miR-124-3p improves mitochondrial function of renal tubular epithelial cells in db/db mice.

Diabetic kidney disease (DKD) is one of the most serious complications of diabetes mellitus (DM) and the main cause of end-stage renal failure. However, the pathogenesis of DKD is complicated. In this study, we found that miR-124-3p plays a key role in regulating renal mitochondrial function and explored its possible mechanism in DKD progression by performing a series of in vitro and in vivo experiments. Decreased expression of miR-124-3p was found in db/db mice compared to db/m mice. Moreover, miR-124-3p down-regulated FOXQ1 by targeting FOXQ1 mRNA 3'-UTR in NRK-52E cells. Also, an increase in FOXQ1 and down-regulation of Sirt4 were found in db/db mouse kidney and renal tubular epithelial cells cultured with high glucose and high lipid. Overexpression of FOXQ1 could further down-regulate the expression of Sirt4 and aggravate the damage of mitochondria. Conversely, the knockdown of the FOXQ1 gene induced Sirt4 expression and partially restored mitochondrial function. To verify the effects of miR-124-3p on Sirt4 and mitochondria, we found that miR-124-3p mimics could up-regulate Sirt4 and inhibit ROS production and MitoSOX, thus restoring the number and morphology of mitochondria. These results showed that under high-glucose and high-lipid conditions, the down-regulation of miR-124-3p induces FOXQ1 in renal tubular epithelial cells, which in turn suppresses Sirt4 and leads to mitochondrial dysfunction, promoting the development of DKD.

Alpha lipoamide inhibits diabetic kidney fibrosis via improving mitochondrial function and regulating RXRα expression and activation.

Previous studies have shown mitochondrial dysfunction in various acute kidney injuries and chronic kidney diseases. Lipoic acid exerts potent effects on oxidant stress and modulation of mitochondrial function in damaged organ. In this study we investigated whether alpha lipoamide (ALM), a derivative of lipoic acid, exerted a renal protective effect in a type 2 diabetes mellitus mouse model. 9-week-old db/db mice were treated with ALM (50 mg·kg-1·d-1, i.g) for 8 weeks. We showed that ALM administration did not affect blood glucose levels in db/db mice, but restored renal function and significantly improved fibrosis of kidneys. We demonstrated that ALM administration significantly ameliorated mitochondrial dysfunction and tubulointerstitial fibrotic lesions, along with increased expression of CDX2 and CFTR and decreased expression of ß-catenin and Snail in kidneys of db/db mice. Similar protective effects were observed in rat renal tubular epithelial cell line NRK-52E cultured in high-glucose medium following treatment with ALM (200 µM). The protective mechanisms of ALM in diabetic kidney disease (DKD) were further explored: Autodock Vina software predicted that ALM could activate RXRα protein by forming stable hydrogen bonds. PROMO Database predicted that RXRα could bind the promoter sequences of CDX2 gene. Knockdown of RXRα expression in NRK-52E cells under normal glucose condition suppressed CDX2 expression and promoted phenotypic changes in renal tubular epithelial cells. However, RXRα overexpression increased CDX2 expression which in turn inhibited high glucose-mediated renal tubular epithelial cell injury. Therefore, we reveal the protective effect of ALM on DKD and its possible potential targets: ALM ameliorates mitochondrial dysfunction and regulates the CDX2/CFTR/ß-catenin signaling axis through upregulation and activation of RXRα. Schematic figure illustrating that ALM alleviates diabetic kidney disease by improving mitochondrial function and upregulation and activation of RXRα, which in turn upregulated CDX2 to exert an inhibitory effect on ß-catenin activation and nuclear translocation. RTEC renal tubular epithelial cell. ROS Reactive oxygen species. RXRα Retinoid X receptor-α. Mfn1 Mitofusin 1. Drp1 dynamic-related protein 1. MDA malondialdehyde. 4-HNE 4-hydroxynonenal. T-SOD Total-superoxide dismutase. CDX2 Caudal-type homeobox transcription factor 2. CFTR Cystic fibrosis transmembrane conductance regulator. EMT epithelial mesenchymal transition. α-SMA Alpha-smooth muscle actin. ECM extracellular matrix. DKD diabetic kidney disease. Schematic figure was drawn by Figdraw ( www.figdraw.com ).

Atorvastatin Restores PPARα Inhibition of Lipid Metabolism Disorders by Downregulating miR-21 Expression to Improve Mitochondrial Function and Alleviate Diabetic Nephropathy Progression.

