Arq Bras CardiolabcArquivos Brasileiros de CardiologiaArq. Bras. Cardiol.0066-782X1678-4170Sociedade Brasileira de Cardiologia - SBChmxyfz4mS6rHqMJb47NtZMpS0066-782X2018001600571S0066-782X201800501110110.5935/abc.20180144Original ArticleGenetic Testing and Pregnancy Outcome Analysis of 362 Fetuses with Congenital Heart Disease Identified by Prenatal UltrasoundLuoShiyuMengDahuaLiQifeiHuXuehuaChenYuhuaHeChunXieBoboSheShangyangLiYingfengFuChunyunMaternal and Child Health Hospital of Guangxi Zhuang Autonomous RegionGuangxiChinaMaternal and Child Health Hospital of Guangxi Zhuang Autonomous Region, Guangxi - ChinaMailing Address: Chunyun Fu, N°.59 Xiangzhu Road, Nanning, Guangxi - China. E-mail: fuchunyun2008@sina.com, 948178849@qq.com
Author contributions
Conception and design of the research: Fu C; Acquisition of data: Meng D, Hu X, Xie B; Analysis and interpretation of the data: Luo S, Li,Q, Chen Y, He C, Xie B, She S, Li Y; Statistical analysis: Meng D, Chen Y, He C; Obtaining financing: Meng D; Writing of the manuscript: Luo S, Li,Q; Critical revision of the manuscript for intellectual content: She S, Li Y, Fu C.
Potential Conflict of Interest
No potential conflict of interest relevant to this article was reported.
200820181020181114571577160720171104201811042018This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.AbstractBackground:
Congenital heart defects (CHD), as the most common congenital anomaly, have been reported to be associated with chromosomal abnormalities. Currently, patients with CHD are routinely offered karyotyping and chromosomal microarray (CMA) testing, but the genotype-phenotype relationship has not yet been fully established.
Objective:
To determine the type and frequency of chromosomal abnormalities in fetuses with CHD and to analyze pregnancy outcomes of fetuses with heart abnormalities caused by different genetic factors.
Methods:
A total of 362 cases of CHD were enrolled from 2009 to 2016. Detailed ultrasound and laboratory examinations, including karyotyping and CMA, were performed. Outcome was obtained from discharge summaries.
Results:
Of the 362 fetuses, 220 were found with an isolated CHD, and 142 had CHD with extracardiac anomaly. Among these 362 fetuses, 140 were identified with a genetic cause, including 111 cases with aneuploidy, 10 cases with abnormality of chromosomal structure by karyotyping and 19 cases with pathogenic or likely pathogenic copy-number variations (CNVs) by CMA. The detection rate is close to 38.7%. Only one (identified as trisomy 18 syndrome) in 140 positive cases resulted in perinatal death, with the others being induced. The remaining 222 cases had negative results for both genetic testing and of these cases, 56 resulted in induced labor, and 77 had natural childbirth or caesarean births. The pregnancy outcome of the remaining 89 cases was uncertain.
Conclusions:
Karyotyping and CMA are effective and accurate prenatal genetic techniques for identifying fetal chromosomal abnormalities associated with cardiac defects, and this can assist clinical doctors to perform appropriate genetic counselling with regard to the etiology and outcome of CHD.
Keywords:Heart Defects, CongenitalChromosome DisordersSpectral KaryotypingPregnancyFetusUltrasonographyResearch and Development Project of Guangxi Medical and Health TechnologyS201309_04S201611Guangxi Natural Science Foundation Program2016GXNSFCA380001Science and Technology project of Guangxi Zhuang Autonomous Region14124004-1-8This study was partly funded by the Research and Development Project of Guangxi Medical and Health Technology (S201309_04 and S201611), Guangxi Natural Science Foundation Program (2016GXNSFCA380001) and the Science and Technology project of Guangxi Zhuang Autonomous Region (gui-ke-gong 14124004-1-8).Introduction
Congenital heart disease (CHD), one of the most common birth defects, affecting approximately 1 in 100 live births.1-3 With the availability of advanced surgical techniques, normal or near normal cardiac function can be restored after surgical treatment of most types of CHDs ranging from simple ventricular septal defects (VSD) to more complex cardiovascular abnormalities. However, the long-term prognosis of a small, but significant number of CHD fetuses is usually complicated by severe extracardiac abnormalities, such as developmental delay and mental retardation. There is increasing evidence that genetic factors influence the development of most types of CHD,4-6 but the precise genetic basis of most CHD cases remains not fully understood. Current ultrasound technologies are able to detect most of CHD. However, it is difficult for physicians to make a comprehensive assessment of fetuses with CHD merely based on the evidence of prenatal ultrasound, as well as to manage the course of established pregnancy.7 Therefore, genetic testing is now highly recommended for fetuses with CHD.
