Abstract：
Could epidermal growth factor-like domain 7 (EGFL7) be a factor involved in the preparation of the endometrium for implantation and could its dysregulation be implicated in poor reproductive outcomes?
EGFL7 is highly expressed in the endothelium and glandular epithelium throughout the menstrual cycle; it is upregulated by stromal cells in secretory phase and appears strongly reduced in endometrial biopsies and isolated stromal cells of women with unexplained recurrent pregnancy loss (uRPL) and recurrent implantation failure (RIF).
The secreted factor EGFL7, originally identified as a gene primarily expressed in endothelial cells, is also expressed by the mouse blastocyst and by mouse and human trophoblast cells. It regulates trophoblast migration and invasion by activating NOTCH1 signaling. NOTCH1 has been demonstrated to play a fundamental role in endometrial receptivity and its dysregulation may be involved in selected pregnancy complications characterized by altered endometrial receptivity, such as uRPL.
This is an exploratory study for which 84 endometrial biopsies were collected from normally fertile women, as well as from women with uRPL and RIF.
Samples were collected from women in both the proliferative and secretory phases of the menstrual cycle and stratified into three sub-groups according to the patient clinical history: 20 fertile women (8 in proliferative and 12 in secretory phase), 41 women with uRPL (6 in proliferative and 35 in secretory phase), and 27 women with RIF (8 in proliferative and 19 in secretory phase). Immunohistochemistry, real-time PCR, and western blot analyses were performed to study the expression of EGFL7 and NOTCH1, as well as the NOTCH target genes.
Analysis of spatial and temporal distribution of EGFL7 in endometrial biopsies from fertile women revealed higher levels of EGFL7 in samples from the secretory phase compared to proliferative phase. The expected expression of EGFL7 in endothelial cells was shown as well as the novel, not previously reported, expression in endometrial glands and stromal cells. EGFL7 was significantly reduced in the endometrium of women with uRPL and RIF in the secretory phases and this was associated with a downregulation of the NOTCH1 signaling pathway. Human recombinant EGFL7 was able to activate the NOTCH1 signaling pathway in endometrial stromal cells (EndSCs) obtained from fertile women but not in cells from uRPL or RIF patients. EndSCs from fertile women and decidualized in vitro for three days showed an upregulation of EGFL7 expression, whereas cells obtained from women with uRPL and RIF and decidualized in vitro did not.
This study was conducted with a relatively small number of patient samples. Although results are highly reproducible and consistent, additional observations from multicentric cohorts would strengthen the relevance of the data. Moreover, this is an in vitro study, which might only partially represent the in vivo conditions.
Our results demonstrate for the first time that EGFL7 is new player involved in decidualization and provide new insights into the pathophysiology of selected implantation defects and early pregnancy complications. Our studies have revealed that alterations in EGFL7 expression and the consequent dysregulation of NOTCH signaling are potential underlying causes of RIF and uRPL. Our results might have therapeutic relevance, as the EGFL7/NOTCH pathway may represent a potential target for medical intervention.
This study has been supported by the Grant for Fertility Innovation 2017 (Merck KGaA). There are no competing interests to disclose.
Not applicable.
EGFL7 is highly expressed in the endothelium and glandular epithelium throughout the menstrual cycle; it is upregulated by stromal cells in secretory phase and appears strongly reduced in endometrial biopsies and isolated stromal cells of women with unexplained recurrent pregnancy loss (uRPL) and recurrent implantation failure (RIF).
The secreted factor EGFL7, originally identified as a gene primarily expressed in endothelial cells, is also expressed by the mouse blastocyst and by mouse and human trophoblast cells. It regulates trophoblast migration and invasion by activating NOTCH1 signaling. NOTCH1 has been demonstrated to play a fundamental role in endometrial receptivity and its dysregulation may be involved in selected pregnancy complications characterized by altered endometrial receptivity, such as uRPL.
