Lin Qi,1,2,* Yong Yang,1,* Xiaofang Xie,1 Liping Xiang,3 Shuyun Zhang,3 Xue Li,3 Ling Zhou,3 Dan Zhao,1 Ningzheng Dong,2,4,5 Haifang Zhang,1 Hong Zhang3 

1Department of Clinical Laboratory, the second Affiliated Hospital of Soochow University, Suzhou, Jiangsu, 215000, People’s Republic of China; 2State Key Laboratory of Radiation Medicine and Protection, Soochow University, Suzhou, Jiangsu, 215123, People’s Republic of China; 3Department of Gynaecology and Obstetrics,The second Affiliated Hospital of Soochow University, Suzhou, Jiangsu, 215000, People’s Republic of China; 4Cyrus Tang Hematology Center, Collaborative Innovation Center of Hematology, Suzhou, Jiangsu, 215000, People’s Republic of China; 5MOH Key Laboratory of Thrombosis and Hemostasis, Jiangsu Institute of Hematology, Medical College, the First Affiliated Hospital of Soochow University, Soochow University, Suzhou, Jiangsu, 215000, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Haifang Zhang, Department of Clinical Laboratory, The Second Affiliated Hospital of Soochow University, Gusu District, Suzhou, Jiangsu, 215000, People’s Republic of China, Tel +86 18896808917, Email haifangzhang@suda.edu.cn Hong Zhang, Department of Gynaecology and Obstetrics, The Second Affiliated Hospital of Soochow University, Gusu District, Suzhou, Jiangsu, 215000, People’s Republic of China, Tel +86 13814882907, Email Szzhanghong126@126.com

Background: Recurrent spontaneous abortion, a common condition in reproductive medicine, often arises from complex factors and lacks specific treatments. While studies have associated folate metabolism abnormalities with poor embryonic development, research on methionine metabolism, particularly homocysteine levels, has been limited.
Methods: In this study, we analyzed blood samples from women with RSA and those with normal pregnancies.
Results: We observed elevated homocysteine levels in women with RSA, which were correlated with increased total plasma microparticles, endothelial microparticles (EMPs), and free plasma DNA. Furthermore, we exposed human umbilical vein endothelial cells (HUVECs) to varying concentrations of homocysteine (0, 0.5, 1, 2, 4, 8 mmol/L). We found that higher homocysteine levels exacerbated its cytotoxic effects on HUVECs. Flow cytometry revealed that homocysteine compromised cell membrane integrity, enhanced membrane permeability, and promoted EMPs release. Our findings suggest that elevated homocysteine levels in women with RSA may induce significant endothelial cell apoptosis, leading to endothelial dysfunction and an increase in released microparticles and free DNA, potentially contributing to miscarriages.
Conclusion: This study may contribute to understanding and exploring the underlying mechanisms of unexplained recurrent spontaneous abortion (URSA).

Keywords: unexplained recurrent spontaneous abortion, homocysteine, microparticles, vascular endothelial cells, endothelium-derived microparticles, plasma DNA