[Prenatal diagnosis and genetic analysis of two fetuses with Wolf-Hirschhorn syndrome].

OBJETIVE: To explore the prenatal ultrasound phenotype and genetic basis of two fetuses with Wolf-Hirschhorn syndrome (WHS). METHODS: A retrospective analysis was conducted on the ultrasound imaging data of two fetuses suspected for WHS at the Prenatal Diagnostic Center of Qingyuan People's Hospital in July 2017 and August 2019, respectively. Amniotic fluid samples of the two fetuses were subjected to chromosomal karyotyping and chromosomal microarray analysis (CMA). This study was approved by the Qingyuan People's Hospital (Ethics No. IRB-2022-064). RESULTS: Prenatal ultrasound examination of the two fetuses had consistently revealed WHS-associated traits including intrauterine growth restriction (IUGR), craniofacial abnormalities and cardiovascular anomalies. Karyotyping analysis suggested that both fetuses had harbored cryptic chromosomal translocations involving partial deletion of 4p. And parental verification revealed that it was de novo for fetus 1 and paternal for fetus 2. CMA has confirmed that fetus 1 had an approximately 8.7 Mb deletion at 4p16.3p16.1 and a 6.8 Mb duplication at 8p23.1p23.1, whilst fetus 2 had a 20.05 Mb deletion at 4p16.3p15.31 and a 7.66 Mb duplication at 9p24.3p24.1. The karyotype of fetus 1 was determined as 46,XN,der(4)t(4;8)(p16.1;p23.1)dn.arr[hg19]4p16.3p16.1(68345_8721580)×1, 8p23.3p23.1(158048_6933745)×3, and that of fetus 2 was determined as 46,XN,der(4)t(4;9)(p15.3;p24)pat.arr[hg19]4p16.3p15.31(68345_20116061)×1, 9p24.3p24.1(208454_7868292)×3. CONCLUSION: The 4p deletion is probably the main cause for the WHS phenotype in both fetuses. WHS should be suspected when IUGR, renal anomalies, craniofacial and cardiovascular abnormalities are detected upon prenatal ultrasound screening.

miR-138-5p Inhibits Vascular Mimicry by Targeting the HIF-1α/VEGFA Pathway in Hepatocellular Carcinoma.

Tumour vascular mimicry (VM) is the process by which new blood vessels are formed by tumour cells rather than endothelial cells. An increasing number of studies have revealed that the VM process is associated with cancer progression and metastasis. MiR-138-5p has been reported to act as a tumour suppressor in many cancers. However, the role and underlying mechanism of miR-138-5p in hepatocellular carcinoma (HCC) VM remain unclear. In this study, VM density was detected by CD31/periodic acid-Schiff double staining in HCC clinical specimens. We found that miR-138-5p expression correlated strongly and negatively with microvessel density. Additionally, the miR-138-5p mimic or inhibitor decreased or increased, respectively, tube formation capacity in HepG2 and Hep3B cells. Consistent with this finding, miR-138-5p repressed vessel density in vivo. Moreover, miR-138-5p targeted hypoxia-inducible factor 1α (HIF-1α) and regulated the expression of HIF-1α and vascular endothelial growth factor A (VEGFA), which are established classical master regulators for angiogenesis. Consistent with these findings, the HIF-1α inhibitor CAY10585 effectively blocked HCC cell VM and VEGFA expression. In conclusion, miR-138-5p inhibits HepG2 and Hep3B cell VM by blocking the HIF-1α/VEGFA pathway. Therefore, miR-138-5p may serve as a useful therapeutic target for miRNA-based HCC therapy.

Prenatal genetic testing in 19 fetuses with corpus callosum abnormality.