Karyotyping has been the mainstream diagnostic method for detecting chromosomal abnormalities associated with CHD.8 For CHD cases in prenatal diagnosis, chromosomal anomalies are estimated to be as high as 22.9,10 Now, chromosomal microarray (CMA) has become the first tier technique in fetal structural anomalies detected by ultrasonography.11,12 The advent of CMA technology has allowed genome-wide searches of submicroscopic chromosomal deletions or duplications in the genome, known as copy-number variations (CNVs). CNV is a form of structural variation in the genome: specifically, it is a type of duplication or deletion that has an influence in the base pairs,13 and CNVs play an important role in generating necessary variation in the population and disease phenotypes.14 Recent studies have shown that a substantial proportion of CHD patients were detected with pathogenic CNVs,15,16 and the syndromic or isolated CHD patients were found with multiple recurrent CNV loci, such as 22q11.2 (the DiGeorge syndrome region), 7q11.23, 8p23.1, 9q34.3, and 1q21.1.17-19
At present, only a few studies have reported genetic testing among large groups of fetuses with CHD in China,20-24 the genotype-phenotype relationship has not yet been fully established. The Laboratory of Genetics and Metabolism from the Maternal and Child Health Hospital in Guangxi is one of the largest Perinatal Diagnostic centers in South China. This study aimed to analyze the chromosomal abnormalities and pregnancy outcomes in 362 fetuses with CHD.
MethodsSubjects
Fetal ultrasound anatomy scans were routinely performed for pregnant women at the Prenatal Diagnosis Center of Guangxi Zhuang Autonomous Region in China. The anatomy scans were conducted between 20 and 28 weeks of gestation by senior sonographers using GE E8 ultrasound machines (General Electric Healthcare, USA). If CHD was suspected, the echocardiography was subsequently performed for confirmation.
A total of 8,430 pregnancies between June 2012 and June 2016 were screened for fetal cardiac defects, and 362 fetuses were identified with CHD. The Medical Ethics Committee of the Guangxi Maternal and Child Health Hospital approved the study protocol (Approval no.160220), and the parents of all selected fetuses with CHD gave their written consent.
Testing of SNP microarray
All samples of amniotic fluid or fetal cord blood were collected from the pregnant women, and genomic DNA was extracted using the QIAamp DNA Blood Mini Kit (Qiagen, Germany) according to the manufacturer's protocol. SNP (Single Nucleotide Polymorphism) microarray testing was performed using Illumina HumanCytoSNP-12 v2.1 BeadChip (Illumina, USA). The laboratory policy at the time of testing was not to report well-established polymorphisms, CNVs that do not contain genes and CNVs smaller than 0.20 Mb. However, stretches of homozygosity larger than 10 Mb were reported.
Karyotyping
All samples of amniotic fluid or fetal cord blood were used to perform G-banding according to the standard procedure as described previously.25
ResultsClinical data
Among the 8,430 pregnancies, 362 cases of CHD were diagnosed using fetal echocardiography, for a frequency of 4.2%. The mean age of the pregnant women was 31.1 ± 5.1 years, and the mean gestational week at diagnosis was 24.4 ± 3.8 weeks.
The 5 most common types of CHD were, in order, ventricular septal defect (51.9%, 188/362), persistent left superior vena cava (13.0%, 47/362), endocardial cushion defects (0.9%, 33/362), single umbilical artery (0.9%, 32/362) and right-sided aortic arch (0.8%, 29/362).
