Third trimester predictors of interventional timing and accuracy of fetal anticipatory guidance in tetralogy of Fallot: A multi-center study.

OBJECTIVE: The objective was to evaluate and improve accuracy of anticipatory counseling regarding neonatal intervention for prenatally diagnosed tetralogy of Fallot (TOF) by assessing new and previously published predictors of neonatal intervention. METHODS: This is a multi-center, retrospective study from three centers of 112 fetal TOF patients undergoing third trimester fetal echocardiograms from 2004 to 2017. Additional cardiac defects requiring neonatal intervention were excluded. Fetal echocardiographic, clinical, and consultation data were compared between neonatal and late intervention. Optimal echocardiographic values were determined. RESULTS: Twenty-six infants (23%) required neonatal intervention. Those infants had significantly different pulmonary valve (PV) z-scores, PV:aortic valve (AoV) ratios, PV:AoV z-score differences (absolute difference between z-scores), and increased likelihood of abnormal ductal flow. Counseling during fetal echocardiogram regarding interventional timing was accurate for 50% needing neonatal intervention and 86% undergoing late intervention (P = .002). The best neonatal intervention predictors were PV:AoV ratio of <0.6 and counseling for neonatal intervention. PV:AoV z-score difference ≥5 provided 89% negative predictive value for excluding patients from neonatal repair. CONCLUSIONS: Third trimester fetal echocardiograms can predict interventional timing. The best predictors of neonatal intervention are PV:AoV ratio <0.6, PV:AoV z-score difference ≥5, and cardiologist counseling that neonatal intervention was likely.

scaRNA1 Levels Alter Pseudouridylation in Spliceosomal RNA U2 Affecting Alternative mRNA Splicing and Embryonic Development.

The heart is the first major organ to develop during embryogenesis and must receive proper spatiotemporal signaling for proper development. Failure of proper signaling between the first and second heart fields at twenty days gestation contributes to the generation of a congenital heart defect. The most common cyanotic congenital heart defect is tetralogy of Fallot (TOF) which requires surgical intervention in the first year of life. In right ventricular tissue of infants born with TOF, the levels of scaRNA1 are reduced and mRNA splicing is dysregulated. In this study, we investigate a method of quantifying pseudouridylation levels in relation to scaRNA1 levels in spliceosomal RNA U2 in three different groups of samples: right ventricular (RV) tissue of infants born with TOF versus RV tissue from normally developing infants, scaRNA1 knockdown in primary normal cardiomyocytes derived from normally developing infants, and scaRNA1 overexpression in primary cells derived from RV tissue from infants born with TOF. We hypothesize that the amount of pseudouridylation is dependent on scaRNA1 level, compromising spliceosomal function and therefore, contributing to the generation of a congenital heart defect. Our results revealed a statistically significant decrease of pseudouridylation levels in the right ventricular tissue of infants born with TOF compared to the controls. Knocking down the scaRNA1 levels in normal primary cardiomyocytes resulted in a statistically significant decrease of pseudouridylation. Finally, an overexpression of scaRNA1 in TOF primary cells resulted in an increase in pseudouridylation levels, but it did not achieve statistical significance. Our previous research provided an association between scaRNA levels, alternative splicing, and development. Here, we demonstrate that pseudouridylation levels in spliceosomal RNA U2 is dependent on the expression level of scaRNA1. Although further investigation is needed, we believe that scaRNA expression regulates biochemical modifications to spliceosomal RNAs, adjusting the fidelity of the spliceosome, allowing for controlled alternative splicing of mRNA that is important in embryonic development. If validated, this is an underappreciated mechanism that is critical for regulating proper embryonic development.

Tetralogy of Fallot With Absent Pulmonary Valve and Nonconfluent Pulmonary Arteries: A Management Conundrum.

Tetralogy of Fallot with absent pulmonary valve syndrome is a rare form of congenital heart disease. Among the different variations with this rare anomaly is nonconfluent pulmonary artery branches with anomalous origin of the left pulmonary artery from the ductus arteriosus. The authors present one such case which was diagnosed prenatally to have tetralogy of Fallot with absent pulmonary valve and identified postnatally to have nonconfluent pulmonary artery branches in addition. We discuss the conundrum of respiratory management in this patient pre- and postoperatively due to a unique ventilation perfusion mismatch problem, which varies between the two lungs.

