Motor patterns of patients with spinal muscular atrophy suggestive of sensory and corticospinal contributions to the development of locomotor muscle synergies.

Complex locomotor patterns are generated by combination of muscle synergies. How genetic processes, early sensorimotor experiences, and the developmental dynamics of neuronal circuits contribute to the expression of muscle synergies remains elusive. We shed light on the factors that influence development of muscle synergies by studying subjects with spinal muscular atrophy (SMA, types II/IIIa), a disorder associated with degeneration and deafferentation of motoneurons and possibly motor cortical and cerebellar abnormalities, from which the afflicted would have atypical sensorimotor histories around typical walking onset. Muscle synergies of children with SMA were identified from electromyographic signals recorded during active-assisted leg motions or walking, and compared with those of age-matched controls. We found that the earlier the SMA onset age, the more different the SMA synergies were from the normative. These alterations could not just be explained by the different degrees of uneven motoneuronal losses across muscles. The SMA-specific synergies had activations in muscles from multiple limb compartments, a finding reminiscent of the neonatal synergies of typically developing infants. Overall, while the synergies shared between SMA and control subjects may reflect components of a core modular infrastructure determined early in life, the SMA-specific synergies may be developmentally immature synergies that arise from inadequate activity-dependent interneuronal sculpting due to abnormal sensorimotor experience and other factors. Other mechanisms including SMA-induced intraspinal changes and altered cortical-spinal interactions may also contribute to synergy changes. Our interpretation highlights the roles of the sensory and descending systems to the typical and abnormal development of locomotor modules.NEW & NOTEWORTHY This is likely the first report of locomotor muscle synergies of children with spinal muscular atrophy (SMA), a subject group with atypical developmental sensorimotor experience. We found that the earlier the SMA onset age, the more the subjects' synergies deviated from those of age-matched controls. This result suggests contributions of the sensory/corticospinal activities to the typical expression of locomotor modules, and how their disruptions during a critical period of development may lead to abnormal motor modules.

Comprehensive Analysis of Congenital Adrenal Hyperplasia Using Long-Read Sequencing.

BACKGROUND: Congenital adrenal hyperplasia (CAH) is an autosomal recessive disorder that has been included in newborn screening programs. Current approaches to gene testing for CAH are facing challenges because of the complexity of the CYP21A2 locus and genetic heterogeneity of the disease. METHODS: A comprehensive analysis of CAH (CACAH) combining long-range locus-specific PCR and long-read sequencing (LRS) was developed to perform full sequence analysis of 5 common CAH candidate genes, including CYP21A2, CYP11B1, CYP17A1, HSD3B2, and StAR. In a blind retrospective study, the clinical utility of CACAH was evaluated in 37 samples by comparing to standard CAH testing using multiplex ligation-dependent probe amplification (MLPA) plus Sanger sequencing. RESULTS: Of the 37 clinical samples, a total of 69 pathogenic variants were identified, comprising 65 CYP21A2 variants, 2 HSD3B2 variants, and 2 CYP17A1 variants. For CYP21A2, the most frequent variant was c.518T > A (29.2%), followed by c.293-13C/A > G (21.5%). Compared with the current CAH testing using MLPA plus Sanger sequencing, the CACAH assay showed 100% specificity and 100% sensitivity, and precisely determined the junction sites of deletions/insertions and cis-trans configuration of multiple variants without analyzing family samples. Moreover, CACAH identified a case carrying 2 copies of CYP21A1 with the c.1451_1452delinsC variant on the same chromosome, which was not confirmed by MLPA plus Sanger sequencing. CONCLUSION: LRS-based CACAH can determine all genotypes of CAH accurately and reliably in one assay, presenting a comprehensive approach for CAH genetic diagnosis and carrier screening.

Loss of PIGK function causes severe infantile encephalopathy and extensive neuronal apoptosis.

