Genetic Test And Pregnancy Outcome Between Fetuses With Cervical Cystic Hygroma And Fetuses With Increase Nuchal Translucency During Early Pregnancy

Objective 1.To explore the genetic test results,combined structural malformations and postnatal outcomes of increase nuchal translucency（NT）fetuses with NT≥5 mm.2.To explore the genetic test results,combined structural malformations and postnatal outcomes of cystic hygroma（CH）fetuses with NT≥5 mm.3.By comparing the genetic test results,combined structural malformation and postnatal outcomes between increased NT fetuses and CH fetuses,to provide a theoretical basis for the establishment of standard prenatal diagnosis process.Method 1.Patients visiting our center from September 2017 to December 2020 were collected.The Selection criteria is NT measurement was: Chinese pregnant woman with Single pregnancy;performed at 11-13+6 weeks;NT measurement is strictly standardized,the picture quality is high;NT measured value ≥ 5.00 mm;Pregnant women receive prenatal diagnostic testing at our center.Sequential genetic tests including karyotype analysis,chromosome microarray analysis（CMA）and whole exome sequencing analysis（WES）were performed according to the samples.Case grouping: NT ≥5.0 mm in the increased NT group,and no separation was found in the median sagittal plane;In CH group,NT ≥5.0 mm was observed,and the septum was visible in the median sagittal plane.2.Provide adequate prenatal counseling to pregnant women,including preoperative counseling for invasive prenatal diagnosis,pre-test and post-test counseling for various techniques,and sign written informed consent.Pregnant women choose genetic testing independently,and there are three testing strategies: Only chromosome karyotype analysis technology was selected,only CMA test was selected,and chromosome karyotype analysis technology +CMA test was selected.For fetuses with negative genetic tests,those who met the indications for WES and the pregnant woman asked WES to test were further tested.3.Karyotype analysis was performed according to the technical requirements of G-banding chromosome karyotype;Fetal and parental genomic DNA was extracted using QIAGEN DNA extraction kit,and the whole process was extracted according to the standard requirements;The CMA experiment was conducted in strict accordance with the standard procedures provided by Affymetrix,using the Cytoscan HD chip,and the results were interpreted in strict accordance with the ACMG 2019 Guidelines;The center extracts DNA and sends it to the company to complete the experiment,and the company returns the original data.The next generation sequencing analysis team of the center conducts independent data analysis.Finally,the mutation sites were identified by Sanger sequencing technology.4.Clinical data were retrieved from local databases（ultrasound workstation,prenatal diagnosis system,outpatient system,inpatient system）,and clinical data were followed up by telephone for pregnant women who did not visit our hospital.Postnatal follow-up was conducted 1-15 months after the expected date of birth,mainly including pregnancy outcome,obvious appearance abnormalities after birth,relevant imaging examination and results after birth,postnatal growth and development and movement,etc.5.Statistical analysis was performed using SPSS software.P < 0.05 was considered statistically significant.Result 1.A total of 152 patients were enrolled,including 89 in the increased NT group and 63 in the CH group.There were no significant differences in maternal age（P=0.291,Z=-1.057）and fetal CRL（P=0.494,Z=-0.684）between the two groups,but fetal NT thickness（8.0 mm）in CH group was significantly higher than that in increased NT group（6.2 mm）（P < 0.001,Z=-4.876）.2.Of the 63 cases in CH group,there were 35 cases（55.6%）with positive genetic test results,including 31 cases（59.2%）with chromosomal disease,1 case（1.6%）with pathogenic/likely pathogenic CNV,2 case（3.2%）with chimerism,and 1 case（1.6%）with structural abnormality.The first three cases of chromosomal diseases were 45,X（20 cases,31.7%）,trisomy 18（6 cases,9.5%）,and trisomy 21（5 cases,7.9%）.Of the 28 fetuses negative for conventional genetic test,2 fetuses received WES test,all（100%）were detected with disease-related gene mutation.Case C6714 was detected with likely pathogenicity mutation of KCNT1 gene c.1420C>T（p.Arg474Cys）,KCNT1 gene mutation can cause early infant epileptic encephalopathy type 14 and nocturnal frontal lobe epilepsy type 5.Likely pathogenic mutation of SPTAN1 gene was detected in case c.3109G>A（p.E1037K）,SPTAN1 gene mutation can lead to early type 5 epileptic encephalopathy in infants.Genetic test was negative in 26 fetuses（41.6%）,and structural abnormality was found in 6 fetuses（23.1%）by ultrasonography