Department of Obstetrics and Gynaecology, University Hospital, Ludwig-Maximilians-University, 81377 Munich, Germany.
Laboratory for Functional Genome Analysis LAFUGA, Gene Center, Ludwig-Maximilians -University, 81377 Munich, Germany.
EBioMedicine. 2019 Mar;41:610-622. doi: 10.1016/j.ebiom.2019.02.027. Epub 2019 Mar 1.
We recently demonstrated the increased abundance of anti-trophoblast antibodies (ATAB) in sera of patients with unexplained recurrent miscarriages (uRM). Further, the ATAB-positive sera bound to JEG-3 human choriocarcinoma cells in vitro, resulting in decreased productions of β-human chorionic gonadotropin (β-hCG) and progesterone in these cells. However, the specific antigenic epitopes of ATAB have remained unknown. Therefore, it was the aim of this study to determine specific targets of ATAB in uRM patients.
Potential targets of ATAB were analyzed by 2-dimensional difference gel electrophoresis (2D-DIGE) and mass spectrometry, and thereby identifying α-Enolase (ENO1). ATAB targeting of ENO1 was further confirmed in a competitive binding assay. Levels of anti-ENO1 antibodies as well as β-hCG and progesterone were quantified with enzyme-linked immunosorbent assay (ELISA). Additionally, expression of ENO1 was analyzed in first trimester placentas by immunohistochemistry and immunofluorescence analysis.
We here identified ENO1 as a prominent target of ATAB. Serum levels of anti-ENO1 antibodies were increased in ATAB-positive compared to ATAB-negative patients. Further, increased expression of ENO1 and its co-expression with β-arrestin was found in the extra villous trophoblasts of uRM patients in first trimester placentas. In vitro, anti-ENO1 antibodies decreased the secretion of β-hCG and progesterone in JEG-3 and primary human villous trophoblast cells.
Serum anti-ENO1 antibodies might be an autoimmune biomarker for uRM. Targeting the formation of anti-ENO1 antibodies or inhibition of ENO1 expression could potentially represent therapeutic strategies for these patients. FUND: All authors declare no conflict of interest. Yao Ye was supported by the China Scholarship Council. Hellen Ishikawa-Ankerhold and Christian Schulz were supported by the SFB914, projects Z01 and A10. None of the rest authors has any conflict of interest to declare.
我们最近发现不明原因复发性流产（uRM）患者血清中抗滋养层抗体（ATAB）的丰度增加。此外，ATAB 阳性血清在体外与 JEG-3 人绒癌细胞结合，导致这些细胞中β-人绒毛膜促性腺激素（β-hCG）和孕酮的产生减少。然而，ATAB 的特定抗原表位仍然未知。因此，本研究旨在确定 uRM 患者 ATAB 的特定靶标。
通过二维差异凝胶电泳（2D-DIGE）和质谱分析潜在的 ATAB 靶标，从而鉴定α-烯醇酶（ENO1）。在竞争结合测定中进一步证实了 ATAB 对 ENO1 的靶向作用。用酶联免疫吸附试验（ELISA）定量测定抗 ENO1 抗体以及β-hCG 和孕酮的水平。此外，通过免疫组织化学和免疫荧光分析分析妊娠早期胎盘中 ENO1 的表达。
我们在这里将 ENO1 鉴定为 ATAB 的一个重要靶标。与 ATAB 阴性患者相比，ATAB 阳性患者血清中抗 ENO1 抗体的水平升高。此外，在妊娠早期 uRM 患者的绒毛外滋养层中发现 ENO1 及其与β-抑制素的共表达增加。在体外，抗 ENO1 抗体减少 JEG-3 和原代人绒毛滋养层细胞中β-hCG 和孕酮的分泌。
血清抗 ENO1 抗体可能是 uRM 的自身免疫生物标志物。针对抗 ENO1 抗体的形成或抑制 ENO1 表达可能是这些患者的潜在治疗策略。
所有作者均声明无利益冲突。姚烨得到中国国家留学基金委的资助。 Hellen Ishikawa-Ankerhold 和 Christian Schulz 得到 SFB914 项目 Z01 和 A10 的资助。其余作者均无利益冲突声明。
