[Analysis of clinical phenotypes and MMACHC gene variants in 65 children with Methylmalonic acidemia and homocysteinemia].

OBJECTIVE: To carry out Sanger sequencing for MMACHC gene variants among 65 Chinese pedigrees affected with combined methylmalonic aciduria and homocysteinemia, and summarize their genetic and clinical characteristics and prognosis. METHODS: Clinical characteristics of the 65 children identified with Methylmalonic acidemia and homocysteinemia at the Children's Hospital Affiliated to Zhengzhou University (Zhengzhou Children's Hospital) from April 2017 to April 2022 were selected as the study subjects. Potential variants of the MMACHC gene were detected by direct sequencing of the PCR products. RESULTS: The median age of the 65 children was 3 months (14 days to 17 years old). These included 28 cases (43.08%) from neonatal screening, 11 cases (16.92%) with a history of jaundice, and 9 cases (13.85%) with various degrees of anemia. The main clinical symptoms included development delay, slow growth, epilepsy, hydrocephalus, lethargy, feeding difficulty, regression or decline in motor ability, recurrent respiratory infections, anemia, jaundice, respiratory and heart failures, hydrocephalus, limb weakness, and hypertension. Blood and urine tandem mass spectrometry screening has revealed increase of methylmalonic acid, propionyl carnitine, propionyl carnitine/acetylcarnitine ratio, and propionyl carnitine/free carnitine ratio to various extents, and blood homocysteine was increased in all patients. The detection rate of genetic variants was 98.46% (128/130), and in total 22 types of MMACHC gene variants were detected. The most common ones have included c.609G>A (W203X) (58/128), c.658-660del (K220del) (19/128), and c.80A>G (Q27A) (16/128). Two novel variants have been identified, namely c.565C>T (p.R189C) and c.624_ 625delTG (p.A208Afs), which were respectively predicted as likely pathogenic (PM2_Supporting+PM3+PP2+PP3) and pathogenic (PVS1+PM2_Supporting+PM3+PP2) based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). Exon 4 had the highest frequency for the detection. CONCLUSION: Identification of MMACHC gene variants has confirmed the diagnosis in the children, among which the c.609G>A variant has the highest frequency. Discovery of the new variants has enriched the mutational spectrum of the MMACHC gene.

[Analysis of NF1 gene variants among thirteen patients with neurofibromatosis type 1].

OBJECTIVE: To detect variants of NF1 gene among thirteen patients with neurofibromatosis type 1. METHODS: Genomic DNA was extracted from peripheral blood samples of the patients. High-throughput sequencing was employed to detect potential variants of the NF1 and NF2 genes. RESULTS: Thirteen pathogenic variants were identified among the patients, which included one NF1 deletion, three missense variants, three nonsense variants and six frameshifting variants. Among these, 10 variants have been associated with neurofibromatosis type 1. c.4180A>T (p.Asn1394Tyr), c.4217dupT (p.Leu1406fs) and c.1753dupT(p.Leu585Phefs*3) were unreported previously. Based on the guidelines of the American College of Medical Genetics and Genomics, c.4180A>T (p.Asn1394Tyr) was predicted to be likely pathogenic (PS2+PM1+PM2+PP2), while c.4217dupT (p.Leu1406fs) and c.1753dupT (p.Leu585Phefs*3) were predicted to be pathogenic (PVS1+PS2+PM2). CONCLUSION: Variants of the NF1 gene probably underlay the disease among these children. Above findings have enriched the the spectrum of NF1 gene variants.

[Analysis of a Chinese pedigree with autosomal dominant Charcot-Marie-Tooth disease type 2A2A].

OBJECTIVE: To explore the genetic basis of a pedigree affected with peroneal muscular atrophy. METHODS: Neuroelectrophysiological examination and whole exome sequencing were carried out for the proband, a six-year-and-ten-month-old boy. Suspected variant was verified in his family members through Sanger sequencing. Bioinformatic analysis was carried to predict the conservation of amino acid sequence and impact of the variant on the protein structure and function. RESULTS: Electrophysiological examination showed demyelination and axonal changes of motor and sensory nerve fibers. A heterozygous missense c.1066A>G (p. Thr356Ala) variant was found in exon 11 of the MFN2 gene in the proband and his mother, but not in his sister and father. Bioinformatic analysis using PolyPhen-2 and Mutation Taster software predicted the variant to be pathogenic, and that the sequence of variation site was highly conserved among various species. Based no the American College of Medical Genetics and Genomics standards and guidelines, the c.1066A>G (p. Thr356Ala) variant of MFN2 gene was predicted to be likely pathogenic (PS1+ PM2+ PP3+ PP4). CONCLUSION: The heterozygous missense c.1066A>G (p.Thr356Ala) variant of the MFN2 gene probably underlay the disease in the proband, and the results have enabled genetic counseling and prenatal diagnosis for this family.

