Sperm apoptosis in nonpregnant luteal phase sera after in vitro fertilization as assessed by comparative genomic hybridization

Toxicity in serum has been reported in cases of recurrent spontaneous abortions and endometriosis. The null hypothesis was that serum toxicity was not involved in failed pregnancies after in vitro fertilization procedures. The objective was to expose donor sperm to pregnant versus nonpregnant patient sera and analyze for sperm DNA damaging effects using a novel comparative genomic hybridization method. Luteal phase sera (N = 21 cases) were drawn one week after embryo transfer. Colloid-washed donor sperm were incubated (48 h, 37°C, 5% CO₂ in air) in 0% or 50% sera. Single-stranded DNA (ssDNA) of control sperm were stained in Hoechst 33342 and hybridized to Sybr Gold-stained ssDNA of sera-treated sperm. Image analyses were performed and fluorescent intensities analyzed. Nonpregnant patient sera (57% of cases) were associated with DNA fragmentation (64.4 ± 8.8 pixels; mean ± S.E.M.) when compared with pregnant patient sera (106.3 ± 8.4 pixels). There were no differences in the sera of biochemical (108.2 ± 15.3) versus clinical pregnancy cases (105.3 ± 11.4). The results suggest that nonpregnant patient sera contained factor(s) that cause DNA fragmentation leading to pregnancy losses.