Phenotype and genotype spectra of a Chinese cohort with nephronophthisis-related ciliopathy.

BACKGROUND: Nephronophthisis-related ciliopathies (NPHP-RC) account for the majority of cases of monogenetically caused end-stage renal disease (ESRD) in children. Exploring the correlation between the phenotype and genotype of NPHP-RC is helpful for early diagnosis and management. We investigated the phenotype and genotype spectra of NPHP-RC in a Chinese multicentre cohort. METHODS: Crosss-ectional and longitudinal data of 60 patients from 57 families with pathogenic NPHP-RC gene mutations distributed in 22 regions of China were collected into a unified, anonymous database. The mean observation time of this cohort was 3.5±3.1 years. RESULTS: Mutations in NPHP1 and NPHP3 were the most common genetic defects. Overall, 45% of patients presented with isolated nephronophthisis (NPH), and 55% exhibited the extrarenal phenotype, which frequently involved the liver (41.7%, n=25), central nervous system (26.7%, n=16), eyes (26.7%, n=16) and skeletal system (11.7%, n=7). Accidental detection of elevated serum creatinine and non-specific symptoms caused by chronic kidney disease occurred in 65% of patients. Patients carrying NPHP1 mutations mainly presented with isolated NPH (90%, 18/20) and progressed to ESRD at a mean age of 12.9±0.5 years. The mean age of ESRD onset in the non-NPHP1 group was lower than that in the NPHP1 group (6.2±1.4 years, p<0.001), especially for patients carrying NPHP3 mutations (3.1±1.2 years), showing a heterogeneous phenotype characterised by Bardet-Biedl syndrome (12.5%, n=5), Joubert syndrome (7.5%, n=3), COACH syndrome (2.5%, n=1), Mainzer-Saldino syndrome (2.5%, n=1), short-rib thoracic dysplasia (2.5%, n=1) and unclassified symptoms (32.5%, n=13). CONCLUSIONS: The Chinese Children Genetic Kidney Disease Database registry characterised the spectrum of the phenotype and genotype of NPHP-RC in the Chinese population. NPHP1 and NPHP3 were the most common pathogenic genes. Rapid progression to ESRD and liver involvement were noted in patients with NPHP3 mutations.

STARD3NL inhibits the osteogenic differentiation by inactivating the Wnt/ß-catenin pathway via binding to Annexin A2 in osteoporosis.

Osteoporosis is one of the leading forms of systemic diseases related to bone metabolism in the world. STARD3 N-terminal like (STARD3NL) showed robust association with osteoporosis-related traits. Yet, the molecular functional mechanisms of STARD3NL in osteoblasts is still obscure. In this study, we demonstrated a high level of STARD3NL expression in the bone tissues from the patients with low bone mass and ovariectomized (OVX)-induced osteoporotic mice. We identified Stard3nl as a potent factor that negatively and positively regulates osteoblast differentiation and cell proliferation, respectively. Furthermore, inhibition of Stard3nl induced ß-catenin gene expression and the nuclear translocation of ß-catenin, as well as Wnt signalling activities, contributing to the activation of Wnt/ß-catenin signalling. Mechanistic studies revealed that Stard3nl bound with Annexin A2 (Anxa2) to suppress ß-catenin expression, resulting into the suppression of Wnt signalling and downstream osteogenic differentiation. Moreover, adeno-associated virus 9 (AAV9)-mediated silencing of Stard3nl reversed bone loss in OVX-induced osteoporotic mice by the injection into the knee joints. Collectively, our study revealed that Stard3nl suppressed osteogenesis via binding with Anxa2, resulting into the inactivation of Wnt signalling. It also highlights the preventive and therapeutic potential of STARD3NL as a specific and novel target for osteoporotic patients.

PARD3 gene variation as candidate cause of nonsyndromic cleft palate only.

