O-197 Vertical transmission of maternal mitochondrial DNA via extracellular vesicles (EVs) modulates embryo bioenergetics during the periconceptional period
Authors: Vilella F , Bolumar D , Moncayo-Arlandi J , Gonzalez-Fernandez J , Ochando A , Moreno I , Marin C , Diez A , Simon C 
Published: June 2023

Abstract

Study question
Does maternal endometrial mitochondrial (mt) DNA cargo of EVs modulate embryo bioenergetics during embryo implantation?

Summary answer
We demonstrate the vertical transmission of maternal mtDNA within endometrial-derived EVs and their uptake by the trophoblast which reduces mitochondrial respiration and ATP production.

What is known already
The release and uptake of membrane-enclosed compartments with specific cargos, commonly known as EVs, represents a novel cell-to-cell communication mechanism in physiological and pathogenic conditions. EVs are generally classified into three populations based on their biogenetic pathways, composition, and physical characteristics: apoptotic bodies (ABs), microvesicles (MVs), and exosomes (EXOs). Among other contents, EVs contain single- and double-stranded DNA, with their relative abundance varying depending on the cell and vesicle type. The vertical transmission of EV-associated DNA has been proposed as a novel genetic material transfer mechanism that may impact genome evolution and tumorigenesis.

Study design, size, duration
Prospective observational multicenter analysis in which EVs were obtained from endometrial fluid from healthy donors aged 18–35 years (n = 10) during the receptive phase of their natural cycle and under hormonal replacement therapy in pre-receptive (P + 2), receptive (P + 5), post-receptive (P + 8) stages (n = 13).

Participants/materials, setting, methods
Endometrial EVs isolated using ultracentrifugation and classified according to `parameters obtained from electron microscopy, Western blotting, and size distribution analysis. DNA copy number identified using high throughput sequencing. EV-associated DNA tagged with 5-ethynyl-2’-deoxyuridine was followed by confocal imaging after co-incubation of EVs with murine embryos (n = 200). ATP levels assessed using the FLASC luciferase reporter system, and the Seahorse XFE96 extracellular flux analyzer used to measure embryo oxygen consumption rate (OCR) (n = 400).

Main results and the role of chance
The human endometrium secretes all three EV types - ABs, MVs, and EXOs - into the human endometrial fluid. Deep sequencing revealed that EVs encapsulated nuclear and mtDNA. When analyzing endometrial biopsies, we observed the reduced mtDNA content of endometrial cells and the activation of mitochondrial clearance mechanisms, which coincided with the time of embryo implantation together with specific enrichment in endometrial MVs secreted during the periconceptional period. EVs were internalized and DNA was transferred to the cytoplasm and nuclei of trophectoderm of murine embryos. We analyzed ATP concentrations in murine embryos and found a significant reduction in ATP levels following the coculture of embryos with a combination of all EVs types compared to control embryos cultured without endometrial EVs (p &lt; 0.001). Finally, we demonstrated a reduction in the OCR in embryos treated with endometrial EVs obtained during the receptive phase compared with the pre-receptive phase. In conclusion, maternal EVs modulate the bioenergetics of the preimplantation embryo by increasing the embryo's metabolic rate and oxygen consumption during the periconceptional period.

Limitations, reasons for caution
These results were obtained using a combination of a human endometrial model and a murine embryo model.

Wider implications of the findings
Our results suggest that the vertical transmission of maternal mtDNA encapsulated within EVs to the trophectoderm might energetically assist the preimplantation embryo through the implantation process.

Trial registration number
N/A