| Objective: Through the expression of zinc transporter family(Zinctranspoter,Zn T)in different tissues and organs of mice,the dominant protein members were screened,and the expression of zinc transporter in tissues and spermatozoa was explored.Through the construction of obese mouse model and detection of clinical asthenospermia samples,to explore the influencing factors of regulating zinc transporter expression,and finally provide new ideas for the diagnosis and treatment of male infertility.Methods: First of all,we used normal C57 mice as experimental subjects to detect the m RNA expression level of Zn T family members in different organs and tissues by realtime PCR technique,and selected Zn T6 with high expression level and difference in epididymal head and caudal epididymal tissues as the target protein for further study.At the same time,immunofluorescence and immunoblotting techniques were used to detect the distribution and protein expression of Zn T6 protein in mouse epididymal head,tail tissue,epididymal head sperm and tail sperm.Secondly,we established a model of obese mice induced by high-fat diet,and detected the fluorescence and protein expression of Zn T6 protein in epididymal head,tail tissue,epididymal head sperm and tail sperm of obese mice by immunofluorescence and immunoblotting.Finally,the clinical semen samples were collected and divided into normal group and obesity-induced asthenospermia group,and the protein differences of Zn T6 in sperm and seminal plasma were detected respectively.Results: 1.Ten members of Zn T family were expressed in various organs and tissues of normal mice,among which Zn T6 was highly expressed in the head and tail of mouse epididymis,and gradually increased from head to tail,which was used as a protein for further study.2.The distribution of Zn T6 protein in the epithelial cells of the head and tail of the epididymis was mainly located in the top cytoplasm of the epithelial cells,and the fluorescence increased gradually from the head to the tail.In the head and tail of epididymis,the spermatozoa were mainly located in the neck and middle segment of the epididymis,and the fluorescence increased gradually.3.In the obese mouse model established by high-fat diet,the body weight,blood lipid,liver and testicular morphology of obese mice showed that the obese mouse model was successfully established.At the same time,there was no significant difference in the distribution of Zn T6 protein between obese mice and normal control mice,but the expression of Zn T6 in obese mice was lower than that in normal control mice in both epididymal head and tail of epididymis.The results of protein immunoblotting technique are consistent with those of indirect fluorescence staining.The fluorescence intensity of Zn T6 on the neck and middle segment of epididymal tail spermatozoa in obese mice was lower than that in normal control mice.4.The clinical semen samples were divided into obesity-induced asthenospermia group and normal group.The distribution of Zn T6 in the two groups was similar to that in mouse sperm,mainly in the neck and middle segment of human sperm tail,and the fluorescence in asthenospermia was weaker than that in normal sperm.Western blotting showed that the expression of Zn T6 protein in seminal plasma of asthenospermia was decreased.Conclusions: 1.The expression of Zn T members is different,among which the expression of Zn T6 in epididymis is higher,and there is a significant difference in the expression of Zn T6 in the head and tail of epididymis.Zn T6 is one of the main carriers of zinc transport in epididymis.2.It is clear that obesity can cause changes in the expression of Zn T6 protein in animal experiments,and there is a correlation between Zn T6 protein and obesity.3.It is clear that there is a correlation between the expression of Zn T6 protein and obesity-induced asthenospermia in patients with obesity-induced asthenospermia.4.There is a negative correlation between Zn T6 zinc transporter and asthenospermia caused by obesity. |
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