Expression Of TDP-43 And SPANX Proteins In Sperm And Their Correlation With AID Pregnancy Outcome

At present,infertile couples account for about 15%of couples of childbearing age in the world,of which up to 50%are caused by male factors.The cause of male infertility is complex,and the number of infertility is increasing year by year.With the decline in environmental quality and lifestyle changes,male reproductive health problems have been seriously affected.The main factors leading to male infertility are endocrine diseases,abnormal gonad development,genetic factors,gonad or accessory gonadal infection,abnormal sperm DNA,unhealthy lifestyles such as smoking,staying up late,alcoholism and heavy metal pollution in the environment.However,about 40%of male infertility patients have unknown etiology and pathogenesis.At present,the choice of clinical diagnosis of male infertility and assisted reproductive technology treatment methods still depend on routine semen analysis,but only rely on routine semen analysis to make judgments on male fertility and guide and prognostic assessment of assisted reproductive technology..Because in the clinical diagnosis and treatment process,on the one hand,there are many male patients with normal semen parameters,but having difficulty in fertility;on the other hand,the semen indicators of some normal fertile men also have abnormal phenomena.Therefore,the analysis of routine parameters of semen cannot be used as a standard for judging male fertility.In order to make up for the lack of routine semen parameters in the clinical diagnosis of male infertility,some scholars try to find other detection methods to judge male fertility,such as:sperm zona binding test,DNA fragmentation,in vitro penetration test,oxidative stress test and top Body reaction test,etc.However,these laboratory tests have not made satisfactory progress as clinical tests.Therefore,it is necessary to further explore more accurate indicators to guide the clinical evaluation and diagnosis and treatment of male fertility.In recent years,research on proteins related to male fertility is gradually deepening.The research on the transactive response DNA-binding protein of 43(TDP-43)and the sperm X chromosome nuclear binding sperm protein(The sperm protein associated with the nucleus on the X chromosome,SPANX)has become more and more It is getting more and more attention.In order to further explore the precise indicators of clinical diagnosis of male fertility and study the pathogenesis of male infertility,this study used Western Blot method to detect SPANX and TDP-in sperm of sperm from frozen and high-fertility groups of semen volunteers 43 protein expression,to explore its relationship with male fertility.At the same time,the K-means cluster analysis method was used to statistically analyze the relationship between the clinical pregnancy outcomes of 487 AID cycles and the expression of TDP-43 and SPANX protein in sperm,thereby providing a new basis for the clinical evaluation,diagnosis and treatment of male fertility.PurposeTo explore the relationship between sperm TDP-43 and SPANX protein and male fertility,and to provide new targets for the research and evaluation of male fertility and the clinical diagnosis and treatment of male infertility.Materials and Methods 1 ObjectiveA total of 30 semen samples from the high fertility group and 21 semen samples from the low fertility group were collected from the human sperm bank in Henan Province from January 1,2009 to December 31,2017.TDP-43 and SPANX protein were tested.Inclusion criteria for semen samples of high fertility group:frozen semen of volunteers was used in clinical treatment for up to 8 AID cycles and all five women were conceived within 8 cycles;inclusion criteria for semen samples of low fertility group:for frozen semen of volunteers After 12 cycles of AID in clinical treatment,none of them conceived a woman.All frozen semen specimens used for AID treatment in the sperm bank comply with the "Basic Standards and Technical Specifications of Human Sperm Bank".Collected the medical records of 487 clinical cycles of the above 51 cases of semen specimens of the sperm from the Reproductive Medicine Center of the Third Affiliated Hospital of Zhengzhou University during the same period and the age of the recipient,anti-Mullerian hormone(AMH)、Antral follicle count(AFC),endometrial thickness and body mass index(BMI).Recipient inclusion criteria:①younger than 35 years old;②endometrial thickness≥ 8 mm;③no history of adverse pregnancy;④at least one side of the fallopian tube is unobstructed.Exclusion criteria:①Endometrial diseases,such as endometriosis,endometrial polyps,etc.;②Polycystic ovary syndrome;③pelvic adhesions;④Endocrine diseases;⑤Ovarian diseases;⑥tubal incompetence;⑦chronic Diseases such as high blood pressure,diabetes and other diseases that affect the outcome of pregnancy.2 Methods2.1 Western blot testsWestern Blot method was used to detect the expression of TDP-43 and SPANX protein in high and low fertility groups.2.2 K-means clustering analysisAccording to the detection results of TDP-43 and SPANX protein in sperm of high fertility group and low fertility group,51 volunteers were divided into TDP-43 and SPANX protein high expression group and low using K-means clustering analysis method.Two groups of expression groups.Correlation analysis of pregnancy outcomes,basic conditions of recipients(recipe age,AMH,AFC,endometrial thickness,BMI)and corresponding semen parameters of the corresponding 487 AID clinical cycles in two groups.3 Statistics analysisSPSS20.0 data was used for statistical analysis.The measurement data in the result is indicated by.Data analysis between the two groups used two independent sample t tests,with α=0.05 as the test level.Result1 The expression of TDP-43 protein in high and low fertility groupsThe relative expression level of TDP-43 protein in sperm of high fertility group(0.93±0.16)was higher than that of low fertility group(0.78±0.14),with statistical difference(P<0.05).2 The expression of SPANX protein in high and low fertility groupsThe relative expression level of SPANX protein in sperm of high fertility group(0.96±0.17)was higher than that of low fertility group(0.80±0.15),with statistical difference(P<0.05).3 High and low fertility group clinical cycle number,conception number and average cycle pregnancy rateThe amount of clinical cycles in the high and low fertility groups was 235 and 252,and the number of conceptions was 150 and 0,respectively.The average cycle pregnancy rate in the high fertility group was 63.83%(150/235)higher than the low fertility group 0%(0/252),which was statistically different(P<0.05).4 Correlation analysis between TDP-43 protein expression and AID clinical average cycle pregnancy rateThe relative expression level of TDP-43 protein in sperm in high expression group(n=28)(1.05±0.08)was higher than that in low expression group(n=23)(0.73±0.10),with statistical difference(P<0.05);The clinical average cycle pregnancy rate of 39.84%(100/251)in TDP-43 protein high expression group in sperm was higher than that in low expression group 21.19%(50/236),with statistical difference(P<0.05).There was no statistically significant difference in age,AMH,AFC,endometrial thickness,and BMI between the two groups of recipient women(P>0.05).5 Correlation analysis between SPANX protein expression and AID clinical average cycle pregnancy rateThe relative expression level of SPANX protein in sperm in high expression group(n=31)(1.07±0.11)was higher than that in low expression group(n=20)(0.75±0.09),with statistical difference(P<0.05);The average clinical pregnancy rate in the high expression group was 41.82%(115/275)higher than that in the low expression group 16.51%(35/212),with statistical difference(P<0.05).The age,AMH,AFC,endometrial thickness,and BMI of the two groups of recipient women were not statistically different(P>0.05).ConclusionsThe expression of TDP-43 and SPANX protein in sperm in the high fertility group was significantly higher than that in the low fertility group,and the clinical average cycle pregnancy rate of TDP-43 and SPANX protein high expression group was significantly higher than that of the low expression group,it can be inferred The expression of TDP-43 and SPANX protein in semen is related to male fertility.TDP-43 and SPANX proteins are expected to become potential markers for clinical evaluation and diagnosis of male fertility.