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Taiwanese Journal of Obstetrics & Gynecology 53 (2014) 129e132

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Taiwanese Journal of Obstetrics & Gynecology

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Research Letter

Prenatal diagnosis and molecular cytogenetic characterization ofmosaicism for a small supernumerary marker chromosome derivedfrom chromosome 15

Chih-Ping Chen a,b,c,d,e, f,*, Ming Chen g,h, i, Yi-Ning Su j, Schu-Rern Chern b, Peih-Shan Wuk,Shun-Ping Chang g,h, Yu-Ling Kuo l, Wen-Lin Chen a, Wayseen Wang b,m

aDepartment of Obstetrics and Gynecology, Mackay Memorial Hospital, Taipei, TaiwanbDepartment of Medical Research, Mackay Memorial Hospital, Taipei, TaiwancDepartment of Biotechnology, Asia University, Taichung, Taiwand School of Chinese Medicine, College of Chinese Medicine, China Medical University, Taichung, Taiwane Institute of Clinical and Community Health Nursing, National Yang-Ming University, Taipei, TaiwanfDepartment of Obstetrics and Gynecology, School of Medicine, National Yang-Ming University, Taipei, TaiwangDepartment of Medical Research, Center for Medical Genetics, Changhua Christian Hospital, Changhua, TaiwanhDepartment of Genomic Medicine, Center for Medical Genetics, Changhua Christian Hospital, Changhua, TaiwaniDepartment of Obstetrics and Gynecology, Changhua Christian Hospital, Changhua, TaiwanjDepartment of Obstetrics and Gynecology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, TaiwankGene Biodesign Co. Ltd, Taipei, TaiwanlDepartment of Obstetrics and Gynecology, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, TaiwanmDepartment of Bioengineering, Tatung University, Taipei, Taiwan

a r t i c l e i n f o

Article history:Accepted 6 December 2013

A 34-year-old, primigravid woman underwent amniocentesis at18 weeks of gestation because of advanced maternal age. Cytoge-netic analysis of the cultured amniocytes revealed mosaicism for asmall supernumerary marker chromosome (sSMC) and a karyotypeof 47,XY,þmar[15]/46,XY[5]. Among 20 colonies of culturedamniocytes, 15 colonies had a karyotype of 47,XY,þmar, while theother five colonies had a karyotype of 46,XY. The parental karyo-types were normal. Level II ultrasound findings were unremark-able. She underwent repeat amniocentesis at 29weeks of gestation.Array comparative genomic hybridization (aCGH) on DNA extractedfrom the uncultured amniocytes obtained from 10 mL of amnioticfluid was performed using NimbleGen ISCA Plus Cytogenetic Array(Roche NimbleGen, Madison, WI, USA). aCGH revealed no genomicimbalance in the pericentromeric euchromatic regions of all 24chromosomes. Cytogenetic analysis of cultured amniocytesrevealed a karyotype of 47,XY,þmar[12]/46,XY[10]. In 12/22

Conflicts of interest: The authors have no conflicts of interest to declare.* Corresponding author: Department of Obstetrics and Gynecology, Mackay

Memorial Hospital, 92, Section 2, Chung-Shan North Road, Taipei, Taiwan.E-mail address: [email protected] (C.-P. Chen).

http://dx.doi.org/10.1016/j.tjog.2013.12.0041028-4559/Copyright � 2014, Taiwan Association of Obstetrics & Gynecology. Published

separated colonies of cultured amniocytes, a karyotype of47,XY,þmar (Fig. 1) was noted, while the other 10 colonies had akaryotype of 46,XY. Fluorescence in situ hybridization (FISH) wasperformed on cultured amniocytes using the probes of AquariusSatellite Enumeration (Cytocell Inc., Adderbury, Oxfordshire, UK)and Vysis Prader-Willi/Angelman Region (Abbott Inc., Abbott Park,IL, USA). The probes used included CEP15 (D15Z4, 15p11.1-q11.1;D15Z1, 15p11.2), CEP1/5/19, CEP6, CEP7, CEP10, CEP18, CEP13/21,and CEP14/22 (Cytocell), and LSI SNRPN (15q11.2) and LSI PML(15q15) (Abbott). FISH analysis revealed that the sSMC was positivefor D15Z4 (Fig. 2) and negative for D15Z1, SNRPN, and PML (Fig. 3).FISH analysis also revealed a negative result for CEP1/5/19, CEP6,CEP7, CEP10, CEP18, CEP13/21, and CEP14/22. The sSMCwas derivedfrom chromosome 15 without involvement of the Prader-Willi/Angelman region. The karyotype at repeat amniocentesis was47,XY,þmar.ish der(15)(D15Z4þ, D15Z1-, SNRPN-, PML-)[12]/46,XY[10]. Methylation analysis of the Prader-Willi/Angelman criticalregion (PWACR) by the methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) kit of SALSA MS-MLPAME028-B1 Prader-Willi syndrome/Angelman syndrome (PWS/AS)probemix (MRC-Holland bv. Amsterdam, The Netherlands) using

by Elsevier Taiwan LLC. All rights reserved.

