Letter to the Editor
Rapid diagnosis of trisomy 18 using uncultured amniocytes in late second
trimester in a pregnancy with fetal congenital heart defects, arthrogryposis,
omphalocele and mega cisterna magna
Chih-Ping Chen a,b,c,d,e,f,g*, Yi-Yung Chen a, Jui-Der Liou h,i, Schu-Rern Chern c, Peih-Shan Wu j, Jun-Wei Su a,k, Meng-Shan Lee a and Wayseen Wang b,l
a Department of Obstetrics and Gynecology, Mackay Memorial Hospital, Taipei, Taiwan b Department of Medical Research, Mackay Memorial Hospital, Taipei, Taiwan
c Department of Medicine, Mackay Medical College, New Taipei City, Taiwan d Department of Biotechnology, Asia University, Taichung, Taiwan
e School of Chinese Medicine, College of Chinese Medicine, China Medical University, Taichung, Taiwan f Institute of Clinical and Community Health Nursing, National Yang-Ming University, Taipei, Taiwan g Department of Obstetrics and Gynecology, School of Medicine, National Yang-Ming University, Taipei,
Taiwan
h College of Medicine, Chang Gung University, Taoyuan, Taiwan
i Department of Obstetrics and Gynecology, Chang Gung Memorial Hospital, Taipei, Taiwan j Gene Biodesign Co. Ltd, Taipei, Taiwan
k Department of Obstetrics and Gynecology, China Medical University Hospital, Taichung, Taiwan l Department of Bioengineering, Tatung University, Taipei, Taiwan
* Correspondence to: Chih-Ping Chen, MD
Department of Obstetrics and Gynecology, Mackay Memorial Hospital 92, Section 2, Chung-Shan North Road, Taipei, Taiwan
Tel: +886-2-25433535; Fax: +886-2-25433642, +886-2-25232448 E-mail: [email protected]
A 33-year-old, gravida 3, para 2, woman was referred to the hospital for genetic counseling at 26 weeks of gestation because of fetal anomalies. Prenatal ultrasound revealed intrauterine growth restriction, an atrioventricular septal defect, omphalocele, arthrogryposis of bilateral hands and mega cisterna magna (Fig. 1). The fetal biometry was equivalent to 24 weeks. About 33 mL amniotic fluid was aspirated, of which 10 mL was used for array comparative genomic hybridization (aCGH) analysis using uncultured amniocytes, 5 mL for quantitative fluorescent polymerase chain reaction (QF-PCR) analysis using uncultured amniocytes, and 16 mL for conventional cytogenetic analysis using cultured amniocytes. The aCGH investigation using whole-genome ISCA Plus Cytogenetic array (Roche NimbleGen, Madison, WI, USA) on uncultured amniocytes showed the result of trisomy 18 [arr cgh 18p11.32q23 (1 – 7,810,771,248)3] (Fig. 2). QF-PCR assays revealed a 1:2 ratio of two parental alleles in chromosome 18-specific polymorphic DNA markers. The result was consistent with a homologous duplication of chromosome 18 and the diagnosis of trisomy 18 (Fig. 3). Conventional cytogenetic analysis using cultured amniocytes revealed a karyotype of 47,XY,+18 (Fig. 4). The pregnancy was continued to 32 weeks of gestation, and a dead malformed fetus was delivered with facial dysmorphisms, omphalocele and arthrogryposis of the hands.
Molecular cytogenetic technologies such as interphase fluorescence in situ hybridization (FISH), multiplex ligation-dependent probe amplification (MLPA), aCGH and QF-PCR for rapid aneuploidy diagnosis (RAD) without the need of cell cultures have been well described [1-5]. In the present case, application of aCGH and QF-PCR on uncultured amniocytes has replaced cord blood sampling for RAD. Amniocentesis in late gestation is easier and less invasive than cord blood sampling. The present case shows that aCGH and QF-PCR using uncultured amniocytes are useful alternatives to cordocentesis for RAD in late gestation.
The present case manifested an atrioventricular septal defect, arthrogryposis, omphalocele, mega cistern magna on prenatal ultrasound. Congenital heart defects (CHD), omphalocele, arthrogryposis and mega cistern magna have been associated with trisomy 18 [6-9].
Snijders et al [10] observed CHD in 52% of the fetuses with trisomy 18 (n = 137). Chaoui et al [11] found chromosomal abnormalities in 22.7% (46/203) of the fetuses with CHD including trisomy 18 (n = 15), trisomy 21 (n = 13), trisomy 13 (n = 5), 45,X (n = 5), triploidy (n = 2) and
associated with a high risk of aneuploidy up to 49.1%-57.9% [12-13]. Delisle et al [12] reported aneuploidy in 57.9% (22/38) of the fetuses with an atrioventricular septal defect including trisomy 21 (n = 19), trisomy 18 (n = 1), trisomy 13 (n = 1) and mosaic trisomy 19q (n = 1). Huggon et al [13] reported aneuploidy in 49.1% (107/218) of the fetuses with an atrioventricular septal defect including trisomy 21 (n = 86), trisomy 18 (n = 13), trisomy 13 (n = 4) and other rearrangements (n = 4).
Mega cisterna magna has been observed in 6.7% (6/89) of the fetuses with trisomy 18 [6]. Chen [6] observed central nervous system (CNS) anomalies in 22.5% (20/89) of the fetuses with trisomy 18 including holoprosencephaly (n = 5), neural tube defects (n = 2), hydrocephalus (n = 2), Dandy-Walker malformation (n = 5) and mega cisterna magna (n = 6).