PIGK gene, encoding a key component of glycosylphosphatidylinositol (GPI) transamidase, was recently reported to be associated with inherited GPI deficiency disorders (IGDs). However, little is known about the specific downstream effects of PIGK on neurodevelopment due to the rarity of the disease and the lack of in vivo study. Here, we described 2 patients in a Chinese family presented with profound global developmental delay, severe hypotonia, seizures, and postnatal progressive global brain atrophy including hemisphere, cerebellar and corpus callosum atrophy. Two novel compound heterozygous variants in PIGK were identified via genetic analysis, which was proved to cause significant decrease of PIGK protein and reduced cell surface presence of GPI-APs in the patients. To explore the role of Pigk on embryonic and neuronal development, we constructed Pigk knock-down zebrafish and knock-in mouse models. Zebrafish injected with a small dose of morpholino oligonucleotides displayed severe developmental defects including small eyes, deformed head, curly spinal cord, and unconsumed yolk sac. Primary motor neuronal dysplasia and extensive neural cell apoptosis were further observed. Meanwhile, the mouse models, carrying the two variants respectively homologous with the patients, both resulted in complete embryonic lethality of the homozygotes, which suggested the intolerable effect caused by amino acid substitution of Asp204 as well as the truncated mutation. Our findings provide the in vivo evidence for the essential role of PIGK during the embryonic and neuronal development. Based on these data, we propose a basis for further study of pathological and molecular mechanisms of PIGK-related neurodevelopmental defects.

Molecular diagnosis for 55 fetuses with skeletal dysplasias by whole-exome sequencing: A retrospective cohort study.

Skeletal dysplasias (SDs) are common birth defects, but they are difficult to diagnose accurately according to only the limited phenotypic information available from ultrasound during the pregnancy. To evaluate the application of whole-exome sequencing (WES) and expand the data in the prenatal molecular diagnosis of fetuses with SDs, we collected 55 fetuses with SDs based on ultrasonographic features. WES of the fetuses or parent-fetus trio were subjected to sequential tests and produced a diagnostic yield of 64% (35/55). 65% (11/17) of families with a history of adverse pregnancies were diagnosed, 16 genes were involved and 37 different pathogenic or likely pathogenic variants were identified, including 14 novel variants, which were first reported in this study. De novo variants were identified in 21 cases (60%, 21/35) among the fetuses with a genetic diagnosis. The pathogenicity of two novel splice-site variants was confirmed by constructing minigene in vitro. Our results revealed that WES can provide new evidence for the relationship between the genotype and phenotype of fetuses with SDs, as well as broaden the mutation spectrum of detected genes, which is significant for prenatal diagnosis and genetic counseling.

Biochemical and genetic characteristics of 40 neonates with carnitine deficiency. / 40ä¾‹è‚‰ç¢±ç¼ºä¹ç—‡æ–°ç”Ÿå„¿çš„ç”ŸåŒ–å’Œé—ä¼ å­¦ç‰¹å¾.

OBJECTIVES: Primary carnitine deficiency (PCD) is a rare fatty acid metabolism disorder that can cause neonatal death. This study aims to analyze carnitine levels and detect SLC22A5 gene in newborns with carnitine deficiency, to provide a basis for early diagnosis of PCD, and to explore the relationship between carnitine in blood and SLC22A5 genotype. METHODS: A total of 40 neonates with low free carnitine (C0<10 µmol/L) in blood were the subjects of the study. SLC22A5 gene was detected by Sanger sequencing to analyze the value of carnitine, the results of gene test and their relationship. RESULTS: A total of 15 variants of SLC22A5 gene were detected, including 11 pathogenic or likely pathogenic variants and 4 variants of uncertain significance. There were 5 new mutations: c.288delG (p.G96fsX33), c.744_745insTCG (p.M258_L259insS), c.752A>G (p.Y251C), c.495 C>A (p.R165E), and c.1298T>C (p.M433T). We found 14 PCD patients including 2 homozygous mutations and 12 heterozygous mutations, 14 with 1 mutation, and 12 with no mutation among 40 children. The C0 concentration of children with SLC22A5 gene homozygous or complex heterozygous mutations was (4.95±1.62) µmol/L in the initial screening, and (3.90±1.33) µmol/L in the second screening. The C0 concentration of children with no mutation was (7.04±2.05) µmol/L in the initial screening, and (8.02±2.87) µmol/L in the second screening. There were significant differences between children with homozygous or compound heterozygous mutations and with no mutation in C0 concentration of the initial and the second screening (both P<0.05), as well as between children with truncated mutation and with untruncated mutation in C0 concentration of the initial screening (P=0.022). CONCLUSIONS: There are 5 new mutations which enriched the mutation spectrum of SLC22A5 gene. C0<5 µmol/L is highly correlated with SLC22A5 gene homozygous or compound heterozygous mutations. Children with truncated mutation may have lower C0 concentration than that with untruncated mutation in the initial screening.