Placental DNA methylation changes in detection of tetralogy of Fallot.

OBJECTIVES: To determine whether the methylation level of cytosine nucleotides in placental DNA can be used to predict tetralogy of Fallot (TOF) and provide insights into the developmental mechanism of this condition. METHODS: Tissue sections were obtained from formalin-fixed paraffin-embedded specimens of placental tissue obtained at birth from eight cases with non-chromosomal, non-syndromic TOF and 10 unaffected newborns. The Illumina Infinium HumanMethylation450 BeadChip assay was used to measure cytosine ('CpG' or 'cg') methylation levels at loci throughout the placental genome. Differential methylation was assessed by comparing the ß-values (a measure of the extent of cytosine methylation) for individual CpG loci in fetuses with TOF vs in controls. The most discriminating CpG sites were determined based on a preset cut-off of ≥ 2.0-fold change in the methylation level. The predictive accuracy of CpG loci with significant methylation changes for TOF was determined by the area under the receiver-operating-characteristics curve (AUC). A false-discovery-rate (FDR) P-value < 0.05 was used to define a statistically significant difference in the methylation level. Ingenuity Pathway Analysis (IPA) (Qiagen) was used to identify gene pathways that were significantly overexpressed, and thus altered, in TOF cases compared with controls. RESULTS: We found a total of 165 significantly differentially methylated CpG loci in TOF cases compared with controls, in 165 separate genes. These biomarkers demonstrated from fair to excellent individual predictive accuracy for TOF detection, with AUCs ≥ 0.75 (FDR P-value < 0.001 for all). The following CpG loci (gene) had the highest predictive accuracy: cg05273049 (ARHGAP22; AUC = 1.00; 95% CI, 1.00-1.00), cg02540011 (CDK5; AUC = 0.96; 95% CI, 0.87-1.00), cg08404201 (TRIM27; AUC = 0.95; 95% CI, 0.84-1.00) and cg00687252 (IER3; AUC = 0.95; 95% CI, 0.84-1.00). IPA revealed over-representation (dysregulation) of 14 gene pathways involved in normal cardiac development, including cardiomyocyte differentiation via bone morphogenetic protein receptors, cardiac hypertrophy signaling and role of nuclear factor of activated T cells in cardiac hypertrophy. Cardiac hypertrophy is an important feature of TOF. CONCLUSIONS: Analysis of placental DNA cytosine methylation changes yielded accurate markers for TOF detection and provided mechanistic information on TOF development. Our work appears to confirm the central role of epigenetic changes and of the placenta in the development of TOF. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Prenatal diagnosis of tetralogy of Fallot associated with pulmonary artery sling: Two case reports.

Tetralogy of Fallot (TOF) is a common condition accounting for 10%-20% of all fetal cyanotic congenital heart disease cases. Pulmonary artery sling (PAS), or aberrant left pulmonary artery, is a rare congenital cardiovascular malformation. Approximately 58%-83% of PAS is associated with other cardiovascular malformations, TOF being rarest. The diagnosis of PAS is generally incidental or made at autopsy. Cases of prenatal diagnoses of TOF associated with PAS have not yet been reported. Here, we report two cases of TOF associated with PAS diagnosed prenatally in our hospital.

Genetic anomalies in fetuses with tetralogy of Fallot by using high-definition chromosomal microarray analysis.