[Clinical and genetic analysis of a case with infantile Parkinsonism with motor delay due to tyrosine hydroxylase deficiency].

OBJECTIVE: To explore the clinical characteristics and genetic variants in a child with tyrosine hydroxylase-deficient infantile Parkinsonism with motor delay. METHODS: Clinical feature of the patient was summarized. Genomic DNA was extracted from peripheral blood samples taken from the child and her family members. All exons of GCH1, TH and SPR genes were subjected to targeted capture and next-generation sequencing. Suspected variants were verified by Sanger sequencing. RESULTS: The child could not sit alone at 7 month and 11 days. Physical examination suggested motor retardation and hypotonia, limb stiffness, head nodding, slight torticollis, and language and intellectual developmental delays. She developed involuntary shaking of limbs at 3 month old, which lasted approximately 10 seconds and aggregated with excitement and before sleeping. Cranial MRI revealed widening of subarachnoid space on the temporomandibular and particularly temporal sides. Genetic testing revealed that she has carried a nonsense c.457C>T (p.R153X) variant, which was known to be pathogenic, and a novel missense c.720C>G (p.I240M) variant of the TH gene. The two variants were derived from her father and mother, respectively. CONCLUSION: The child was diagnosed as tyrosine hydroxylase-deficient infantile Parkinsonism with motor delay due to compound heterozygous variants of the TH gene. Above finding has enriched the spectrum of TH gene variants.

Genomic and biological characterization of a pandemic norovirus variant GII.4 Sydney 2012.

Genogroup II, genotype 4 noroviruses (GII.4 NoVs) are a leading cause of epidemic and sporadic acute non-bacterial gastroenteritis worldwide. In this study, we isolated a GII.4 NoV strain (designated 2015HN08) from a kid presenting with acute gastroenteritis and determined its near-complete genome sequence. We then performed sequence analysis by comparing this strain with the prototypical GII.4 strain. Virus-like particles (VLPs) derived from the major capsid protein (VP1) were expressed by using a recombinant-baculovirus expression system, and monoclonal antibodies (mAbs) were produced to compare changes in antigenic or histo-blood group antigens (HBGAs) binding sites with the previously characterized GII.4 NoV strain (JZ403). The genome of 2015HN08 was 7559 nucleotides (nt) long, excluding the poly(A) tail. Genotyping analysis indicated that this strain was a Sydney 2012 variant. In comparison with the prototype Sydney 2012 strain, there were 74, 35, and 16 differences in nucleotide sequences in ORF1, OFR2, and OFR3, causing 7, 10, and 6 amino acid (aa) changes, respectively. Expression of VP1 led to successful assembly of VLPs, as demonstrated by electron microscopy. Screening of hybridoma cell supernatants with an in vitro VLP-HBGAs binding blockade assay led to the identification of a cell clone 3G10 that exhibited HBGA-blocking effects. This mAb also exhibited blocking effects against JZ403 strain, suggesting maintenance of the antigenic site and/or HBGAs binding sites between the two strains. In summary, we determined the near-complete genome sequence of a GII.4 Sydney 2012 variant and produced an mAb with blocking effects that might be useful in evaluating the evolution of current Sydney 2012 NoV strains.

LIN28A gene polymorphisms modify neuroblastoma susceptibility: A four-centre case-control study.

