Double-edged sword: effects of human sperm reactive oxygen species on embryo development in IVF cycles
Authors: Liu Jin , Zhu Keheng , Xu Shiyan , Tu Wenjiao , Lin Xiaotan , Su Youpeng , Huang Rong , Deng Yuao , Liu Yu 
Published: January 2023

Abstract
Background
The exact role of sperm reactive oxygen species (ROS) in early embryo development has yet to be fully identified, and most of existing research did not differentiate female infertility factors, ignoring the importance of oocyte quality in embryo development and the large differences in oocyte quality in women with infertility of different etiologies. And there has been no relevant report on whether different types of sperm ROS have distinct effects on embryo development. This study aimed to study the impact of selected sperm ROS, namely, sperm mitochondrial ROS (mROS) and hydrogen peroxide, on human embryo development after conventional in vitro fertilization (IVF) cycles in patients with normo-ovulatory infertility vs. anovulatory infertility.

Methods
This was a prospective investigation including 393 couples underwent IVF cycles, among whom 90 patients had anovulatory infertility and 303 patients had normo-ovulatory infertility in a public university-affiliated in vitro fertilization center. Sperm mROS and hydrogen peroxide testing were performed by flow cytometry and analyzed for their relationship with embryo development indices on days 1–6 after IVF. Multivariate logistic regression analysis was used to control for female potential confounders. The nonlinear effects of sperm ROS on embryo development were analyzed by the Restricted cubic spline (RCS) method.

Results
1. Multivariate linear logistic regression analysis showed that high proportion of mROS positive sperm improved the 2PN rate (OR = 1.325, 95% CI: 1.103–1.595), day 3 embryo utilization rate (OR = 1.362, 95% CI: 1.151–1.614) and good-quality day 3 embryo rate (OR = 1.391, 95% CI: 1.089–1.783) in patients with anovulatory infertility. High percentage of sperm mROS and hydrogen peroxide had adverse effects on cleavage-stage embryo and blastocyst development in patients with normo-ovulatory infertility. 2. For patients with polycystic ovarian syndrome (PCOS) anovulatory infertility, there were significant distinct effects on embryo development indices between sperm mROS and hydrogen peroxide, and the increased rate of sperm mROS improved the good-quality day 3 embryo rate (OR = 1.435, 95% CI: 1.045–1.981); however, high percentage of sperm hydrogen peroxide reduced the blastocyst utilization rate (OR = 0.555, 95% CI: 0.353–0.864) and the good-quality blastocyst rate (OR = 0.461, 95% CI: 0.292–0.718). 3. Multivariate RCS analysis revealed that sperm ROS had a nonlinear (such as a parabolic curve) effect on embryo development in patients with anovulatory infertility (P < 0.05), and either greatly increased or greatly decreased affected cleavage-stage embryo and blastocyst development. The effects of sperm ROS in patients with normo-ovulatory infertility were both linear and nonlinear.

Conclusions
These findings indicate that contrary effects of sperm mROS on embryo development depending on whether patients treated with IVF cycles had normal ovulation. Regardless of whether the patients ovulated normally, increased sperm hydrogen peroxide rate damaged blastocyst development. It is necessary to evaluate male sperm ROS levels and the female ovulatory state to determine an individualized intervention plan before starting cycles, as this may be beneficial for infertile couples.