Nonsyndromic cleft palate only (NSCP) is a common congenital malformation worldwide. In this study, we report a three-generation pedigree with NSCP following the autosomal-dominant pattern. Whole-exome sequencing and Sanger sequencing revealed that only the frameshift variant c.1012dupG [p. E338Gfs*26] in PARD3 cosegregated with the disease. In zebrafish embryos, ethmoid plate patterning defects were observed with PARD3 ortholog disruption or expression of patient-derived N-terminal truncating PARD3 (c.1012dupG), which implicated PARD3 in ethmoid plate morphogenesis. PARD3 plays vital roles in determining cellular polarity. Compared with the apical distribution of wild-type PARD3, PARD3-p. E338Gfs*26 mainly localized to the basal membrane in 3D-cultured MCF-10A epithelial cells. The interaction between PARD3-p. E338Gfs*26 and endogenous PARD3 was identified by LC-MS/MS and validated by co-IP. Immunofluorescence analysis showed that PARD3-p. E338Gfs*26 substantially altered the localization of endogenous PARD3 to the basement membrane in 3D-cultured MCF-10A cells. Furthermore, seven variants, including one nonsense variant and six missense variants, were identified in the coding region of PARD3 in sporadic cases with NSCP. Subsequent analysis showed that PARD3-p. R133*, like the insertion variant of c.1012dupG, also changed the localization of endogenous full-length PARD3 and that its expression induced abnormal ethmoid plate morphogenesis in zebrafish. Based on these data, we reveal PARD3 gene variation as a novel candidate cause of nonsyndromic cleft palate only.

Downregulation of MEG3 and upregulation of EZH2 cooperatively promote neuroblastoma progression.

Neuroblastoma (NB), an embryonic tumour originating from sympathetic crest cells, is the most common extracranial solid tumour type in children with poor overall prognosis. Accumulating evidence has demonstrated the involvement of long non-coding RNA (lncRNA) in numerous biological processes and their associations with embryonic development and multiple diseases. Ectopic lncRNA expression is linked to malignant tumours. Previous studies by our team indicate that MEG3 attenuates NB autophagy through inhibition of FOXO1 and epithelial-mesenchymal transition via the mTOR pathway in vitro. Moreover, MEG3 and EZH2 negatively regulate each other. In present study, we first collected 60 NB tissues and 20 adjacent tissues for Quantitative real-time polymerase chain reaction (Q-PCR) experiments and performed clinical correlation analysis of the results. At the same time, nude mice were used for subcutaneous tumour formation to detect the effect of MEG3 in vivo. Two NB cell lines, SK-N-AS and SK-N-BE(2)C, were overexpressed MEG3 and rescued with EZH2 and then were subjected to proliferation, migration, invasion, apoptosis and autophagy experiments. RNA-binding protein immunoprecipitation (RIP) and Co-Immunoprecipitation (Co-IP) experiments were performed to explore the molecular mechanism of MEG3 and EZH2 interaction. Q-PCR revealed that MEG3 expression was negatively correlated with INSS stage and risk grade of NB. Moreover, MEG3 overexpression was associated with inhibition of NB growth in vivo. MEG3 exerted an anti-cancer effect via stimulatory effects on EZH2 ubiquitination leading to its degradation. Conversely, EZH2 interacted with DNMT1 and HDAC1 to induce silencing of MEG3. The EZH2 inhibitor, DZNep, and HDAC inhibitor, SAHA, displayed synergistic activity against NB. Combined treatment with DZNep and SAHA inhibited proliferation, migration and invasion of NB through suppression of the PI3K/AKT/mTOR/FOXO1 pathway. In conclusion, downregulation of MEG3 and upregulation of EZH2 forms a feedback loop that concertedly promotes the development of NB. Combined blockage of EZH2 and HDAC1 with the appropriate inhibitors may therefore present an effective treatment strategy for NB cases with low MEG3 and high EZH2 expression.

A novel adiponectin receptor agonist (AdipoAI) ameliorates type 2 diabetes-associated periodontitis by enhancing autophagy in osteoclasts.