BACKGROUND: Corpus callosum abnormality (CCA) can lead to epilepsy, moderate severe neurologic or mental retardation. The prognosis of CCA is closely related to genetic etiology. However, copy number variations (CNVs) associated with fetal CCA are still limited and need to be further identified. Only a few scattered cases have been reported to diagnose CCA by whole exome sequencing (WES). METHODS: Karyotyping analysis, copy number variation sequencing (CNV-seq), chromosomal microarray analysis (CMA) and WES were parallelly performed for prenatal diagnosis of 19 CCA cases. RESULTS: The total detection rate of karyotyping analysis, CMA (or CNV-seq) and WES were 15.79% (3/19), 21.05% (4/19) and 40.00% (2/5), respectively. Two cases (case 11 and case 15) were diagnosed as aneuploidy (47, XY, + 13 and 47, XX, + 21) by karyotyping analysis and CNV-seq. Karyotyping analysis revealed an unknown origin fragment (46,XY,add(13)(p11.2)) in case 3, which was further confirmed to originate from p13.3p11.2 of chromosome 17 by CNV-seq. CMA revealed arr1q43q44 (238923617-246964774) × 1(8.04 Mb) in case 8 with a negative result of chromosome karyotype. WES revealed that 2 of 5 cases with negative results of karyotyping and CNV-seq or CMA carried pathogenic genes ALDH7A1 and ARID1B. CONCLUSION: Parallel genetic tests showed that CNV-seq and CMA are able to identify additional, clinically significant cytogenetic information of CCA compared to karyotyping; WES significantly improves the detection rate of genetic etiology of CCA. For the patients with a negative results of CNV-seq or CMA, further WES test is recommended.

Genetic testing in fetuses with isolated agenesis of the corpus callosum.

PURPOSE: The objectives of this study were to explore genetics pathogenesis of isolated agenesis of corpus callosum (ACC) and assess the utility of chromosomal microarray analysis (CMA) for genetic diagnosis of isolated ACC. METHODS: We analyzed the genomes of 16 fetuses with isolated ACC using Afymetrix CytoScan HD arrays and conducted further bioinformatic analysis for one proband fetus with an abnormal copy number variation (CNV). RESULTS: Of the 16 fetal samples examined, two (12.5%) had pathogenic CNVs and three (18.75%) had variants of unknown significance. Two cases, case 2 and case 9, were found to have pathogenic CNVs. Bioinformatic analyses indicated that the CNV of one fetus (case 9) contained 115 annotated coding genes, five of which (SLC6A5, BDNF, ELP4, PAX6, and SLC1A2) have been associated with neurodevelopment. Three of these genes (SLC1A2, BDNF, and PAX6) may play a key role in ACC development. GO cluster analysis of the implicated genes revealed strong representations of protein binding and metal ion binding functions. KEGG pathway analysis pointed to four pathways: longevity regulating pathway, amyotrophic lateral sclerosis, cocaine addiction, and autophagy-animal. CONCLUSIONS: BDNF, SLC1A2, and PAX6 may be involved in the development of isolated ACC. CMA is a feasible technology for prenatal diagnosis of isolated ACC.

Analysis of the clinical features of pericentric inversion of chromosome 9.

OBJECTIVE: The pericentric inversion of chromosome 9 (inv9) is one of the most common structural balanced chromosomal variations, and it is considered to be a normal population variant. The aim of this study was to re-evaluate the clinical impact of patients with inv9. METHODS: We studied the karyotypes from 4853 patients at a single center and retrospectively reviewed their clinical data. RESULTS: There were 67 inv9 patients among 2988 adults, and 62 of them showed different clinical features, including male and female infertility, oligoasthenozoospermia, and azoospermia. Thirty-one cases of inv9 were found in 1865 fetuses, including two cases in chorionic villus (6.90%) and 29 in amniotic fluid (1.67%), but there were no cases in umbilical cord blood. The rates of fetal phenotype abnormal and adverse pregnancy outcome with inv9 in the chorionic villus were 100.00% (2/2), while only 17.24% (5/29) in the amniotic fluid showed abnormalities, among which 60.00% (3/5) had adverse pregnancy outcomes. CONCLUSIONS: Although there is no clear evidence that inv9 is pathogenic, the genetic counseling on inv9 in early pregnancy and adults needs to be given more attention.

Diagnostic cytogenetic testing following positive noninvasive prenatal screening results of sex chromosome abnormalities: Report of five cases and systematic review of evidence.

