| Artificial insemination technology is widely used in sheep breeding industry in order to effectively improve reproductive efficiency and facilitate the preservation of good seed and cross breeding.However,after semen is extracted,it is not conducive to survival in vitro,which greatly limits the utilization rate of semen.Therefore,improving the quality of semen preservation has become a key link in artificial insemination technology.At present,motherwort alkali has been proved to have antioxidant effect in myocardium and osteoblast cells.As an antioxidant,there are few reports on semen quality improvement of Motherwort.Objective: This research was conducted to comprehensively evaluate the effects of Leonuri on sheep semen preservation by measuring semen quality,antioxidant capacity,m RNA relative expression of related apoptotic genes and conception rate,so as to provide theoretical basis for the preparation of diluents more suitable for sheep semen preservation.Methods: In this research,20,40,60,80,100 μmol/m L of Leonuri were added into the diluent of dorang sheep semen at room temperature,low temperature and cryopreservation respectively.The semen quality indexes such as sperm viability,acrosome integrity and plasma membrane integrity,as well as antioxidant indexes such as T-AOC level,GSH-Px activity,SOD activity,CAT activity and MDA content were detected during the process of semen preservation at room temperature,low temperature and cryopreservation of Dorang sheep,so as to determine the optimal concentration of Leonuri.The m RNA relative expression levels of Fas,Caspase-3,Bax and Bcl-2 related apoptotic genes in the optimal concentration group were measured at room temperature,low temperature and cryopreservation compared with 0μmol/m L group,and the effects of Leonurus alkali on apoptotic genes in sheep semen preservation were explored;Finally,the conception rate of semen with the optimal concentration of Leonuri alkali was detected after semen cryopreservation and cryopreservation,and the effect of adding Leonuri alkali on the practical application of semen preservation was judged.Results: The results showed that the semen viability,acrosomal integrity and plasma membrane integrity of dorang sheep preserved at room temperature,low temperature and cryopreservation were significantly increased by adding Leonuri in preservation dilution compared with 0μmol/m L group(P<0.05);The activities of SOD,GSH-Px and CAT in semen were significantly increased,T-AOC level was increased and MDA content was decreased(P<0.05);Among them,in room temperature storage,the best preservation effect was achieved at the addition concentration of 60μmol/m L(P<0.05),and in low temperature storage,the best preservation effect was achieved at the addition concentration of 40μmol/m L(P<0.05).In cryopreservation,leonurus alkaloids at 60μmol/m L had the best preservation effect(P<0.05);In addition,compared with 0μmol/m L group,the m RNA relative expression levels of sperm-related apoptotic genes Fas,Caspase-3 and Bax in the optimal concentration of Leonurus alkali group were significantly decreased,while the relative expression levels of anti-apoptotic gene Bcl-2 were significantly increased(P < 0.05);At the same time,the sperm conception rate of 40μmol/m L group in cryopreservation was 75%,reaching the average level of fresh sperm vaginal sperm transfusion,and the sperm conception rate of 60μmol/m L group in cryopreservation was 60%,reaching the lowest level of fresh sperm vaginal sperm transfusion.Conclusion: In conclusion,this research confirmed that adding Leonuri alkali into semen dilution can effectively improve the preservation effect of room temperature,low temperature and cryo-preservation.The semen quality of dorang sheep stored at room temperature was significantly improved by adding 60μmol/m L Leonurus alkaloid,which enhanced its antioxidant capacity and had anti-apoptotic effect;Adding40 μmol/m L leonuri alkali during cryopreservation can significantly improve the semen quality of dorang sheep cryopreservation,enhance its antioxidant capacity,and have anti-apoptotic effect,and make its conception rate reach the average level of fresh semen vaginal semen delivery;The semen quality of dorang sheep cryopreservation was significantly improved by adding leonuri with 60 μmol/m L concentration during cryopreservation,and its antioxidant capacity and anti-apoptotic effect were improved.The conception rate of dorang sheep cryopreservation reached the lowest average level of vaginal semen delivery of fresh semen. |
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