UTERINE CALCITONIN DURING IMPLANTATION

本研究计划的长期目标是探索降钙素（CT）调节胚胎着床的分子基础，降钙素是一种调节钙稳态的肽类激素。 CT的表达诱导大鼠子宫腺上皮在着床前阶段的妊娠和关闭一旦植入完成。 CT在人子宫内膜中的表达仅限于月经周期的中期分泌期（第19-24天），与推定的着床窗口密切重叠。 这些结果表明，CT可能作为一个重要的调节信号在子宫着床过程中。 该提案的具体目标是：1.确定植入期间CT调节的功能事件。 给药的反义寡脱氧核苷酸（ODNs），针对CT mRNA，到着床前阶段子宫的子宫CT mRNA的稳态水平的显着抑制的结果。 这种干预也伴随着植入胚胎数量的严重减少。 这些结果表明，种植障碍可能是由反义ODN阻断CT基因表达的直接表型后果。 通过胚胎移植实验检测反义ODN诱导的CT缺陷对（a）子宫容受性和（B）胚胎着床能力的影响。 2.探讨CT在人子宫内膜转化细胞系石川和原代培养人子宫内膜上皮细胞中的信号转导途径。 CT通过特定的细胞表面受体作用于靶细胞。 CT受体的表达在着床前子宫内膜上皮中也明显升高。将在子宫内膜上皮细胞的石川和原代培养物中研究CT通过其受体激活的第二信使途径。 3.确定介导CT细胞作用的基因。 CT刺激人子宫内膜细胞c-fos mRNA表达，抑制骨桥蛋白mRNA表达。 为了进一步了解CT如何影响胚胎-子宫相互作用，将通过消减克隆鉴定其在靶细胞中的表达响应于这种激素而被调节的其他基因，并确定其在月经周期期间在人类子宫内膜中的时空表达。 拟议的研究将提供有价值的见解，潜在的事件链的分子机制，链接的瞬时表达CT在子宫腺的胚胎-子宫内膜的相互作用在植入过程中的控制。

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.