[Prenatal diagnosis for 30 women carrying a FMR1 mutation].

OBJECTIVE: To determine the CGG repeat number and methylation status of FMR1 gene for fetuses whose mothers have carried a FMR1 mutation. METHODS: For 30 pregnant women, the fetal CGG repeat number was determined with a GC-rich PCR system by using chorionic villus, amniotic fluid or umbilical blood samples. The methylation status of the FMR1 gene was confirmed with Southern blotting. RESULTS: In total 30 prenatal diagnoses were performed for 29 carriers of FMR1 gene mutations and 1 with FMR1 gene deletion mosaicism. Three fetuses were found to carry premutations, 9 were with full mutations and 1 with mosaicism of premutation and full mutations. Eighteen fetuses were normal. CONCLUSION: Considering the genetic complexity of Fragile X syndrome (FXS), single method may not suffice accurate determination of their genetic status. The pitfalls and technical limitations of protocols requires adoption of personalized strategy for its prenatal diagnosis.

The Caenorhabditis elegans gene mfap-1 encodes a nuclear protein that affects alternative splicing.

RNA splicing is a major regulatory mechanism for controlling eukaryotic gene expression. By generating various splice isoforms from a single pre-mRNA, alternative splicing plays a key role in promoting the evolving complexity of metazoans. Numerous splicing factors have been identified. However, the in vivo functions of many splicing factors remain to be understood. In vivo studies are essential for understanding the molecular mechanisms of RNA splicing and the biology of numerous RNA splicing-related diseases. We previously isolated a Caenorhabditis elegans mutant defective in an essential gene from a genetic screen for suppressors of the rubberband Unc phenotype of unc-93(e1500) animals. This mutant contains missense mutations in two adjacent codons of the C. elegans microfibrillar-associated protein 1 gene mfap-1. mfap-1(n4564 n5214) suppresses the Unc phenotypes of different rubberband Unc mutants in a pattern similar to that of mutations in the splicing factor genes uaf-1 (the C. elegans U2AF large subunit gene) and sfa-1 (the C. elegans SF1/BBP gene). We used the endogenous gene tos-1 as a reporter for splicing and detected increased intron 1 retention and exon 3 skipping of tos-1 transcripts in mfap-1(n4564 n5214) animals. Using a yeast two-hybrid screen, we isolated splicing factors as potential MFAP-1 interactors. Our studies indicate that C. elegans mfap-1 encodes a splicing factor that can affect alternative splicing.

The survival motor neuron gene smn-1 interacts with the U2AF large subunit gene uaf-1 to regulate Caenorhabditis elegans lifespan and motor functions.