during pregnancy,1 case had multiple malformations.Three structural malformations were detected in CH fetuses,including cardiac malformations（19.2%,5 cases）,thoracic/peritoneal effusion（3.8%,1 case）,and skeletal malformations（3.8%,1 case）.No abnormality was found in the 20 cases by fetal genetic examination and ultrasound screening during pregnancy.The pregnancy outcomes were fetal arrest or stillbirth in 3 cases（15.0%）,induced labor in 7 cases （35.0%）,and pregnancy in 4 cases（20.0%）before the due date of delivery.The remaining 6 cases（30.0%）were born,and 2 cases were abnormal after birth,among which 1 case was diagnosed with Pirot syndrome after birth and died due to dyspnea.Another case was born prematurely due to intrauterine distress and psychomotor retardation.The other 4 fetuses showed no obvious developmental retardation or mental retardation after birth.3.Among 89 cases in NT thickening group,50 cases（56.2%）with positive genetic test results,including 41 cases（46.1%）with chromosomal disease,6 cases（6.7%）with pathogenic/likely pathogenic CNV,2 cases（2.2%）with chimerism,and 1 case（1.1%）with structural abnormality.The first three cases of chromosomal diseases were trisomy 21 syndrome（16 cases,18.0%）,trisomy 18 syndrome（12 cases,13.5%）and 45,X syndrome（10 cases,11.2%）.Of the 39 fetuses（43.8%）negative for conventional genetic testing,4 fetuses were tested for WES and 2 fetuses（50.0%）were detected with disease-related gene mutations.FOXC2 gene mutationc.122₁23ins GACA（p.Tyr41Ter）was detected in case C6512,FOXC2 gene mutation can lead to lymphedema-double eyelash syndrome;LZTR1 gene mutation was detected in case C6488,c.741C>A（p.Ser247Arg）and c.1349G>A（p.Gly450Asp）,LZTR1 gene can cause Noonan syndrome and schwannomatosis.Among the 37 cases（41.6%）with negative genetic test,11 cases（29.7%）were found with structural abnormality by ultrasonography during pregnancy,include 4 had multiple malformations.Among all ultrasonic abnormalities,the top three were cardiac malformation（6 cases,16.2%）,thoracic/peritoneal effusion（4 cases,10.8%）and skeletal malformation（3 cases,8.1%）.Genetic tests and ultrasound screening during pregnancy showed no abnormalities in 26 cases,the results of pregnancy were as follows: 1 miscarriage（3.8%）,3 induced labor（11.5%）,9 pregnancies（34.6%）before the expected date of delivery,and the remaining 13（50.0%）had no abnormal results in postnatal clinical follow-up.4.Statistical analysis showed that there was no significant difference in the incidence of fetal routine genetic testing between the two groups(P= 0.813,X2= 0.025 for karyotype analysis;When choosing CMA,P= 0.9（21）（24）（11）X（17）（28）（15）（13）（15）（15）（17）（8）（26）A total of 4 cases in the increased NT group and 2 cases with positive results（50%）were selected for WES test,and 2 cases with positive results（100%）were selected for WES test in the CH group.However,due to the small sample size of WES,the statistical results may be biased.When the genetic test was negative,there was no significant difference in the probability of fetuses with ultrasonic structural malformations between the two groups（P= 0.558,X2= 0.343）.When chromosomal and sonographic abnormalities were excluded,there was a statistically significant difference between increased NT and adverse pregnancy outcomes in CH fetuses（7.1% VS 55.6%,P=0.018）.Conclusion 1.For CH fetuses with NT≥5.0mm and fetuses with NT thickening,the risk of chromosomal abnormalities and structural malformation were significantly increased.In clinical practice,invasive prenatal diagnosis and comprehensive ultrasonography should be recommended for fetal cases with NT≥5.0mm.2.In the process of prenatal genetic diagnosis of CH fetuses with NT≥5.0mm and Increased NT fetuses,karyotype analysis or karyotype analysis + CMA should be carried out first,and chromosomal diseases and genomic diseases should be excluded.Further WES testing can be performed for CH fetuses with normal karyotype analysis and/or genomic disease detection,or for Increased NT fetuses with abnormal sonographic structure.3.The most common chromosomal abnormalities in CH fetuses were 45,X;the most common chromosomal abnormality in fetus with NT thickening was trisomy 21 syndrome.4.The most common congenital structural abnormalities associated with CH fetuses with NT≥5.0mm and Increased NT fetuses were abnormalities of the cardiovascular system,thoracic/abdominal effusion,and skeletal system.5.Under the premise of no abnormal results of a series of genetic tests and no abnormal findings of prenatal ultrasound scan,the probability of good clinical prognosis was about 92.9% for fetuses with NT thickening（NT≥5mm）,and 44.4% for fetuses with CH.The clinical prognosis of fetuses with NT thickening was better than that of fetuses with CH.