| Objective:In this study,ATP,hyaluronidase and ATP + hyaluronidase were added to the seminal plasma of asthenospermia respectively to compare the effects of exogenous addition of enzyme on semen quality.The changes of mitochondria,acrosome and other structures in sperm cells were observed under transmission electron microscope.Combined with the previous research of the research group,the effects of exogenous addition of ATP and hyaluronidase on semen quality were discussed.Enzyme linked immunosorbent assay(ELISA)was used to detect the expression of AMP dependent protein kinase(AMPK)in the seminal plasma of asthenospermia,so as to explore the effect of energy changes in the seminal plasma of asthenospermia on sperm,and to find a new way to improve the quality of semen of asthenospermia.Methods:(1)Methods: 103 samples were collected from the outpatients of Andrology Department of reproductive center of Affiliated Hospital of Youjiang Medical University for Nationalities from November 2019 to may 2020.The semen quality was detected by sac semen quality analyzer.Each sample was detected at least 3 times,including 42 normal semen and 61 asthenospermia They were divided into three groups,100 UL in each group was placed in the EP tube,and ATP,hyaluronidase and ATP + hyaluronidase(ATP: hyaluronidase= 1:1)were added at the ratio of 1:1 respectively.The semen quality was analyzed again.(2)The ultrastructural changes of mitochondria and acrosome were observed by transmission electron microscope.(3)According to the inclusion criteria,178 cases of semen were collected from the andrology clinic of reproductive center of Affiliated Hospital of Youjiang Medical University for Nationalities from November 2019 to August 2020,including 89 cases of asthenospermia and 89 cases of normal semen.The AMPK content in seminal plasma was determined by ELISA.(2)Results:(1)Sperm forward movement(PR): in asthenospermia group and normal group,after exogenous addition of ATP,hyaluronidase,ATP + hyaluronidase,sperm forward movement(PR)was weakened,P < 0.05,the difference was statistically significant.Compared with ATP + hyaluronidase group,the sperm forward motility(PR)decreased after exogenous ATP addition,P < 0.05,with statistical significance;compared with ATP +hyaluronidase group,the sperm forward motility(PR)decreased after exogenous ATP addition,P < 0.05,with statistical significance;compared with ATP + hyaluronidase group,the sperm forward motility(PR)decreased after exogenous ATP addition,P < 0.05,with statistical significance;compared with ATP + hyaluronidase group,the sperm forward motility(PR)decreased after exogenous ATP addition,with statistical significance There was no significant difference in forward motor(PR)between the two groups(P > 0.05).(2)Total sperm activity(PR + NP): in asthenospermia group and normal group,after exogenous addition of ATP,hyaluronidase,ATP + hyaluronidase,the total sperm activity(PR + NP)decreased,P < 0.05,the difference was statistically significant.Compared with the ATP +hyaluronidase group,the total sperm activity(PR + NP)decreased after exogenous addition of ATP,P < 0.05,the difference was statistically significant;compared with the ATP +hyaluronidase group,the total sperm activity(PR + NP)decreased after exogenous addition of ATP alone,P < 0.05,the difference was statistically significant;compared with the ATP +hyaluronidase group,the total sperm activity(PR + NP)decreased after exogenous addition of ATP,P < 0.05,the difference was statistically significant;compared with the ATP +hyaluronidase group,the total sperm activity(PR + NP)decreased There was no significant difference in PR + NP(P > 0.05).(3)Transmission electron microscope: the results showed that after adding ATP,hyaluronidase and ATP + hyaluronidase to asthenospermia and normal semen,the ultrastructure changes of sperm were as follows: the sperm was edematous in varying degrees,the cell membrane area was damaged and dissolved in varying degrees,the organelles were free,the whole body was disintegrated,and the head was irregular.The nuclei were locally sunken,and the chromatin was homogeneous,in which nuclear vesicles and nuclear membrane were visible.The acrosome is small,the structure is fuzzy,and the dense area disappears.The results showed that the structure of sperm tail swelled,mitochondria swelled,the matrix in the membrane became thin,cristae reduced,disappeared,and the matrix overflowed;the thickness of peripheral dense fibers was uneven,and the structure was fuzzy;the number of double microtubules decreased,some double microtubules disappeared,and the structure of sub microtubules disappeared;the structure of double microtubules in the central microtubule was fuzzy or disappeared.(4)Results: the content of AMPK in seminal plasma of asthenospermia was significantly higher than that of normal seminal plasma(P < 0.05).Conclusion:(1)In asthenospermia,exogenous addition of ATP,hyaluronidase and ATP+ hyaluronidase decreased sperm forward motility(PR)and total sperm activity(PR + NP);(2)In asthenospermia,exogenous addition of ATP decreased sperm forward motility(PR)and total sperm activity(PR + NP),and it was weaker than exogenous addition of hyaluronidase and exogenous addition of ATP + hyaluronidase In asthenospermia;(3)exogenous addition of ATP,exogenous addition of hyaluronidase,exogenous addition of ATP + hyaluronidase,sperm forward movement(PR)and sperm total activity(PR + NP)decreased,which may be related to sperm and sperm ultrastructure changes and functional damage.(4)AMPK existed in seminal plasma of asthenospermia,and the content of AMPK in seminal plasma of asthenospermia was significantly higher than that of normal group. |
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