This is an exploratory study for which 84 endometrial biopsies were collected from normally fertile women, as well as from women with uRPL and RIF.
Samples were collected from women in both the proliferative and secretory phases of the menstrual cycle and stratified into three sub-groups according to the patient clinical history: 20 fertile women (8 in proliferative and 12 in secretory phase), 41 women with uRPL (6 in proliferative and 35 in secretory phase), and 27 women with RIF (8 in proliferative and 19 in secretory phase). Immunohistochemistry, real-time PCR, and western blot analyses were performed to study the expression of EGFL7 and NOTCH1, as well as the NOTCH target genes.
Analysis of spatial and temporal distribution of EGFL7 in endometrial biopsies from fertile women revealed higher levels of EGFL7 in samples from the secretory phase compared to proliferative phase. The expected expression of EGFL7 in endothelial cells was shown as well as the novel, not previously reported, expression in endometrial glands and stromal cells. EGFL7 was significantly reduced in the endometrium of women with uRPL and RIF in the secretory phases and this was associated with a downregulation of the NOTCH1 signaling pathway. Human recombinant EGFL7 was able to activate the NOTCH1 signaling pathway in endometrial stromal cells (EndSCs) obtained from fertile women but not in cells from uRPL or RIF patients. EndSCs from fertile women and decidualized in vitro for three days showed an upregulation of EGFL7 expression, whereas cells obtained from women with uRPL and RIF and decidualized in vitro did not.
This study was conducted with a relatively small number of patient samples. Although results are highly reproducible and consistent, additional observations from multicentric cohorts would strengthen the relevance of the data. Moreover, this is an in vitro study, which might only partially represent the in vivo conditions.
Our results demonstrate for the first time that EGFL7 is new player involved in decidualization and provide new insights into the pathophysiology of selected implantation defects and early pregnancy complications. Our studies have revealed that alterations in EGFL7 expression and the consequent dysregulation of NOTCH signaling are potential underlying causes of RIF and uRPL. Our results might have therapeutic relevance, as the EGFL7/NOTCH pathway may represent a potential target for medical intervention.
