SNP and DNA methylation analyses of a monozygotic twins discordant for complete endocardial cushion defect: a case report.

The exact cause of complete endocardial cushion defect (ECD) is still unknown. This report describes a unique pair of monozygotic twins (MZ twins) discordant for ECD. The chromosome karyotyping analysis revealed normal karyotype of 46, XY, 16qh+ and mat in both MZ twins. A genome-wide analysis of DNA using the Affymetrix SNP 6.0 revealed identical genotyping of single nucleotide polymorphisms (SNPs) and copy number variations (CNVs). An extensive methylation assay was carried out by NimbleGen 3 × 720 K CpG Island Plus RefSeq Promoter Arrays to analyze the potential epigenetic differences. The DNA methylation profiles of the affected twin seemed increased compared with that of the unaffected twin. However, further validation of Notch1 promoter hypermethylation and six top-ranked differentially methylated CpG sites by sodium bisulfate modification and methylation-specific PCR, failed to reveal consistent methylation differences between the twins. Other relevant factors, such as heritability and penetrance of the condition that place the MZ twins near to a threshold for ECD or variations in local epigenetic events in the twins' heart tissues, are probably responsible for the phenotypic discordance.

Co-culture induces expression of female primordial germ cell-specific genes in human Wharton's jelly-derived mesenchymal stem cells.

OBJECTIVE: To detect mRNA and protein expression of meiosis-specific genes in human umbilical cord mesenchymal stem cells (hUMSCs) in an in vitro co-culture microenvironment with mouse primordial germ cells (PGCs), and to further explore the effective potential of hUMSCs to differentiate into PGCs. METHODS: HUMSCs were obtained from human Wharton's jelly fragments by adherent culture. PGCs were derived from 12.5 days post-coitum (dpc) BalbC mice. Then hUMSCs were co-cultured with PGCs in Matrigel, inside or outside of a culture chamber, respectively. The changes in morphology and cytogenetic characteristics were observed. SCP3 and DDX4 expression in hUMSCs were detected and analyzed using immunofluorescence staining. Oct-4, Stra8, Zp3 and Dmc1 gene expressions in PGCs, hUMSCs, and hUMSCs after co-culture with PGCs were analyzed by real time reverse transcription-polymerase chain reaction. RESULTS: Both hUMSCs and PGCs expressed Oct-4 at different degrees. After co-culture with PGCs, hUMSCs became rounded and showed AKP activity. HUMSCs suspension-cultured in Matrigel or adherent cultured with cell chamber significantly expressed Stra8, DMC1, SCP3 and DDX4 genes. CONCLUSION: HUMSCs can be induced to express PGC-specific genes Stra8 and DMC1, spermatogonium/oogonium-specific genes SCP3 and DDX4 that predict directed differentiation potential into early germ cells at a molecular level.

Endoglin is necessary for angiogenesis in human ovarian carcinoma-derived primary endothelial cells.

Endoglin (CD105, END) is upregulated in proliferating endothelial cells, suggesting potential therapeutic properties. However, it is not clear whether endoglin mediates an enhanced proliferative rate or may be upregulated as part of a negative feedback loop. To gain insights into context-dependent and cell type-dependent regulatory effects of endoglin, we studied its role properties in human ovarian carcinoma-derived endothelial cells (ODMECs). We isolated and cultured primary ODMECs from epithelial ovarian carcinoma tissue. ODMECs had higher expression of endoglin and VEGFR-2, and also exhibited enhanced spontaneous formation of vessel-like structures in vitro. Transfection of siRNA targeting endoglin in ODMECs cells resulted in the reduction of the proliferation and tube formation. These results indicate that a subset of ODMECs display abnormal angiogenic properties and this phenotype was blocked by decreasing endoglin levels, suggesting endoglin is essential for stimulating angiogenesis, and targeting it may be an attractive approach to anti-angiogenesis therapy for ovarian carcinoma.

Karyotypic and molecular genetic changes associated with fetal cardiovascular abnormalities: results of a retrospective 4-year ultrasonic diagnosis study.

OBJECTIVE: To investigate the incidence of aneuploidy in fetuses with congenital heart defects (CHDs) and to further identify submicroscopic changes and global DNA methylation levels as potential biomarkers in complex CHD cases. METHODS: Fetuses at high risk for birth defects or with obvious sonographic anomalies were recruited at the Prenatal Diagnosis Center and Ultrasonic Diagnosis Center. Elective fetal karyotyping and DNA copy number and promoter methylation analyses were carried out following parental consent. G-banded karyotyping was performed to detect fetal aneuploidy. Copy number variations (CNVs) were detected using the Affymetrix SNP Array 6.0 and validated by real time PCR. Global DNA methylation analyses were conducted using a Roche NimbleGen Human DNA Methylation 3x720K Array, and DNA methylation differences were assayed by a Sequenom MassARRAY EpiTYPER. RESULTS: Conventional karyotyping identified 30 cases with aneuploidy in 179 CHD fetuses. Various CNVs were found in two aneuploid fetuses and in five euploid CHD fetuses. Verified segmental deletion or duplications were not directly associated with cardiovascular malformations except in DAAM1 and GATA6. Verifiable aberrant DNA methylation could not be identified in three complex CHD fetuses. CONCLUSIONS: In this study, Trisomy 18, Trisomy 21 and 45,XO were the most common aneuploidies identified in CHD fetuses. In the affected samples, only DAAM1 deletion and GATA6 amplification could be associated with cardiovascular biological processes.

Deletion of a single-copy DAAM1 gene in congenital heart defect: a case report.

BACKGROUND: With an increasing incidence of congenital heart defects (CHDs) in recent years, genotype-phenotype correlation and array-based methods have contributed to the genome-wide analysis and understanding of genetic variations in the CHD population. Here, we report a copy number deletion of chromosomal 14q23.1 in a female fetus with complex congenital heart defects. This is the first description of DAAM1 gene deletion associated with congenital heart anomalies. CASE PRESENTATION: Compared with the control population, one CHD fetus showed a unique copy number deletion of 14q23.1, a region that harbored DAAM1 and KIAA0666 genes. CONCLUSIONS: Results suggest that the copy number deletion on chromosome 14q23.1 may be critical for cardiogenesis. However, the exact relationship and mechanism of how DAAM1 and KIAA0666 deletion contributes to the onset of CHD is yet to be determined.