Evoked Potentials ; Social Isolation ; Attention ; Humans

γ-aminobutyric acid can be produced by a one-step enzymatic reaction catalyzed by glutamic acid decarboxylase. The reaction system is simple and environmentally friendly. However, the majority of GAD enzymes catalyze the reaction under acidic pH at a relatively narrow range. Thus, inorganic salts are usually needed to maintain the optimal catalytic environment, which adds additional components to the reaction system. In addition, the pH of solution will gradually rise along with the production of γ-aminobutyric acid, which is not conducive for GAD to function continuously. In this study, we cloned the glutamate decarboxylase LpGAD from a Lactobacillus plantarum capable of efficiently producing γ-aminobutyric acid, and rationally engineered the catalytic pH range of LpGAD based on surface charge. A triple point mutant LpGAD^{S24R/D88R/Y309K} was obtained from different combinations of 9 point mutations. The enzyme activity at pH 6.0 was 1.68 times of that of the wild type, suggesting the catalytic pH range of the mutant was widened, and the possible mechanism underpinning this increase was discussed through kinetic simulation. Furthermore, we overexpressed the Lpgad and Lpgad^{S24R/D88R/Y309K} genes in Corynebacterium glutamicum E01 and optimized the transformation conditions. An optimized whole cell transformation process was conducted under 40 ℃, cell mass (OD₆₀₀) 20, 100 g/L l-glutamic acid substrate and 100 μmol/L pyridoxal 5-phosphate. The γ-aminobutyric acid titer of the recombinant strain reached 402.8 g/L in a fed-batch reaction carried out in a 5 L fermenter without adjusting pH, which was 1.63 times higher than that of the control. This study expanded the catalytic pH range of and increased the enzyme activity of LpGAD. The improved production efficiency of γ-aminobutyric acid may facilitate its large-scale production.
Glutamate Decarboxylase/genetics* ; Lactobacillus plantarum/genetics* ; Catalysis ; gamma-Aminobutyric Acid ; Hydrogen-Ion Concentration ; Glutamic Acid

L-arabinose isomerase (L-AI) is the key enzyme that isomerizes D-galactose to D-tagatose. In this study, to improve the activity of L-arabinose isomerase on D-galactose and its conversion rate in biotransformation, an L-arabinose isomerase from Lactobacillus fermentum CGMCC2921 was recombinantly expressed and applied in biotransformation. Moreover, its substrate binding pocket was rationally designed to improve the affinity and catalytic activity on D-galactose. We show that the conversion of D-galactose by variant F279I was increased 1.4 times that of the wild-type enzyme. The K_m and k_cat values of the double mutant M185A/F279I obtained by superimposed mutation were 530.8 mmol/L and 19.9 s^-1, respectively, and the catalytic efficiency was increased 8.2 times that of the wild type. When 400 g/L lactose was used as the substrate, the conversion rate of M185A/F279I reached a high level of 22.8%, which shows great application potential for the enzymatic production of tagatose from lactose.
Galactose/metabolism* ; Limosilactobacillus fermentum/genetics* ; Lactose ; Hexoses/metabolism* ; Aldose-Ketose Isomerases/genetics* ; Hydrogen-Ion Concentration

Neurodevelopmental disorders (NDDs) in children are a group of chronic developmental brain disorders caused by multiple genetic or acquired causes, including disorders of intellectual development, developmental speech or language disorders, autism spectrum disorders, developmental learning disorders, attention deficit hyperactivity disorder, tic disorders, and other neurodevelopmental disorders. With the improvement in the research level and the diagnosis and treatment techniques of NDDs, great progress has been made in the research on NDDs in children. This article reviews the research advances in NDDs, in order to further improve the breadth and depth of the understanding of NDDs in children among pediatricians.
Humans ; Child ; Neurodevelopmental Disorders/therapy* ; Autism Spectrum Disorder/therapy* ; Attention Deficit Disorder with Hyperactivity

Atomoxetine is the first non-stimulant drug for the treatment of children and adults with attention deficit hyperactivity disorder (ADHD), and its safety and efficacy show significant differences in the pediatric population. This article reviews the genetic factors influencing the pharmacokinetic differences of atomoxetine from the aspect of the gene polymorphisms of the major metabolizing enzyme CYP2D6 of atomoxetine, and then from the perspective of therapeutic drug monitoring, this article summarizes the reference ranges of the effective concentration of atomoxetine in children with ADHD proposed by several studies. In general, there is an association between the peak plasma concentration of atomoxetine and clinical efficacy, but with a lack of data from the Chinese pediatric population. Therefore, it is necessary to establish related clinical indicators for atomoxetine exposure, define the therapeutic exposure range of children with ADHD in China, and combine CYP2D6 genotyping to provide support for the precision medication of atomoxetine.
Adult ; Child ; Humans ; Adrenergic Uptake Inhibitors/therapeutic use* ; Atomoxetine Hydrochloride/therapeutic use* ; Attention Deficit Disorder with Hyperactivity/genetics* ; Cytochrome P-450 CYP2D6/therapeutic use* ; Drug Monitoring ; Genetic Testing ; Propylamines/therapeutic use* ; Treatment Outcome