This study has been supported by the Grant for Fertility Innovation 2017 (Merck KGaA). There are no competing interests to disclose.
Not applicable.
摘要：
目的：表皮生长因子样结构域7（EGFL7）可能是子宫内膜植入准备的一个因素，它的失调可能与不良的生殖结局有关吗？
结论：EGFL7在整个月经周期中在内皮和腺体上皮中高度表达；在子宫内膜分泌细胞减少，并在子宫内膜间质失败中出现强烈的复发）。
背景：分泌因子EGFL7最初被鉴定为主要在内皮细胞中表达的基因，也由小鼠胚泡以及小鼠和人滋养层细胞表达。它通过激活NOTCH1信号调节滋养细胞迁移和侵袭。NOTCH1已被证明在子宫内膜容受性中起着重要作用，其失调可能与某些以子宫内膜容受性改变为特征的妊娠并发症有关。例如uRPL。
方法：这是一项探索性研究，从正常生育的妇女中收集了84个子宫内膜活检，以及具有uRPL和RIF的女性。
方法：从月经周期的增殖期和分泌期的女性中收集样本，并根据患者的临床病史分为三个亚组：20名可育女性（增殖期8名，分泌期12名），41例uRPL女性（6例处于增殖期，35例处于分泌期），和27名女性RIF（增殖期8名，分泌期19名）。免疫组织化学,实时PCR，进行蛋白质印迹分析以研究EGFL7和NOTCH1以及NOTCH靶基因的表达。
结果：对来自可育妇女的子宫内膜活检组织中EGFL7的空间和时间分布的分析显示，与增殖期相比，分泌期的样本中EGFL7的水平更高。显示了EGFL7在内皮细胞中的预期表达以及新的，以前没有报道过，在子宫内膜腺体和基质细胞中表达。在分泌期uRPL和RIF女性的子宫内膜中EGFL7显著降低，这与NOTCH1信号通路的下调有关。人重组EGFL7能够激活从可育妇女获得的子宫内膜基质细胞（EndSCs）中的NOTCH1信号通路，但不能激活uRPL或RIF患者的细胞。来自可育女性的EndSCs和蜕膜化3天显示EGFL7表达上调，而从患有uRPL和RIF并在体外蜕膜化的女性获得的细胞则没有。
结论：本研究使用相对少量的患者样本进行。尽管结果具有很高的可重复性和一致性，多中心队列的额外观察将加强数据的相关性.此外，这是一项体外研究，这可能仅部分代表体内条件。
结论：我们的研究结果首次表明，EGFL7是参与蜕膜化的新参与者，并为选定的种植缺陷和早期妊娠并发症的病理生理学提供了新的见解。我们的研究表明，EGFL7表达的改变和随之而来的NOTCH信号的失调是RIF和uRPL的潜在根本原因。我们的结果可能有治疗意义,EGFL7/NOTCH途径可能是医学干预的潜在目标。
背景：这项研究得到了2017年生育创新补助金（MerckKGaA）的支持。没有竞争利益可披露。
背景：不适用。
结论：EGFL7在整个月经周期中在内皮和腺体上皮中高度表达；在子宫内膜分泌细胞减少，并在子宫内膜间质失败中出现强烈的复发）。
背景：分泌因子EGFL7最初被鉴定为主要在内皮细胞中表达的基因，也由小鼠胚泡以及小鼠和人滋养层细胞表达。它通过激活NOTCH1信号调节滋养细胞迁移和侵袭。NOTCH1已被证明在子宫内膜容受性中起着重要作用，其失调可能与某些以子宫内膜容受性改变为特征的妊娠并发症有关。例如uRPL。
方法：这是一项探索性研究，从正常生育的妇女中收集了84个子宫内膜活检，以及具有uRPL和RIF的女性。
方法：从月经周期的增殖期和分泌期的女性中收集样本，并根据患者的临床病史分为三个亚组：20名可育女性（增殖期8名，分泌期12名），41例uRPL女性（6例处于增殖期，35例处于分泌期），和27名女性RIF（增殖期8名，分泌期19名）。免疫组织化学,实时PCR，进行蛋白质印迹分析以研究EGFL7和NOTCH1以及NOTCH靶基因的表达。
结果：对来自可育妇女的子宫内膜活检组织中EGFL7的空间和时间分布的分析显示，与增殖期相比，分泌期的样本中EGFL7的水平更高。显示了EGFL7在内皮细胞中的预期表达以及新的，以前没有报道过，在子宫内膜腺体和基质细胞中表达。在分泌期uRPL和RIF女性的子宫内膜中EGFL7显著降低，这与NOTCH1信号通路的下调有关。人重组EGFL7能够激活从可育妇女获得的子宫内膜基质细胞（EndSCs）中的NOTCH1信号通路，但不能激活uRPL或RIF患者的细胞。来自可育女性的EndSCs和蜕膜化3天显示EGFL7表达上调，而从患有uRPL和RIF并在体外蜕膜化的女性获得的细胞则没有。
结论：本研究使用相对少量的患者样本进行。尽管结果具有很高的可重复性和一致性，多中心队列的额外观察将加强数据的相关性.此外，这是一项体外研究，这可能仅部分代表体内条件。
结论：我们的研究结果首次表明，EGFL7是参与蜕膜化的新参与者，并为选定的种植缺陷和早期妊娠并发症的病理生理学提供了新的见解。我们的研究表明，EGFL7表达的改变和随之而来的NOTCH信号的失调是RIF和uRPL的潜在根本原因。我们的结果可能有治疗意义,EGFL7/NOTCH途径可能是医学干预的潜在目标。
背景：这项研究得到了2017年生育创新补助金（MerckKGaA）的支持。没有竞争利益可披露。
背景：不适用。
