The effect of peritoneal fluid from patients with endometriosis on mitochondrial function and development of early mouse embryos

Abstract

Background: Peritoneal fluid (PF) from patients with endometriosis can inhibit early embryo development via probable functional changes of embryo mitochondria in the early stage of embryo development. The purpose of this study was to determine the effect of PF from patients with endometriosis on mitochondrial function and development of early mouse embryos.

Methodology/principal findings: PF was collected from patients with infertility and endometriosis, infertility due to tubal factors, and normal control subjects, and the level of NO was measured. Early murine embryos were then cultured with PF from normal control subjects, those with endometriosis, and with human tubal fluid (HTF), respectively. Cleavage and blastulation rates, mitochondrial DNA (mtDNA) copy numbers, adenosine triphosphate (ATP) level, and mitochondrial membrane potential (ΔΨm) of the different groups were compared. The NO level in the PF of patients with endometriosis was significantly greater than in those without endometriosis and control patients. The embryos cultures with PF from patients with endometriosis had a lower cleavage rate and blastulation rate, and higher ATP and ΔΨm level at the 2- and 4-cell stages. No significant difference was found in mtDNA copies among the 3 groups.

Conclusions/significance: PF from patients with endometriosis can inhibit early embryo development via probable functional changes of embryo mitochondria in the early stage of embryo development. Understanding the effects of PF on embryo development may assist in developing new methods of treatment for infertility.

Figure 1. Cleavage (A) and blastulation (B) rates at different concentrations of sodium nitroprusside.

^a Indicates a significant difference between the given group and HTF-control group.

Figure 2. Cleavage (A) and blastulation (B) rates of HTF-control group and groups cultured with peritoneal fluid from patients with endometriosis (PF-E) and those without endometriosis (PF-control).

^a Indicates a significant difference between the given group and HTF-control group. ^bIndicates a significant difference between the PF-control and PF-E groups.

Figure 3. ATP level (A), mtDNA copies (B), and ΔΨ_m(C) of HTF-control group and groups cultured with peritoneal fluid from patients with endometriosis (PF-E) and those without endometriosis (PF-control) at different phases.

^a Indicates a significant difference between the given group and HTF-control group. ^bIndicates a significant difference between the PF-control and PF-E groups.

Figure 4. JC-1 staining of different stage early embryos in the HTF-control, PF-control, and PF-E groups.

Rows, from top to bottom, correspond to 2-cell, 4-cell, morula, and blastocyst stages, respectively. Columns, from left to right, correspond to the HTF-control (A, D, G, and J), PF-control (B, E, H, and K), and PF-E (C, F, I, and L) groups, respectively.

Figure 5. Electron microscopy of 2-cell and 4-cell embryos in the HTF-control, PF-control, and PF-E groups.

A) 2-cell embryos in the HTF-control group. The mitochondria were round with few cristae (17,500×). B) 4-cell embryos in the HTF-control group. Several mitochondria with rich cristae were seen (24,000×). C) 2-cell embryos in the PF-control group. Most mitochondria were round with few cristae, and a few mitochondria had a transverse crest (24,000×). D) 4-cell embryos in the PF-control group. Mitochondria with transverse cristae were increased (24,000×). E) 2-cell embryos in the PF-E group. Mitochondria with lamellar transverse cristae were increased significantly (24,000×). F) 4-cell embryos in the PF-E group. Multiple mitochondria with rich transverse cristae were seen (24,000×). (The single arrow indicates mitochondria rich in transverse cristae, and the double arrow indicates the Golgi apparatus.)