BACKGROUND: The etiology of TOF is complex and the genesis of TOF has been associated with environmental factors and genetic disorders, including chromosomal anomalies, aneuploidies, 22q11.2 deletion and single-gene disease. Previous literatures have shown that a chromosome alteration in about 30% patients with TOF and recently published articles reported that 22q11.2 deletion syndrome accounts for 16% cases with TOF diagnosed postnatally. CMA now is considered as gold standard for detecting genetic anomalies in fetuses with congenital malformations. CMA could detect a 6.6-25% incremental yield of CNVs in CHDs. The aim of this study was to assess the genetic anomalies in fetal tetralogy of Fallot (TOF) by using high-definition CMA. METHODS: This retrospective study reviewed all the fetuses diagnosed with TOF between 2013 and 2018. Prenatal ultrasongraphic findings, including cardiac angle, and the findings of CMA using Affymetrix CytoScan HD array were collected. RESULTS: Ninety-six fetuses with TOF and known genetic results were enrolled. Right aortic arch was the most common associated anomalies (22.9%). One fetus with trisomy 18, one with 46, XX, t (7;10)(q36;q22), one with 47, XYY and five with trisomy 21 were identified. Clinically significant CNVs occurred in 6.8% and uncertain significant CNVs in 3.4% fetal TOF with normal karyotype. A total of four cases with 22q11.2 microdeletion and two fetuses with Yq11.223q11.23 microduplication have been identified. Genetic anomalies, including chromosomal aberrations and pathogenic CNVs, were significantly higher in the TOF with extracardiac anomaly group than in the TOF without extracardiac anomaly group (P = 0.005). Abnormal cardiac angle was noticed in 24.0% fetal TOF. Genetic anomalies were more common in the TOF with abnormal cardiac angle than with normal cardiac angle (P = 0.001). On the other hand, abnormal cardiac angle was noticed in 64.3% fetal TOF with genetic anomalies while abnormal cardiac angle occurred in 17.1% fetal TOF with normal genetic results (P = 0.001). CONCLUSIONS: Genetic testing should be offered, specially using microarray analysis, for the fetal TOF with abnormal cardiac angle or extracardiac defects.

Tetralogy of Fallot with absent pulmonary valve-When the ductus is present: A case of isolated branch pulmonary artery and review of literature.

Tetralogy of Fallot/Absent Pulmonary Valve (TOF/APV) has been classically associated with the absence of a patent ductus arteriosus (PDA). We present a rare case of APV in TOF with a discontinuous left pulmonary artery (LPA) that was suspected during fetal echocardiogram. Postnatal echocardiogram confirmed the origin of a hypoplastic LPA from the PDA. Despite an aneurysmal (right pulmonary artery) (RPA), axial imaging demonstrated widely patent tracheobronchial system with no evidence of bronchial compression. Clinically, the child required only minimal respiratory support. Genetic testing was positive for 22 q11deletion, commonly associated with this lesion. Surgery consisted of unifocalization of the discontinuous LPA with placement of a valved pulmonary homograft during complete repair of this lesion. Our case highlights the importance of prenatal detection, to aid in the prompt initiation of prostaglandins so as to ensure early rehabilitation of the left lung. Inability to visualize one of the branch pulmonary arteries (PA's) and a PDA on fetal echocardiogram in TOF/APV must raise suspicion for an eccentric branch PA with ductal origin.

Proteomics analysis indicated the protein expression pattern related to the development of fetal conotruncal defects.

Abnormal development of embryonic conus arteriosus could lead to conotruncal defects in fetal heart, and increase the incidence of fetal congenital heart disease. Tetralogy of Fallot (TOF) is one of the most common forms of congenital heart disease. It may be helpful for us to solve this clinical problem through exploring the molecular mechanisms of development in embryonic congenital heart disease. Proteomics has attracted much attention in understanding the development of human diseases during the past decades. However, there is still little information about the relationship between protein expression pattern and TOF. In this study, we aimed to explore the potential linkage of proteomics and TOF development. Briefly, 121 differentially expressed proteins were identified from a TOF group, compared with a control group. The expression levels of 34 of these proteins were significantly different (>1.5 absolute fold change, p < 0.05) between the two groups. Gene ontology (GO) and pathway analysis showed that these proteins were mainly associated with carbon metabolism, biosynthesis of antibodies, positive regulation of transcription from RNA polymerase II promoter, nucleus, ATP binding, and so on. The ingenuity pathway analysis (IPA) results indicated that 435 of upstream regulators were identified of these differentially expressed proteins, which might be involved in the development of TOF. Data of string analysis showed the protein-protein interaction network among the differentially expressed proteins and regulators, which are related to TOF. In conclusion, our study explored the protein expression pattern of TOF, which might provide new insights into understanding the mechanism of TOF development and afford potential targets for TOF diagnosis and therapy.

Association Between Tetralogy of Fallot and Tracheobronchial Branching Abnormalities: A New Clue for Pathogenesis?

