益母草碱对绵羊精液低温保存效果的影响

doi:10.16431/j.cnki.1671-7236.2023.07.020

摘要/Abstract

摘要： 【目的】探究益母草碱对绵羊精液低温保存效果的影响，为实际生产中制备适宜的绵羊精液低温保存稀释液提供理论依据。【方法】采用假阴道法采集3只多浪羊公羊精液，在精液低温保存稀释液中分别添加0（对照）、20、40、60、80和100 μmol/mL益母草碱，检测精液低温保存过程中精子活率、顶体完整率和质膜完整率等精液品质指标以及总抗氧化能力（T-AOC）、谷胱甘肽过氧化物酶（GSH-Px）、超氧化物歧化酶（SOD）和过氧化氢酶（CAT）活性及丙二醛（MDA）含量等抗氧化指标，筛选精液保存过程中最适益母草碱浓度；添加最适浓度益母草碱后，利用实时荧光定量PCR法检测精液低温保存过程中TNF受体超家族成员6（Fas）、胱天蛋白酶3（Caspase-3）、B淋巴细胞瘤-2（Bcl-2）和B淋巴细胞瘤-2关联X蛋白（Bax）等相关凋亡基因表达量；采用子宫颈口内输精方式完成两次输精，统计母羊受胎率。【结果】与对照组相比，低温保存48 h，40 μmol/mL益母草碱组精子活率、质膜完整率、顶体完整率及T-AOC水平、SOD、GSH-Px、CAT活性均显著提高（P<0.05），MDA含量显著降低（P<0.05）；低温保存144 h，40 μmol/mL益母草碱组各项指标与其他益母草碱添加组相比差异显著（P<0.05）。实时荧光定量PCR结果显示，与对照组相比，40 μmol/mL益母草碱组Fas、Caspase-3和Bax基因表达量显著降低（P<0.05），Bcl-2基因表达量显著升高（P<0.05）。添加40 μmol/mL益母草碱的低温保存精液输精后，母羊受胎率为75%，达到鲜精阴道输精的平均水平。【结论】低温保存精液稀释液中添加40 μmol/mL益母草碱后可明显改善低温保存多浪羊精液品质，提升其抗氧化能力，同时起到抗凋亡作用；含40 μmol/mL益母草碱低温保存72 h的精液输精后母羊受胎率为75%，达到了鲜精阴道输精平均水平。

关键词: 益母草碱; 低温保存; 精液品质; 抗氧化能力; 受胎率

Abstract: 【Objective】 This study was aimed to explore the effect of leonurine on the cryopreservation effect of sheep semen, and provide theoretical basis for the preparation of appropriate diluent for cryopreservation of sheep semen in practical production.【Method】 Semen of three Duolang sheep were collected by pseudoginosis method.0 (control), 20, 40, 60, 80 and 100 μmol/mL leonurine was added to semen cryopreserved diluent, respectively.Semen quality indexes such as sperm viability, acrosome integrity and plasma membrane integrity, total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) activities, and malondialdehyde (MDA) content were detected during semen cryoppreservation.The optimal concentration of leonuri during semen preservation was screened.After the optimal concentration of leonuri was added, the expression of TNF receptor superfamily member 6 (Fas), Caspase-3, B lymphoblastoma-2 (Bcl-2) and B lymphoblastoma-2-associated X protein (Bax) during cryopreserved semen were detected by Real-time quantitative PCR.The conception rate of ewe was calculated using the method of two intraventricular insemination.【Result】 Compared with control group, the sperm viability, plasma membrane integrity, acrosome integrity, T-AOC level, SOD, GSH-Px and CAT activities of 40 μmol/mL leonurine group were significantly increased (P<0.05), while the content of MDA was significantly decreased (P<0.05). Cryopreservation for 144 h, the indexes of 40 μmol/mL leonurine group were significantly different from those of other leonurine groups (P<0.05).Real-time quantitative PCR results showed that compared with control group, the expressions of Fas, Caspase-3 and Bax genes in 40 μmol/mL leonurine group were significantly decreased (P <0.05), while the expression of Bcl-2 gene was significantly increased (P<0.05).【Conclusion】 In this study, adding 40 μmol/mL leonurine to cryopreservation semen diluent could significantly improve the quality of cryopreservation of Duolang sheep semen, enhance the antioxidant capacity, and played an anti-apoptotic role.The conception rate of ewes with 40 μmol/mL leonurine cryopreservation for 72 h was 75%, which reached the average level of vaginal transfusion of fresh semen.

Key words: leonurine; cryopreservation; semen quality; antioxidant capacity; conception rate

中图分类号:

S814.3

李智星, 卿育洪, 李志远, 曹少奇, 高新梅, 曾维斌, 王静, 胡广东. 益母草碱对绵羊精液低温保存效果的影响[J]. 中国畜牧兽医, 2023, 50(7): 2801-2810.

LI Zhixing, QING Yuhong, LI Zhiyuan, CAO Shaoqi, GAO Xinmei, ZENG Weibin, WANG Jing, HU Guangdong. Effects of Leonurine on Cryopreservation of Sheep Semen[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(7): 2801-2810.

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