BACKGROUND: Follow-up cytogenetic analysis has been recommended for cases with positive noninvasive prenatal screening (NIPS) results. This study of five cases with numerical and structural sex chromosomal abnormalities (SCA) and a review of large case series of NIPS provided guidance to improve prenatal diagnosis for SCA. METHODS: Following positive NIPS results for SCA, karyotype analysis, chromosomal microarray analysis (CMA), fluorescence in situ hybridization (FISH), and locus-specific quantitative PCR were performed on cultured amniocytes, chorionic villi cells, and stimulated lymphocytes. Review of large case series was performed to evaluate the NIPS positive rate, follow-up rate of cytogenetic analysis, positive predictive value (PPV) for major types of SCA, and relative frequencies of subtypes of major SCA. RESULTS: Of the five cases with positive NIPS for SCA, case 1 showed a mosaic pattern of monosomy X and isodicentric Y; case 2 showed a mosaic pattern of monosomy X confined to the placenta; cases 3 and 4 had an isochromosome of Xq, and case 5 showed a derivative chromosome 14 from a Yq/14p translocation of maternal origin. Review of literature showed that mean positive rate of NIPS for SCA was 0.61%, follow-up rate of cytogenetics analysis was 76%, and mean PPV for SCA was 48%. Mosaic patterns and structural rearrangements involving sex chromosomes were estimated in 3%-20% and 3% of SCA cases, respectively. CONCLUSION: These five cases further demonstrated the necessity to pursue follow-up cytogenetic analysis to characterize mosaic patterns and structural abnormalities involving sex chromosomes and their value for prenatal genetic counseling. A workflow showing the performance of current NIPS and cytogenetic analysis for SCA was summarized. These results could facilitate an evidence-based approach to guide prenatal diagnosis of SCA.

Genetic and sex hormone analysis of infertile men.

OBJECTIVE: Genetic defects and endocrine-related factors are the leading causes of male infertility. This study was performed to analyze the genetic characteristics and sex hormone levels in different types of male infertility. METHODS: A total of 423 men with infertility underwent genetic and sex hormone analysis at The Sixth Affiliated Hospital of Guangzhou Medical University. RESULTS: The incidences of abnormal karyotypes in patients with male infertility, azoospermia, and oligoasthenozoospermia were 6.94%, 22.40%, 15.09%, respectively. Among men with azoospermia, Klinefelter syndrome (47,XXY) was identified in 60.71% (17/28) of those with abnormal karyotypes. Additionally, the levels of follicle-stimulating hormone and human luteinizing hormone were significantly higher in men with azoospermia showing abnormal karyotypes than in men of the other study groups. The serum testosterone level in men with azoospermia showing abnormal karyotypes was lower than that in men of the other study groups. CONCLUSIONS: Azoospermia is closely associated with chromosome abnormalities. The levels of testosterone, human luteinizing hormone, and follicle-stimulating hormone in men with azoospermia showing abnormal karyotypes provide a clinical reference for genetic counseling and assisted reproduction.

The Effect of Freezing on Non-invasive Prenatal Testing.

Plasma cryopreservation is unavoidable in China, due to technical specifications requiring storage of additional plasma at -80 degrees for three years. However, the effect of freezing on non-invasive prenatal testing (NIPT) is still uncertain. We collected 144 euploid pregnant samples, 22 on trisomy 21, 4 on trisomy 13, and 3 on trisomy 18, by massively parallel sequencing before and after freezing. Compared with the success rate of 100% of fresh samples, the detection success rates of trisomy 21, trisomy 13 and euploidy in frozen samples by NIPT were 95.45%, 75% and 95.14%, respectively. Of these, 9 cases of frozen sample sequencing failed, with 8 cases being due to high GC content. The chromosome 21 (chr21) z-value of the frozen trisomy 21 samples was lower than that of fresh samples. Meanwhile, freezing reduced the male positive foetal cell-free DNA (cfDNA) fraction, which was accompanied by an increase in the Unimap-GC level in the massively parallel sequencing data and a decrease in the Unique reads/Total reads ratio. Laboratory freezing reduced the chr21 z-value of foetal trisomy 21, which can be explained by a reduction in the foetal cfDNA fraction and effective Unique reads for NIPT analysis. The Unimap-GC content of the serum samples after freezing was higher, which can lead to failure of NIPT detection.

[Prenatal diagnosis of a fetus with trisomies of 11q23.3q25 and 22q11.1q11.21].

OBJECTIVE: To explore the phenotype and pathogenesis of a fetus with a rare chromosomal abnormality. METHODS: The fetus was analyzed by clinical prenatal ultrasonography, G-banding karyotyping and next generation sequencing (NGS). RESULTS: Prenatal ultrasonography of the fetus showed Dandy-Walker syndrome, growth restriction, and right-heart system dysplasia. The fetus had a chromosomal karyotype of 47,XY,t(11;22)(q23.3;q11.2),+der(22)t(11;22). Duplication of 11q23.3q25 and 22q11.1q21 were also detected by NGS. The chromosomal translocation carried by the fetus was derived from his father. CONCLUSION: Duplications of chromosome 11q23.3q25 and 22q11.1q11.21 segments probably underlie the Dandy-Walker syndrome, growth restriction, and hypoplasia of the right heart system in the fetus.