BACKGROUND: In our practice, we noticed an increased frequency of tracheobronchial branching abnormalities (TBAs) in patients with tetralogy of Fallot (ToF). This study aimed to determine whether an association exists between congenital TBAs and ToF with or without pulmonary atresia. METHODS AND RESULTS: The frequency of TBAs on chest computed tomography was assessed in 55 patients with ToF without pulmonary atresia, 34 patients with ToF with pulmonary arteria, and 100 control patients. We then looked for a possible association between TBAs and pulmonary artery branch hypoplasia, the presence of major aortopulmonary collateral arteries, and the presence of the chromosome 22q11 deletion. TBAs were significantly more frequent in patients with ToF with or without pulmonary atresia than in the control group (any TBAs, 21% versus 2% [P<0.001]; bronchial situs anomalies, 6% versus 0% [P=0.002]; right tracheal bronchus, 4% versus 0% [P=0.04]; left eparterial bronchus, 8% versus 0% [P=0.005]); and tended to be more frequent in those with ToF without pulmonary atresia than in those with ToF with pulmonary atresia (any TBAs, 27% versus 12% [P=0.11]; left eparterial bronchus, 13% versus 0% [P=0.04]). TBAs were readily multiple (8 patients of 19 with TBA) and concerned essentially the upper lobes. TBAs were not associated with pulmonary branch hypoplasia, major aortopulmonary collateral arteries, or the chromosome 22q11 deletion. CONCLUSIONS: We demonstrated a significantly increased frequency of tracheobronchial abnormalities in patients with ToF with or without pulmonary atresia compared with a control group. These results suggest an interaction between abnormalities in conotruncal septation and tracheobronchial branching and may provide a new clue to the pathogenesis of conotruncal heart diseases.

Main pulmonary artery cross-section ratio is low in fetuses with tetralogy of Fallot and ductus arteriosus-dependent pulmonary circulation.

OBJECTIVES: This study aimed to determine fetal echocardiographic features of tetralogy of Fallot in association with postnatal outcomes. METHODS: The Z-scores of the main and bilateral pulmonary arteries and the aorta were measured, and the following variables were calculated in 13 fetuses with tetralogy of Fallot: pulmonary artery-to-aorta ratio and main pulmonary artery cross-section ratio - the main pulmonary artery diameter squared divided by the sum of the diameter squared of the left and right pulmonary arteries. Fetuses were classified as having ductus arteriosus-dependent or ductus arteriosus-independent pulmonary circulation. RESULTS: We included two infants with pulmonary atresia and six infants with ductus-dependent pulmonary circulation, who underwent systemic-to-pulmonary shunt surgeries at ⩽1 month of age. The Z-scores of the main pulmonary artery and the pulmonary artery-to-aorta ratio in fetuses with ductus-dependent pulmonary circulation were lesser than those in fetuses with ductus independence, but not significantly. The main pulmonary artery cross-section ratio in fetuses with ductus dependence was significantly lesser (0.65±0.44 versus 1.56±0.48, p<0.005). Besides, the flow of the ductus arteriosus was directed from the aorta to the pulmonary artery in the ductus arteriosus-dependent group during the fetal period. CONCLUSIONS: The main pulmonary artery cross-section ratio was the most significant variable for predicting postnatal outcomes in fetuses with tetralogy of Fallot.

Fetal MRI detects early alterations of brain development in Tetralogy of Fallot.

OBJECTIVE: Prenatal imaging has identified alterations of brain growth in fetuses with congenital heart disease. However, little is known about the timing of altered brain development and its occurrence in specific congenital heart disease subgroups. This magnetic resonance imaging study aimed to identify early (median, 25 gestational weeks [GW]) changes in fetal total brain (TBV), gray matter (GMV), and subcortical brain (SBV) volumes in Tetralogy of Fallot (TOF) cases in utero. STUDY DESIGN: Fetal magnetic resonance imaging (1.5 Tesla) was performed in 24 fetuses who were diagnosed with TOF and 24 normal age-matched control fetuses (20-34 GW). TBV, GMV, SBV, intracranial cavity, cerebellar, ventricular, and external cerebrospinal fluid volumes were quantified by manual segmentation based on coronal T2-weighted sequences. Mixed model analyses of variance and t-tests were conducted to compare cases and control fetuses. RESULTS: TBV was significantly lower (P < .001) in early (<25 GW) and late TOF cases. Both GMV (P = .003) and SBV (P = .001) were affected. The GMV-to-SBV ratio declined in fetuses with TOF (P = .026). Compared with normal fetuses, ventricular volume was increased (P = .0048). External cerebrospinal fluid was enlarged in relation to head size (P < .001). Intracranial cavity volume (P = .314) and cerebellar volume (P = .074) were not significantly reduced in fetuses with TOF. CONCLUSION: TOF is associated with smaller volumes of gray and white matter and enlarged cerebrospinal fluid spaces. These changes are present at ≤25 GW and indicate altered fetal brain growth in this pathophysiologic entity during early stages of human brain development.

