Format

Send to

Choose Destination
Hum Reprod. 2013 Oct;28(10):2652-60. doi: 10.1093/humrep/det314. Epub 2013 Jul 25.

Mitochondrial DNA content in embryo culture medium is significantly associated with human embryo fragmentation.

Author information

1
Center of Physiopathology of Human Reproduction, Unit of Obstetrics and Gynecology, IRCSS A.O.U. San Martino IST.

Abstract

STUDY QUESTION:

Is the amount of cell-free DNA released by human embryos into culture medium correlated with embryo morphological features?

SUMMARY ANSWER:

The mitochondrial DNA (mtDNA) content of culture medium is significantly associated with the fragmentation rate on Days 2 and 3 of embryo development, whether the oocyte came from women ≤ 35 or >35 years old.

WHAT IS KNOWN ALREADY:

Cellular fragmentation is often utilized as one of the morphological parameters for embryo quality assessment. The amount of cellular fragments is considered to be an important morphological parameter for embryo implantation potential. It has been hypothesized that fragments are apoptotic bodies or anuclear cytoplasmatic pieces of blastomeres, although no definitive conclusion has been drawn about their pathogenesis.

STUDY DESIGN, SIZE, DURATION:

Human fertilized oocytes were individually cultured from Day 1 to Days 2 and 3. A total of 800 samples (166 spent media from Day 2 and 634 from Day 3) were enrolled into the present study.

PARTICIPANTS/MATERIALS, SETTING, METHODS:

Double-stranded DNA (dsDNA) was quantified in 800 spent embryo culture media by Pico Green dye fluorescence assay. After DNA purification, genomic DNA (gDNA) and mtDNA were profiled by specific quantitative PCR. Statistical analyses defined correlations among DNA contents, embryo morphology and maternal age.

MAIN RESULTS AND THE ROLE OF CHANCE:

Different independent tests confirmed the presence of DNA into embryo culture medium and, for the first time, we demonstrate that both gDNA and mtDNA are detectable in the secretome. The amount of DNA is larger in embryos with bad quality cleavage compared with high-grade embryos, suggesting that the DNA profile of culture medium is an objective marker for embryo quality assessment. In particular, DNA profiles are significantly associated with fragmentation feature (total dsDNA: P = 0.0010; mtDNA; P = 0.0247) and advanced maternal age.

LIMITATIONS, REASONS FOR CAUTION:

It is necessary to establish whether DNA profiling of spent embryo culture medium is a robust onsite test that can improve the prediction of blastulation, implantation and/or pregnancy rate.

WIDER IMPLICATIONS OF THE FINDINGS:

The approach we are proposing may provide a novel, non-invasive, objective tool for embryo quality grading. The correlation between a high mtDNA concentration and the fragmentation rate of embryos is suggestive that fragments are mainly anuclear cytoplasmatic debris arising during cleavage. Therefore, blastomere shaping as an early event during in vitro development may play a homeostatic role and be related to embryo competence.

STUDY FUNDING/COMPETING INTEREST:

This project was funded by Merck Serono (Grant for Fertility Innovation 2011). The sponsor had no role in study design, data collection, data analysis, data interpretation and writing of the paper. Authors declare no conflicts of interest.

TRIAL REGISTRATION NUMBER:

ClinicalTrials.gov Identifier: NCT01397136.

KEYWORDS:

IVF; cell-free DNA; embryo fragmentation; human embryos; mitochondrial DNA

PMID:
23887072
DOI:
10.1093/humrep/det314
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Silverchair Information Systems
Loading ...
Support Center