Spinal muscular atrophy (SMA), the most frequent human congenital motor neuron degenerative disease, is caused by loss-of-function mutations in the highly conserved survival motor neuron gene SMN1. Mutations in SMN could affect several molecular processes, among which aberrant pre-mRNA splicing caused by defective snRNP biogenesis is hypothesized as a major cause of SMA. To date little is known about the interactions of SMN with other splicing factor genes and how SMN affects splicing in vivo. The nematode Caenorhabditis elegans carries a single ortholog of SMN, smn-1, and has been used as a model for studying the molecular functions of SMN. We analyzed RNA splicing of reporter genes in an smn-1 deletion mutant and found that smn-1 is required for efficient splicing at weak 3' splice sites. Genetic studies indicate that the defective lifespan and motor functions of the smn-1 deletion mutants could be significantly improved by mutations of the splicing factor U2AF large subunit gene uaf-1. In smn-1 mutants we detected a reduced expression of U1 and U5 snRNAs and an increased expression of U2, U4 and U6 snRNAs. Our study verifies an essential role of smn-1 for RNA splicing in vivo, identifies the uaf-1 gene as a potential genetic modifier of smn-1 mutants, and suggests that SMN-1 has multifaceted effects on the expression of spliceosomal snRNAs.

Application of whole exome sequencing in carrier screening for high-risk families without probands.

Purpose: This study aimed to screen the genetic etiology for the high-risk families including those with an adverse pregnancy history, a history of consanguineous marriages, or a history of genetic diseases, but lack of proband via whole exome sequencing (WES). Methods: 128 individuals from high-risk family were tested by WES. The candidate variants were analyzed according to the ACMG criteria to screen the potential carriers. At-risk couples (ARCs) who harbored the same causative gene were provided with precise fertility guidance to avoid the birth of children with birth defects. Results: The total detection rate was 36.72%, with pathogenic/likely pathogenic (P/LP) variants found in 47 individuals, and variants of uncertain significance (VUS) were found in 34. Among couples with adverse pregnancy history: P/LP variants were found in 38 individuals, and VUS were found in 26, for a detection rate of 34.55%; among members of family history of genetic disease or consanguineous marriages: P/LP variants were found in nine individuals, and VUS were found in 8, for a detection rate of 50.00%. Otherwise, we detected 19 ARCs who both carried P/LP variants in the same gene, with a theoretical offspring prevalence of up to 7.42%. Conclusion: In the absence of probands, carrier screening using WES can provide an efficient tool for screening the molecular etiology of high-risk families.

In vivo effects on intron retention and exon skipping by the U2AF large subunit and SF1/BBP in the nematode Caenorhabditis elegans.

The in vivo analysis of the roles of splicing factors in regulating alternative splicing in animals remains a challenge. Using a microarray-based screen, we identified a Caenorhabditis elegans gene, tos-1, that exhibited three of the four major types of alternative splicing: intron retention, exon skipping, and, in the presence of U2AF large subunit mutations, the use of alternative 3' splice sites. Mutations in the splicing factors U2AF large subunit and SF1/BBP altered the splicing of tos-1. 3' splice sites of the retained intron or before the skipped exon regulate the splicing pattern of tos-1. Our study provides in vivo evidence that intron retention and exon skipping can be regulated largely by the identities of 3' splice sites.

Circ_0033596 depletion ameliorates oxidized low-density lipoprotein-induced human umbilical vein endothelial cell damage.