Etiology
In total, 362 fetuses were diagnosed with CHD. The genetic tests found 111 cases with aneuploidy, 10 cases with abnormality of chromosome structure, and 19 cases with pathogenic or likely pathogenic CNVs (Table 1). The remaining 222 cases showed no abnormal genetic findings. The abnormalities of chromosome numbers consisted of trisomy 18 syndrome (61 cases), trisomy 21 syndrome (31 cases) and trisomy 13 syndrome (19 cases). CMA identified 19 CNVs, including DiGeorge syndrome (8 cases), Jacobsen syndrome (2 cases), Angelman/Prader-Willi syndrome (1 case), 16p11.2-p12.2 microdeletion syndrome (1 case), 16q24-triplication syndrome (1 case), Thrombocytopenia-absent radius (TAR) syndrome (1 case), 3q29 microduplication syndrome (1 case), 22q11 duplication syndrome (1 case), Cri du chat syndrome (1 case) and 2 likely pathogenic CNVs (Table 2).
Genetic testing of 362 fetuses with congenital heart defects
Of the 362 CHD, 181 fetuses were found with single cardiac malformations, and 181 were found with multiple cardiac abnormalities; 220 were found with an isolated CHD; and 142 had CHD with extracardiac anomaly. Table 3 lists the etiology of the various types of fetal cardiac malformations observed.
Genetic detection in different categories of fetuses with congenital heart disease (CHD)
Classification of CHD
Aneuploidy
Abnormality of chromosome structure
CNVs
Other
Single cardiac malformation (n = 181)
40
4
9
128
Multiple cardiac abnormalities (n = 181)
71
6
10
94
Isolated CHD (n = 220)
26
8
14
172
CHD with extracardiac anomaly (n = 142)
85
2
5
50
CNVs: copy-number variations.
Pregnancy Outcomes
Among all 140 cases with a positive genetic testing result, only one woman chose to continue her pregnancy, and the rest of them chose to induce labor. The fetus was diagnosed with trisomy 18 syndrome, presenting difficulties in feeding, and died 4 days after birth. Among the remaining 222 negative cases, 56 were subjected to labor induction, and most of these cases were deemed incurable or had poor prognostic cardiac malformations (including single ventricle, left or right ventricular dysplasia and tetralogy of fallot) or were complicated with extracardiac anomalies (Figure 1).
The patient pathway in the current study. PND: perinatal deaths. CMA: chromosomal microarray; CNVs: copy-number variations; CHD: congenital heart defects.
Mothers of 77 fetuses with mild or curable cardiac malformations chose to maintain their pregnancies. Of these cases, 66 were found with no abnormality after birth, 8 cases needed surgery, one presented delayed development, one was found with clubfoot, one was identified with hypomyotonia, and the pregnancy outcomes of the remaining 89 cases were uncertain (Figure 1).
Discussion
In this study, 362 cases of fetal CHD were identified in a total of 8,430 pregnancies at a single Maternal and Children's hospital from the Southern region of China from June 2012 to June 2016, with an incidence of 4.2%. This incidence was similar to that reported in Xi'an, in Northwestern China,26 and higher than the rate of 2.3% reported in Guangzhou, In southern China.23 Among the 362 CHD fetuses, ventricular septal defect (51.9%, 188/362) and persistent left superior vena cava (13.0%, 47/362) were the most prevalent cardiac abnormalities detected by ultrasound scans.
Many factors such as genetic factors (including chromosomal abnormalities and gene mutations) and risk factors associated with mothers (including the rubella virus, other infections, radiation, drug use and environmental pollution) are reported to be associated with CHD.5-7,27-29 However, the causes of most types of CHD are still poorly understood. In our study, 140 of 362 CHD fetuses were identified with clinically significant chromosomal abnormalities by karyotyping and CMA, with a detection rate of up to 38.7%. The positive rates of genetic testing in this study is far higher than previous reports in Chongqing, China24 and the Netherlands.30 This rate is similar to that of Brazilians.31
Among the 140 chromosomal abnormalities, 111 (79.3%) were aneuploidy, of which trisomy 18 was the most common; 10 (7.1%) cases were abnormality of chromosome structure; and 19 (13.6%) cases were pathogenic or likely pathogenic CNVs. It is suggested that aneuploidy is the leading genetic cause of fetuses with CHD in our population. Given that G-banding can only reliably detect structural abnormalities > 10 Mb in size, 11 pathogenic CNVs may be missed by karyotyping but detected by CMA. On this basis, we estimate that the incremental yield of reportable CNVs with less than 10 Mb achieved by CMA was 3.0%.
