| ObjectiveIn the process of preimplantation genetic testing(PGT),intracytoplasmic sperm injection(ICSI)is a common fertilization method,so as to ensure that there is no contamination of paternal(sperms)or maternal(granulosa cells)genetic material in the embryo biopsy products.However,ICSI increases the expense and a series of risks caused by micromanipulation for couples with normal semina.This study aims to demonstrate that conventional in vitro fertilization(IVF)is viable in preimplantation genetic testing for aneuploidy(PGT-A)through clinical data analysis and experimental verification.At first,the clinical outcomes of patients with PGT-A after IVF or ICSI were retrospectively analyzed to explore whether that were affected by two insemination methods.Then an examination method was established to investigate whether the whole genome amplification(WGA)products of embryos from these cases by IVF in PGT-A had been contaminated by parental genetic material,and whether the contamination level had increased and affected the judgements of embryonic euploidy compared with embryo amplified products by ICSI,so as to determine the feasibility of IVF in PGT-A.MethodsPart Ⅰ: 450 patients who underwent PGT-A in Reproduction Center of 900 th Hospital of Joint Logistics Team from January 2015 to December 2019,and received euploid frozen-embryo transfers before April 2020 were studied,including 261 patients underwent IVF(Group IVF-PGT-A)and 189 patients underwent ICSI(Group ICSI-PGTA),whose pregnancy outcomes and the birth indicators of the offspring were analyzed.Meanwhile,the clinical data of patients given regular prenatal examination and delivery in Obstetrics of our hospital in these cases were collected(76 cases),involving 43 patients in Group IVF-PGT-A and 33 patients in Group ICSI-PGT-A,whose comorbidities and complications of pregnancy and abnormal conditions of the fetus and appendages were counted.These data were analyzed to discern whether the effectiveness and safety between two insemination methods in PGT-A is different.Part II: From March 2016 to March 2021,the frozen remaining WGA products of trophectoderm after PGT in Reproduction Center of 900 th Hospital of Joint Logistics Team were studied.Group IVF was 29 patients who received euploid frozen-embryo transfers,which were tested by PGT-A after IVF,and childbirth successively(29 blastocysts);Group ICSI was 13 patients who got PGT after ICSI(36 blastocysts).The ASA chips(Infinium Asian Screening Array-24 v1.0 Bead Chip,Illumina,California)were used to test these WGA products and blood or saliva samples of these couples for single nucleotide polymorphisms(SNPs).Two strategies,Strategy M(including sites in which maternal heterozygote and paternal homozygote),and Strategy F(including sites in which paternal heterozygote and maternal homozygote)were used to calculate the heterozygous sites ratios of embryos respectively,which are the same without contamination theoretically,in order to analyze whether the potential contamination ratio of the parental genetic material in the Group IVF and Group ICSI is consistent,so as to find whether it will have different influence on the results.ResultsPart Ⅰ: There is no difference in rate of pregnancy,implantation,live birth,or abortion between groups(P>0.05).The ectopic pregnancy rate in Group ICSI-PGT-A was significantly higher than that in Group IVF-PGT-A(P>0.05),but its clinical significance needs to be further expanded.Meanwhile,though the chromosome abnormity rate of abortion in Group ICSI-PGT-A was significantly higher than the other group(P<0.05).But the resolution of abortion detections is 100 Kb,and that of PGT is 10 Mb.The lengths of all the chromosome abnormity fragments of abortions are shorter than 3Mb,as well as the limited cases(3 cases in Group IVF-PGT-A and 4 cases in Group ICSI-PGT-A),so the detection error of PGT-A is not considered.In addition,there is no difference in the incidences of gestational diabetes mellitus,hypertensive disorder of pregnancy,intrahepatic cholestasis of pregnancy,idiopathic thrombocytopenia,placenta previa,abnormal placental morphology,placenta accreta,oligohydramnios,abnormal fetal position,premature rupture of membranes,fetal growth restriction,fetal distress,premature birth,macrosomia,low birth weight infants,very low birth weight infants and birth defects between groups(P>0.05).Part II: There is no significant difference in the call rates of SNPs between groups(P>0.05).In Group ICSI,there are statistic differences in heterozygous sites ratios of embryos between Chromosome 1-22 by these strategies(P<0.05),and that in Chromosome 19 and 22 are lower(P<0.05).In Group IVF,there is no statistic difference in heterozygous sites ratios among all chromosomes by Strategy M(P>0.05),but Chromosome 22 is lower than others by Strategy F(P<0.05).Except for Chromosome 1,4,9 and 19,there is no statistic difference in total variation of heterozygous sites ratios between groups by Strategy M(P>0.05),and that by Strategy F is no statistic difference(P>0.05).But heterozygous sites ratios in Group IVF is higher than that in Group ICSI(P<0.05),which shows the existence of difference between WGA methods.Difference of heterozygous sites ratios is not found between two strategies in each chromosome and group(P>0.05).All these demonstrate that there was no maternal contamination in Group ICSI,and no dominating paternal or maternal contamination in Group IVF,but whether the equal proportion of parental contamination exists can not be excluded,though which seems unlikely.ConclusionPart Ⅰ:(1)There is no effect on the incidences of comorbidities and complications of pregnancy,abnormal conditions of the fetus and appendages and the birth indicators of the offspring after the use of IVF and ICSI in PGT-A.(2)There is no difference in clinical outcomes was observed when selecting embryos to transfer after IVF or ICSI in PGT-A,so the results obtained by PGT-A after IVF were as reliable as ICSI.Part Ⅱ:(1)Compared with ICSI,IVF did not increase the parental contamination during the amplification and detection of embryonic genomes.(2)IVF can be recommended for patients who are diagnosed as non-male infertility and preparing for PGT-A. |
