HUMAN TROPHECTODERM BIOPSY AND SECRETION OF CHORIONIC-GONADOTROPIN

We have previously developed a technique of trophectoderm biopsy to obtain cells from human blastocysts for preimplantation genetic diagnosis. To determine whether this technique affects the subsequent development of the blastocyst, 45 manipulated blastocysts were observed from days 3 to 14 in culture, the amount of human chorionic gonadotrophin (HCG) secreted by each embryo was measured and these results were compared with those of 26 non-manipulated controls documented in a previous study. A slit was made in the zona pellucida opposite the inner cell mass in 18 of the 45 blastocysts. This increased the rate of hatching but the other morphological changes up to day 14 were similar to those seen in the non-manipulated controls. There was no difference in the mean cumulative HCG secretion by these zona-slit controls (149.8 +/- 45.7 mIU/ml) compared to the non-manipulated controls (146.2 +/- 23.7 mIU/ml). A slit was also made in the zona pellicida of the other 27 blastocysts. Approximately 12-18 h later, in 18 blastocysts a biopsy of the herniating trophectoderm cells (5-30) was performed. The rate of hatching and adherence to the culture dish was similar to the non-manipulated controls. The mean cumulative HCG secretion decreased significantly (57.5 +/- 16.2 mIU/ml, P < 0.01) after the biopsy procedure. However, if a small biopsy was performed (< 10 cells removed) the decrease in HCG secretion (87.6 +/- 24.8 mIU/ml) was not significant, whereas when a large biopsy was performed (> 10 cells), HCG levels fell to 19.9 +/- 9.1 mIU/ml. The amount of HCG secreted by the blastocyst after biospy was inversely proportional to the number of trophectoderm cells removed. Moreover, the removal of < 10 cells does not appear to impair the further development of the blastocyst. This biopsy method can provide sufficient cells to allow replicate analysis necessary for reliable genetic diagnosis.