| Objective:By studying the difference of ACE gene polymorphisms in non-obstructive azoospermia patients and normal male patients,ligase typing technology was used for gene sequencing and typing to determine the ACE gene polymorphism and nonobstructive.The association between the occurrence of spermatogenesis may provide a theoretical and experimental basis for the establishment of certain mechanisms that can reveal the occurrence of non-obstructive azoospermia and the genetic diagnosis of non-obstructive azoospermia that can be applied in clinical practice.Methods:Selected from February 2013 to February 2016 for "male infertility" of the Department of Reproductive and Prenatal Diagnosis at the First Hospital of Jilin University.Males of Han nationality in the three northeastern provinces were diagnosed with "non-obstructive azoospermia",as an experimental group.A total of121 patients who met the screening criteria were included.Selected volunteers who donated sperm from the human sperm bank of Jilin Province from September 2013 to November 2015 and routinely analyzed semen density and sperm motility by semen were used as control groups.A total of 256 volunteers meeting the screening criteria were included.Two groups of subjects completed a professional questionnaire and met the inclusion criteria: normal testicular examination,normal karyotype analysis of G-banding in peripheral blood,and no reported microdeletions of AZF gene in peripheral blood.Excluding recent history of high fever,long-term radiation exposure history,history of mumps,history of major systemic diseases,history of genital trauma,history of radiotherapy and chemotherapy,severe varicocele(3 degrees),karyotypic abnormalities,and AZF microdeletions on the Y chromosome Past history and current medical history.The semen from the experimental group and the control group were processed and analyzed,and the results were recorded.The level of reproductive hormones was detected and analyzed using a commercial assay kit;routine karyotype analysis was performed and recorded for all subjects;the polymerase chain reaction was performed.Detection of whether the subject's Y chromosome AZF gene is missing;the use of peripheral blood genomic DNA extraction kit according to the standard procedure to extract the subject's peripheral blood DNA,PCR sequencing typing technology to identify genetic polymorphic loci,detection and genotyping The agarose electrophoresis was used to detect the PCR product.The 3730 sequencer was used to detect the genotyping results by electrophoresis.The haplotype analysis software was used to analyze the SNPs on the same chromosome.Statistical analysis software SPSS19.0 was used for statistical analysis.Results:1.In this study,121 patients were involved in the NOA experimental group and 256 in the normal control group.The average age of the NOA group was significantly higher than that of the control group(30.00±5.69 vs 25.35±5.51);the BMI index NOA group was significantly higher than the control group(25.31±5.13 vs22.89±3.83);the sperm p H NOA group was significantly lower than the control group(7.48± 0.41 vs 7.51±0.06);sperm density in NOA group was significantly lower than that in control group(0 vs 66.88±11.11);semen volume in NOA group was significantly lower than in control group(2.74±1.49 vs 3.69±1.29);serum FSH NOA group was significantly higher The control group(19.42±11.62 vs3.35±1.58);serum LH NOA group was significantly higher than the control group(9.17±5.20 vs 4.96±2.05);serum PRL NOA group was significantly higher than the control group(409.28±215.19 vs 305.11±168.37);The serum E2 NOA group was significantly higher than the control group(35.46±25.32 vs34.21±11.85);the serum T NOA group was significantly lower than the control group(12.46±6.90 vs 17.72±6.43);the berry fructose NOA group was significantly higher than the control group(66.47±57.96 vs 20.40±12.88);?-glucosidase NOA group was significantly higher than the control group(41.25±43.95 vs 30.32±18.99);seminal zinc NOA group was significantly higher than the control group(7.45±5.31 vs 2.91±2.88);serum The B NOA group was significantly lower than the control group(51.90±84.52 vs.200.93±71.27).There was no statistical difference between p H and E2 in the two groups,and there were significant differences in other indicators.2.This study binary logistic regression analysis method was applied to two groups of subjects minimum genotype frequency and allele frequency(Minor allele frequency,MAF)data such as correlation analysis,and genotype in accordance with Hardy Weinberg equilibrium(Hardy-Weinberg equilibrium,HWE)and "age" factors as covariate in correction,the results are as follows: the ACE gene loci rs4316,rs4331 and rs4343 MAF value in NOA group were 63.20% and0.355 PHWE value;The MAF values in the control group were 62.70%,and the PHWE value was 0.089,and the p value of the group was 0.889,with no statistical difference.The MAF value in the NOA group was 59.10%,the PHWE value was 0.397,the MAF value of the control group was 60.02%,the PHWE value was 0.381,and the p value of the group was 0.781,with no statistical difference.In addition,ACE c.81C>T(rs4316),ACE c.471A>G(rs4331),ACE c.606G>A(rs4343)and ACE c.1665T> c(rs4362)showed no statistical difference in genotype distribution in NOA group and control group(p<0.05).3.This study applied binary logistic regression analysis of two groups of subjects were four candidate SNPs loci show hidden model with NOA correlation analysis,and to "age" relevant correction factors for the covariate,the results are as follows:In the dominant model,ACE c.81C>T(rs4316),ACE c.471 a >G(rs4331),and the p value of ACE c.606G>A(rs4343)were all 0.070(OR=1.933,95%CI=0.948-3.941),and the p value of the ACE c.1665T> c(rs4362)was0.352(OR=1.368,95%CI=0.706-2.650),and there was no statistical difference between the groups.The results showed that there was no statistical correlation between the dominant model and NOA.In the implicit model,ACE c.81C>T(rs4316),ACE c.471 a >G(rs4331),and the p value of the ACE c.606G>A(rs4343)were all 0.867(OR=1.045,95%CI=0.625-1.746),and the p value of the ACE c.1665T> c(rs4362)was 0.887(OR=0.963,95%CI=0.568-1.631),and there was no statistical difference between the groups.The results showed that there was no statistical correlation between the recessive model and NOA.4.ACE c.81C> T(rs4316),ACE c.471A> G(rs4331),ACE c.606G> A(rs4343),ACE c.1665T> C(rs4362)on chromosome 17,formed an overlay 11 kb haplotype block with haplotypes TGAC,CAGT,TGAT.The P values between the three haploids were all greater than 0.05,suggesting that TGAC,CAGT,and TGAT are not related to NOA.5.Genotype frequencies of SNPs in the normal and abnormal reproductive hormone groups: ACE c.81C>T(rs4316),ACE c.471A>G(rs4331),and ACE c.606G>A(rs4343)p values in the FSH group All were 0.340;in the LH group,the p-values were 0.659;in the E2 group,the p-values were both 0.986;in the PRL group,the p values were 0.408;in the inhibin B group,the p-values were 0.479,p>0.05,and there was no statistical difference.The rs4316,rs4331 and rs4343 polymorphisms were not associated with abnormal levels of FSH,LH,PRL,E2 and inhibin B.ACE c.1665T>C(rs4362)in the FSH group p=0.592;LH group p=0.490;E2group p=0.851;PRL group p=0.646;inhibin B group p=0.224;p=0.129,p>0.05,no statistical difference,suggesting that the rs4362 polymorphism is not associated with abnormal levels of FSH,LH,PRL,E2,T and inhibin B.The frequencies of ACE c.81C>T(rs4316),ACE c.471A>G(rs4331)and ACE c.606G>A(rs4343)in the abnormal T group were significantly lower than those in the normal group(29.2% vs 43.3)%,p=0.038),with statistical difference,suggesting that the rs4316,rs4331 and rs4343 polymorphisms were positively correlated with the decrease of T hormone levels.6.Seminal plasma biochemical parameters SNPs genotype frequency comparison between normal and abnormal groups: ACE c.81C>T(rs4316),ACE c.471A>G(rs4331)and ACE c.606G>A(rs4343)in the refined fructose The p-values in the group were 0.954;the p-values in the ?-glucosidase group were 0.778;the pvalues in the seminal plasma zinc group were both 0.920;p>0.05.There was no statistical difference,suggesting that the rs4316,rs4331,and rs4343 polymorphisms were associated with The abnormal levels of quercitose,alpha glucosidase and seminal plasma zinc were not related.The p values of ACE c.1665T>C(rs4362)in the querglucose group were 0.925;the p-values in the ?-glucosidase group were both 0.708;the p values in the seminal plasma zinc group were all 0.881,p>0.05,with no statistical difference suggesting that rs4362 locus polymorphism is not related to abnormity of seminal fructose,alpha glucosidase and seminal plasma zinc levels.Conclusions:1.ACE rs4316,rs4331,rs4343,rs4362 site polymorphism has no significant correlation with non-obstructive azoospermia.2.The abnormal expression of ACE rs4316,rs4331,rs4343,rs4362 was positively correlated with the decrease of the reproductive hormone T level in NOA patients.There was no correlation between FSH,LH,PRL,E2,Inhibin B and the biochemical indices.ACE c.1665T>C(rs4362)site was not associated with the abnormal biochemical index of seminal plasma. |
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