BACKGROUND: Previous data have shown that circ_0033596 is involved in the pathogenesis of atherosclerosis (AS). The study aims to reveal the detailed mechanism of circ_0033596 in AS. METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with oxidized low-density lipoprotein (ox-LDL) to establish an AS cell model. Quantitative real-time polymerase chain reaction and western blot were implemented to detect the expression of circ_0033596, miR-637, growth factor receptor bound protein2 (GRB2), BCL2-associated x protein (Bax) and B-cell lymphoma-2 (Bcl-2). Cell viability, proliferation, apoptosis and tube formation were investigated by cell counting kit-8, EdU assay, flow cytometry and tube formation assay, respectively. The production of interleukin (IL-6) and tumor necrosis factor-α (TNF-α) was evaluated by enzyme-linked immunosorbent assay. Oxidative stress was evaluated by lipid peroxidation malondialdehyde assay kit and superoxide dismutase activity assay kit. Dual-luciferase reporter assay, RNA pull-down assay and RIP assay were performed to identify the associations among circ_0033596, miR-637 and GRB2. RESULTS: The expression of circ_0033596 and GRB2 was significantly increased, while miR-637 was decreased in the blood of AS patients and ox-LDL-induced HUVECs compared with controls. Ox-LDL treatment inhibited HUVEC viability, proliferation and angiogenic ability and induced cell apoptosis, inflammation and oxidative stress, while these effects were attenuated after circ_0033596 knockdown. Circ_0033596 interacted with miR-637 and regulated ox-LDL-induced HUVEC damage by targeting miR-637. In addition, GRB2, a target gene of miR-637, participated in ox-LDL-induced HUVEC injury by combining with miR-637. Importantly, circ_0033596 activated GRB2 by interacting with miR-637. CONCLUSION: Circ_0033596 depletion protected against ox-LDL-induced HUVEC injury by miR-637/GRB2 pathway, providing a therapeutic target for AS.

Diagnostic accuracy of different ECG-based algorithms in wide QRS complex tachycardia: a systematic review and meta-analysis.

OBJECTIVE: Several ECG-based algorithms have been proposed to enhance the effectiveness of distinguishing Wide QRS complex tachycardia (WCT), but a comprehensive comparison of their accuracy is still lacking. This meta-analysis aimed to assess the diagnostic precision of various non-artificial intelligence ECG-based algorithms for WCT. DESIGN: Systematic review with meta-analysis. DATA SOURCES: Electronic databases (PubMed, MEDLINE, the Cochrane Library, and Web of Science) are searched up to May 2022. ELIGIBILITY CRITERIA FOR SELECTING STUDIES: All studies reporting the diagnostic accuracy of different ECG-based algorithms for WCT are included. The risk of bias in included studies is assessed using the Cochrane Collaboration's risk of bias tools. DATA EXTRACTION AND SYNTHESIS: Two independent reviewers extracted data and assessed risk of bias. Data were pooled using random-effects model and expressed as mean differences with 95% CIs. Heterogeneity was calculated by the I2 method. The Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool was applied to assess the internal validity of the diagnostic studies. RESULTS: In total, 467 studies were identified, and 14 studies comprising 3966 patients were included, involving four assessable ECG-based algorithms: the Brugada algorithm, Vereckei-pre algorithm, Vereckei-aVR algorithm and R wave peak time of lead II (RWPT-II) algorithm. The overall sensitivity was 88.89% (95% CI: 85.03 to 91.86), with a specificity of 70.55% (95% CI: 62.10 to 77.79) and a diagnostic OR (DOR) of 19.17 (95% CI: 11.45 to 32.10). Heterogeneity of the DOR was 89.1%. The summary sensitivity of each algorithm was Brugada 90.25%, Vereckei-pre 94.80%, Vereckei-aVR 90.35% and RWPT-II 78.15%; the summary specificity was Brugada 64.02%, Vereckei-pre 75.40%, Vereckei-aVR 60.88% and RWPT-II 88.30% and the summary DOR was Brugada 16.48, Vereckei-pre 60.70, Vereckei-aVR 14.57 and RWPT-II 27.00. CONCLUSIONS: ECG-based algorithms exhibit high sensitivity and moderate specificity in diagnosing WCT. A combination of Brugada or Vereckei-aVR algorithm with RWPT-II could be considered to diagnose WCT. PROSPERO REGISTRATION NUMBER: CRD42022344996.

A de novo heterozygous POU3F3 genotype for the p.(Q214*) variant in a fetus with transient isolated bilateral mild ventriculomegaly: a case report and review of the literature.