Atorvastatin is a classical lipid-lowering drug. It has been reported to have renoprotective effects, such as reducing urinary protein excretion and extracellular matrix aggregation. The present study aimed to investigate the specific mechanism of action of Atorvastatin in type 1 diabetic mice (T1DM) in inhibiting renal tubular epithelial cell injury following treatment with high glucose and high fat. The anti-injury mechanism of Atorvastatin involved the inhibition of miR-21 expression and the upregulation of the transcription and expression of its downstream gene Peroxisome proliferator-activated receptors-α(PPARα). An increase in blood glucose and lipid levels was noted in the T1DM model, which was associated with renal fibrosis and inflammation. These changes were accompanied by increased miR-21 levels, downregulation of PPARα and Mfn1 expressions, and upregulation of Drp1 and IL6 expressions in renal tissues. These phenomena were reversed following the administration of Atorvastatin. miR-21 targeted PPARα by inhibiting its mRNA translation. Inhibition of miR-21 expression or Fenofibrate (PPARα agonist) administration prevented the decrease of PPARα in renal tubular epithelial cells under high glucose (HG) and high fat (Palmitic acid, PA) conditions, alleviating lipid metabolism disorders and reducing mitochondrial dynamics and inflammation. Consistent with the in vivo results, the in vitro findings also demonstrated that mRTECs administered with Atorvastatin in HG + PA increased PPARα expression and restored the normal expression of Mfn1 and Drp1, and effectively increasing the number of biologically active mitochondria and ATP content, reducing ROS production, and restoring mitochondrial membrane potential following Atorvastatin intervention. In addition, these effects were noted to the inhibition of FN expression and tubular cell inflammatory response; however, in the presence of miR-21mimics, the aforementioned effects of Atorvastatin were significantly diminished. Based on these observations, we conclude that Atorvastatin inhibits tubular epithelial cell injury in T1DM with concomitant induction of lipid metabolism disorders by a mechanism involving inhibition of miR-21 expression and consequent upregulation of PPARα expression. Moreover, Atorvastatin regulated lipid metabolism homeostasis and PPARα to restore mitochondrial function. The results emphasize the potential of Atorvastatin to exhibit lipid-regulating functions and non-lipid effects that balance mitochondrial dynamics.

Senescent renal tubular epithelial cells activate fibroblasts by secreting Shh to promote the progression of diabetic kidney disease.

Introduction: Diabetic kidney disease (DKD) is one of the complications of diabetes; however, the pathogenesis is not yet clear. A recent study has shown that senescence is associated with the course of DKD. In the present study, we explored whether senescent renal tubular cells promote renal tubulointerstitial fibrosis by secreting Sonic hedgehog (Shh) which mediates fibroblast activation and proliferation in DKD. Methods: A 36-week-old db/db mice model and the renal tubular epithelial cells were cultured in high glucose (HG, 60 mmol/L) medium for in vivo and in vitro experiments. Results: Compared to db/m mice, blood glucose, microalbuminuria, serum creatinine, urea nitrogen, and UACR (microalbuminuria/urine creatinine) were markedly increased in db/db mice. Collagen III, monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) were also increased in db/db mice kidneys, suggesting fibrosis and inflammation in the organ. Moreover, the detection of SA-ß-galactosidase (SA-ß-Gal) showed that the activity of SA-ß-Gal in the cytoplasm of renal tubular epithelial cells increased, and the cell cycle inhibition of the expression of senescence-related gene cell cycle inhibitor p16 INK4A protein and p21 protein increased, indicating that renal fibrosis in db/db mice was accompanied by cell senescence. Furthermore, Shh is highly expressed in the injured renal tubules and in the kidney tissue of db/db mice, as detected by enzyme-linked immunosorbent assay (ELISA). The results of immunofluorescence staining showed increased positive staining for Shh in renal tubular epithelial cells of db/db mice and decreased positive staining for Lamin B1, but increased positive staining for γH2A.X in cells with high Shh expression; similar results were obtained in vitro. In addition, HG stimulated renal tubular epithelial cells to secrete Shh in the supernatant of the medium. D-gal treatment of renal tubular epithelial cells increased the protein levels of Shh and p21. We also found enhanced activation and proliferation of fibroblasts cultured with the supernatant of renal tubular epithelial cells stimulated by HG medium but the proliferative effect was significantly diminished when co-cultured with cyclopamine (CPN), an inhibitor of the Shh pathway. Discussion: In conclusion, HG induces renal tubular epithelial cell senescence, and the secretion of senescence-associated proteins and Shh mediates inflammatory responses and fibroblast activation and proliferation, ultimately leading to renal fibrosis.

[Application of SNP-array technology in the genetic analysis of pediatric patients with growth retardation].