OBJECTIVES: To study the mediating role of emotion regulation strategies between symptoms and anxiety problems in children with attention deficit hyperactivity disorder (ADHD). METHODS: Convenience cluster sampling was performed to select 435 eligible children with ADHD from 10 211 students in 10 elementary schools in Kashgar of Xinjiang Uygur Autonomous Region. SNAP-IV Assessment Scale, Emotion Regulation Questionnaire for Children and Adolescents, and Conners Parent Symptoms Questionnaire were used to obtain relevant information. The Spearman correlation analysis and the moderated mediation method were used for data analysis. RESULTS: For the children with ADHD, the score of cognitive reappraisal was negatively correlated with the scores of symptoms and anxiety problems (r_s=-0.254 and -0.270 respectively, P<0.001), and the score of symptoms was positively correlated with the score of anxiety problems (r_s=0.329, P<0.001). The analysis of mediating effect showed that cognitive reappraisal played a partial mediating role between symptoms and anxiety problems, with a 95% confidence interval of 0.008-0.027, and the mediating effect accounted for 16.10% of the total effect. In addition, sex played a regulatory role in this mediation model (P<0.001). CONCLUSIONS Cognitive reappraisal plays a mediating role between symptoms and anxiety problems in children with ADHD, while sex regulates the relationship of symptoms with cognitive reappraisal and anxiety problems in these children.
Adolescent ; Humans ; Child ; Attention Deficit Disorder with Hyperactivity ; Emotional Regulation ; Anxiety ; Anxiety Disorders ; Surveys and Questionnaires

OBJECTIVE: To investigate the preparation of decellularized small intestinal submucosa (dSIS) sponge scaffolds with chelated strontium (Sr) ions at different pH values, and to select the appropriate pH values for synthesizing Sr/dSIS scaffolds using the physicochemical properties and biocompatibility of the scaffolds as evaluation indexes. METHODS: (1) Sr/dSIS scaffolds preparation and grouping: After mixing dSIS solution and strontium chloride solution in equal volumes, adjusting pH of the solution to 3, 5, 7, and 9 respectively, porous scaffolds were prepared by freeze-drying method after full reaction at 37℃, which were named Sr/dSIS-3, -5, -7, and -9 respectively, and the dSIS scaffolds were used as the control group. (2) Physicochemical property evaluation: The bulk morphology of the scaffolds was observed in each group, the microscopic morphology analyzed by scanning electron microscopy, and the porosity and pore size determined, the surface elements analyzed by energy spectroscopy, the structure of functional groups analyzed by infrared spectroscopy, the chelation rate determined by atomic spectrophotometry, the water absorption rate detected by using specific gravity method, and the compression strength evaluated by universal mechanical testing machine.(3) Biocompatibility evaluation: The cytotoxicity and proliferative effect to bone mesenchymal stem cells (BMSCs) of each group were evaluated by Calcein-AM/PI double staining method. RESULTS: Scanning electron microscopy showed that the scaffolds of each group had an interconnected three-dimensional porous structure with no statistical difference in pore size and porosity. Energy spectrum analysis showed that strontium could be detected in Sr/dSIS-5, -7 and -9 groups, and strontium was uniformly distributed in the scaffolds. Functional group analysis further supported the formation of chelates in the Sr/dSIS-5, -7 and -9 groups. Chelation rate analysis showed that the Sr/dSIS-7 group had the highest strontium chelation rate, which was statistically different from the other groups (P < 0.05). The scaffolds in all the groups had good water absorption. The scaffolds in Sr/dSIS-5, -7 and -9 groups showed significantly improved mechanical properties compared with the control group (P < 0.05). The scaffolds in all the groups had good biocompatibility, and the Sr/dSIS-7 group showed the best proliferation of BMSCs. CONCLUSION When pH was 7, the Sr/dSIS scaffolds showed the highest strontium chelation rate and the best proliferation effect of BMSCs, which was the ideal pH value for the preparation of the Sr/dSIS scaffolds.
Tissue Scaffolds/chemistry* ; Biocompatible Materials ; Strontium/pharmacology* ; Ions ; Hydrogen-Ion Concentration ; Tissue Engineering/methods* ; Porosity

