Altered BER and increased oxidative stress in DS fetal liver. (A) UDG expression. RNA was isolated and reverse transcribed from DS and non-DS (NDS) fetal liver tissues as described in “Fetal liver tissue.” Median gene expression levels were determined by quantitative real-time RT-PCR and normalized to GAPDH expression. (B) Uracil accumulation in DNA. DNA was isolated from DS and non-DS fetal tissue and analyzed for uracil incorporation as described in “Uracil detection.” Relative uracil levels were determined and median values are presented. (C) DNA polymerase β expression. RNA was isolated and reverse transcribed from DS and non-DS fetal liver tissue as described in “Base excision repair activity.” Median gene expression levels were determined by quantitative real-time RT-PCR and normalized to GAPDH expression. (D) Base excision repair activity. Nuclear proteins were isolated, and BER capacity was determined as described in “Methods.” BER capacity is calculated as the percentage of probe repaired (16-mer/30-mer), and median values are presented. (E) F2 isoprostane detection. F2 isoprostanes were measured by gas chromatography–mass spectrometry in fetal liver tissues as described in “Fetal liver tissue.” Median values are presented.
