# Oleic Acid Improves Goat Sperm Quality by Enhancing the MBOAT2/ACSL3 Pathway to Attenuate Ferroptosis

## TL;DR

Oleic acid improves goat sperm quality during cold storage by reducing cell death called ferroptosis and protecting against oxidative damage.

## Contribution

This study reveals a novel mechanism by which oleic acid protects sperm through the MBOAT2/ACSL3 pathway, offering new strategies for preserving sperm quality.

## Key findings

- 1 mM oleic acid significantly improved sperm motility, membrane integrity, and mitochondrial activity during cold storage.
- Oleic acid reduced oxidative stress markers like ROS, MDA, and lipid peroxidation in stored sperm.
- Oleic acid suppressed iron accumulation and inhibited ferroptosis in a model of RSL3-induced damage.

## Abstract

Exposure to low temperatures during goat sperm preservation induces oxidative damage and triggers ferroptosis, a regulated form of cell death, thereby impairing sperm quality and fertility. This study investigated whether oleic acid (OA), a naturally occurring monounsaturated fatty acid, can mitigate these effects during chilled storage. Supplementation of semen extenders with varying concentrations of OA revealed that 1 mM OA significantly enhanced sperm motility, membrane and acrosome integrity, and mitochondrial activity after preservation at 4 °C. Also, OA reduced oxidative stress markers—including reactive oxygen species (ROS), malondialdehyde (MDA), and lipid peroxidation—and modulated the expression of ferroptosis-associated proteins (ACSL3, ACSL4, GPX4, and SLC7A11). Moreover, OA suppressed iron accumulation and alleviated ferroptosis in an RSL3-induced damage model. These findings suggest that OA exerts potent cytoprotective effects by modulating lipid metabolism and inhibiting ferroptosis, rather than functioning solely as an antioxidant. This study provides new insights into improving sperm chilled protocols and may have broader implications for assisted reproductive technologies in livestock breeding and genetic resource conservation.

Exposure to low temperatures during goat sperm preservation induces oxidative damage and triggers ferroptosis, a regulated form of cell death, thereby impairing sperm quality and fertility. This study aims to elucidate the protective mechanism of oleic acid (OA) on goat sperm during cryopreservation through the regulation of ferroptosis pathways. Different concentrations of OA (0.125 mM, 0.25 mM, 5 mM, 1 mM, and 2 mM) were added to a yolk-containing extender. Following chilled storage at 4 °C, sperm motility parameters, membrane and acrosome integrity, mitochondrial membrane potential, and other physiological parameters were systematically evaluated. Oxidative damage was analyzed by quantifying ferrous ion (Fe2+), malondialdehyde (MDA), lipid peroxidation (LPO), and reactive oxygen species (ROS). Western blotting was employed to assess the expression of key regulators of the ferroptosis pathway, including ACSL3, ACSL4, GPX4 and SLC7A11. The results showed that sperm motility in the 1 mM OA treatment group was significantly higher than that in the control group (p < 0.05), with improved membrane and acrosome integrity, as well as significant mitigation of mitochondrial damage. Moreover, OA effectively suppressed ferroptosis by reducing Fe2+ accumulation and inhibiting MDA and LPO production. Mechanistically, OA may regulate membrane phospholipid composition by competitively incorporating monounsaturated phosphatidyl ethanolamine, thereby reducing ACSL4-mediated lipid peroxidation. These results suggest that oleic acid improves sperm motility by inhibiting ferroptosis during liquid storage at 4 °C.