Complex multiple cardiac malformations have poor prognosis and heavily affect the quality of life of surviving infants, but cases such as mild tetralogy of fallot have a reasonable outcome after surgery, as well as a good prognosis. In our study, ultrasonic results of some fetuses with CHD caused by aneuploidy only displayed mild cardiac malformations, although complex CHD combined with extra cardiac defects were more common in these cases. Besides, some symptoms such as mental disability cannot be found by prenatal ultrasound. In these cases, the results of genetic testing is of great importance, because this situation is easily ignored by patients and clinicians. However, several negative cases featured complex CHD and extra cardiac defects after karyotype and CMA testing, and these cases provide an important clue for the study of other factors that lead to CHD.
Several limitations should be considered in the study when reviewing these findings. Firstly, a comprehensive analysis of all known CHD associated genes was not carried out. Secondly, the inheritance of CNVs in some cases with likely pathogenicity was not identified.
Conclusion
Karyotyping and CMA analysis was conducted in 362 CHD fetuses, and it was found that 38.7% of CHD fetuses had a positive genetic testing result. Aneuploidy is the major cause of CHD fetuses in our population. The combination of ultrasonic detection and genetic testing can effectively diagnose fetuses with cardiac malformations and extra cardiac defects, thus providing valuable information to the clinician and patients.
Sources of Funding
This study was partly funded by the Research and Development Project of Guangxi Medical and Health Technology (S201309_04 and S201611), Guangxi Natural Science Foundation Program (2016GXNSFCA380001) and the Science and Technology project of Guangxi Zhuang Autonomous Region (gui-ke-gong 14124004-1-8).
Study Association
This study is not associated with any thesis or dissertation work.
Ethics approval and consent to participate
This study was approved by the Ethics Committee of the Guangxi Maternal and Child Health Hospital under the protocol number 160220. All the procedures in this study were in accordance with the 1975 Helsinki Declaration, updated in 2013. Informed consent was obtained from all participants included in the study.
Acknowledgements
The authors would like to thank the families for their participation in this study.
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Pediatr Cardiol. 2010;31(8):1146-50.BelluccoFTBelangeroSIFarahLMMachadoMVCruzAPLopesLMInvestigating 22q11.2 deletion and other chromosomal aberrations in fetuses with heart defects detected by prenatal echocardiographyPediatr Cardiol.201031811461150Artigo OriginalAnálise de Teste Genético e de Desfecho da Gestação de 362 Fetos com Cardiopatia Congênita Identificados por Ultrassom Pré-NatalLuoShiyu1MengDahua1LiQifei1HuXuehua1ChenYuhua1HeChun1XieBobo1SheShangyang1LiYingfeng1FuChunyun1Maternal and Child Health Hospital of Guangxi Zhuang Autonomous Region, Guangxi - ChinaCorrespondência: Chunyun Fu, N°. 59 Xiangzhu Road, Nanning, Guangxi - China. E-mail: fuchunyun2008@sina.com, 948178849@qq.com
Contribuição dos autores
Concepção e desenho da pesquisa: Fu C; Obtenção de dados: Meng D, Hu X, Xie B; Análise e interpretação dos dados: Luo S, Li,Q, Chen Y, He C, Xie B, She S, Li Y; Análise estatística: Meng D, Chen Y, He C; Obtenção de financiamento: Meng D; Redação do manuscrito: Luo S, Li,Q; Revisão crítica do manuscrito quanto ao conteúdo intelectual importante: She S, Li Y, Fu C.
Potencial conflito de interesses
Declaro não haver conflito de interesses pertinentes.
ResumoFundamento:
As cardiopatias congênitas (CCs) são as anomalias congênitas mais comuns, e têm sido associadas a anormalidades cromossômicas. Atualmente, a cariotipagem e a análise cromossômica por microarray (CMA) são oferecidas rotineiramente aos pacientes, mas a relação genótipo-fenótipo ainda não foi totalmente estabelecida.
