Heterozygous Seryl-tRNA Synthetase 1 Variants Cause Charcot-Marie-Tooth Disease.

OBJECTIVE: Despite the increasing number of genes associated with Charcot-Marie-Tooth (CMT) disease, many patients currently still lack appropriate genetic diagnosis for this disease. Autosomal dominant mutations in aminoacyl-tRNA synthetases (ARSs) have been implicated in CMT. Here, we describe causal missense mutations in the gene encoding seryl-tRNA synthetase 1 (SerRS) for 3 families affected with CMT. METHODS: Whole-exome sequencing was performed in 16 patients and 14 unaffected members of 3 unrelated families. The functional impact of the genetic variants identified was investigated using bioinformatic prediction tools and confirmed using cellular and biochemical assays. RESULTS: Combined linkage analysis for the 3 families revealed significant linkage (Zmax LOD = 6.9) between the genomic co-ordinates on chromosome 1: 108681600-110300504. Within the linkage region, heterozygous SerRS missense variants segregated with the clinical phenotype in the 3 families. The mutant SerRS proteins exhibited reduced aminoacylation activity and abnormal SerRS dimerization, which suggests the impairment of total protein synthesis and induction of eIF2α phosphorylation. INTERPRETATION: Our findings suggest the heterozygous SerRS variants identified represent a novel cause for autosomal dominant CMT. Mutant SerRS proteins are known to impact various molecular and cellular functions. Our findings provide significant advances on the current understanding of the molecular mechanisms associated with ARS-related CMT. ANN NEUROL 2023;93:244-256.

Whole exome sequencing reveals a broader variant spectrum of Charcot-Marie-Tooth disease type 2.

Charcot-Marie-Tooth disease type 2 (CMT2) is a clinically and genetically heterogeneous inherited neuropathy. Although new causative and disease-associated genes have been identified for CMT2 in recent years, molecular diagnoses are still lacking for a majority of patients. We here studied a cohort of 35 CMT2 patients of Chinese descent, using whole exome sequencing to investigate gene mutations and then explored relationships among genotypes, clinical features, and mitochondrial DNA levels in blood as assessed by droplet digital PCR. We identified pathogenic variants in 57% of CMT2 patients. The most common genetic causes in the cohort were MFN2 mutations. Two patients with typical CMT phenotype and neuromyotonia were detected to harbor compound heterozygous variations in the HINT1 gene. In conclusion, our work supports that the molecular diagnostic rate of CMT2 patients can be increased via whole exome sequencing, and our data suggest that assessment of possible HINT1 mutations should be undertaken for CMT2 patients with neuromyotonia.

Melanin-Like Nanoquencher on Graphitic Carbon Nitride Nanosheets for Tyrosinase Activity and Inhibitor Assay.

Graphitic C3N4 (g-C3N4) nanosheets are a type of emerging graphene-like carbon-based nanomaterials with high fluorescence and large specific surface areas that hold great potential for biosensor applications. However, current g-C3N4 based biosensors have prevailingly been limited to coordination with metal ions, and it is of great significance to develop new designs for g-C3N4 nanosheets based biosensors toward biomarkers of general interest. We report the development of a novel g-C3N4 nanosheet-based nanosensor strategy for highly sensitive, single-step and label-free detection of tyrosinase (TYR) activity and its inhibitor. This strategy relies on the catalytic oxidation of tyrosine by TYR into melanin-like polymers, which form a nanoassembly on the g-C3N4 nanosheets and quench their fluorescence. This strategy was demonstrated to provide excellent selectivity and superior sensitivity and to enable rapid screening for TYR inhibitors. Therefore, the developed approach might create a useful platform for diagnostics and drugs screening for TYR-based diseases including melanoma cancer.

Design, Synthesis, and Properties of Silicon-Containing meta-Diamide Insecticides.

Silicon-containing compounds are sporadically used in crop protection and drug discovery and have demonstrated to increase the biological efficacy as well as to reduce toxicity, improve physicochemical properties, and favorably impact the environmental profile. As part of our research, we have investigated the application of bioisosteric silicon replacements in meta-diamide insecticides and studied the biological activity and molecular properties of the corresponding novel compounds. At all meaningful structural elements of the meta-diamides, silicon-containing substituents were introduced and synthetic methodology was developed for their syntheses. As the most promising compound, silicon-containing meta-diamide II-18 emerged, which exhibits a very low LC50 value of 2.00 mg/L against Mythimna separata and compares well to the reference compounds 28 (LC50 = 0.17 mg/L) and II-20 (LC50 = 0.27 mg/L). Our research on silicon-containing crop protection compounds once again confirmed that the biological activity can be beneficially affected by the insertion of silicone substituents and that the introduction of well-chosen silicone motifs is an excellent strategy for agrochemical research.