BACKGROUND AND OBJECTIVE: Type 2 diabetes (T2D)-associated periodontitis is severe and refractory in many cases. Considered an inflammatory disease, T2D predisposes to periodontitis by increasing whole-body inflammation. One mechanism of increased inflammation is thatT2D is mediated by loss of production or function of the anti-inflammatory hormone adiponectin. In our previous report, AdipoRon, an adiponectin receptor agonist, and AdipoAI, a newly discovered, more specific agonist, attenuated T2D-associated inflammation by inhibiting osteoclastogenesis and LPS-induced endotoxemia. Autophagy plays an important role during osteoclast differentiation and function. The impact of AdipoAI on osteoclast function and autophagy involved in osteoclastogenesis is not known. Here, we compare AdipoRon and AdipoAI potency, side effects and mechanism of action in T2D-associated periodontitis. METHODS: The RAW 264.7 cell line was used for in vitro studies. We analyzed the potential cytotoxicity of AdipoAI using the CCK-8 assay. The anti-osteoclastogenic potential of AdipoAI was studied by real-time qPCR and tartrate-resistant acid phosphatase staining. The actions of AdipoAI involved in autophagy were tested by real-time qPCR, western blot and immunofluorescence staining. In the diet-induced obesity model of T2D, we investigated the impact of AdipoAI on fasting blood glucose, alveolar bone loss, and gingival inflammation in mice with experimental periodontitis. RESULTS: AdipoRon inhibited osteoclastogenesis and AdipoAI inhibited osteoclastogenesis at lower doses than AdipoRon without any cytotoxicity. In DIO mice with experimental periodontitis, AdipoAI reduced mouse body weight in 14 days, reducing fasting glucose levels, alveolar bone destruction, osteoclast number along the alveolar bone surface, and decreased the expression of pro-inflammatory factors in periodontal tissues. AdipoAI and AdipoRon also enhanced LC3A/B expression when cultured with RANKL.3-Methyladenine, a known autophagy inhibitor, decreased LC3A/B expression and reversed the inhibition of osteoclastogenesis during AdipoAI treatment. CONCLUSIONS: Our results demonstrate that AdipoAI ameliorates the severity of T2D-associated periodontitis by enhancing autophagy in osteoclasts at lower doses than AdipoRon without demonstrable side effects. Thus, AdipoAI has pharmaceutical potential for treating diabetes-associated periodontal disease.

Advances in the Genetics of Congenital Ptosis.

Congenital ptosis, a birth defects presents at birth or by 1 year of age, is characterized by the drooping of the upper eyelid. Either in isolation (nonsyndromic) or with many different systemic disorders (syndromic). The estimated prevalence of ptosis (congenital and acquired) ranges from 0.79 to 1.99 per 10,000 people in different populations, and it is more prevalent in males. The underlying pathogenesis of congenital ptosis is myogenic and neurogenic, related to the development of muscles and nerves. Although most cases are sporadic, there are familial transmission characteristics, including autosomal dominant, recessive mode, and X-linkage inheritance patterns. Moreover, some forms are due to chromosomal aberrations and mutations and deletions in mitochondrial DNA. Genes involved in simple congenital ptosis (SCP) are ZFHX4 and COL25A1. The clinical aspects of various syndromes involving congenital ptosis are partly caused by single-gene mutations. However, the pathogenesis of congenital ptosis is not fully understood. We review the reported epidemiology, genetics, and clinical features of congenital ptosis and associated syndromes here.

ARID1B/SUB1-activated lncRNA HOXA-AS2 drives the malignant behaviour of hepatoblastoma through regulation of HOXA3.

It has been becoming increasingly evident that long non-coding RNAs (lncRNAs) play important roles in various human cancers. However, the biological processes and clinical significance of most lncRNAs in hepatoblastoma (HB) remain unclear. In our previous study, genome-wide analysis with a lncRNA microarray found that lncRNA HOXA-AS2 was up-regulated in HB. Stable transfected cell lines with HOXA-AS2 knockdown or overexpression were constructed in HepG2 and Huh6 cells, respectively. Our data revealed knockdown of HOXA-AS2 increased cell apoptosis and inhibited cell proliferation, migration and invasion in HB. Up-regulation of HOXA-AS2 promoted HB malignant biological behaviours. Mechanistic investigations indicated that HOXA-AS2 was modulated by chromatin remodelling factor ARID1B and transcription co-activator SUB1, thereby protecting HOXA3 from degradation. Therefore, HOXA-AS2 positively regulates HOXA3, which might partly demonstrate the involvement of HOXA3 in HOXA-AS2-mediated HB carcinogenesis. In conclusion, HOXA-AS2 is significantly overexpressed in HB and the ARID1B/HOXA-AS2/HOXA3 axis plays a critical role in HB tumorigenesis and development. These results might provide a potential new target for HB diagnosis and therapy.

