研究開発・学会発表

 

Introduction
Some embryos show only one pronucleus (1PN) after IVF and ICSI. Possible causes are parthenogenetic activation, male and female pronuclear fusion, and asynchronous pronuclear appearance. The chromosome of parthenogenetic activated embryos will be haploid (n), and the chromosome of other 1PN embryos might be diploid (2n). In this study, we analyzed the chromosomes of blastocysts (BLs) that developed from 1PN embryos after IVF and ICSI.

Material and Methods
　The percentages of 1PN embryos in 6972 ova after IVF (IVF 1PN) and in 2739 ova after ICSI (ICSI 1PN) were examined.  The percentages of BLs formed from these 1PN embryos were also investigated. The chromosomes of 41 BLs developed from IVF 1PN and 11 BLs developed from ICSI 1PN were analyzed by fluorescence in situ hybridization (FISH). The probes used in FISH were CEP 18/ CEP X/ CEP Y (Vysis Co.). Chromosomal abnormalities in IVF 1PN and ICSI 1PN were compared. The percentages of embryos with Y chromosome in IVF 1PN and ICSI 1PN were also determined.
 
Results
The percentage of ICSI 1PN was 7.3%, which was significantly higher than that of IVF 1PN (4.7%) (P<0.001). The percentage of BLs formed from IVF 1PN was 33.5%, which was significantly higher than that formed from ICSI 1PN (16.0%). The results of chromosome analysis showed that 84.6%, 2.6%, 7.7% and 5.1% of the BLs derived from IVF 1PN and 45.5%, 9.1%, 27.3% and 18.2% of the BLs derived from ICSI 1PN were 2n, n, mosaic of n and 2n, and chaotic mosaic BLs, respectively. The ratio of 2n BLs derived from IVF was significantly higher than that of ICSI 1PN (P<0.05). Y chromosome was detected in 54.8% of 2n BLs derived from IVF 1PN and in 40.0% of 2n BLs derived from ICSI 1PN. No 1PN BLs with chromosomal abnormality had Y chromosome.
All of the BLs derived from IVF 1PN that had developed to more than Grade 3 on day5 were 2n. Of the BLs derived from IVF 1PN that were less than Grade 3 on day 5, 66.7% were 2n and 33.3% showed chromosomal abnormality. The percentage of 2n BLs that were more than Grade 3 was significantly higher than that of 2n BLs that were less than Grade 3 (P<0.01). On the other hand, in the BLs derived from ICSI 1PN, the percentages of 2n BLs in these two groups were not significantly different.

Conclusions
　ICSI 1PN had a low blastocyst formation rate compared with that of IVF 1PN, and ICSI 1PN had a high rate of chromosomal abnormality compared with that of IVF 1PN. The mechanism of ICSI 1PN formation might terefore be different from that of IVF 1PN.
Y chromosome was not detected in the BLs derived from IVF 1PN and ICSI 1PN that had chromosomal abnormality. Therefore, these BLs were thought to be grown through parthenogenesis activation.  The results also suggest that parthenogenesis activation occurs more frequently in ICSI 1PN than in IVF 1PN. All of the BLs derived from IVF 1PN that had grown to more than Grade 3 on day5 were 2n BLs. These BLs might therefore be useful for embryo transfer. On the other hand, only 50% of the BLs derived from ICSI 1PN that had grown to more than Grade 3 on day5 were 2n BLs. We should therefore be more careful with the clinical use of BLs derived from ICSI 1PN.