The prenatal prevalence of isolated ventriculomegaly is 0.039%-0.087%. Most isolated mild ventriculomegaly (MV) fetuses (>90%) have a favorable prognosis. However, 5.6% to 7.9% of fetuses with isolated MV have adverse neurodevelopmental outcomes. In this study, we reported the first case of prenatal Snijders Blok-Fisher syndrome (OMIM: #618604) caused by a truncating variant of POU3F3 (OMIM: *602480) in a fetus with transient isolated bilateral MV. The results of karyotype analysis, chromosomal microarray analysis, and TORCH infection evaluation for the fetus were all negative. However, a de novo likely pathogenic nonsense variant of NM_006236.3 (POU3F3): c.640C > T [rs1254251078] p.(Q214*) was identified by whole-exome sequencing (WES). Despite sufficient genetic counseling, the mother refused to undertake further brain magnetic resonance imaging (MRI) and decided to keep the fetus. She gave birth to a male infant through a full-term vaginal delivery. With a long-term follow-up, the infant unfortunately gradually presented with delayed motor development. The postnatal brain MRI of the proband showed dysplasia of the corpus callosum and ventriculomegaly. Considering the high probability of misdiagnosis for such cases, we further summarized the prenatal phenotypes from 19 reported patients with variants in POU3F3. The results revealed that 14 patients displayed a normal prenatal ultrasonographic manifestation, while only approximately 26.32% of fetuses showed MV or cysts without structural deformity. Thus our findings expand the variant spectrum of POU3F3 and suggest the importance of undertaking WES and brain MRI when the fetus has isolated bilateral MV.

The exploration of genetic aetiology and diagnostic strategy for 321 Chinese individuals with intellectual disability.

BACKGROUND: Intellectual disability is a heterogeneous neurodevelopmental disorder with complex genetic architectures. Different sequential methodologies are usually applied to identify the genetic aetiologies of ID patients. METHODS: We collected 321 consecutive ID patients. All patients underwent karyotyping, while 293 and 164 cases further received copy number variation sequencing (CNV-seq) and whole-exome sequencing (WES). The updated WES technology can detect CNVs simultaneously. The diagnostic data from 137 patients who received WES and CNV-seq were used to define the approach that could be recommended as the first-tier test. RESULTS: WES obtains the highest diagnostic yield of 50% (82/164), compared with karyotyping (7.79%, 25/321) and CNV-seq (19.80%, 58/293). Among the variants detected by WES, 66.67% (44/66) de novo and 57.58% (38/66) novel pathogenic/likely pathogenic (P/LP) variants were identified in patients with ID. Besides, 24 out of 25P/LP CNVs discovered by CNV-seq can also be accurately identified using WES in 137 patients who received WES and CNV-seq. Thus, genetic abnormalities found through karyotyping, CNV-seq, and WES can be completely detected by combined karyotyping and WES. CONCLUSIONS: This study illustrates the genetic aberrations of a Chinese ID cohort and expands the mutation spectrum of ID-related genes. Compared with the conventional diagnostic strategy, a combination of karyotype analysis and WES could be recommended as the first-tier diagnostic strategy for ID patients.

Functional identification of two novel variants and a hypomorphic variant in ASS1 from patients with Citrullinemia type I.

Background: Citrullinemia type I (CTLN1) is a rare autosomal recessive inborn error of the urea cycle caused by mutations in the gene encoding the arginosuccinate synthetase (ASS1) enzyme. Classic CTLN1 often manifests with acute hyperammonemia and neurological symptoms. Molecular genetic testing is critical for patient diagnosis. Methods: Three unrelated families with clinically suspected CTLN1 were included in this study. Potential pathogenic variants were identified using whole exome sequencing (WES) and validated using Sanger sequencing. Western blotting, quantitative PCR, immunofluorescent staining, and ELISA were used to assess functional changes in candidate ASS1 variants. Results: Five variants were identified, two of which were novel, and one has been reported, but its pathogenicity was not validated. The novel variant c.649-651del (p.P217del) and the 5'UTR variant (c.-4C>T) resulted in a decrease in ASS1 expression at both the protein and transcription levels. The other novel variant, c.1048C>T (p.Q350*), showed a marked decrease in expression at the protein level, with the formation of truncated proteins but an increased transcription. Both c.649_651del (p.P217del) and c.1048C>T (p.Q350*) showed a highly significant reduction in enzyme activity, while c.-4C>T had no effect. Conclusion: We identified two novel variants and a hypomorphic non-coding variant in ASS1 and validated the pathogenicity using functional studies. Our findings contribute to expanding the spectrum of ASS1 variants and understanding the genotype-phenotype relationships of CTLN1.