Chen [14] reported aneuploidy in 36.1% (415/1148) of the fetuses with prenatally detected omphalocele. Among the 415 fetuses with omphalocele and aneuploidy, 66.7% (277/415) had trisomy 18, 17.3% (72/415) had trisomy 13 and 6.3% (26/415) had trisomy 21. Snijders et al [10] observed omphalocele in 31% of the fetuses with trisomy 18 (n = 137). Chen [7] observed omphalocele in 13.5% of the fetuses with trisomy 18 (n = 89).
Trisomy 18 has been associated with arthrogryposis, pre-axial upper limb reduction, overlapping fingers, rocker-bottom feet and talipes. Snijders et al [10] observed abnormalities of the hands and feet in 72% of the fetuses with trisomy 18 (n = 137). Chen [8] observed arthrogryposis in 25.8% (23/89) of the fetuses with trisomy 18. In his observation, the wrist was more frequent than the ankle, and the left side was more frequent than the right side in cases with asymmetry.
In conclusion, prenatal diagnosis of CHD, arthrogryposis, omphalocele and mega cisterna magna should raise a suspicion of aneuploidy. We suggest that aCGH and QF-PCR using uncultured amniocytes are useful for RAD in pregnancy with fetal anomalies detected in late gestation.
Acknowledgements
This work was supported by research grant NSC-99-2628-B-195-001-MY3 from the National Science Council and MMH-E-101-04 from Mackay Memorial Hospital, Taipei, Taiwan.
References
1. Chen C-P, Su Y-N, Tsai F-J, Chern S-R, Hsu C-Y, Huang M-C, et al. Rapid genome-wide aneuploidy diagnosis using uncultured amniocytes and array comparative genomic hybridization in pregnancy with abnormal ultrasound findings detected in late second and third trimesters. Taiwan J Obstet Gynecol 2010; 49: 120-3.
2. Chen C-P, Su Y-N, Wu P-C, Lee C-C, Pan C-W, Wang W. Rapid aneuploidy diagnosis by array comparative genomic hybridization using uncultured amniocytes in a pregnancy with fetal nuchal edema and mild ascites. J Med Ultrasound 2011; 19: 64-9.
3. Chen C-P, Su Y-N, Lin S-Y, Chang C-L, Wang Y-L, Huang J-P, et al. Rapid aneuploidy diagnosis by multiplex ligation-dependent probe amplification and array comparative genomic hybridization in pregnancy with major congenital malformations. Taiwan J Obstet Gynecol 2011; 50: 85-94.
4. Chen C-P, Hsu C-Y, Chern S-R, Wu P-S, Su J-W, Lee C-C, et al. Rapid diagnosis of monosomy X using uncultured amniocytes in amniotic fluid and cultured lymphocytes in cystic fluid in a pregnancy with fetal cystic hygroma and hydrops. J Med Ultrasound 2012; 20: 129-32.
5. Chen C-P, Su Y-N, Chang C-L, Chen Y-Y, Su J-W, Chern S-R, et al. Rapid aneuploidy diagnosis by multiplex ligation-dependent probe amplification using uncultured amniocytes in pregnancy with major fetal structural abnormalities. Taiwan J Obstet Gynecol 2012; 51: 123-8.
6. Chen C-P. Central nervous system anomalies associated with fetal trisomy 18. Prenat Diagn 2005; 25: 419-21.
7. Chen C-P. Omphalocele and congenital diaphragmatic hernia associated with fetal trisomy 18. Prenat Diagn 2005; 25: 421-3.
8. Chen C-P. Arthrogryposis of the wrist and ankle associated with fetal trisomy 18. Prenat Diagn 2005; 25: 423-5.
9. Chen C-P. Prenatal sonographic features of the fetuses in trisomy 13 pregnancies (III). Taiwan J Obstet Gynecol 2009; 48: 342-9.
10.Snijders RJM, Farrias M, von Kaisenberg C, Nicolaides KH. Fetal abnormalities. In: Snijders RJM, Nicolaides KH, eds. Ultrasound Markers for Fetal Chromosomal Defects. New York: Parthenon Publishing Group, 1996; 1-62.
11.Chaoui R, Körner H, Bommer C, Göldner B, Bierlich A, Bollmann R. Prenatal diagnosis of heart defects and associated chromosomal aberrations. Ultraschall Med 1999; 20: 177-84. [German].
12.Delisle MF, Sandor GG, Tessier F, Farquharson DF. Outcome of fetuses diagnosed with atrioventricular septal defect. Obstet Gynecol 1999; 94: 763-7.
13.Huggon IC, Cook AC, Smeeton NC, Magee AG, Sharland GK. Atrioventricular septal defects diagnosed in fetal life: associated cardiac and extra-cardiac abnormalities and outcome. J Am Coll Cardiol 2000; 36: 593-601.
14.Chen C-P. Chromosomal abnormalities associated with omphalocele. Taiwan J Obstet Gynecol 2007; 46: 1-8.
Figure Legends
Fig. 1. Prenatal ultrasound at 26 weeks of gestation shows (A) an atrioventricular septal defect, (B) an omphalocele, (C) arthrogryposis of the wrist and (D) mega cisterna magna.
Fig. 2. (A) Whole genome view and (B) chromosomal view of array comparative genomic hybridization analysis on uncultured amniocytes show a duplication of chromosome 18 (arrows), consistent with the diagnosis of trisomy 18.
Fig. 3. Representative electrophoretograms of quantitative fluorescent polymerase chain reaction assays at short tandem repeat markers specific for chromosome 18q. Two peaks (144 bp: 152 bp) at D18S535 and two peaks (186 bp: 166 bp) at D18S878 of equal fluorescence activity from different parental alleles at a ratio of 1:2 on uncultured amniocytes indicate a homologous duplication of chromosome 18. The findings are consistent with the diagnosis of trisomy 18 resulting from meiosis II non-disjunction or post-zygotic mitotic non-disjunction.
