Rongyu Chen

Next-generation sequencing analysis of DUOX2 in 192 Chinese subclinical congenital hypothyroidism (SCH) and CH patients

BACKGROUND Defects in the human dual oxidase 2 (DUOX2) gene are reported to be one of the major causes of congenital hypothyroidism (CH). This study was set to examine the DUOX2 mutation spectrum and prevalence among Chinese CH and subclinical congenital hypothyroidism (SCH) patients and to define the relationships between DUOX2 genotypes and clinical phenotypes. METHODS Peripheral venous blood samples were collected from 192 CH/SCH patients in Guangxi Zhuang Autonomous Region of China. All exons and their exon-intron boundary sequences of the 11 known CH associated genes including DUOX2 were screened by next-generation sequencing (NGS). RESULTS NGS analysis of DUOX2 revealed 18 rare non-polymorphic variants in 57 CH/SCH patients. Sequencing of other CH candidate genes in the 57 patients revealed 2 thyroglobulin (TG) variants. All variants included 11 known mutations, 8 novel variants in DUOX2 and one novel variant in TG, among which three variants p.K530X, p.L1343F and p.R683L are highly recurrent in our patient cohort. 35 (83%) of the 42 patients with one or two DUOX2 pathogenic variants turned out to be SCH or transient congenital hypothyroidism (TCH), whereas 13 (87%) of the 15 patients with three or more DUOX2 pathogenic variants are associated with permanent congenital hypothyroidism (PCH). The accumulation of defects in DUOX2 contribute to the more severe disease regarding thyroid stimulating hormone (TSH) levels, free thyroxine (FT4) levels and initial dose of l-thyroxine (L-T4). CONCLUSION Our study expanded the mutational spectrum of the DUOX2 and TG genes and provided the best estimation of the DUOX2 mutation rate (29%) for CH/SCH patients in Guangxi Zhuang Autonomous Region of China. Most one or two DUOX2 pathogenic variants turned out to be SCH or TCH, whereas patients with three or more DUOX2 pathogenic variants were mostly associated with PCH. The coexistence of multiple pathogenic variants may have contributed to the severity of the hypothyroid condition.

Mutation screening of the TPO gene in a cohort of 192 Chinese patients with congenital hypothyroidism.

Objectives Defects in the human thyroid peroxidase (TPO) gene are reported to be one of the causes of congenital hypothyroidism (CH) due to dyshormonogenesis. The aim of this study was to examine the TPO mutation spectrum and prevalence among patients with CH in the Guangxi Zhuang Autonomous Region of China and to define the relationships between TPO genotypes and clinical phenotypes. Methods Blood samples were collected from 192 patients with CH in the Guangxi Zhuang Autonomous Region, China and genomic DNA was extracted from peripheral blood leucocytes. All exons of the 10 common CH-associated genes including TPO together with their exon-intron boundaries were screened by next-generation sequencing (NGS). The effect of the novel TPO mutation was investigated by ‘in silico’ studies. Results NGS analysis of TPO in 192 patients with CH revealed 3 different variations in 2 individuals (2/192, 1%). Sequencing other CH candidate genes in the patients with TPO variants revealed that patient 1 was homozygous for c.2422delT TPO mutation combined with double heterozygous DUOX2 pathogenic variants (p.R683L/p.L1343F) and patient 2 was triallelic for TPO pathogenic variants (p.R648Q/p.T561M/p.T561M). The present study identified a novel TPO variation c.1682C>T/p.T561M; and four known mutations: c.2422delT/p.C808Afs×24 and c.1943C>T/p.R648Q in TPO, c.2048G>T/p.R683L and c.4027C>T/p.L1343F in DUOX2. Conclusions Our study indicated that the prevalence of TPO mutations was 1% among studied Chinese patients with CH. More than two variations in one or more CH-associated genes can be found in a single patient, and may, in combination, affect the phenotype of the individual. A novel TPO variation c.1682C>T/p.T561M was found, thereby expanding the mutational spectrum of the gene.