A homozygous variant in INTS11 links mitosis and neurogenesis defects to a severe neurodevelopmental disorder.

The INTS11 endonuclease is crucial in modulating gene expression and has only recently been linked to human neurodevelopmental disorders (NDDs). However, how INTS11 participates in human development and disease remains unclear. Here, we identify a homozygous INTS11 variant in two siblings with a severe NDD. The variant impairs INTS11 catalytic activity, supported by its substrate's accumulation, and causes G2/M arrest in patient cells with length-dependent dysregulation of genes involved in mitosis and neural development, including the NDD gene CDKL5. The mutant knockin (KI) in induced pluripotent stem cells (iPSCs) disturbs their mitotic spindle organization and thus leads to slow proliferation and increased apoptosis, possibly through the decreased neurally functional CDKL5-induced extracellular signal-regulated kinase (ERK) pathway inhibition. The generation of neural progenitor cells (NPCs) from the mutant iPSCs is also delayed, with long transcript loss concerning neurogenesis. Our work reveals a mechanism underlying INTS11 dysfunction-caused human NDD and provides an iPSC model for this disease.

VSIG4 regulates macrophages polarization and alleviates inflammation through activating PI3K/AKT and inhibiting TLR4/NF-κB pathway in myocardial ischemia-reperfusion injury rats.

Background: Myocardial infarction is the primary cause of high disability and mortality in patients with cardiovascular disease worldwide. The pathological process of myocardial ischemia/reperfusion (I/R) may trigger harmful inflammatory response and ultimately lead to serious cardiac dysfunction. The mechanism of myocardial repair post myocardial infarction has not been fully elucidated. The present study speculated that VSIG4 is related to the regulation of heart injury. Methods: The myocardial I/R injury model was established in Sprague-Dawley (SD) rats. Before I/R operation, the viral solution containing AAV-NC or AAV-VSIG4 was intravenously injected into rats. Cardiac function indicators, mRNA expression, the apoptosis ratio of cardiomyocytes, myocardial infarct area, phenotype polarization of macrophage, and the protein expression of apoptosis or macrophage phenotype were measured. Results: Myocardial I/R injury decreased the expression of VSIG4 and subsequently triggered myocardial apoptosis. The induction of AAV-VSIG4 produced a protective effect on general cardiac function and attenuated the I/R-induced cellular apoptosis in rats. Moreover, VSIG4 signaling might potentially modulate macrophage M1/M2-related inflammatory disorders via activation of PI3K/AKT and inhibition of TLR4/NF-κB expression. Conclusion: In summary, the present study provided evidence that VSIG4 had cardiac protective role in myocardial I/R injury. More importantly, enhanced VSIG4 expression inhibited M1 polarization of macrophages by blocking TLR4/NF-κB activation, subsequently suppressing cardiomyocyte apoptosis. This finding provides vital insights into the role of VSIG4 in I/R injury and may provide a new target for I/R therapy.

Whole-exome sequencing identifies genetic variants of hearing loss in 113 Chinese families.

