title: Raltitrexed's effect on the development of neural tube defects in mice is associated with DNA damage, apoptosis, and proliferation.
abstract: The causal metabolic pathway and the underlying mechanism between folate deficiency and neural tube defects (NTDs) remain obscure. Thymidylate (dTMP) is catalyzed by thymidylate synthase (TS) using the folate-derived one-carbon unit as the sole methyl donor. This study aims to examine the role of dTMP biosynthesis in the development of neural tube in mice by inhibition of TS via a specific inhibitor, raltitrexed (RTX). Pregnant mice were intraperitoneally injected with various doses of RTX on gestational day 7.5, and embryos were examined for the presence of NTDs on gestational day 11.5. TS activity and changes of dUMP and dTMP levels were measured following RTX treatment at the optimal dose. DNA damage was determined by detection of phosphorylated replication protein A2 (RPA2) and γ-H2AX in embryos with NTDs induced by RTX. Besides, apoptosis and proliferation were also analyzed in RTX-treated embryos with NTDs. We found that NTDs were highly occurred by the treatment of RTX at the optimal dose of 11.5 mg/kg b/w. RTX treatment significantly inhibited TS activity. Meanwhile, dTMP was decreased associated with the accumulation of dUMP in RTX-treated embryos. Phosphorylated RPA2 and γ-H2AX were significantly increased in RTX-treated embryos with NTDs compared to control. More apoptosis and decreased proliferation were also found in embryos with NTDs induced by RTX. These results indicate that impairment of dTMP biosynthesis caused by RTX led to the development of NTDs in mice. DNA damage and imbalance between apoptosis and proliferation may be potential mechanisms.
authors.affiliation: Department of Biochemistry and Molecular Biology, Shanxi Medical University, Taiyuan, 030001, China.
authors.forename: Yanting
authors.initials: Y
authors.lastname: Dong
authors.forename: Xiuwei
authors.initials: X
authors.lastname: Wang
authors.forename: Jianlin
authors.initials: J
authors.lastname: Zhang
authors.forename: Zhen
authors.initials: Z
authors.lastname: Guan
authors.forename: Lin
authors.initials: L
authors.lastname: Xu
authors.forename: Jianhua
authors.initials: J
authors.lastname: Wang
authors.forename: Ting
authors.initials: T
authors.lastname: Zhang
authors.forename: Bo
authors.initials: B
authors.lastname: Niu
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects*; D004249:DNA Damage; D003856:Deoxyuracil Nucleotides / Q000378:metabolism; D004622:Embryo, Mammalian / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005260:Female; D005493:Folic Acid Antagonists / Q000008:administration & dosage / Q000633:toxicity; D005865:Gestational Age; D006657:Histones / Q000378:metabolism; D007274:Injections, Intraperitoneal; D008297:Male; D008810:Mice, Inbred C57BL; D009436:Neural Tube Defects / Q000139:chemically induced / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011247:Pregnancy; D021122:Protein Subunits / Q000378:metabolism; D011799:Quinazolines / Q000008:administration & dosage / Q000633:toxicity*; D051716:Replication Protein A / Q000378:metabolism; D013876:Thiophenes / Q000008:administration & dosage / Q000633:toxicity*; D013938:Thymidine Monophosphate / Q000378:metabolism; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D003856:Deoxyuracil Nucleotides; D005493:Folic Acid Antagonists; D006657:Histones; D021122:Protein Subunits; D011799:Quinazolines; D051716:Replication Protein A; D013876:Thiophenes; C475961:gamma-H2AX protein, mouse; D013938:Thymidine Monophosphate; C007267:2'-deoxyuridylic acid; D013940:Thymidylate Synthase; C068874:raltitrexed