Study And Analysis Of Chromosomal Mosaicism In Interventional Prenatal Diagnosis

Karyotype analysis is the gold standard in prenatal diagnosis and plays a key role in diagnosing fetal chromosomal abnormalities.One of the difficulties associated with this process is chromosomal mosaicism,which refers to the presence of two or more different populations of cells in one individual.Amniocentesis and chorionic villus sampling are employed in prenatal diagnosis.The sources of cells used for diagnosis are diverse.Amniotic fluid cells are derived from shedding of cells by multiple germ layers of the fetus and contain a very small amount of extraembryonic tissue from the amniotic cavity.Villi cells are derived from Leydig cells of the chorionic mesoderm and therefore are derivatives of the extraembryonic tissue.Methods of karyotype analysis also involve in vitro cell culture,and there is a risk of in vitro chromosomal structural rearrangement and absence of chromosome segregation.In addition,the process of film production cannot be completely automated.The above-mentioned issues are challenges that are associated with accurate diagnosis of fetal chromosomal mosaicism.Currently,the laboratory evaluation method is mainly based on the mosaicism grading method described by Hsu.However,differences exist with respect to the type and level of each fetal mosaicism,specific chromosomes involved,and choice of puncture methods;these differences make accurate diagnosis a complex process.Thus,this grading method is not absolute and requires specific analysis for certain cases.In this study,the issue of chromosomal mosaicism in prenatal diagnosis was investigated,and its clinical diagnosis and treatment were discussed in two sections.Aim: To explore the distinction between true mosaicism and pseudomosaicism of amniotic fluid and chorionic villi karyotype in prenatal diagnosis,and to provide a basis for proper clinical treatment of mosaicism.Methods: 1.Pregnant women who underwent karyotype diagnosis using amniotic fluid or chorionic villus sampling owing to various high risk factors in our hospital from January 2013 to June 2016 were selected.Based on the mosaicism grading principles revised by Hsu,mosaicism was assessed.In addition,results of fluorescence in situ hybridization(FISH)and umbilical cord blood examination and records of follow-ups were collected.2.To identify maternal cell contamination in villi chromosomal karyotype mosaicism samples in prenatal diagnosis using multiplexed fluorescence labeled STR-PCR.A total of 8 samples with suspected maternal cell contamination were found in the initial chromosomal karyotyping results of villi sampling,and the results suggested that these were tertiary mosaicism and abnormal karyotype/46,XX mosaicism.These 8 samples were included in the study group.Simultaneously,peripheral blood was obtained from the mothers for STR allele comparison.In addition,peripheral blood samples collected from humans with known chromosomal karyotypes,such as 46,XX,46,XY(2 cases for each type)and 47,XY,+21(one case),were used to constitute the control group.Multiplexed fluorescence labeled STR-PCR was employed to test 14 highly hybridized STR loci and 3 non-STR loci used for sex determination,and STR gene polymorphisms were analyzed.Results: 1.Thirteen cases of secondary mosaicisms were detected from 7027 amniotic fluid and 251 villi samples.The detection rates for level Ⅱ mosaicisms were 0.14% and 1.2% in amniotic fluid and villi sampling,respectively.Except 1 case that was not followed up,the remaining 12 cases were followed up and confirmed to be pseudomosaicisms.For the 35 cases of level Ⅲ mosaicism,the detection rates were 0.37% and 3.58% in amniotic fluid and villi sampling,respectively.The 26 level Ⅲ mosaicism cases identified by amniotic fluid sampling were further tested by FISH or umbilical cord blood examination and 24 of these cases were diagnosed as true mosaicism(92.3%).Level Ⅲ mosaicisms were mainly mosaicisms in the sex chromosomes,followed by mosaicisms in chromosomes 21,18,and 13,and translocation mosaicisms,which were the least common.In contrast,in case of level Ⅱ mosaicisms the most common types were translocation mosaicisms and non-21,18,13,X,and Y chromosomal mosaicisms.The percentage of mosaicisms with a proportion difference of ≥20% between FISH and karyotype analysis was 29.6%(8 cases/27 cases).2.Maternal cell contamination was not detected in the 5 samples of the control group,while maternal cell contamination was detected in only 1 sample in the study group.Conclusion:1.The grading method of Hsu plays an important role in distinguishing between true and false mosaicisms in the amniotic fluid and chorionic villus.2.Level Ⅲ mosaicism in amniotic fluid were most true mosaicism.The proportion of Level Ⅲ mosaicism in villi samples was on the high side and there was the possibility of maternal cell contamination.3.True mosaicisms were mainly in the sex chromosomes,and translocation mosaicisms were the least common.In contrast,the most common types of pseudomosaicisms were translocation mosaicisms and non-21,18,13,X,and Y chromosomal mosaicisms.4.FISH results from uncultured cells were more representative of the true situation of mosaicisms in the sample.5.Multiplexed fluorescence labeled STR-PCR is a convenient and effective method for the identification of maternal cell contamination in fetal samples and its detection accuracy can be enhanced by comparison with maternal DNA.6.There was maternal cell contamination in the villi samples with mosaicism.Elimination of maternal cell contamination is recommended for villi samples with karyotypes of 46,XX and abnormal/46,XX.