| Objective:Recurrent spontaneous abortion(RSA)is defined as three or more consecutive pregnancy losses.Most of the recurrent spontaneous abortion occur in early abortions,minority in late-term abortion.Recurrent spontaneous abortion is a common disease in department of obstetrics and gynecology,it has adverse impact on maternal physiological and psychological,but also a serious challenge to the clinical workers,so the study of etiology and pathogenesis is extremely important.It suggests that Limited trophoblast invasion is one of the events implicated in the pathogenesis of recurrent spontaneous abortion.Apoptosis,or programmed cell death,is a physiological process necessary for adequate remodeling of maternal deciduas,to control trophoblast invasion and to maintain maternal–fetal immune tolerance in balance.Accordingly,several abroad studies have reported an association between abnormal trophoblast apoptosis and recurrent spontaneous abortion.Studies shows that the FAS/FASL system is one of the most important inducers of apoptosis.Activation of this complex is essential for trophoblast development and invasion and also to protect trophoblasts from being attacked by the maternal immune system.Altered FAS/FASL expression may result in altered apoptosis and ultimately affects both immune response and implantation.Single-nucleotide polymorphisms(SNPs)have been identified in the promoter regions of FAS/FASL genes.It has been showed that the FAS-1377 G/A,FAS-670 A/G and FASL-844 T/C polymorphisms might influence the transcriptional level of the two genes and have an effect on their expression alteration.However,as far as we know,these genetic variants have not been evaluated in women with RSA.This promoted us to conduct this investigation.This study proposed by polymerase chain reaction-multiple SNaPshot SNP classification technique to detect the SNP from the perspective of genetics.The aim of this study was to test whether the three SNPs in the promoter regions of FAS and FASL genes may be associated with the risk of RSA.To discuss the pathogenesis of RSA at the gene level may provide certain theoretical basis for the clinical diagnosis and treatment.To identify the genetic polymorphisms associated with the risk of RSA is likely to be a useful tool to identify high-risk pregnant women,in an effort to carry out preventive intervention and clinical treatment in patients with RSA.Methods:1 This case-control study consisted of 99 women with recurrent spontaneous abortion and 100 healthy women as control.Venous blood(5ml)was drawn from each subject into vacutainer tubes containing EDTA and stored at 4 ?.Genomic DNA was extracted within one week after sampling using proteinase K digestion followed by a salting out procedure.The FAS-1377 G/A,FAS-670 A/G and FASL-844 T/C SNPs were genotyped by polymerase chain reaction-multiple SNaPshot SNP classification technology.2 Statistical analysis was performed using SPSS13.0 software package.The age difference of cases and frequency-matched controls was analyzed by the t-test.Hardy–Weinberg analysis was performed by comparing the observed and expected genotype frequencies in the control group using the chi-square test.Comparison of the FAS and FASL allelotype,genotype,haplotype and combined genotype distribution in endometriosis patients and healthy controls were performed by means of two-sided contingency tables using the chi-square test.Odds ratios(ORs)were calculated and reported within the 95% confidence limits.All statistical tests were two-sided,and P < 0.05 was considered statistically significant.Results:1 FAS/FASL genotypes distribution:The distribution of the FAS-1377 G/A,FAS-670 A/G and FASL-844 T/C genotypes in controls did not significantly deviate from that expected for a Hardy–Weinberg equilibrium(P>0.05);2 The comparison of FAS-1377 G/A(rs2234767)SNP allele and genotype frequencies: The frequencies of the FAS-1377 G/A G and A allele among patients with RSA and healthy controls were 63.1%,36.9% and 74.0%,26.0%,respectively;The distribution of the GG,GA,AA genotypes between patients with RSA(37.4%,51.5%,11.1%)and controls(51.0%,46.0%,3.0%).The frequency of-1377 A alleles was higher in the cases(0.37)than in the controls(0.26)and the difference was statistically significant(P = 0.020),A alleles increased the risk of RSA(OR = 1.66,95% CI = 1.08 2.55).GG genotype of-1377 FAS SNP was considered as the reference.The-1377 AA genotype was more frequent in the cases(11.1%)than in the controls(3.0%)and increased the risk of recurrent miscarriage obviously(OR = 5.05,95% CI = 1.32 19.40);3 The comparison of FAS-670 A/G(rs1800682)SNP allele and genotype frequencies: The frequencies of the FAS-670A/G A and G allele among patients with RSA and healthy controls were 68.2%,31.8% and 66.0%,34.0%,respectively;No significant difference in the FAS-670 A/G allele distribution was shown between patients with RSA and controls(P > 0.05).The distribution of the GG,AG,AA genotypes between patients with RSA(43.4%,49.5%,7.1%)and controls(43.08%,49.0%,8.0%)also had no significant difference(P >0.05).Compared with the AA genotypes,the AG+GG genotypes could not increase the risk of developing RSA,the odds ratio was 0.97(95%CI= 0.55-1.69);4 The comparison of FASL-844T/C(rs763110)SNP allele and genotype frequencies: The frequencies of the FASL-844T/C C and T allele among patients with RSA and healthy controls were 72.2%,27.8% and 73.5%,26.5%,respectively;No significant difference in the FASL-844T/C allele distribution was shown between patients with RSA and controls(P > 0.05).The distribution of the CC,CT,TT genotypes between patients with RSA(50.5%,43.4%,6.1%)and controls(50.0%,47.0%,3.0%)also had no significant difference(P > 0.05).Compared with the CC genotypes,the CT+TT genotypes could not increase the risk of developing RSA,the odds ratio was 1.07(95%CI=0.69-1.66);5 The combined genotypes of FAS/FASL: The FAS-670AG/FASL-844 CC genotypes combination showed highest risk(OR=2.6;95%CI=1.10-61.03).Genotype combinations FAS-670AA/FASL-844 CT was also statistically significant,suggestive of their role in RSA risk(OR=2.6;95%CI=1.06-6.26).Conclusions:1 The results demonstrated that there was significant association of the FAS-1377G/A SNPs with the genetic risk of RSA.A allele may increase the risk of RSA,the AA genotype significantly increase the risk of RSA.2 No significant association of the-670A/G and FASL-844T/C SNPs with the genetic risk of RSA.The FAS-670AG/FASL-844 CC genotypes combination showed highest risk of RSA..Genotype combinations FAS-670AA/FASL-844 CT was also statistically,suggestive of their role in RSA risk. |
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