Morphine promotes tumorigenesis and cetuximab resistance via EGFR signaling activation in human colorectal cancer.

Morphine, a mu-opioid receptor (MOR) agonist, has been extensively used to treat advanced cancer pain. In particular, in patients with cancer metastasis, both morphine and anticancer drugs are given simultaneously. However, evidence showed that morphine might be a risk factor in promoting the tumor's malignant potential. In this study, we report that treatment with morphine could activate MOR and lead to the promotion of proliferation, migration, and invasion in HCT116 and DLD1 colorectal cancer (CRC) cells with time-concentration dependence. Moreover, morphine can also contribute to cetuximab's drug resistance, a targeted drug widely used to treat advanced CRC by inducing the activation of epidermal growth factor receptor (EGFR). The cell phenotype includes proliferation, migration, invasion, and drug resistance, which may be reversed by MOR knockdown or adding nalmefene, the MOR receptor antagonist. Receptor tyrosine kinase array analysis revealed that morphine selectively induced the transactivation of EGFR. EGFR transactivation resulted in the activation of ERK1/2 and AKT. In conclusion, morphine induces the transactivation of EGFR via MOR. It activates the downstream signal pathway AKT-MTOR and RAS-MAPK, increases proliferation, migration, and invasion, and promotes resistance to EGFR inhibitors in a CRC cell line. Furthermore, we verified that EGFR inhibition by cetuximab strongly reversed the protumoral effects of morphine in vitro and in vivo. Collectively, we provide evidence that morphine-EGFR signaling might be a promising therapeutic target for CRC patients, especially for cetuximab-resistant CRC patients.

Recent advances in the epigenetics of bone metabolism.

Osteoporosis is a common form of metabolic bone disease that is costly to treat and is primarily diagnosed on the basis of bone mineral density. As the influences of genetic lesions and environmental factors are increasingly studied in the pathological development of osteoporosis, regulated epigenetics are emerging as the important pathogenesis mechanisms in osteoporosis. Recently, osteoporosis genome-wide association studies and multi-omics technologies have revealed that susceptibility loci and the misregulation of epigenetic modifiers are key factors in osteoporosis. Over the past decade, extensive studies have demonstrated epigenetic mechanisms, such as DNA methylation, histone/chromatin modifications, and non-coding RNAs, as potential contributing factors in osteoporosis that affect disease initiation and progression. Herein, we review recent advances in epigenetics in osteoporosis, with a focus on exploring the underlying mechanisms and potential diagnostic/prognostic biomarker applications for osteoporosis.

Diagnostic and clinical utility of genetic testing in children with kidney failure.

BACKGROUND: Genetic kidney disease is well established as an important cause of pediatric kidney failure, and genetic testing might increase diagnostic accuracy, but evidence is limited. This study was conducted to determine the diagnostic yield and clinical impact of genetic testing for children with kidney failure. METHODS: Patients who were diagnosed with kidney failure before 19 years of age at Children's Hospital of Fudan University from 2009 to 2018 and received next-generation sequencing (NGS) were enrolled. The results for likely pathogenic variants in genes known to cause chronic kidney disease (CKD) were analyzed. RESULTS: A molecular diagnosis was identified in 39.9% (75/188) of children with kidney failure. Specific subtype of clinical category was discerned in 54 (72.0%) patients, kidney disease was reclassified in 7 (9.3%) patients, the unknown etiology of 5 (6.7%) patients was molecularly diagnosed, and the clinical diagnoses of the other 9 (12.0%) patients were confirmed. In addition, genetic diagnosis was considered to have contributed to clinical management, including negating the need for kidney biopsy (26/75, 34.7%), avoiding immunosuppressive therapy (24/75, 32.0%), changing surveillance (48/75, 64.0%), guiding specific treatment (21/75, 28.0%), and guiding peri-transplant management and options for kidney transplantation (12/75, 16.0%). Furthermore, cascade testing was subsequently offered to 34.7% (26/75) of families. CONCLUSIONS: Genetic testing identified a molecular diagnosis in nearly 40% of children with kidney failure. Our results confirm that in children with kidney failure, genetic testing can not only establish a specific molecular diagnosis, but has a significant impact on clinical management.