OBJECTIVE: To explore the value of single nucleotide polymorphism array (SNP-array) for the analysis of pediatric patients with growth retardation. METHODS: One hundred eighty one children with growth retardation were enrolled. DNA was extracted from peripheral samples from the patients, and whole genome copy number variations (CNVs) were detected using Illumina Human Cyto SNP-12. All identified CNVs were further analyzed with reference to databases including ClinGen, ClinVar, DECIPHER, OMIM and DGV as well as comprehensive review of literature from PubMed to determine their pathogenicity. RESULTS: Forty seven patients (26%) with abnormal CNVs were detected, which included 12 known microdeletions/microduplications syndrome (26%), 10 pathogenic non-syndromic CNVs (21%), 3 numerical chromosome aberrations (6%), 3 unbalanced translocations (6%), 4 pathogenic mosaicisms (9%) and 15 cases with unknown clinical significance (32%). After excluding obvious numerical and/or structural chromosomal abnormalities, this study has detected 15 pathogenic microdeletions/microduplications sized 5 Mb or less, which may be missed by routine chromosomal karyotyping. In addition, there were 3 cases with loss of heterozygoisty (LOH) containing known or predicted imprinting genes as well as 2 cases with suspected parental consanguinity. CONCLUSION: SNP-array technology is a powerful tool for the genetic diagnosis of children with growth disorders with advantages of high resolution and improved accuracy.

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.

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.

[A novel compound heterozygous mutation causing 3-methylcrotonyl-CoA carboxylase deficiency].

OBJECTIVE: To explore the molecular mechanism for a boy suspected with 3-methylcrotonyl-CoA carboxylase deficiency by neonatal screening. METHODS: PCR and Sanger sequencing were used to identify potential mutations of MCCC1 and MCCC2 genes. SIFT and Polyphen-2 software was used to predict the effect of variant on the protein function and conservation of the variant across various species. Human Splicing Finder and Swiss-PdbViewer4.1.0 were applied to analyze the possible mechanism of the variant. RESULTS: For the proband, a compound heterozygous mutation was discovered in the MCCC1 gene, namely c.539G>T (p.G180V) and c.704_711del (p.A235Vfs*4), which were inherited from his father and mother, respectively. The two mutations have disrupted the protein conformation, which in turn may impact the function of MCC protein. CONCLUSION: The compound heterozygous mutations of the MCCC1 gene may contribute to the 3-methylcrotonyl-CoA carboxylase deficiency manifested by the patient.

A rare occurrence of two large de novo duplications on 1q42-q44 and 9q21.12-q21.33.

De novo partial distal 1q trisomy is uncommon and mostly occurs in combination with monosomy of another chromosome due to a parental translocation. Distal 1q trisomy co-occurring with another de novo duplication on a separate chromosome is extremely rare. Here, we reported a patient carrying two large de novo interstitial duplications including a 20Mb duplication at 1q42-q44 and a 14.2Mb duplication at 9q21.12-q21.33. The patient presented with features of pre- and postnatal growth retardation, low birth weight, failure to thrive, developmental delay and frequent infection. Her dysmorphic features included macrocephaly, prominent forehead, triangular face, wide fontanelle, hypertelorism, flat nasal bridge, tented mouth, micrognathia, protruding and low-set ears, slender limbs with toe-walking appearance. In addition, she presented with subdural hematoma. The clinical presentations of this patient are mostly consistent with those of distal 1q trisomy syndrome or 9q interstitial duplication. The interstitial 1q trisomy may have contributed to the macrocephaly, prominent forehead and limb abnormalities of our patient. Either or both de novo duplications could have contributed to the features of growth retardation, developmental delay and dysmorphic features including hypertelorism, low-set ears and abnormal nose/nasal bridge.

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

BACKGROUND: Both 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 PRESENTATION: Here 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]). CONCLUSIONS: To 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.

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

ABSTRACT 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.

A novel de novo microdeletion at 17q11.2 adjacent to NF1 gene associated with developmental delay, short stature, microcephaly and dysmorphic features.

BACKGROUND: Microdeletions at 17q11.2 often encompass NF1 gene, is the cause for NF1 microdeletion syndrome. Microdeletion at 17q11.2 without the involvement of NF1 gene is rarely reported. CASE PRESENTATION: Here we reported a patient carrying a novel de novo deletion at 17q11.2 adjacent to NF1 gene, who presented with developmental delay, short stature, postnatal microcephaly, underweight and dysmorphic features including flat facial profile, dolicocephaly, hypertelorism, short philtrum, flat nasal bridge and posteriorly rotated and low set ears. Chromosomal microarray analysis revealed a 1.69 Mb de novo deletion at 17q11.2 adjacent to NF1 gene, which involves 43 RefSeq genes. We compared this with four overlapping deletions at this interval. CONCLUSIONS: A rare de novo microdeletion at 17q11.2 not involving NF1 gene is associated with developmental delay and dysmorphic features. Seven genes, TAOK1, PHF12, NUFIP2, SLC26A4, SEZ6, GIT1 and TRAF4 are possible candidates for the clinical features of our patient. The delineation of this rare deletion and description of associated clinical phenotypes will help to understand the genotype-phenotype correlation of genomic imbalances at this locus.