Objetivo:
Determinar o tipo e a frequência das anomalias cromossômicas em fetos com CC e analisar os desfechos da gestação de fetos com anormalidades cardíacas causadas por diferentes fatores genéticos.
Métodos:
No total, foram admitidos 362 casos de CC entre 2009 e 2016. Ultrassonografia e exames laboratoriais detalhados foram realizados, incluindo cariotipagem e CMA. O resultado foi obtido a partir das folhas de epicrise.
Resultados:
Dos 362 fetos, 220 apresentaram doença coronariana isolada e 142 apresentaram doença coronariana com anomalia extracardíaca. Entre esses 362 fetos, foram identificados 140 com causa genética, incluindo 111 casos com aneuploidia, 10 casos com anormalidade da estrutura cromossômica por cariotipagem e 19 casos com variações no número de cópias (CNVs) patogênicas ou provavelmente patogênicas por CMA. A taxa de detecção é de aproximadamente 38,7%. Apenas um (identificado como síndrome da trissomia do cromossomo 18) em 140 casos positivos resultou em morte perinatal, com as demais sendo induzidas. Os 222 casos restantes tiveram resultados negativos para ambos os testes genéticos e, destes, 56 resultaram em trabalho de parto induzido e 77 tiveram partos naturais ou cesarianas. O desfecho da gravidez dos 89 casos restantes foi incerto.
Conclusões:
A cariotipagem e a CMA são técnicas genéticas pré-natais eficazes e precisas para a identificação de anomalias cromossômicas fetais associadas a defeitos cardíacos, e isso pode ajudar os médicos a realizar aconselhamento genético adequado com relação à etiologia e ao desfecho das cardiopatias congênitas.
Palavras-chave:Cardiopatias CongênitasTranstornos CromossômicosCariotipagem EspectralGravidezFetoUltrassonografiaProjeto de Pesquisa e Desenvolvimento da Tecnologia Médica e de Saúde de GuangxiS201309_04S201611Programa da Fundação de Ciências Naturais de Guangxi2016GXNSFCA380001projeto de Ciência e Tecnologia da Região Autônoma de Guangxi Zhuang14124004-1-8O presente estudo não parcialmente financiado pelo Projeto de Pesquisa e Desenvolvimento da Tecnologia Médica e de Saúde de Guangxi (S201309_04 e S201611), Programa da Fundação de Ciências Naturais de Guangxi (2016GXNSFCA380001) e projeto de Ciência e Tecnologia da Região Autônoma de Guangxi Zhuang (gui-ke-gong 14124004-1-8).Introdução
A cardiopatia congênita (DCC) é um dos defeitos congênitos mais comuns, afetando aproximadamente 1 em cada 100 nascidos vivos.1-3 Com a disponibilidade de técnicas cirúrgicas avançadas, é possível restaurar a função cardíaca normal ou quase normal após tratamento cirúrgico da maioria dos tipos de CC, de simples comunicações interventriculares (VSD) a anormalidades cardiovasculares mais complexas. No entanto, o prognóstico a longo prazo de um número pequeno, mas significativo, de fetos com CC costuma ser complicado por anormalidades extracardíacas graves, como atraso no desenvolvimento e deficiência mental. Há cada vez mais evidências de que fatores genéticos influenciam o desenvolvimento da maioria dos tipos de CC,4-6 mas a base genética precisa da maioria dos casos de CC ainda não é totalmente compreendida. As tecnologias atuais de ultrassom são capazes de detectar a maioria das CCs, mas é difícil para os médicos fazerem uma avaliação abrangente dos fetos com CC com base apenas na evidência da ultrassonografia pré-natal, também dificultando o gerenciamento do curso da gravidez já estabelecida.7 Por isso, o teste genético agora é altamente recomendado para fetos com CC.