The roles of SMYD4 in epigenetic regulation of cardiac development in zebrafish.

SMYD4 belongs to a family of lysine methyltransferases. We analyzed the role of smyd4 in zebrafish development by generating a smyd4 mutant zebrafish line (smyd4L544Efs*1) using the CRISPR/Cas9 technology. The maternal and zygotic smyd4L544Efs*1 mutants demonstrated severe cardiac malformations, including defects in left-right patterning and looping and hypoplastic ventricles, suggesting that smyd4 was critical for heart development. Importantly, we identified two rare SMYD4 genetic variants in a 208-patient cohort with congenital heart defects. Both biochemical and functional analyses indicated that SMYD4(G345D) was pathogenic. Our data suggested that smyd4 functions as a histone methyltransferase and, by interacting with HDAC1, also serves as a potential modulator for histone acetylation. Transcriptome and bioinformatics analyses of smyd4L544Efs*1 and wild-type developing hearts suggested that smyd4 is a key epigenetic regulator involved in regulating endoplasmic reticulum-mediated protein processing and several important metabolic pathways in developing zebrafish hearts.

Hypermethylation-mediated down-regulation of lncRNA TBX5-AS1:2 in Tetralogy of Fallot inhibits cell proliferation by reducing TBX5 expression.

Tetralogy of Fallot (TOF) is the most common complex congenital heart disease (CHD) with uncertain cause. Although long non-coding RNAs (lncRNAs) have been implicated in heart development and several CHDs, their role in TOF is not well understood. This study aimed to investigate how dysregulated lncRNAs contribute to TOF. Using Gene Expression Omnibus data mining, bioinformatics analysis and clinical heart tissue sample detecting, we identified a novel antisense lncRNA TBX5-AS1:2 with unknown function that was significantly down-regulated in injured cardiac tissues from TOF patients. LncRNA TBX5-AS1:2 was mainly located in the nucleus of the human embryonic kidney 293 (HEK293T) cells and formed an RNA-RNA double-stranded structure in the overlapping region with its sense mRNA T-box transcription factor 5 (TBX5), which is an important regulator in heart development. Knock-down of lncRNA TBX5-AS1:2 via promoter hypermethylation reduced TBX5 expression at both the mRNA and protein levels by affecting its mRNA stability through RNA-RNA interaction. Moreover, lncRNA TBX5-AS1:2 knock-down inhibited the proliferation of HEK293T cells. In conclusion, these results indicated that lncRNA TBX5-AS1:2 may be involved in TOF by affecting cell proliferation by targeting TBX5.

[A consensus on the standardization of the next generation sequencing process for the diagnosis of genetic diseases (1) - Procedures prior to genetic testing].

Pre-testing preparation is the basis and starting point of genetic testing. The process includes collection of clinical information, formulation of testing scheme, genetic counseling before testing, and completion of informed consent and testing authorization. To effectively identify genetic diseases in clinics can greatly improve the diagnostic rate of next generation sequencing (NGS), thereby reducing medical cost and improving clinical efficacy. The analysis of NGS results relies, to a large extent, on the understanding of genotype-phenotype correlations, therefore it is particularly important to collect and evaluate clinical phenotypes and describe them in uniform standard terms. Different types of genetic diseases or mutations may require specific testing techniques, which can yield twice the result with half the effort. Pre-testing genetic counseling can help patients and their families to understand the significance of relevant genetic testing, formulate individualized testing strategies, and lay a foundation for follow-up.

[A consensus on the standardization of the next generation sequencing process for the diagnosis of genetic diseases (4) - Report interpretation and genetic counseling].

Clinical genetic testing results are compiled into a standardized report by genetic specialists and provided to clinicians and patients (Should the patient be intellectually disabled or under 18, the report will be provided to his/her parents or legal guardians). The content of genetic testing report should conform to relevant guidelines, industry standards and consensus. The decisions of clinicians will be made based on the report and clinical indications. Genetic counselors should provide post-test counseling to clinicians and patients or their authorized family members. A mechanism of follow-up visit after the genetic testing should be established with informed consent. Data should be shared by clinical institutions and genome sequencing institutions. As findings upon follow-up visit can help with further evaluation of the results, genome sequencing institutions should regularly re-analyze historical and follow-up data, and the updated results should be shared with clinical institutions. All activities involving reporting, genetic counselling, follow-up visiting, and re-analyzing should follow the relevant guidelines and regulations.