Anatomy of the ventricular septal defect in outflow tract defects: similarities and differences.

OBJECTIVE: The study objective was to analyze the anatomy of the ventricular septal defect found in various phenotypes of outflow tract defects. METHODS: We reviewed 277 heart specimens with isolated outlet ventricular septal defect without subpulmonary stenosis (isolated outlet ventricular septal defect, 19); tetralogy of Fallot (71); tetralogy of Fallot with pulmonary atresia (51); common arterial trunk (54); double outlet right ventricle (65) with subaortic, doubly committed, or subpulmonary ventricular septal defect; and interrupted aortic arch type B (17). Special attention was paid to the rims of the ventricular septal defect viewed from the right ventricular side and the relationships between the tricuspid and aortic valves. RESULTS: The ventricular septal defect was always located in the outlet of the right ventricle, between the 2 limbs of the septal band. There was a fibrous continuity between the tricuspid and aortic valves in 74% of specimens with isolated outlet ventricular septal defect, 66% of specimens with tetralogy of Fallot, 39% of specimens with tetralogy of Fallot with pulmonary atresia, 4.6% of specimens with double outlet right ventricle, 1.8% of specimens with common arterial trunk, and zero of specimens with interrupted aortic arch type B (P < .005). When present, this continuity always involved the anterior tricuspid leaflet. CONCLUSIONS: The ventricular septal defect in outflow tract defects is always an outlet ventricular septal defect, cradled between the 2 limbs of the septal band. However, there are some differences regarding the posteroinferior and superior rims of the ventricular septal defect. These differences suggest an anatomic continuum from the isolated outlet ventricular septal defect to the interrupted aortic arch type B rather than distinct physiologic phenotypes, related to various degrees of abnormal rotation of the outflow tract during heart development: minimal in isolated outlet ventricular septal defect; incomplete in tetralogy of Fallot, tetralogy of Fallot with pulmonary atresia, and double outlet right ventricle; absent in common arterial trunk; and excessive in interrupted aortic arch type B.

Development of major aorto-pulmonary collateral arteries in vegf120/120 isoform mouse embryos with tetralogy of fallot.

The degree of right ventricular outflow tract obstruction, pulmonary stenosis (PS) and the development of major aorto-pulmonary collateral arteries (MAPCAs) in patients with tetralogy of Fallot (TOF) is related to clinical outcome. Vegf120/120 mutant mouse embryos develop TOF with various degrees of PS, comparable to humans. We aimed to study the ontogeny of the development of MAPCAs in this mouse model. The development of the right ventricular outflow tract, pulmonary arteries, and ductus arteriosus (DA) and formation of MAPCAs were studied in both wild type as well as Vegf120/120 mice from embryonic day 10.5 until day 19.5. Of the 49 Vegf120/120 embryos, 35 embryos (71%) had ventral displacement of the outflow tract and a subaortic ventricular septal defect. A time-related development in severity of PS to pulmonary atresia (PA) was observed. From embryonic day 12.5, hypoplasia of the DA was seen in 13 (37%) and absent DA in 12 (37%) of these embryos. The 3 (6%) embryos with PA and absent DA developed MAPCAs, after day 15.5. In all, the MAPCAs arose from both subclavian arteries, running posterior in the thoracic cavity, along the vagal nerve. The MAPCAs connected the pulmonary arteries at the site of the hilus. A time-related development of PS to PA can lead, in combination with absent DA, to the development of MAPCAs later in embryonic life as an alternative route for pulmonary perfusion in this mouse model. This finding contributes to a better understanding of the consecutive morphological changes in the development toward MAPCAs in humans.