BACKGROUND: Hearing loss is a group of diseases with high genetic heterogeneity. About 160 genes have been reported to be associated with hereditary hearing loss. METHODS: 113 families with hearing loss were collected, and WES was used to detect SNV, InDel, CNV and mitochondrial gene variants. For some probands with negative WES test results, the copy number of STRC and OTOA were determined by using real-time fluorescence quantitative PCR. RESULTS: Pathogenic or likely pathogenic variants were found in 54 probands, of which 98% (53/54) were SNVs or InDels and 2% (1/54) were CNVs, a positive rate of 48%. 16 families (14%) were detected with candidate variants of uncertain significance. 19 novel pathogenic or likely pathogenic variants and 22 candidate variants of uncertain significance were identified in this study. The most common hearing loss gene in the families was GJB2, accounting for 28% (15/53), followed by SLC26A4 and MYO15A, accounting for 21% (11/53) and 11% (6/53), respectively. Heterozygous gene deletion was detected in 3 probands, including 2 with STRC and 1 with OTOA in 43 families with WES negative test. CONCLUSION: Genetic etiology was clarified in 54 families. All of these findings broadened the mutation spectrum of hearing loss genes, thus providing new variant information for the future diagnosis of patients with hearing loss.

Two novel variants in CEP152 caused Seckel syndrome 5 in a Chinese family.

Background: Seckel syndrome (SCKL) is a rare autosomal recessive inherited disorder, which is mainly characterized by intrauterine and postnatal growth restrictions, microcephaly, intellectual disability, and a typical "bird-head" facial appearance. Here, we aimed to identify the genetic etiology of a family with suspected SCKL. Methods: This study enrolled a Chinese family suspected of SCKL with their detailed family history and clinical data. We performed karyotype analysis, copy number variation sequencing (CNV-seq), and trio whole-exome sequencing (WES) to explore the genetic etiology in the proband. Furthermore, the quantitative real-time polymerase chain reaction (PCR) and reverse transcription-PCR (RT-PCR) were conducted to confirm the pathogenicity of novel variants. Results: The karyotype analysis and CNV-seq were normal in the proband. Two novel variants in CEP152, c.1060C>T (p.Arg354*) and c.1414-14A>G, were identified in the proband through trio-WES. The qPCR results showed that the total CEP152 mRNA expression levels were significantly reduced in c.1060C>T (p.Arg354*) and c.1414-14A>G compared with healthy control individuals. Moreover, aberrant skipping of exon 12 due to the non-canonical splice-site variant was revealed by RT-PCR and Sanger sequencing. Conclusion: Our findings expanded pathogenic variant spectra in SCKL and offered new insights into the pathogenicity of a non-classical splice-site variant in CEP152, which provided additional information for helping the family improve pregnancy plans in the future.

Identification of four novel mutations in BTK from six Chinese families with X-linked agammaglobulinemia.

BACKGROUND: The defect of Bruton's tyrosine kinase (BTK) gene resulted in X-linked agammaglobulinemia (XLA), which is characterized by recurrent bacterial infections, immunodeficiency with low B-cell numbers and immunoglobulin. Diagnosis of XLA depends on clinical phenotype and genetic testing. METHODS: Six unrelated Chinese families with high suspicion of XLA were enrolled in this study. Potential pathogenic variants were detected and validated by Whole Exome Sequencing (WES) and Sanger Sequencing. Western blot, Quantitative PCR (qPCR) analysis and immunofluorescence analysis were used to evaluate the preliminary function of candidate BTK variants. RESULTS: A total of six variants were identified, four of which were not reported before. The novel missense mutation(c.1900 T > G) and deletion(c.897delG) were found that the mutant protein and mRNA expression levels have fallen by Western Blot and qPCR identification. We also constructed minigene expression vector to determine the deletion (c.1751-6_1755delttctagGGGTT) resulting a 35 bp skipping in exon 18. Meanwhile, the break point of gross deletion (Exon2-5) discovered based on WES was confirmed to be located at site ChX:101367539_101376531 through qPCR and Gap-PCR. CONCLUSION: This study makes definitive diagnosis for 6 families with suspected XLA and further expands the spectrum of BTK mutations, providing new information for the diagnosis of the disease.