Next-generation sequencing analysis of TSHR in 384 Chinese subclinical congenital hypothyroidism (CH) and CH patients

BACKGROUND Defects in the human TSHR gene are reported to be one of the causes of CH due to thyroid dysgenesis, the purpose of this study was to examine the TSHR mutation spectrum and prevalence in congenital hypothyroidism (CH) and subclinical congenital hypothyroidism (SCH) patients in the Guangxi Zhuang Autonomous Region of China and to evaluate the genotype-phenotype correlations. METHODS Blood samples were collected from 384 patients including 240 CH and 144 SCH patients in Guangxi, China. Genomic DNA was extracted from peripheral blood leukocytes. All exons of the 11 known CH associated genes including TSHR together with their exon-intron boundaries were screened by next-generation sequencing (NGS). RESULTS NGS analysis of TSHR revealed nine different variants in ten individuals. Six (4.2%) of 144 patients with SCH were found to harbor monoallelic TSHR variants. Four (1.6%) of 240 patients with CH harbored TSHR variants combined with another monoallelic mutation in either DUOX2 or TG gene. The present study identified five novel variants c.1838A>G (p.Y613C), c.1576G>A (p.A526T), c.2087T>G (p.F696C), c.1631G>A (p.G544E) and c.2051C>A (p.A684D) in TSHR, seven known pathogenic variants c.1349G>A (p.R450H), c.326G>A (p.R109Q), c.2066T>G (p.V689G) and c.2272G>A (p.E758K) in TSHR, IVS3+2T>G in TG, and c.1588A>T (p.K530X) and c.2635G>A (p.E879K) in DUOX2. The previously reported hotspot mutation p.R450H was found in only one SCH patient. CONCLUSION The prevalence of TSHR mutations was 1.6% in CH patients and 4.2% in SCH patients in Guangxi Zhuang Autonomous Region of China. Monoallelic TSHR pathogenic variants were associated with SCH, while TSHR pathogenic variants combined with monoallelic mutations in DUOX2 or TG gene might contribute to CH. Our study expands the TSHR mutation spectrum and provides the best estimation of mutation rate for SCH and CH patients in this Chinese population.

Mutation screening of the SLC26A4 gene in a cohort of 192 Chinese patients with congenital hypothyroidism

OBJECTIVE Pendred syndrome (PS) is an autosomal recessive disorder characterised by sensorineural hearing loss and thyroid dyshormonogenesis. It is caused by biallelic mutations in the SLC26A4 gene encoding for pendrin. Hypothyroidism in PS can be present from birth and therefore diagnosed by neonatal screening. The aim of this study was to examine the SLC26A4 mutation spectrum and prevalence among congenital hypothyroidism (CH) patients in the Guangxi Zhuang Autonomous Region of China and to establish how frequently PS causes hearing impairment in our patients with CH. SUBJECTS AND METHODS Blood samples were collected from 192 CH patients in Guangxi Zhuang Autonomous Region, China, and genomic DNA was extracted from peripheral blood leukocytes. All exons of the SLC26A4 gene together with their exon-intron boundaries were screened by next-generation sequencing. Patients with SLC26A4 mutations underwent a complete audiological evaluation including otoscopic examination, audiometry and morphological evaluation of the inner ear. RESULTS Next generation sequencing analysis of SLC26A4 in 192 CH patients revealed five different heterozygous variations in eight individuals (8/192, 4%). The prevalence of SLC26A4 mutations was 4% among studied Chinese CH. Three of the eight were diagnosed as enlargement of the vestibular aqueduct (EVA), no PS were found in our 192 CH patients. The mutations included one novel missense variant p.P469S, as well as four known missense variants, namely p.V233L, p.M147I, p.V609G and p.D661E. Of the eight patients identified with SLC26A4 variations in our study, seven patients showed normal size/location of thyroid gland, and one patients showed a decreased size one. CONCLUSIONS The prevalence of SLC26A4 pathogenic variants was 4% among studied Chinese patients with CH. Our study expanded the SLC26A4 mutation spectrum, provided the best estimation of SLC26A4 mutation rate for Chinese CH patients and indicated the rarity of PS as a cause of CH.