Neuroblastoma ranks the most common seen solid tumour in childhood. Overexpression of LIN28A gene has been linked to the development of multiple human malignancies, but the relationship between LIN28A single nucleotide polymorphisms (SNPs) and neuroblastoma susceptibility is still under debate. Herein, we evaluated the correlation of four potentially functional LIN28A SNPs (rs3811464 G>A, rs3811463 T>C, rs34787247 G>A, and rs11247957 G>A) and neuroblastoma susceptibility in 505 neuroblastoma patients and 1070 controls from four independent hospitals in China. The correlation strengths were determined by using odds ratios (ORs) and corresponding 95% confidence intervals (CIs). Among these SNPs, rs34787247 G>A exhibited a significant association with increased susceptibility in neuroblastoma (GA vs GG: adjusted OR = 1.30, 95% CI = 1.03-1.64; AA vs GG: adjusted OR = 2.51, 95% CI = 1.36-4.64, AA/GA vs GG: adjusted OR = 1.42, 95% CI = 1.12-1.80, AA vs GG/GA: adjusted OR = 2.39, 95% CI = 1.29-4.42). Furthermore, the combined analysis of risk genotypes revealed that subjects carrying three risk genotypes (adjusted OR = 1.64, 95% CI = 1.02-2.63) are more inclined to develop neuroblastoma than those without risk genotype, and so do carriers of 1-4 risk genotypes (adjusted OR = 1.26, 95% CI = 1.01-1.56). Stratification analysis further revealed risk effect of rs3811464 G>A, rs34787247 G>A and 1-4 risk genotypes in some subgroups. Haplotype analysis of these four SNPs yields two haplotypes significantly correlated with increased neuroblastoma susceptibility. Overall, our finding indicated that LIN28A SNPs, especially rs34787247 G>A, may increase neuroblastoma risk.

[Clinical and genetic diagnosis of a pedigree affected with autosomal recessive Alport syndrome].

OBJECTIVE: To explore the genetic basis of a child with chronic kidney disease featuring renal shrinkage and creatinine increase. METHODS: Peripheral venous blood samples were taken from the child, his brother and two parents and subjected to whole exome sequencing. Suspected mutations were verified by Sanger sequencing. Bioinformatic analysis was carried out to predict the influence of mutations on the structure and function of the protein product. RESULTS: High-throughput and Sanger sequencing revealed that the child has carried compound heterozygous mutations of the COL4A4 gene, namely c.4550T>G in exon 47 (inherited from his mother) and c.199C>T in exon 5 (inherited from his father). Neither mutation was reported previously. Bioinformatic analysis showed that both mutations have located in highly conserved regions. The same mutations were not found in his brother. CONCLUSION: The compound heterozygous c.4550T>G and c.199C>T mutations probably underlie the disease in this child. The findings have enriched the mutation spectrum of the COL4A4 gene.

Gastrodin Rescues Autistic-Like Phenotypes in Valproic Acid-Induced Animal Model.

Autism spectrum disorder (ASD) is an immensely challenging developmental disorder characterized by impaired social interaction, restricted/repetitive behavior, and anxiety. GABAergic dysfunction has been postulated to underlie these autistic symptoms. Gastrodin is widely used clinically in the treatment of neurological disorders and showed to modulate GABAergic signaling in the animal brain. The present study aimed to determine whether treatment with gastrodin can rescue valproic acid (VPA) induced autistic-like phenotypes, and to determine its possible mechanism of action. Our results showed that administration of gastrodin effectively alleviated the autistic-associated behavioral abnormalities as reflected by an increase in social interaction and decrement in repetitive/stereotyped behavior and anxiety in mice as compared to those in untreated animals. Remarkably, the amelioration in autistic-like phenotypes was accompanied by the restoration of inhibitory synaptic transmission, α5 GABAA receptor, and type 1 GABA transporter (GAT1) expression in the basolateral amygdala (BLA) of VPA-treated mice. These findings indicate that gastrodin may alleviate the autistic symptoms caused by VPA through regulating GABAergic synaptic transmission, suggesting that gastrodin may be a potential therapeutic target in autism.

O-GlcNAcylation of BMAL1 regulates circadian rhythms in NIH3T3 fibroblasts.

Various physiological processes and behaviors show a circadian rhythm of approximately 24 h, which is crucial in coordinating internal metabolic processes and environmental signals. Post-translational modifications play an important role in regulating circadian core proteins. In this study, we demonstrated that BMAL1 was modified with an O-linked ß-N-acetylglucosamine (O-GlcNAc), which stabilized BMAL1 and enhanced its transcriptional activity. Conversely, inhibition of O-GlcNAcylation resulted in inhibition of circadian rhythms of clock gene expression. Because O-GlcNAcylation is sensitive to the glucose level, such a modification may provide a new mechanism connecting metabolism with circadian rhythms.