中国汉族男性非梗阻性无精子症患者易感基因的相关性研究

发布时间：2018-05-17 12:03

本文选题：不育症 + 非梗阻性无精子症（NOA）　；参考：《安徽医科大学》2014年硕士论文

【摘要】：第一部分中国汉族男性非梗阻性无精子症ETV5，HORMAD1和HORMAD2基因相关性研究目的：探讨ETV5，HORMAD1和HORMAD2基因多态性与非梗阻性无精子症的易感性。方法：应用Sequenom MassArray质谱阵列技术对368例已生育的汉族男性人群(对照组)和361例汉族男性非梗阻性无精子症（实验组）ETV5基因的5个标签单核苷酸多态(single nucleotide polymorphism,SNP)位点(rs12631658, rs6444106, rs7430047,rs7433760, rs9824882)，HORMAD1基因3个SNP位点（rs1336900，rs16840074，rs6694531），HORMAD2基因8个SNP位点（rs8135823，rs11090601，rs4823073，rs5763779， rs718772，rs9620953，rs9625930， rs975704）进行基因型检测。应用Plink1.07软件对数据资料进行统计分析，比较对照组与病例组最小等位基因频率（MAF）及基因型差异，运用Haploview软件对ETV5、HORMAD1和HORMAD2基因进行单体型分析。结果：ETV5基因5个标签SNP，HORMAD13个SNP和HORMAD28个标签SNP的等位基因频率、基因型分布在实验组和对照组均无统计学差异（P＞0.05），进一步的单体型分析亦未显示统计学差异（P＞0.05）。当分析各SNP位点基因型在NOA亚组分布情况，我们发现HORMAD2基因rs718772位点最小等位基因频率（MAF）分布在双侧平均睾丸体积大于10ml组与小于10ml组中有显著性差异（P=0.035）。结论：检测的ETV5基因，HORMAD1和HORMAD2基因SNP位点多态性与汉族男性非梗阻性无精子症的发生可能不相关, HORMAD2基因rs718772位点SNP可能与NOA患者的睾丸发育有关。第二部分CREM信号通路基因单核甘酸多态性与NOA发生的关联性研究目的比较CREM信号通路相关基因CREM、ACT、Kif17b和SPAG8基因17个SNPs（single nucleotide polymorphisms, SNPs）位点的基因型频率、等位基因频率和单体型频率等在特发性非梗阻性无精子症（non-obstructive azoospermia，NOA）患者及正常生育男性间的分布，探讨这4种基因突变是否为NOA的致病因素。同时运用q-RT-PCR方法，研究CREM在NOA（生精阻滞）患者睾丸组织及正常睾丸组织的表达差异，探讨CREM基因在精子发生过程的作用及意义。方法我们选择361例特发性非梗阻性无精子症（NOA）作为实验组，368例精液质量正常的男性作为对照组。采用Sequenome Massarray质谱阵列技术对CREM、ACT、Kif17b和SPAG8基因的17个SNPs位点进行了基因型分型。选取30例生精阻滞患者及30例正常精子发生的睾丸组织，运用q-RT-PCR的方法检测CREM基因的在两组的mRNA表达水平，T检验进行数据统计，P0.05为具有统计学意义。结果CREM基因共检测4个SNP位点（rs4934540，rs2295415，rs11592356，rs1148247），其中rs4934540位点C等位基因频率（24.7%VS34.4%，P=4.9*10-5，OR=0.624,95%CI=(0.497-0.784)，rs2295415位点的G等位基因频率（18.0%VS24.2%，P=0.003，OR=0.686,95%CI=(0.532-0.885)，rs11592356位点G等位基因频率（6.9%VS10%，P=0.036，OR=0.672,95%CI=(0.461-0.978)均在NOA组显著低于正常生育组，提示该位点的变异可能对正常精子的发生具有保护作用，进一步的单体型分析提示TATG（rs493454, rs2295415, rs11592356and rs1148247）单体型的比例在NOA组显著高于正常生育组（P=0.011，OR=1.317，95%CI=1.064-1.631）。而ACT基因9个标签SNP（srs2273621，rs9373985，，rs2252816，rs9398152，rs9398148，rs4839688，rs17056756，rs3798292，rs9486705）、KIF17b3个SNP（rs631357，rs522496，rs2296225）、SPAG8基因1个SNP（rs3763631）基因型分布在实验组及对照组均未见差异。CREM基因在NOA（生精阻滞）组及正常精子发生中睾丸组织的表达用明显差异，CREM基因在NOA（生精阻滞）患者睾丸组织中低表达（P=0.00001）。结论CREM基因多态性可能是中国精子发生障碍患者的遗传易感因素，rs4934540，rs2295415，rs11592356三个位点对精子发生可能起保护作用，而ACT、KIF17b和SPAG8基因单核苷酸多态性与NOA的发生无关。CREM的正常表达对精子的正常发生具有重要的作用，CREM的低表达可能是精子发生障碍的一个重要原因。
[Abstract]:Part one: correlation between ETV5, HORMAD1 and HORMAD2 genes in Chinese Han men with non obstructive azoospermia.
Objective: To investigate the association of ETV5, HORMAD1 and HORMAD2 gene polymorphisms with non obstructive azoospermia.
Methods: the 5 labelling single nucleotide polymorphisms (single nucleotide polymorphism, SNP) loci (rs12631658, rs6444106, rs7430047, rs7433760, rs9824882) of 368 Han male and 361 Han male non obstructive azoospermia (experimental group) ETV5 genes were used by Sequenom MassArray mass spectrometry array technique. The 3 SNP loci of the MAD1 gene (rs1336900, rs16840074, rs6694531), and the 8 SNP loci of the HORMAD2 gene (rs8135823, rs11090601, rs4823073, rs5763779, rs718772, etc.) were used for genotyping. And genotype differences, the haplotypes of ETV5, HORMAD1 and HORMAD2 genes were analyzed by Haploview software.
Results: the allele frequency of ETV5 gene 5 label SNP, HORMAD13 SNP and HORMAD28 label SNP was not statistically different between the experimental group and the control group (P > 0.05), and the further monosomatograph analysis did not show statistical difference (P > 0.05). When the distribution of the SNP loci genotype in the NOA subgroup was analyzed, we found HORM The minimum allele frequency (MAF) of rs718772 locus in AD2 gene was significantly higher in the bilateral mean testicular volume than in the 10ml group and smaller than that in the 10ml group (P=0.035).