PAX8 pathogenic variants in Chinese patients with congenital hypothyroidism.

BACKGROUND The clinical presentation of patients with congenital hypothyroidism (CH) caused by paired box gene 8 (PAX8) pathogenic variants is variable and PAX8 mutation rates differ significantly among different populations. This study was set to examine the PAX8 mutation spectrum and prevalence among patients with CH in Guangxi Zhuang Autonomous Region, China. METHODS Peripheral venous blood samples were collected from the patients. Genomic DNA was extracted from peripheral blood leukocytes. All exons of the 11 known CH associated genes including PAX8 together with their exon-intron boundaries were screened by next-generation sequencing (NGS). Permanent or transient CH was determined using the results of thyroid function tests after temporary withdrawal of L-thyroxine (L-T4) therapy at approximately 2 years of age. RESULTS Next generation sequencing analysis of PAX8 in 378 CH patients revealed five different mutations in nine individuals (two are siblings). The mutations included two known missense variants, namely c.92G>A (p.R31H) and c.91C>T (p.R31C), and one novel missense variant c.68G>T (p.G23V), as well as two novel nonsense variants c.1090C>T (p. R364X) and c.658C>T (p.R220X). The variant c.92G>A (p.R31H) is highly recurrent in our patient cohort but the clinical phenotypes vary greatly among those carrying this variant. PAX8 pathogenic variants were mainly associated with permanent CH. CONCLUSION The prevalence of PAX8 pathogenic variants was 2.38% among patients with CH in Guangxi. Our study expanded the PAX8 mutation spectrum and provided the best estimation of PAX8 mutation rate among CH patients in Guangxi, China.

Mutation screening of the sodium iodide symporter gene in a cohort of 105 China patients with congenital hypothyroidism

OBJECTIVE Dyshormonogenetic congenital hypothyroidism (CH) was reported to be associated with a mutation in the sodium iodide symporter (NIS) gene. The present study was undertaken in the Guangxi Zhuang Autonomous Region of China, to determine the nature and frequency of NIS gene mutations among patients with CH due to dyshormonogenesis. SUBJECTS AND METHODS Blood samples were collected from 105 dyshormonogenetic CH patients in Guangxi Zhuang Autonomous Region, China, and genomic DNA was extracted from peripheral blood leukocytes. All exons of the NIS gene together with their exon-intron boundaries were screened by next-generation sequencing. RESULTS Two silent variations (T221T and T557T) and one missense variation (M435L), as well as two polymorphisms (rs200587561 and rs117626343) were found. CONCLUSIONS Our results indicate that the NIS mutation rate is very low in the Guangxi Zhuang Autonomous Region, China, and it is necessary to study mutations of other genes that have major effects on thyroid dyshormonogenesis and have not as yet been studied in this population.

Next-generation sequencing analysis of twelve known causative genes in congenital hypothyroidism

BACKGROUND Gene variants have been reported to be associated with congenital hypothyroidism (CH), the purpose of this study was to analyze the mutation spectrum and prevalence of 12 known causative genes (TSHR, PAX8, NKX2.1, NKX2.5, FOXE1, DUOX2, TG, TPO, GLIS3, NIS, SLC26A4 and DEHAL1) in CH in China. METHODS Peripheral venous blood samples were collected from the patients. Genomic DNA was extracted from peripheral blood leukocytes. All exons and their exon-intron boundary sequences of the 12 known CH associated genes in 66 CH patients were screened by next-generation sequencing (NGS). RESULTS NGS analysis of 12 known CH associated genes revealed that 32 patients (32/66, 48.5%) were detected to have at least one potentially functional variant. 21, 9, 1, 1, 1 and 1 patients were found to have potential pathogenic variants in DUOX2, TG, PAX8, SLC26A4, TSHR and TPO genes, respectively. Novel variants included one DUOX2 and one TPO missense variants of unknown significance (VUS). CONCLUSION Our study expands the mutation spectrum of DUOX2 and TPO genes. 48.5% CH patients had at least one potential pathogenic variant. We found relatively high frequency of DUOX2 (31.8%) and TG (13.6%) mutations in our cohort.