A cariotipagem tem sido o principal método de diagnóstico para detectar anormalidades cromossômicas associadas a CCs.8 Nos casos de diagnóstico pré-natal de CC, estima-se que existam até 22 anomalias cromossômicas.9,10 Hoje, a análise cromossômica por microarray (CMA) tornou-se a primeira escolha em casos de anomalias estruturais fetais detectadas por ultrassonografia.11,12 O advento da tecnologia da CMA permitiu a busca por deleções ou duplicações cromossômicas submicroscópicas em todo o genoma, conhecidas como variações do número de cópias (CNVs). A CNV é uma forma de variação estrutural no genoma: especificamente, é um tipo de duplicação ou deleção que influencia nos pares de bases,13 desempenhando também um papel importante na geração de variações necessárias nos fenótipos de populações e de doenças.14 Estudos recentes demonstraram que foram detectadas CNVs patogênicas em uma proporção considerável dos pacientes com CC,15,16 e foram encontrados múltiplos locos recorrentes de CNV em pacientes com CCs sindrômicas ou isoladas, como 22q11.2 (região da síndrome de DiGeorge), 7q11.23, 8p23.1, 9q34.3, e 1q21.1.17-19
Até o momento, apenas alguns estudos relataram testes genéticos em grandes grupos de fetos com CC na China,20-24 e a relação genótipo-fenótipo ainda não foi totalmente estabelecida. O Laboratório de Genética e Metabolismo do Hospital de Saúde Materna e Infantil de Guangxi é um dos maiores centros de diagnóstico perinatal no sul da China. Este estudo teve como objetivo analisar as anormalidades cromossômicas e os desfechos da gravidez em 362 fetos com CC.
MétodosSujeitos
Exames de anatomia fetal por ultrassonografia foram realizados rotineiramente em gestantes no Centro de Diagnóstico Pré-natal da Região Autônoma de Guangxi Zhuang, na China. Os exames de anatomia foram realizados entre 20 e 28 semanas de gestação por ultrassonografistas experientes, utilizando máquinas de ultrassom GE E8 (General Electric Healthcare, EUA). Em caso de suspeita de CC, o ecocardiograma era realizado posteriormente para confirmação.
Foram feitos testes para a identificação de defeitos cardíacos fetais em um total de 8.430 gestações entre junho de 2012 e junho de 2016, e 362 fetos com CC foram identificados. O Comitê de Ética Médica do Hospital de Saúde Materno-Infantil de Guangxi aprovou o protocolo do estudo (Aprovação nº. 160220), e os pais de todos os fetos com CC selecionados deram seu consentimento por escrito.
Teste por SNP microarray
Todas as amostras de líquido amniótico ou sangue do cordão umbilical foram coletadas das mulheres grávidas, e o DNA genômico foi extraído utilizando o QIAamp DNA Blood Mini Kit (Qiagen, Alemanha) de acordo com o protocolo do fabricante. O teste por SNP microarray (Polimorfismo de Nucleotídeo Único) foi realizado utilizando o Illumina HumanCytoSNP-12 v2.1 BeadChip (Illumina, EUA). A política laboratorial no momento do teste era a de não relatar polimorfismos bem estabelecidos, CNVs que não continham genes e CNVs menores que 0,20 Mb. No entanto, trechos de homozigose maiores que 10 Mb eram relatados.
Cariotipagem
Todas as amostras de líquido amniótico ou sangue do cordão umbilical foram usadas para realizar o bandeamento G de acordo com o procedimento padrão, conforme descrito anteriormente.25
ResultadosDados clínicos
Entre as 8.430 gestações, 362 casos de CC foram diagnosticados por meio de ecocardiografia fetal, com frequência de 4,2%. A idade média das gestantes foi de 31,1 ± 5,1 anos, e a idade gestacional média do diagnóstico foi de 24,4 ± 3,8 semanas.
Os 5 tipos mais comuns de CC foram, em ordem, comunicação interventricular (51,9%, 188/362), persistência da veia cava superior esquerda (13,0%, 47/362), defeitos do coxim endocárdico (0,9%, 33/362), artéria umbilical única (0,9%, 32/362) e arco aórtico do lado direito (0,8%, 29/362).