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

BACKGROUND: Jacobsen 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 PRESENTATION: We 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. CONCLUSIONS: Our 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.

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.

Persistent and energetic bottom-trapped topographic Rossby waves observed in the southern South China Sea.

Energetic fluctuations with periods of 9-14 days below a depth of 1400 m were observed in the southern South China Sea (SCS) from 5 years of direct measurements. We interpreted such fluctuations as topographic Rossby waves (TRWs) because they obey the dispersion relation. The TRWs persisted from May 24, 2009 to August 23, 2013, and their bottom current speed with a maximum of ~10 cm/s was one order of magnitude greater than the mean current and comparable to the tidal currents near the bottom. The bottom-trapped TRWs had an approximate trapping depth of 325 m and reference wavelength of ~82 km, which were likely excited by eddies above. Upper layer current speed that peaked approximately every 2 months could offer the energy sources for the persistent TRWs in the southern SCS. Energetic bottom-trapped TRWs may have a comparable role in deep circulation to tides in areas with complex topography.

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.

SCSPOD14, a South China Sea physical oceanographic dataset derived from in situ measurements during 1919-2014.

In addition to the oceanographic data available for the South China Sea (SCS) from the World Ocean Database (WOD) and Array for Real-time Geostrophic Oceanography (Argo) floats, a suite of observations has been made by the South China Sea Institute of Oceanology (SCSIO) starting from the 1970s. Here, we assemble a SCS Physical Oceanographic Dataset (SCSPOD14) based on 51,392 validated temperature and salinity profiles collected from these three datasets for the period 1919-2014. A gridded dataset of climatological monthly mean temperature, salinity, and mixed and isothermal layer depth derived from an objective analysis of profiles is also presented. Comparisons with the World Ocean Atlas (WOA) and IFREMER/LOS Mixed Layer Depth Climatology confirm the reliability of the new dataset. This unique dataset offers an invaluable baseline perspective on the thermodynamic processes, spatial and temporal variability of water masses, and basin-scale and mesoscale oceanic structures in the SCS. We anticipate improvements and regular updates to this product as more observations become available from existing and future in situ networks.

[Identification of two novel mutations of MUT gene in a Chinese family affected with isolated methylmalonic acidemia].

OBJECTIVE: To explore the molecular etiology for a Chinese family affected with isolated methylmalonic acidemia (MMA). METHODS: Potential mutations of MUT, MMAA and MMAB genes in the proband were screened by PCR and Sanger sequencing. The pathogenicity of identified mutations was analyzed using Polyphen2, SIFT, HSF, DNAMAN 6.0 and Swiss-PdbViewer4.1.0 software. RESULTS: Two novel mutations of the MUT gene, including c.581C>T (p.P194L) and c.1219A>T (p.N407Y), were discovered in the proband, which were inherited respectively from his mother and father. Bioinformatics analysis suggested that both mutations were damaging. The affected codons P194 and N407, both located in the (beta, alpha) 8 barrel domain and to which the substrate methylmalonyl-CoA is bound, are highly conserved across various species. Both mutations can disrupt the space conformation of its protein product, affecting the function of the MCM protein. CONCLUSION: The novel mutations of MUT gene probably underlie the isolated MMA in this family.

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.

Thyroglobulin gene mutations in Chinese patients with congenital hypothyroidism.

Mutations in Thyroglobulin (TG) are common genetic causes of congenital hypothyroidism (CH). But the TG mutation spectrum and its frequency in Chinese CH patients have not been investigated. Here we conducted a genetic screening of TG gene in a cohort of 382 Chinese CH patients. We identified 22 rare non-polymorphic variants including six truncating variants and 16 missense variants of unknown significance (VUS). Seven patients carried homozygous pathogenic variants, and three patients carried homozygous or compound heterozygous VUS. 48 out of 382 patients carried one of 18 heterozygous VUS which is significantly more often than their occurrences in control cohort (P < 0.0001). Unique to Asian population, the c.274+2T>G variant is the most common pathogenic variant with an allele frequency of 0.021. The prevalence of CH due to TG gene defect in Chinese population was estimated to be approximately 1/101,000. Our study uncovered ethnicity specific TG mutation spectrum and frequency.