Hb H Hydrops Fetalis Syndrome Caused by Association of the − −SEA Deletion and Hb Constant Spring (HBA2: c.427T > C) Mutation in a Chinese Family

Abstract Hb Constant Spring (Hb CS; HBA2: c.427T > C) is an unstable hemoglobin (Hb) variant that results from a nucleotide substitution at the termination codon of the α2-globin gene. Compound heterozygosity for α0-thalassemia (α0-thal) and Hb CS (− −SEA/αCSα) results in Hb H/Hb CS disease, which is generally characterized with mild hemolytic anemia, jaundice, and splenomegaly. Here, we describe one case with Hb H/Hb CS disease that presented with fetal anemia and fetal hydrops, known as Hb H (β4) hydrops fetalis. This is the first report of fetal hydrops caused by association of the − −SEA deletion and the αCSα mutation. Our study highlights the significance of watchful observation using a serial ultrasound method and care of pregnant women who have fetuses found to carry Hb H/Hb CS disease during pregnancy, to guard against the occurrence of fetal hydrops.

Maternal uniparental disomy 14 and mosaic trisomy 14 in a Chinese boy with moderate to severe intellectual disability

BackgroundBoth maternal uniparental disomy 14 (UPD(14)mat) and mosaic trisomy 14 are rare events in live individuals. A combination of the two events in one individual is rarely encountered. Only six live-born cases have so far been reported.Case presentationHere we reported a case of concomitant UPD(14)mat and mosaic trisomy 14 in a 10-year-old Chinese patient. Most clinical features of our patient were consistent with those previous reported for UPD(14)mat cases, which include prenatal and postnatal growth retardation, neonatal hypotonia, feeding difficulty, intellectual disability, truncal obesity, small hands and feet, short stature, and mild facial dysmorphism, but our patient showed more severe intellectual disability and no sign of precocious puberty. SNP array analysis revealed a mixture of chromosome 14 maternal isodisomy with heterodisomy and a low level trisomy mosaicism of whole chromsome 14 in blood and hyperpigmented skin samples, whereas only UPD(14)mat was detected in normal skin sample. Cytogenetic analysis identified one trisomy 14 cell in 100 metaphase of peripheral blood lymphocytes (47,XX, +14[1]/46,XX[99]).ConclusionsTo our knowledge, this is the first case of a patient with UPD(14)mat and mosaic trisomy 14 reported in a Chinese patient. The definitive genetic diagnosis is beneficial for genetic counseling and clinical management of our patient, and for improving our understanding of the genotype-phenotype correlations of concomitant UPD(14)mat and mosaic trisomy 14.

de novo interstitial deletions at the 11q23.3-q24.2 region

BackgroundJacobsen syndrome (JBS) is a contiguous gene deletion syndrome involving 11q terminal deletion. Interstitial deletions at distal 11q are rare and their contributions to the clinical phenotype of JBS are unknown.Case presentationWe presented the chromosome microarray (CMA) data and the clinical features of two individuals carrying a non-overlapping de novo deletion each at the 11q23.3-q24.2 region in an effort to analyze the correlation between region of deletion at 11q and phenotype. Both deletions are likely pathogenic for patient’s condition. The deletion at 11q23.3q24.1 is associated with short stature, relative microcephaly, failure to thrive, hypotonia and sleeping disorder. The deletion at 11q24.2 involves HEPACAM and our patient’s clinical presentation (relative macrocephaly, abnormal MRI, mild developmental delay and seizure) is not inconsistent with Megalencephalic leukoencephalopathy with subcortical cysts 2B.ConclusionsOur finds support the notion that more than one critical region at 11q23.3-qter are responsible for the variable clinical presentation of JBS, thus JBS is a true contiguous gene deletion syndrome where multiple loci contributed to the clinical characteristics of JBS. Small interstitial deletions at 11q23.3-q24.2 and their associated unique features also suggest emerging novel genomic disorders.