Genetic Study On Male Infertility And Female Unexplained Recurrent Spontaneous Abortion In Chinese Population

Part I The association of mitochondrial variations in the ND4 and UUR genes with male asthenospermiaObjective Mitochondrial gene mutations have been reported to be associated with semen quality and sperm motility.The aim of this study was to investigate whether the four mitochondrial genes(ND4,UUR,ATP6 and ATP8)are involved in male asthenospermia.Methods A total of 97 asthenospermia patients and 80 fertile controls were recruited from the Reproductive Medicine Center of the 105 th PLA in this case-control study.Genomic DNA were extracted from the sperm of all participants.In order to find variations,the four mitochondrial DNA genes(ND4,UUR,ATP6 and ATP8)were amplified by using PCR with the gene-specific primers and sequenced.Results For the ND4 gene,we found a total of 64 and 54 nucleotide substitutions in patients and controls,respectively,with no discrepancy in the mutation rates(66.0% vs.67.5%,p>0.05).However,one mutation(g.11084A>G,p.T109A)leading to an amino acid substitution in a highly conserved residue and predicted to be deleterious was detected only in the cases.For the UUR gene,a novel mutation(g.3263C>T)near the anticodon TAA was identified in an asthenospermia patient and was absent from normal controls.It might affect leucine synthesis rate.For the other two genes ATP6 and ATP8,there was no significant difference in frequency between the two groups(P>0.05)and there were no new variations in the cases.Conclusions Our result suggests that the ND4 and UUR genes might be associated with asthenospermia.Part II The association of SLX4 gene variation with male azoospermiaObjective To explore the correlation between SLX4 gene variation and male azoospermia.Methods A total of 40 azoospermia patients were recruited from the Reproductive Medicine Center of the 105 th PLA in the current study and 400 healthy men were used as control.The genomic DNA of the peripheral blood samples of the patients and the control were extracted.All exons and nearby introns of the SLX4 gene were sequenced,the results were compared with the 1000 Genomes Project.The variations found were resequenced in 400 healthy controls,screening for possible pathogenic variations.Bioinformatics was used to predict the possible impact of the variations on protein function and structure.Results In 40 patients with azoospermia,14 exons and nearby introns of SLX4 gene were sequenced.For the variation c.5060C> G(p.Pro1687Arg)found in exon 13 and the variation c.5177C> T(p.Ala1726Val)found in exon 14,they both also found in the healthy controls and might not be associated with azoospermia.In the coding region of the 7th exon and the 8th exon,synonymous mutations c.1740C> T and c.1977G> A both encoded serine.In a patient with azoospermia,a synonymously mutated c.1152A> G was found in the 14 th exon of the SLX4 gene,two heterozygotes were found in the5'UTR region,c DNA.347A>G(rs59311338)and c DNA.283C>G(rs57910835),and another heterozygote was also found in the 3'UTR region of the same patient,c DNA.6244A> G(rs551538592).Conclusions SLX4 gene variation might not be related to male azoospermia.Part III Association of interleukin 1,alpha polymorphism with the risk of unexplained recurrent spontaneous abortionObjective This study is to investigate the association of IL1A(-889C/T)polymorphism with the risk of URSA in Chinese population.Methods 141 cases of Chinese URSA samples and 191 control samples were recruited from the Reproductive Medicine Center of the 105 th PLA in this case-control study.The genotype of IL1a(-889C/T)was analyzed by RFLP and PCR samples were digested by Nco I.The locus carring C allele produced 271 and 29 bp fragments while carring T allele only produced a 300 bp fragment.The difference of IL1a(-889 C / T)polymorphism between case group and control group was compared by ?2.The OR and the 95% CI were used as a measure of the association between the genotype frequency and the URSA.Results There were significantly different frequency in IL1 A polymorphism(-889C/T)between case and control groups(P<0.05).Conclusions IL1A(-889C/T)polymorphism might be a risk factor of URSA in Chinese women.