Deregulated lncRNA expression profile in the mouse lung adenocarcinomas with KRAS-G12D mutation and P53 knockout.

Recent studies have demonstrated that aberrant long non-coding RNAs (lncRNAs) expression are suggested to be closely associated with multiple human diseases, lung cancer included. However, the roles of lncRNAs in lung cancer are not well understood. In this study, we used microarrays to investigate the aberrantly expressed lncRNAs in the mouse lung adenocarcinoma with P53 knockout and the KrasG12D mutation. Results revealed that 6424 lncRNAs were differentially expressed (≥ 2-fold change, P < .05). Two hundred and ten lncRNAs showed more than 8-fold change and conserved across human and were further analysed in the primary mouse lung adenocarcinoma KP cells, which were isolated from the p53 knockout and the KrasG12D mutation mice. Among all the 210 lncRNAs, 11 lncRNAs' expression was regulated by P53, 33 lncRNAs by KRAS and 13 lncRNAs by hypoxia in the primary KP cells, respectively. NONMMUT015812, which was remarkably up-regulated in the mouse lung adenocarcinoma and negatively regulated by the P53 re-expression, was detected to analyse its cellular function. Results showed that knockdown of NONMMUT015812 by shRNAs decreased proliferation and migration abilities of KP cells. Among those aberrantly expressed lncRNAs in the mouse lung adenocarcinoma, NONMMUT015812 was a potential oncogene.

Two novel ANK1 loss-of-function mutations in Chinese families with hereditary spherocytosis.

Hereditary spherocytosis (HS) is the most common inherited haemolytic anaemia disorder. ANK1 mutations account for most HS cases, but pathogenicity analysis and functional research have not been widely performed for these mutations. In this study, in order to confirm diagnosis, gene mutation was screened in two unrelated Chinese families with HS by a next-generation sequencing (NGS) panel and then confirmed by Sanger sequencing. Two novel heterozygous mutations (c.C841T, p.R281X and c.T290G, p.L97R) of the ANK1 gene were identified in the two families respectively. Then, the pathogenicity of the two new mutations and two previously reported ANK1 mutations (c.C648G, p.Y216X and c.G424T, p.E142X) were studied by in vitro experiments. The four mutations increased the osmotic fragility of cells, reduced the stabilities of ANK1 proteins and prevented the protein from localizing to the plasma membrane and interacting with SPTB and SLC4A1. We classified these four mutations into disease-causing mutations for HS. Thus, conducting the same mutation test and providing genetic counselling for the two families were meaningful and significant. Moreover, the identification of two novel mutations enriches the ANK1 mutation database, especially in China.

SLC44A4 mutation causes autosomal dominant hereditary postlingual non-syndromic mid-frequency hearing loss.

Clinical, genetic, and functional investigations were performed to identify the causative mutation in a distinctive Chinese family with postlingual non-syndromic mid-frequency sensorineural hearing loss. Whole-exome sequencing revealed SLC44A4, which encodes the choline transport protein, as the pathogenic gene in this family. In the zebrafish model, downregulation of slc44a4 using morpholinos led to significant abnormalities in the zebrafish inner ear and lateral line neuromasts and contributed, to some extent, to disabilities in hearing and balance. SH-SY5Y cells transfected with SLC44A4 showed higher choline uptake and acetylcholine release than that of cells transfected with mutant SLC44A4. We concluded that mutation of SLC44A4 may cause defects in the Choline- acetylcholine system, which is crucial to the efferent innervation of hair cells in the olivocochlear bundle for the maintenance of physiological function of outer hair cells and the protection of hair cells from acoustic injury, leading to hearing loss.

Novel TRRAP mutation causes autosomal dominant non-syndromic hearing loss.