Etiologia
No total, 362 fetos foram diagnosticados com CC. Os testes genéticos encontraram 111 casos com aneuploidia, 10 casos com anormalidade na estrutura dos cromossomos e 19 casos com CNVs patogênicas ou provavelmente patogênicas (Tabela 1). Os 222 casos restantes não apresentaram achados genéticos anormais. As anormalidades do número de cromossomos consistiram de: síndrome da trissomia do cromossomo 18 (61 casos), síndrome da trissomia do cromossomo 21 (31 casos) e síndrome da trissomia do cromossomo 13 (19 casos). A CMA identificou 19 CNVs, incluindo síndrome de DiGeorge (8 casos), síndrome de Jacobsen (2 casos), síndrome de Angelman/Prader-Willi (1 caso), síndrome de microdeleção 16p11.2-p12.2 (1 caso), síndrome de triplicação 16q24 (1 caso), síndrome de Trombocitopenia e Ausência de Rádio (TAR) (1 caso), síndrome de microduplicação 3q29 (1 caso), síndrome de duplicação 22q11 (1 caso), síndrome de Cri-du-Chat (1 caso) e 2 CNVs patogênicas (Tabela 2).
Testes genéticos realizados em 362 fetos com cardiopatia congênita
Etiologia
Classificação
Números
Aneuploidia (111, 30,7%)
Trissomia 18
61
Trissomia 21
31
Trissomia 13
19
Anormalidade da estrutura dos cromossomos (10, 2,8%)
Das 362 CC, foram observadas malformações cardíacas únicas em 181 fetos, e 181 apresentaram múltiplas anormalidades cardíacas; 220 apresentaram uma CC isolada, e 142 apresentavam CC com anomalia extracardíaca. A Tabela 3 lista a etiologia dos vários tipos de malformações cardíacas fetais observadas.
Detecção genética em diferentes categorias de fetos com cardiopatia congêntita (CC)
Classificação da CC
Aneuploidia
Anormalidade da estrutura dos cromossomos
CNVs
Outros
Malformação cardíaca única (n = 181)
40
4
9
128
Múltiplas anormalidades cardíacas (n = 181)
71
6
10
94
CC isolada (n = 220)
26
8
14
172
CC com anomalia extracardíaca (n = 142)
85
2
5
50
CNVs: variações no número de cópias.
Desfechos da Gravidez
Entre os 140 casos com resultados positivos nos testes genéticos, apenas uma mulher optou por continuar a gravidez, e o restante optou por induzir o parto. O feto foi diagnosticado com síndrome da trissomia do cromossomo 18, apresentando dificuldades na alimentação, e morreu 4 dias após o nascimento. Entre os 222 casos negativos restantes, 56 foram submetidos à indução do parto, e a maioria desses casos foi considerada incurável ou apresentava malformações cardíacas com prognósticos desfavoráveis (incluindo ventrículo único, displasia ventricular esquerda ou direita e tetralogia de Fallot) ou complicadas por anomalias extracardíacas (Figura 1).
Evolução dos pacientes no presente estudo. OPN: óbitos perinatais. CMA: análise cromossômica por microarray; CNVs: ariações no número de cópias; CC: cardiopatia congêntita
As mães de 77 fetos com malformações cardíacas leves ou curáveis optaram por manter a gravidez. Destes casos, 66 foram encontrados sem anormalidade após o nascimento, 8 casos necessitaram de cirurgia, um apresentou atraso no desenvolvimento, um foi encontrado com pé torto, um foi identificado com hipomiotonia e os desfechos da gravidez dos 89 casos restantes foram incertos (Figura 1).
Discussão
Neste estudo, 362 casos de cardiopatia congênita fetal foram identificados em um total de 8.430 gestações em um único hospital materno-infantil da região sul da China, de junho de 2012 a junho de 2016, com uma incidência de 4,2%. Esta incidência foi semelhante à relatada em Xi'an, no noroeste da China,26 e superior à taxa de 2,3% relatada em Guangzhou, no sul da China.23 Entre os 362 fetos com CC, a comunicação interventricular (51,9%, 188/362) e a persistente da veia cava superior esquerda (13,0%, 47/362) foram as anormalidades cardíacas mais prevalentes detectadas por ultrassonografia.
