Study Of The Relationship Between TEKT3、SMCP And Idiopathic Asthenozoospermia

Background and Objective In recent years, the prevalence of infertility is increasing year by year, infertility is a global reproductive health problem, becoming a global medical and social problem of human health and development. According to the World Health Organization’s research, the proportion of couples in the world is to reach 10%-15%, in infertility patients, male infertile factors accounted for 50%. Therefore, to assess the relationship between male fertility and diagnosis and effective treatment of male infertility that has important clinical significance to male reproductive health. Because of the male semen quality problems in the male infertility have many causes, including spermatogenesis, sperm morphology abnormalities, low sperm motility, which low sperm activity is the most important reason, clinically known as asthenozoospermia(Asthenozoospermia, AZS). Sperm motility mainly refers to the ability of sperm forward movement. According to the World Health Organization(World Health Organization, WHO) the fifth edition of the semen analysis standards manual, in clinical semen analysis, sperm progressive motility(PR) is lower than 32%, sperm concentration is more than or equal to 15×106/ml is asthenozoospermia.Clinically, the common causes of low sperm motility are smoking, alcohol drinking, incomplete semen liquefaction, male varicocele, and autoimmune factors and so on. Because of the numerous and complicated etiology, the pathogenesis of the asthenozoospermia is not clear, therefore, it is of great significance to further study the occurrence and development of the mechanism of the treatment of the disease. The pathogenesis of asthenozoospermia mainly include: abnormal signal transduction pathway, abnormal mitochondrial energy metabolism, sperm tail structure and functional abnormalities. The mature mammalian sperm is mainly constituted by the head and tail, sperm tail also known as sperm flagella, mammalian spermatozoa before to the movement is mainly depend on the swing of the tail flagellum, outer dense fibers and fibrous sheath of the cytoskeleton is the swing of the sperm’s tail, the flagellum of main material basis, sperm morphology and structure of abnormal sperm activity and sperm motility disorder. The sperm tail mainly includes four parts: neck piece, middle piece, and principal piece, end piece, each part of sperm movement plays a crucial role. The middle piece of the sperm tail is complex, and the main structure is the spindle microtubules, the peripheral dense fibers and the mitochondrial sheath. The previous studies have found that more than 300 sperm related genes have been confirmed to be related to the incidence of male infertility. In order to further reveal the pathogenesis of asthenozoospermia. In this study, reverse transcriptase polymerase chain reaction(RT-PCR) and Western blot technique was used to detect the expression of TEKT3 and SMCP in the sperm of normal men and asthenozoospermia patients to preliminarily study the pathogenesis role of TKET3 and SMCP in idiopathic asthenozoospermia, in-depth understanding of asthenozoospermia and provide a theoretical basis for and to further elucidate the asthenozoospermia pathogenesis and clinical treatment provide experimental and theoretical basis.Materials and methods 1 ObjectiveA total of 100 participants were enrolled in our study: 50 infertile men with idiopathic asthenozoospermia, aged 20-40 years; and 50 age-matched controls with normal semen parameters. They were all recruited from June 2014 to October 2014 from the Department of Reproductive Medicine Center and Human Sperm Bank of the third affiliated hospital of the Zhengzhou University. All the infertile patients were incapable of impregnating their wives for at least 12 months, despite having unprotected sexual intercourse. Subjects collected semen before abstinence 3-7 days,analyze the semen according to the WHO the fifth semen laboratory analysis manual until semen liquefaction completely, and the computer-assisted semen analysis system of continuous testing three times for the same specimen. The information collected from study subjects included age, height, personal background, lifestyle, reproductive status, genetic risk factors and medical history. The exclusion criteria were as follows:(1) abnormal semen liquefaction,(2) semen samples of patients with pyospermia, varicocele, teratozoospermia, leukocytospermia, male reproductive system abnormalities or smoking history. All patients with asthenozoospermia were confirmed by at least three consecutive routine semen analyses on ejaculated semen. This study was approved by the ethics review board of the third affiliated hospital of the Zhengzhou University, Zhengzhou, China. All study subjects signed written informed consent before semen sample collection and personal information acquisition. 2 Methods 2.1 RT-PCR RT-PCR method was taken to determine the relative expression of TEKT3 m RNA and SMCP m RNA in the sperm of the two different groups. 2.2 Western blot The expression of sperm TEKT3 protein and SMCP protein in the two different groups were determined by Western-blot method.3 Statistics analysis Using SPSS17.0 statistical software for statistical analysis of the results data, and the results of measurement data in mean ± SD. For statistical analysis, two independent unpaired Student’s t-test was used to compare the difference in sperm TEKT3 and SMCP expression level between idiopathic asthenozoospermia patients and normal controls. Pearson correlation coefficient(r) was used to describe the correlation between the expression of TEKT3 and SMCP and the ability of sperm forward movement. A value of P<0.05 was considered statistically significant.Results 1 Expression levels of TEKT3 m RNA in the two groups’ sperm The relative expression levels of TEKT3 m RNA in the two groups were: 0.82 ± 0.03、0.60 ± 0.13, and the TEKT3 m RNA relative expression level was significant lower in the sperm of patients with idiopathic asthenozoospermia compared to the normal control male group(t= 9.61, P=0.001). 2 Expression levels of TEKT3 protein in the two groups’ sperm The relative expression levels of TEKT3 protein in the two groups were: 0.78 ± 0.03、0.57 ± 0.12, and the TEKT3 relative expression level of Tektin3 protein was obviously lower in the sperm of patients with idiopathic asthenozoospermia compared to the normal control male group(t= 10.09, P=0.001). 3 Expression levels of SMCP m RNA in the two groups’ sperm SMCP m RNA in the two groups of relative expression quantity respectively: 0.93 ± 0.44、0.27 ± 0.11, compared the two groups of sperm SMCP m RNA relative expression and found that idiopathic asthenozoospermia sperm SMCP m RNA expression levels were significantly decreased(t = 10.29, P = 0.001).SMCP protein in the sperm of the two groups relative expression were respectively: 1.02 ± 0.27、0.59 ± 0.23, compared the sperm relative expression of SMCP protein between the two groups, found that in idiopathic asthenozoospermia patients sperm SMCP protein expression level was significantly reduced.(t = 8.45, P = 0.002) 5 Correlation between the expression of TEKT3 and SMCP and the forward movement of sperm in the two groups Sperm motility and TEKT3 expression levels(r=718, P=0.001; r=0.729, P=0.001) and SMCP expression levels(r=0.705, P=0.013; P=0.001, r=0.716) were correlated. 4 Expression levels of SMCP protein in the two groups’ spermConclusion In the sperm of idiopathic asthenozoospermia patients, TEKT3, SMCP expression levels were positively correlated with sperm motility, the lower expression levels of TEKT3 and SMCP may related to the decrease of sperm progressive motility, which may be involved in the occurrence of asthenozoospermia.