Hereditary non-syndromic hearing loss is the most common inherited sensory defect in humans. More than 40 genes have been identified as causative genes for autosomal dominant non-syndromic hearing loss (ADNSHL), but there are many other candidate genes that remain to be discovered. We aimed to identify the causative gene mutation for post-lingual progressive ADNSHL in a Chinese family. Whole-exome sequencing, bioinformatic analysis, and Sanger sequencing were used to verify the co-segregation of a novel pathogenic variant (NM_ 001244580, c.511C>T, p.Arg171Cys) in the TRansformation/tRanscription domain-Associated Protein gene associated with hearing loss in a three-generation Chinese family with ADNSHL). Additionally, three more novel variants of transformation/transcription domain associated protein (TRRAP) were detected in 66 sporadic cases of hearing loss. Morpholino oligonucleotides knockdown and clustered regularly interspaced short palindromic repeats/Cas9 knockout zebrafish were constructed to validate the genetic findings. Knockdown or knockout of TRRAP resulted in significant defects in the inner ear of zebrafish, indicating that TRRAP plays an important role in inner ear development. In conclusion, TRRAP (NM_ 001244580, c.511C>T, p.Arg171Cys) co-segregated with hearing loss in a Chinese family with ADNSHL, and TRRAP deficiency caused hearing disability in zebrafish, suggesting TRRAP is a gene associated with ADNSHL.

Genetic spectrum of renal disease for 1001 Chinese children based on a multicenter registration system.

To explore the approaches and diagnostic yield of genetic testing for renal disease in children, we describe the genotype and phenotype of the national cohort of children with renal disease from 13 different regions of China recruited from 2014 to 2018 by building up the multicenter registration system (Chinese Children Genetic Kidney Disease Database, CCGKDD). Genetic diagnosis was confirmed in 42.1% of our cohort of 1001 pediatric patients with clinical suspicion of a genetic renal disease. Of the 106 distinct monogenetic disorders detected, 15 accounted for 60.7% of genetic diagnoses. The diagnostic yield was 29.1% in steroid resistant nephritic syndrome (SRNS), 61.4% in cystic renal disease, 17.0% in congenital anomalies of the kidney and urinary tract (CAKUT), 62.3% in renal tubular disease/renal calcinosis, and 23.9% for chronic kidney disease (CKD) 3 to 5 stage with unknown origin. Genetic approaches of target gene sequence (TGS), singleton whole-exome sequencing (WES) and trio-WES were performed with diagnostic rates of 44.8%, 36.2%, and 42.6%, respectively. The early use of trio-WES could improve the diagnostic rate especially in renal tubular disease and calcinosis. We report the genetic spectrum of Chinese children with renal disease. Establishment of the CCGKDD will improve the genetic work on renal disease.

A brief measure of predeath grief in dementia caregivers: the Caregiver Grief Questionnaire.

OBJECTIVE: The study of predeath grief is hampered by measures that are often lengthy and not clearly differentiated from other caregiving outcomes, most notably burden. We aimed to validate a new 11-item Caregiver Grief Questionnaire (CGQ) assessing two dimensions of predeath grief, namely relational deprivation and emotional pain. DESIGN: Cross-sectional survey. SETTING: Community and psychogeriatric clinics. PARTICIPANTS: 173 Alzheimer (AD) caregivers who cared for relatives with different degrees of severity (63 mild, 60 moderate, and 50 severe). MEASUREMENTS: Besides the CGQ, measures of caregiver burden and depressive symptoms, and care-recipients' neuropsychiatric symptoms and functional impairment were assessed. RESULTS: Confirmatory factor analysis supported the hypothesized 2-factor over the 1-factor model, and both subscales were only moderately correlated with burden. Two-week test-retest reliabilities were excellent. Caregivers for mild AD reported less grief than those caring for more severe relatives. Z tests revealed significantly different correlational patterns for the two dimensions, with emotional pain more related to global burden and depressive symptoms, and relational deprivation more related to care-recipients' functional impairment. Both dimensions were mildly correlated with neuropsychiatric symptoms (especially disruptive behaviors and psychotic symptoms) of the care-recipient. CONCLUSIONS: Results supported the reliability and validity of the two-dimensional measure of predeath grief. As a brief measure, it can be readily added to research instruments to facilitate study of this important phenomenon along with other caregiving outcomes.