Muitos fatores, como fatores genéticos (incluindo anomalias cromossômicas e mutações genéticas) e fatores de risco associados às mães (incluindo o vírus da rubéola, outras infecções, radiação, uso de drogas e poluição ambiental) estão associados à CC.5-7,27-29 No entanto, as causas da maioria dos tipos de CC ainda são pouco compreendidas. Em nosso estudo, 140 dos 362 fetos com CC apresentaram anormalidades cromossômicas clinicamente significativas por cariotipagem e CMA, com uma taxa de detecção de até 38,7%. As taxas positivas dos testes genéticos neste estudo são muito maiores do que as dos relatórios anteriores em Chongqing, China24 e Holanda.30 Essa taxa é semelhante à dos brasileiros.31
Entre as 140 anormalidades cromossômicas, 111 (79,3%) eram aneuploidias, das quais a trissomia do cromossomo 18 era a mais comum; 10 casos (7,1%) foram de anormalidade da estrutura cromossômica; e 19 casos (13,6%) foram CNV patogênicas ou provavelmente patogênicas. Sugere-se que a aneuploidia é a principal causa genética de fetos com CC em nossa população. Dado que o bandeamento G só pode detectar com segurança anormalidades estruturais > 10 Mb de tamanho, 11 CNVs patogênicas podem não ser identificadas por cariotipagem, mas detectadas pela CMA. Com base nesse fator, é possível estimar que o rendimento incremental de CNVs reportáveis com menos de 10 Mb alcançado pela CMA foi de 3,0%.
Malformações cardíacas múltiplas complexas têm prognóstico desfavorável e afetam fortemente a qualidade de vida dos bebês sobreviventes, mas casos como tetralogia de Fallot leve têm um resultado razoável após cirurgia, e um bom prognóstico. No presente estudo, os resultados ultrassônicos de alguns fetos com CC causada por aneuploidia mostraram apenas malformações cardíacas leves, embora CCs complexas combinadas com defeitos extracardíacos tenham sido mais comuns nesses casos. Além disso, alguns sintomas, como a deficiência mental, não podem ser verificados pela ultrassonografia pré-natal. Nestes casos, os resultados do teste genético são de grande importância, porque esta situação é facilmente ignorada pelos pacientes e médicos. No entanto, vários casos negativos apresentaram CC complexa e defeitos extracardíacos após o cariótipo e o teste de CMA, e esses casos fornecem uma pista importante para o estudo de outros fatores que levam à CC.
Várias limitações no estudo devem ser consideradas ao revisar seus achados. Primeiramente, não foi realizada uma análise abrangente de todos os genes associados a CCs conhecidas. Em segundo lugar, não foi identificada a hereditariedade de CNVs em alguns casos com patogenicidade provável.
Conclusão
Foram realizadas a cariotipagem e a análise de CMA em 362 fetos com CCs, e verificou-se que 38,7% dos fetos com CCs tiveram resultado positivo nos testes genéticos. A aneuploidia é a principal causa de fetos com CC na nossa população. A combinação da detecção ultrassônica e dos testes genéticos pode diagnosticar efetivamente fetos com malformações cardíacas e defeitos extracardíacos, fornecendo informações valiosas ao médico e aos pacientes.
Fontes de financiamento
O presente estudo não parcialmente financiado pelo Projeto de Pesquisa e Desenvolvimento da Tecnologia Médica e de Saúde de Guangxi (S201309_04 e S201611), Programa da Fundação de Ciências Naturais de Guangxi (2016GXNSFCA380001) e projeto de Ciência e Tecnologia da Região Autônoma de Guangxi Zhuang (gui-ke-gong 14124004-1-8).
Vinculação acadêmica
Não há vinculação deste estudo a programas de pós-graduação.
Aprovação ética e consentimento informado
Este estudo foi aprovado pelo Comitê de Ética do Hospital de Saúde Materno-Infantil de Guangxi sob o número de protocolo 160220. Todos os procedimentos envolvidos nesse estudo estão de acordo com a Declaração de Helsinki de 1975, atualizada em 2013. O consentimento informado foi obtido de todos os participantes incluídos no estudo.
Agradecimentos
Os autores agradecem às famílias por sua participação neste estudo.