Intracytoplasmic sperm injection may increase the risk of imprinting defects

Abstract

In germ cells and the early embryo, the mammalian genome undergoes widespread epigenetic reprogramming. Animal studies suggest that this process is vulnerable to external factors. We report two children who were conceived by intracytoplasmic sperm injection (ICSI) and who developed Angelman syndrome. Molecular studies, including DNA methylation and microsatellite and quantitative Southern blot analysis, revealed a sporadic imprinting defect in both patients. We discuss the possibility that ICSI may interfere with the establishment of the maternal imprint in the oocyte or pre-embryo.

Figure 1

Molecular analysis. A, DNA was digested with XbaI and NotI and was hybridized with a probe for SNRPN. A normal control subject (C) and the parents of patient 1 (M1, mother; F1, father) have a methylated maternal band (top of lane) and an unmethylated paternal band (bottom of lane) of equal intensity. Patients with AS and Prader-Willi syndrome (PWS) show only a single unmethylated or methylated band, respectively. Patient 2 shows an unmethylated band only. Patient 1 shows a very faint methylated band, in addition to the unmethylated band (increased intensity). Similar results were obtained by methylation-specific PCR (data not shown). B, IC deletion analysis. DNA was digested with EcoRV and was hybridized with a probe for the AS element of the IC (IC1/3; upper band) (Schumacher et al. 1998) and a probe for the RB1 gene on chromosome 13 (lower band). The relative hybridization intensities, as determined by scanning densitometry, are given below each lane. A deletion patient (DEL) has a reduced signal ratio. The two patients described in the present report (1 and 2) have a ratio similar to that of a normal control subject (i.e., no IC deletion). Similar results were obtained in a second, independent experiment (data not shown).