Telomere DNA deficiency is associated with development of human embryonic aneuploidy

Abstract

Aneuploidy represents the most prevalent form of genetic instability found in human embryos and is the leading genetic cause of miscarriage and developmental delay in newborns. Telomere DNA deficiency is associated with genomic instability in somatic cells and may play a role in development of aneuploidy commonly found in female germ cells and human embryos. To test this hypothesis, we developed a method capable of quantifying telomere DNA in parallel with 24-chromosome aneuploidy screening from the same oocyte or embryo biopsy. Aneuploid human polar bodies possessed significantly less telomere DNA than euploid polar bodies from sibling oocytes (-3.07 fold, P = 0.016). This indicates that oocytes with telomere DNA deficiency are prone to aneuploidy development during meiosis. Aneuploid embryonic cells also possessed significantly less telomere DNA than euploid embryonic cells at the cleavage stage (-2.60 fold, P = 0.002) but not at the blastocyst stage (-1.18 fold, P = 0.340). The lack of a significant difference at the blastocyst stage was found to be due to telomere DNA normalization between the cleavage and blastocyst stage of embryogenesis and not due to developmental arrest of embryos with short telomeres. Heterogeneity in telomere length within oocytes may provide an opportunity to improve the treatment of infertility through telomere-based selection of oocytes and embryos with reproductive competence.

Conflict of interest statement

RTS is a founding partner of the Reproductive Medicine Associates of New Jersey (RMANJ), and all the authors are employed by RMANJ.

Figures

Figure 1. Valdiation of a novel assay for telomere DNA relative quantitation from limited amounts of starting material.

(A) The telomere DNA assay amplification product produced a single peak upon dissociation curve analysis demonstrating specificity on purified genomic (g) DNA from a large quantity of cells, and from whole genome amplified (WGA) DNA from picogram quantities of purified gDNA. (B) Quantities of telomere DNA in various cancer cell lines relative to MCF-7 cells either reported in the literature or determined with the assay developed in this study. In each case, the Pearson correlation coefficient (R2) of the relative quantitation profiles are shown and illustrate the maintenance of relative abundance after characterization of picogram quantities of starting material.

Figure 2. Example results of SNP microarray based 24 chromosome aneuploidy screening in human oocytes and preimplantation embryos that were also characterized for telomere DNA content.

(A) SNP microarray based chromosome specific copy number graphs of euploid and aneuploid sibling polar bodies from polar body specific patients 2, 5, 8, and 9 (Table 1). (B) SNP microarray based copy number graphs of euploid and aneuploid sibling blastomeres from blastomere specific patients 5, 6, 7, and 9 (Table 2). (C) Photographs of cleavage stage sibling embryos from the 9 blastomere specific patients analyzed in the present study (Table 2). (D) SNP microarray based copy number graphs of euploid and aneuploid sibling blastocyst stage embryos from trophectoderm specific patients 1, 7, 8, and 9 (Table 3).

Figure 3. Mean quantities (±SEM) of telomere DNA found in aneuploid relative to euploid polar bodies, blastomeres, and trophectoderm biopsies.

P-values from paired analysis of sibling samples within each patient are shown and illustrate the significant decrease in relative quantity in aneuploid polar bodies and blastomeres but not trophectoderm.

Figure 4. Telomere DNA in aneuploid blastocysts.

(A) Diagram illustrating two potential mechanisms by which aneuploid blastocysts could obtain normalized quantities of telomere DNA. (B) Results of testing the potential for “correction” to explain normalized quantities of telomere DNA in aneuploid blastocysts. A significant increase (P<0.05) was found in telomere DNA quantity in trophectoderm from blastocysts relative to the 1st polar body from the same oocyte. A significant increase was not found in blastomeres from cleavage stage embryos. This suggests that telomere DNA is reset between the cleavage and blastocyst stage of embryogenesis in humans. (C) Results of testing the potential for “selection” to explain normalized quantities of telomere DNA in aneuploid blastocysts. Significance was not reached when comparing sibling arrested and developmentally competent embryos (P = 0.29). This suggests that embryos with short telomeres aren't more prone to developmental arrest between the cleavage and blastocyst stage of embryogenesis in humans.