Effects Of Single Nucleotide Polymorphism Of Susceptible Genes On Spermatogenic Failure

Objectives: Non-obstructive azoospermia is the most severe type of male infertility, and it exhibits spermatogenic arrest in the spermatogenesis stage. On the basis of previous researches and knockout mice models, we picked out 5 candidate genes(RNF8,BRDT,USF1,GTF2A1 Land OR2W3) which play important roles in spermatogenesis, then in the Chinese Han population respectively, we investigated the relationships between the minor allele frequency(MAF), genotype frequency and haplotype of 23 single nucleotide polymorphisms(SNPs) from five spermatogenesis-related genes(RNF8, BRDT, USF1, GTF2A1 L and OR2W3) and non-obstructive azoospermia(NOA) by using sequencing, and explored the association between the SNPs and clinical data(testis volume and follicle-stimulating hormone level) in the NOA group, in this way, we could find out the underlying genetic factors of non-obstructive azoospermia in Chinese Han male population. Methods: We collected the blood samples of 361 non-obstructive azoospermia patients in Chinese Han population and 368 normal people in Chinese Han population as the experimental group and the control group, and extract their DNA from whole blood samples using DNA extraction kit(QIAGEN) according to the instruction book, then using the Sequenom Mass Array technology to analyze the SNPs rs104669, rs195379, rs195434, rs2284922, rs2284924, rs9380661, rs2269058, rs195432, rs169005 in RNF8 gene, rs10783071, rs1156281, rs11165906, rs2183036, rs17576636 in BRDT gene, rs2516838, rs1556259, rs2774276 in USF1 gene, rs1916838, rs11677854, rs10432667, rs13024367 in GTF2A1 L gene and rs10888267, rs11204546 in OR2W3 gene. Using Plink 1.07 software to analyze the outcomes to compare the distributions of the minor allele frequency of the tag SNPs between the experimental group and the control group,and applying the Haploview software and SPSS software to analyze the association between the genotype and the clinical data. Results: The results showed that the minor allele frequency of rs104669 and rs2516838 between the experimental group and the control group showed big differences(p=0.038, OR=1.326; p=0.020, OR=1.436). The haplotypic data showed that the haplotype AC(rs104669, rs195432) of RNF8 gene could reduce the risk of NOA, while the haplotype TC(rs104669, rs195432), ATCATCTCT(rs2269058, rs2284924, rs9830661, rs169005, rs195379, rs2284922, rs104669, rs195432, rs195434) and TCATCTC(rs2284924, rs9830661, rs169005, rs195379, rs2284922, rs104669, rs195432) could add the risk of NOA(p=0.0031, OR=1.693; p=0.0086, OR=4.502; p=0.0085, OR=1.630). In addition, the haplotype TCG(rs1556259, rs2516838, rs2774276) of USF1 gene could also add the risk of NOA. The association analysis between the genotype and clinical data demonstrated that rs195432, rs196534 and rs2284922 of RNF8 gene were associated with smaller testis volume(p=0.030; p=0.025; p=0.034). Also, rs11204546 of GTF2A1 L gene and rs11677854 of OR2W3 gene were associated with follicle-stimulating hormone(FSH)(p=0.004; p=0.018). Conclusions: Our results provided a new insight into susceptibility of RNF8, BRDT, USF1, GTF2A1 L and OR2W3 genes in non-obstructive azoospermia. Among the 23 SNPs of interest, only rs104669 of RNF8 gene and rs2516838 of USF1 gene showed associations with non-obstructive azoopermia of Chinese Han population. Furthermore, clinically, rs1556259, rs2516838 and rs2774276 of RNF8 gene were associated with testis volume. The SNPs rs11204546 of GTF2A1 L gene and rs11677854 of OR2W3 gene were associated with FSH level, and because of the importance of follicle-stimulating hormone level in sperm extraction, they might be additional molecule predictive markers for assessing the treatment of NOA patients.