Conclusion: the detection of ETV5 gene, HORMAD1 and HORMAD2 gene polymorphisms may not be related to the occurrence of non obstructive azoospermia in Han men. The rs718772 locus SNP of the HORMAD2 gene may be related to the testicular development of NOA patients.
The second part is the association between CREM signal pathway gene mononuclear nucleotide polymorphism and the occurrence of NOA.
Objective to compare the genotype frequencies of the 17 SNPs (single nucleotide polymorphisms, SNPs) loci of CREM, ACT, Kif17b and SPAG8 genes, and the distribution of allelic frequencies and haplotype frequencies between the allele frequencies and the haplotype frequencies of the CREM signaling pathway related genes in idiopathic non obstructive azoospermia (non-obstructive azoospermia) and normal fertile men. Whether the 4 kinds of gene mutations are the pathogenic factors of NOA, and the difference in the expression of CREM in the testis and normal testicular tissues of the patients with NOA (spermatogenesis block) by using the q-RT-PCR method, and to explore the role and significance of the CREM gene in the process of spermatogenesis.
Methods we selected 361 cases of idiopathic non obstructive azoospermia (NOA) as the experimental group and 368 men with normal semen quality as the control group. The genotyping of 17 SNPs loci of CREM, ACT, Kif17b and SPAG8 genes was performed by Sequenome Massarray mass spectrometry. 30 cases of spermatogenesis block and 30 normal spermatozoa were selected. The testis tissue of the newborn was detected by q-RT-PCR. The expression level of CREM gene in the two groups was detected by T, and the data were statistically analyzed by T test.
Results 4 SNP loci (rs4934540, rs2295415, rs11592356, rs1148247) were detected in the CREM gene, and the frequency of rs4934540 loci C allele frequencies (24.7%VS34.4%, P=4.9*10-5, OR=0.624,95%CI= (0.497-0.784), and allele frequencies of the loci. 6.9%VS10%, P=0.036, and OR=0.672,95%CI= (0.461-0.978) were significantly lower in the NOA group than in the normal growth group, suggesting that the mutation of the loci may have protective effects on normal sperm. Further haplotype analysis suggests that the proportion of TATG (rs493454, rs2295415, rs11592356and rs1148247) in the NOA group is significantly higher than that in the normal growth group (P=). 0.011, OR=1.317,95%CI=1.064-1.631) and ACT gene 9 label SNP (srs2273621, rs9373985, rs2252816, rs9398152, rs9398148, rs4839688, rs17056756, rs3798292, rs9486705). There was a significant difference in the expression of testicular tissue between normal group and normal spermatogenesis group. The expression of CREM gene was low in testicular tissue of NOA (spermatogenesis arrest) (P=0.00001).
Conclusion CREM gene polymorphism may be a genetic predisposing factor in Chinese spermatogenesis disorders. The three loci of rs4934540, rs2295415, and rs11592356 may play a protective role in spermatogenesis, while the positive expression of ACT, KIF17b and SPAG8 single nucleotide polymorphisms is not related to the occurrence of NOA, and it has an important role in the normal occurrence of sperm. The low expression of CREM may be an important cause of spermatogenesis disorders.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R698

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