Evaluation of SORD mutations as a novel cause of Charcot-Marie-Tooth disease.

Biallelic mutations in the sorbitol dehydrogenase (SORD) encoding gene were recently identified as a common genetic cause in autosomal-recessive CMT patients. Here, we investigated the clinical, genetic, and electrophysiological characteristics of three CMT patients with biallelic SORD mutations from a Chinese cohort. Two patients harbored c.757delG (p.A253Qfs*27) homozygous mutations, and one patient carried both c.757delG (p.A253Qfs*27) and c.625C>T (p.R209X) compound heterozygous mutations. Interestingly, the two patients homozygous for the c.757delG mutation exhibited positive responses for pinprick test. In conclusion, we confirmed SORD mutations as causative for CMT and further expanded the mutational and phenotypic spectrum of SORD-related CMT.

[Inclination of crown and tooth longitudinal axis in cephalometric analysis of normal occlusions].

OBJECTIVE: We measured and analyzed the angle between the longitudinal axis of incisor crown and tooth to provide a reference for orthodontists for selecting orthodontic methods and evaluating treatment results. METHODS: A total of 120 participants were included according to the criteria of Andrews' six keys, and cephalometric radiograph under the instructions of modified natural head position acquirement method was performed. The angles of maxillary incisor crown longitudinal axis, tooth longitudinal axis, occlusion plane (OP), and true vertical (TV) plane were measured, as well as mandibular incisors. RESULTS: As for maxillary incisors, the angle between crown longitudinal axis and TV plane, OP, and tooth longitudinal axis were 11.72°±4.71°, 73.29°±5.69°, and 20.04°±3.71°, respectively. For mandibular incisors, the angle between crown longitudinal axis and TV plane, OP, and tooth longitudinal axis were 16.03°±5.40°, 81.76°±4.81°, and 14.82°±4.01°, respectively. For the maxillary incisor, the angles between crown longitudinal axis and tooth longitudinal axis were mainly within 15° to 25°, whereas those for mandibular incisors were within 10° to 20°. CONCLUSIONS: The longitudinal axis inclinations of the maxillary and mandibular incisor crown and of the incisor tooth need to be considered when cephalometric radiographs are used for treatment planning or for evaluating the treatment result.

Optimization on condition of glycyrrhetinic acid liposome by RSM and the research of its immunological activity.

The aim of this study is to prepare glycyrrhetinic acid liposome (GAL) and optimize the preparation condition and to investigate further whether liposome could promote the immunological activity of glycyrrhetinic acid (GA). GAL was prepared using a film-dispersion method and the preparation conditions of GAL were optimized with response surface methodology (RSM). Moreover, GAL prepared under the optimal preparation conditions was added into chicken's T and B lymphocytes in vitro. The optimal preparation conditions for GAL by response surface methodology was as follows: ratio 9:1, soybean phospholipid cholesterol (w/w) 2.5:1 and water bath temperature 31 °C. Under these conditions, the experimental encapsulation efficiency of GAL was 83.46 ± 0.55%, which was close with the predicted value. Therefore, the optimized preparation condition is very reliable. The results showed that GAL could significantly promote T and B lymphocytes proliferation singly or synergistically with PHA and LPS and the concentration of immunoglobulins G (IgG) and immunoglobulins M (IgM). These results indicated that liposome could significantly improve the immunological activity of GA and drug action of GA. GAL demonstrates the significant immunological activity, which provides the theoretical basis for the further experiment in vivo.

Modulus of elasticity of human periodontal ligament by optical measurement and numerical simulation.

OBJECTIVE: To determine the elastic modulus of the periodontal ligament (PDL). MATERIALS AND METHODS: This study was carried out on eight human maxillary jaw segments containing central incisors. Displacements were measured under load using a laser sensing system, electronic speckle pattern interferometry (ESPI). Subsequently, finite element models presenting the same individual geometry as the respective autopsy material were developed by the software of Mimics and Ansys, based on scanning data from micro computed tomography (micro CT), to simulate tooth mobility numerically under the same force systems as were used in the experiment. Numerical force/deflection curves obtained from the models were fitted to the experimental curves by repeatedly calculating theoretical tooth deflections and varying the elasticity parameters of the human PDL. RESULTS: A bilinear material parameter set was assumed to simulate tooth deflections. Mean values of E1  =  0.04 MPa, E2  =  0.16 MPa, and ultimate strain of ε12  =  7.3% were derived for the elastic behavior of the PDL. CONCLUSION: Force/deflection curves from the measurements showed a significant nonlinear behavior of elastic stiffness of the PDL. A bilinear material parameter set was suitably assumed to be a description of nonlinear properties of the PDL.

