Detection of pyrimethamine-induced DNA damage in mouse embryo and maternal organs by the modified alkaline single cell gel electrophoresis assay

Abstract

We studied the embryonic and maternal genotoxicity of pyrimethamine (PYR), a potent teratogen and folate antagonist, using alkaline single cell gel electrophoresis (SCG, or Comet) assay as modified by us (we used isolated nuclei instead of isolated cells). ICR mice were treated on the 13th day of pregnancy with a single oral dose of 50 mg PYR/kg. Six maternal organs (liver, kidney, lung, brain, spleen, bone marrow), maternal and fetal placentas, and two embryos were taken 6 and 16 h after treatment; the embryos were divided into head and body portions. Each sample was minced, homogenized gently, and centrifuged. The nuclei from the precipitates were used. PYR induced DNA damage in all maternal organs except spleen and bone marrow 6 h after administration. The DNA damage in all the affected organs was less at 16 h than at 6 h, and that of the kidney and brain returned to control level at 16 h. PYR also induced DNA damage in maternal and fetal placentas and embryos that was detected at 6 and 16 h, with greater damage at 6 h. Co-treatment of folinic acid calcium salt (FNA, 10 mg/kg ip), a reduced active folate form, prevented the PYR-induced DNA damage in all target tissues examined 6 h after treatment. These data indicate that the observed embryonic and maternal DNA damage caused by PYR may be related to folate deficiency, and that the modified alkaline SCG assay can be used to predict fetal/embryonic genotoxicity in vivo, in addition to the organ-specific maternal genotoxicity.