Introduction Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by isolated thrombocytopenia. ITP during pregnancy represents the most prevalent and clinically challenging cause of significant thrombocytopenia, which can lead to bleeding complications during the first and second trimesters. However, although the majority of research has focused on maternal and fetal outcomes, the pathogenesis of ITP during pregnancy remains incompletely understood. 17β-Estradiol is a critically important hormone during pregnancy, and 17β-estradiol levels increase significantly after conception. Platelets are released by megakaryocytes (MKs) via cytoplasmic extensions called proplatelets, which require profound changes in microtubule and actin organization. The demarcation membrane system (DMS) is a unique and extensive membrane system formed during the maturation of MKs. The DMS is functionally essential for proplatelet formation (PPF) in MKs. The primary objective of this study was to explore the role of 17β-estradiol in the pathogenesis of ITP during pregnancy.

Methods We established mouse models of the following conditions: healthy controls (HCs), healthy pregnancy, ITP, and ITP in pregnancy. Then, we monitored platelet dynamics and isolated bone marrow MKs for RNA sequencing (RNA-seq) analysis. CD34+ hematopoietic stem cells were also isolated from the bone marrow of ITP patients and cultured in vitro with thrombopoietin (TPO), stem cell factor (SCF), varying concentrations of 17β-estradiol, and either estrogen receptor β (ERβ) or estrogen receptor α (ERα) antagonists. Subsequent analyses included RNA-seq, flow cytometry, confocal microscopy, and transmission electron microscopy (TEM).

Results Compared with those in the HC and healthy pregnancy groups, the ITP and ITP in pregnancy groups presented significantly lower platelet counts, with mean values of 866 × 10⁹/L, 760 × 10⁹/L, 449 × 10⁹/L, and 311 × 10⁹/L (P<0.001), respectively. Flow cytometry analysis revealed a significant decrease in the proportions of both CD42+ (P<0.001) MKs and megakaryocyte progenitors (MKPs) (P<0.05) in the bone marrow of ITP in pregnancy mice. GSEA of bone marrow MK RNA-seq data from murine models revealed significant negative enrichment of gene sets related to the Golgi membrane, actin monomer binding, platelet activation, and the regulation of hemopoiesis in pregnant mice with ITP.

Under high-concentration 17β-estradiol treatment, bone marrow CD34+ hematopoietic stem cell-derived MKs from ITP patients exhibited impaired maturation and polyploidization. This was manifested by decreased proportions of CD42+ (P<0.001) cells and reduced CD42 (P<0.05) mean fluorescence intensity (MFI), along with an increased percentage of 2N MKs. GSEA of the RNA-seq data revealed significant negative enrichment of gene sets associated with the trans-Golgi network, actin cytoskeleton, actin binding, and blood coagulation under high-concentration 17β-estradiol treatment. Differential gene expression analysis revealed significantly decreased expression of the CORO7-PAM16 gene under high-concentration 17β-estradiol treatment. Western blot (WB) analysis demonstrated that high-concentration 17β-estradiol significantly suppressed CORO7 expression in CD34+ hematopoietic stem cells derived MKs. Confocal microscopy and TEM revealed that high concentrations of 17β-estradiol impaired the structure of the Golgi apparatus by suppressing CORO7 expression, consequently disrupting PPF in MKs and reducing platelet production.

In vitro culture of ITP patient-derived bone marrow CD34+ hematopoietic stem cells with TPO, SCF, and high-concentration 17β-estradiol combined with either the ERα antagonist MPP or the ERβ antagonist PHTPP revealed that 17β-estradiol primarily suppresses CORO7 expression via ERα, consequently impairing PPF and reducing platelet production, as demonstrated by flow cytometry and confocal microscopy analyses.

Conclusions Our results suggest that thrombocytopenia in pregnancy may be associated with elevated 17β-estradiol levels. In vitro findings revealed that high physiological concentrations of 17β-estradiol suppress CORO7 expression in MKs, leading to structural alterations in the Golgi apparatus, impaired PPF, and a consequent reduction in platelet production. These findings partially expand the understanding of the pathogenesis of ITP during pregnancy.

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2025
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