Gas vesicles (GVs) are gas-filled protein nanostructures that can regulate the buoyancy of microorganisms such as cyanobacteria and archaea. Recent studies have shown that GVs have the potential to be used as ultrasound molecular imaging probes in disease diagnosis and treatment. However, the mechanism of the inflation and deflation of GVs remains unclear, which hampers the preservation of GVs and gas replacement. In the present study, the environmental pH value was found to be an important factor in regulating the inflation and deflation of GVs. It can not only regulate the inflation and deflation of GVs in vivo to make Microcystis sp. cells present distinct levitation state, but also regulate the inflation and deflation of purified GVs in vitro, and the regulation process is reversible. Our results may provide a technical support for the large-scale production and preservation of biosynthetic ultrasound molecular imaging probes, especially for gas replacement to meet different diagnostic and therapeutic needs, and would facilitate the application of biosynthetic ultrasound molecular imaging probes.
Cyanobacteria ; Proteins/chemistry* ; Nanostructures/chemistry* ; Molecular Imaging ; Hydrogen-Ion Concentration

The hydrolysis of xylo-oligosaccharides catalyzed by β-xylosidase plays an important role in the degradation of lignocellulose. However, the enzyme is easily inhibited by its catalytic product xylose, which severely limits its application. Based on molecular docking, this paper studied the xylose affinity of Aspergillus niger β-xylosidase An-xyl, which was significantly differentially expressed in the fermentation medium of tea stalks, through cloning, expression and characterization. The synergistic degradation effect of this enzyme and cellulase on lignocellulose in tea stems was investigated. Molecular docking showed that the affinity of An-xyl to xylose was lower than that of Aspergillus oryzae β-xylosidase with poor xylose tolerance. The K_i value of xylose inhibition constant of recombinant-expressed An-xyl was 433.2 mmol/L, higher than that of most β-xylosidases of the GH3 family. The K_m and V_max towards pNPX were 3.6 mmol/L and 10 000 μmol/(min·mL), respectively. The optimum temperature of An-xyl was 65 ℃, the optimum pH was 4.0, 61% of the An-xyl activity could be retained upon treatment at 65 ℃ for 300 min, and 80% of the An-xyl activity could be retained upon treatment at pH 2.0-8.0 for 24 h. The hydrolysis of tea stem by An-xyl and cellulase produced 19.3% and 38.6% higher reducing sugar content at 2 h and 4 h, respectively, than that of using cellulase alone. This study showed that the An-xyl mined from differential expression exhibited high xylose tolerance and higher catalytic activity and stability, and could hydrolyze tea stem lignocellulose synergistically, which enriched the resource of β-xylosidase with high xylose tolerance, thus may facilitate the advanced experimental research and its application.
Aspergillus niger/genetics* ; Xylose/metabolism* ; Molecular Docking Simulation ; Xylosidases/genetics* ; Cellulases ; Tea ; Hydrogen-Ion Concentration ; Substrate Specificity

β-glucosidase has important applications in food, pharmaceutics, biomass conversion and other fields, exploring β-glucosidase with strong adaptability and excellent properties thus has received extensive interest. In this study, a novel glucosidase from the GH1 family derived from Cuniculiplasma divulgatum was cloned, expressed, and characterized, aiming to find a better β-glucosidase. The amino acid sequences of GH1 family glucosidase derived from C. divulgatum were obtained from the NCBI database, and a recombinant plasmid pET-30a(+)-CdBglA was constructed. The recombinant protein was induced to express in Escherichia coli BL21(DE3). The enzymatic properties of the purified CdBglA were studied. The molecular weight of the recombinant CdBglA was 56.0 kDa. The optimum pH and temperature were 5.5 and 55 ℃, respectively. The enzyme showed good pH stability, 92.33% of the initial activity could be retained when treated under pH 5.5-11.0 for 1 h. When pNPG was used as a substrate, the kinetic parameters K_m, V_max and K_cat/K_m were 0.81 mmol, 291.99 μmol/(mg·min), and 387.50 s^-1 mmol^-1, respectively. 90.33% of the initial enzyme activity could be retained when CdBglA was placed with various heavy metal ions at a final concentration of 5 mmol/L. The enzyme activity was increased by 28.67% under 15% ethanol solution, remained unchanged under 20% ethanol, and 43.68% of the enzyme activity could still be retained under 30% ethanol. The enzyme has an obvious activation effect at 0-1.5 mol/L NaCl and can tolerate 0.8 mol/L glucose. In conclusion, CdBglA is an acidic and mesophilic enzyme with broad pH stability and strong tolerance to most metal ions, organic solvents, NaCl and glucose. These characteristics may facilitate future theoretical research and industrial production.
beta-Glucosidase ; Sodium Chloride ; Temperature ; Glucose ; Ethanol/chemistry* ; Ions ; Hydrogen-Ion Concentration ; Enzyme Stability ; Substrate Specificity