Effect of Lactoferrin on Sperm Capacitation in Dorper Sheep

Fei Mao^1,2Junliang Zhao^3,4Yifan Zhao¹Lu Chen^1,2Bin Hou^1,2Zhenxu Xu^1,2Feifei Zhao¹Jie Xing^1,2Yali Wang^1,2Xuedong Ding^1,2Puguo Hao^1,2Caiyun Wang^1*

• ¹College of Veterinary Medicine, Inner Mongolia Agricultural University, Hohhot, China
• ²Ordos Vocational College of Ecological Environment, Ordos 017100, China., Ordos, China
• ³Hetao College, Bayan Nur, China
• ⁴Inner Mongolia Key Laboratory of Vector- borne Zoonotic Infectious Disease, Department of Agriculture, College of Hetao, Bayan Nur city, Inner Mongolia Autonomous Region, China., Bayan Nur city, China

To explore the application of lactoferrin (LTF) in sperm in vitro fertilization, this study focused on investigating its effect on sheep sperm capacitation; first, proteomic mass spectrometry was used to screen for differentially expressed proteins between fresh sperm and capacitated sperm, then Dorper ram sperm were cultured in a capacitation medium, from which proteins and RNA were extracted to verify the expression of LTF before and after capacitation, and immunofluorescence localization technology was employed to confirm the presence of LTF in sperm, and finally, different concentrations of LTF (0 μg/mL, 0.1 μg/mL, 10 μg/mL, 100 μg/mL, and 500 μg/mL) were added to the capacitation medium, with sperm capacitation rate and motility evaluated at 0 h, 2 h, and 4 h of culture, respectively. The results showed that there were 203 differentially expressed proteins in sheep sperm before and after capacitation, among which 27 proteins were upregulated and 176 proteins were downregulated after capacitation; LTF was expressed both before and after sperm capacitation, with a significant difference in expression levels (p < 0.05), and it was mainly localized in the head and tail of capacitated sperm; in addition, although adding 100 μg/mL LTF to the capacitation medium reduced sperm motility, it promoted sperm capacitation. In conclusion, adding 100 μg/mL LTF to the capacitation medium can effectively promote sheep sperm capacitation while moderately reducing sperm motility, and this study is the first to clarify the expression dynamics and regulatory effects of LTF during sheep sperm capacitation, providing experimental evidence for optimizing the formula of capacitation medium in the sheep in vitro fertilization system.