[To investigate pathogen of hand, foot and mouth disease in Shenzhen in 2008].

OBJECTIVE: To investigate EV71 and CA16 pathogen of HFMD in Shenzhen in 2008, and to provide the evidence for the prevention and treatment HFMD. METHOD: Using RT-PCR technology to detect the EV71 and CoxA16 genes of 307 samples HFMD; sequencing the purified PCR products from 14 samples. Using ClustalW2 online analysis software for sequence and phylogenetic analysis of enterovirus 71. RESULT: Percentage of positive EV71 from different samples is shown as follows respectively: positive EV71 from stool samples is 24.4% (75/307), from throat swab--7.8% (24/307), from peripheral blood--12.5% (1/8). Percentage of positive CoxA16 is shown as follows respectively: positive EV71 from stool samples is 13.8% (28/203), from throat swab-11.0% (20/181). Among all the 307 samples, three are positive for both EV71 and CoxA16. EV71 and CoxA16 are not detected in the samples of cerebrospinal fluid.Comparative analysis of nucleotide sequences of EV71 with those of strains BrCr and 11 deposited in GenBank demonstrated numerous disparities from 8 samples, but residue 595 from 2 samples and residue 658 from 1 sample are variable. The phylogenetic analysis based on VP1 region demonstrates that strains from 2 samples has the nearest genetic relationship with anhui strains, the farthest with BrCr and SHH02-6, SHZH02-40, SHZH03-58 strains, also strains from other 12 samples have the farthest genetic relationship with them. The genotypes A, B and C were classified as proposed by Brown et al. (1999). The EV71 from 14 samples were the member of genotype C. CONCLUSION: EV71 among the pathogen of HFMD in Shenzhen in 2008 was majority. These EV71 may belong to the same genegroup with Anhui predominant strains.

[Expression of recombinant VP1 protein of enterovirus 71 and development of serological assay for detection of EV71 infection].

OBJECTIVE: To obtain a recombinant purified Enterovirus 71 VPI protein and establishment of an early, rapid and accurate serological ELISA (enzyme-linked immunosorbent assay) for detection of EV71 infection. METHODS: VP1 gene was amplified by PCR and clonel into pET-21b (+) vector, the positive recombinant plasmid were transformed into E. coli BI21(DE3), and was induced with IPTG, the recombinant protein by SDS-PAGE and Western Blot assays. Finally, the recombinant purified VP1 protein was used as a coated antigen for detection of serum anti-IgM and IgG against EV71 by ELISA. RESULTS: The purified VP1 was obtained, and it can be recognized by sera of patients with EV71 infection associated with hand-foot-mouth disease. The A values of anti-EV71 IgM and IgG were significantly elevated as compared to healthy objects and HFMD patients without EV71 infection (P < 0.05). The sensitivity and specificity of IgM to EV71 were 73% and 77% compared with the RT-PCR results, respectively;and those of IgG being 82% and 83%, respectively. CONCLUSIONS: The recombinant protein VP1 was produced and purified, and it was proved to have a good antigenicity and could be used to develop a serological diagnosis kit for EV71 infection in the future.

[Study of the clinical and laboratory features of hand-foot-mouth disease].

OBJECTIVE: To study the clinical and laboratory features of the mild and severe hand-foot-mouth diseases (HFMD) in Shenzhen in 2008. METHODS: 145 cases were observed in East-Lake Hospital and Shenzhen Children's Hospital. Of the 145 cases, 124 mild cases and 21 severe cases were involved.All the clinical data and laboratory findings were collected and summarized. After collection of the acute and convalescent consecutive stools and peripheral bloods from the patients with HFMDI, EV71 genes were amplified from these samples by RT-PCR. Enterovirus 71 were cultured and isolated using Vero cell line and R&D cell line. RESULTS: The WBC counts and blood glucose levels of the severe cases were significantly elevated, but the ages of the severe ones significantly decreased compared with those of the mild cases (P < 0.05). EV71 genes could be detected by RT-PCR with 35% positive rate in mild cases and 67% in severe cases. The EV71 gene detection rate of the severe cases was significantly increased in contrast to that of the mild ones. The EV71 were isolated and cultured from the stools of 9 patients, one specimens from the dead's stool. Two severe cases died of neurogenic pulmonary edema and brain-stem encephalitis. CONCLUSIONS: EV71 mainly contributes to HFMD and is responsible for death of some severe cases. High fever, less rash, elevated white blood cell counts and blood glucose concentrations as well as age less than 4 years old should be used for prediction of severe cases.