Fig. 1. The G-banded karyotype of 47,XY,þmar. mar ¼ marker chromosome.

C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 53 (2014) 129e132130

the DNA extracted from uncultured amniocytes excluded unipa-rental disomy (UPD) 15 (Fig. 4). The parents decided to continue thepregnancy. At 40 weeks of gestation, a healthy male baby wasdelivered with a body weight of 3004 g and a karyotype of47,XY,þmar[21]/46,XY[19] in cord blood.

sSMCs occur in 0.075% of prenatal fetal cases and may or maynot be associated with phenotypic abnormalities, depending on theorigin of the chromosome and the presence of euchromatic mate-rials [1e4]. Prenatal diagnosis of sSMCs demands genetic coun-seling and requires molecular cytogenetic techniques to identifythe nature of the sSMC [5e15]. In cases with an sSMC, about 70% arede novo, 70% are derived from acrocentric chromosomes, and 70%present no phenotypic effects [1e4,16]. In a study of 112 patientswith constitutional SMCs ascertained by FISH, Crolla et al [17] foundthat 68% (76/112) were from the acrocentric chromosomes of 13/21,14, 15, and 22, and among these acrocentric SMCs, 51% (39/76) werederived from chromosome 15, indicating a high frequency of 35%(39/112) for SMC(15) in all SMCs with known chromosomal origins.

Fig. 2. Fluorescence in situ hybridization (FISH) using the probe of D15Z4 (15p11.1-q11.1; spectrum green) shows a positive hybridization signal on the marker chromo-some (mar).

Attention should be paid to prenatal diagnosis of an sSMCderived from chromosome 15with regard to the involvement of thePWACR at 15q11-q13, and a differential diagnosis of the inv dup(15)or idic(15) syndrome (tetrasomy 15q), chromosome 15q11-q13duplication syndrome and maternal UPD 15 included. The invdup(15) or idic(15) syndrome (tetrasomy 15q) is caused by an SMCinvolving the inverted duplication of proximal chromosome 15containing the PWACR, and is characterized by muscle hypotonia,developmental delay, intellectual disability, and autistic behavior[18]. Chromosome 15q11-q13 duplication syndrome (OMIM608636) is characterized by clinical features of autism, mentalretardation, ataxia, and epilepsy caused by 15q11q13 duplication[19e26]. Prenatal diagnosis of an sSMC(15) even without

Fig. 3. Fluorescence in situ hybridization (FISH) using the probes of D15Z1 (15p11.2;spectrum green), LSI SNRPN (15q11.2; spectrum red) and LSI PML (15q15; spectrumred) shows absence of the green and red signals on the marker chromosome (mar).

Fig. 4. Methylation analysis using SALSA MS-MLPA ME028-B1 PWS/AS probemix shows no evidence of uniparental disomy (UPD) 15: (A) fetus (uncultured amniocytes); (B)negative control (wild type); (C) positive control [Prader-Willi syndrome (PWS) UPD type]; (D) positive control [Angelman syndrome (AS) UPD type].

C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 53 (2014) 129e132 131

involvement of the PWACR should also test for UPD 15. Liehr et al[27] reported PWS with a karyotype of 47,XY,þmin(15)(pter/q11.1:) and maternal heterodisomic UPD 15, andsuggested the necessity to exclude the presence of a UPD in pre-natal diagnosis of a de novo sSMC.

Human chromosome 15q11-q14 region contains six clusters ofchromosome 15 low-copy repeat (LCR15) duplicons referred to asBP1wBP6, which mediate chromosomal rearrangements leading to

translocations, deletions, duplications, triplications, and sSMCs[28e36]. The clinically relevant sSMC(15) contains euchromatic 15qmaterial, especially the PWACR between BP2 and BP3. The 5.9-Mb15q proximal region between BP2 and BP3 contains the PWACR andincludes the genes ofMKRN3,MAGEL2,NDN, SNRPN, UBE3A, ATP10A,GABRB3, OCA2, and HERC2. In a review of 32 cases with thesSMC(15), Eggermann et al [37] found that SMC(15) with euchro-matic content causes mental and psychomotor retardation,

C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 53 (2014) 129e132132

whereas SMC(15) without euchromatic content has no influence onthe carrier’s phenotype, but is associated with a high incidenceamong infertile males. In a study of 20 patients with clinicallyrelevant SMC(15) with triplicated 15pter to BP3 or15pter/BP4::BP5/pter, Kleefstra et al [36] found consistentphenotypic abnormalities with a high prevalence of autisticbehavior, attention problems, aggressive behavior, anxiety, andsleeping problems in these patients. The present case belongs tothe clinically irrelevant sSMC(15) that contains only heterochro-matin and 15p material.

In conclusion, molecular genetic technologies such as FISH,aCGH, andmethylation analysis are useful for rapid exclusion of theinvolvement of the PWACR and UPD 15 in prenatally detectedmosaic sSMC(15) at amniocentesis.

Acknowledgments

This work was supported by research grants NSC-99-2628-B-195-001-MY3 and NSC-101-2314-B-195-011-MY3 from the Na-tional Science Council and MMH-E-102-04 from Mackay MemorialHospital, Taipei, Taiwan.

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