Cardiovascular Z-scores in fetuses with tetralogy of Fallot.

OBJECTIVE: To establish formulae for the calculation of fetal cardiovascular Z-scores based on femur length (FL), and to compare cardiovascular parameters between fetuses with tetralogy of Fallot (TOF) and normal fetuses in order to assess their value for the prenatal diagnosis of TOF. METHODS: A total of 329 normal fetuses and 43 fetuses with TOF were included in this study. Eleven cardiovascular dimensions were measured offline after cardiac spatiotemporal image correlation volume acquisition. Normal cardiovascular Z-score formulae were constructed for these measurements based on FL by performing a standard regression analysis followed by weighted regression of absolute residual values. The following ratios were calculated: right ventricular internal diameter (RVID) to left ventricular internal diameter (LVID) and pulmonary artery diameter (PA) to aorta diameter (Ao). Subsequently, all parameters were compared between the normal and TOF groups. RESULTS: Formulae for calculating Z-scores for the 11 cardiovascular dimensions were constructed. FL was significantly correlated with all cardiovascular dimensions assessed (r = 0.816-0.944, P < 0.001 for all). RVID, Ao, PA, aortic arch isthmus, and ductus arteriosus mean Z-scores and PA/Ao ratio were statistically significantly different between normal fetuses and those with TOF. In the TOF group, all Ao Z-scores (43/43) were > + 2 and all PA/Ao ratios (43/43) were below the normal 95% range. However, only 48.8% (21/43) of the PA Z-scores were < -2. CONCLUSIONS: The cardiovascular Z-score formulae developed can provide a quantitative basis for the prenatal diagnosis of TOF. Aortic dilatation and abnormal PA/Ao ratio may be markers for the antenatal diagnosis of TOF.

Midgestation fetal pulmonary annulus size is predictive of outcome in tetralogy of fallot.

BACKGROUND: Surgical management of tetralogy of Fallot (TOF) is increasingly moving toward valve-sparing approaches rather than transannular patch (TAP). We evaluate whether fetal pulmonary valve (PV) size is predictive of postnatal course and surgical approach in TOF. METHODS: In this retrospective study, fetal and postnatal demographic, clinical, and echocardiographic data on 66 patients diagnosed prenatally with TOF were collected. We compared those with midgestation PV z-score > -3.5 to those with z-score ≤-3.5. We analyzed fetal and postnatal PV size and growth and outcomes between groups RESULTS: Gestational age at first fetal echo was 23 weeks (range 18-28). PV diameter and z-score on midgestation echo were 3.5 mm (1.3-6.0) and -2.8 (-0.5 to -6.0) respectively. Patients with PV z-score ≤ -3.5 on first fetal echo had smaller PV diameter (4.5 vs. 5.0 mm, P = .047) and PV z-score (-3.8 vs. -2.8, P < .001) in late gestation and at time of surgery (6.0 mm vs. 7.0 mm, P = .01; z-score = -2.9 vs. -1.7, P = .007). Similarly, those with smaller fetal PV z-score had smaller main and branch pulmonary arteries at time of surgery. PV growth rate over gestation was similar between groups, while after-birth PV growth rate was lower in those with smaller PV (0 mm/month vs. 0.6 mm/month, P = .002). Those with smaller pulmonary valve were more likely to be cyanotic (P = .05), to undergo surgery at <1 month (P < .01), and to have a TAP repair (P = .01). Among patients undergoing valve-sparing repair, those with smaller PV underwent more reinterventions for residual valvar PS (P < .01). CONCLUSION: Midgestation fetal PV size is predictive of postnatal PV and PA size in TOF. Midgestation PV size has implications for timing and type of surgical management as well as for need for reintervention in valve-sparing repair patients and is therefore important to consider in prenatal counseling for TOF fetuses.

Left main coronary atresia in a child with absent pulmonary valve syndrome: early entrapment of an innocent bystander.

Atresia of the left main coronary artery is a rare anomaly that, if left untreated, has an unfavorable outcome. We hereby report left main coronary artery atresia in a child with tetralogy of Fallot with absent pulmonary valve and discuss the possible developmental basis of the association.