Advances in medicinal research on tetrodotoxin / 药学实践杂志

Tetrodotoxin (TTX) is a neurotoxin found in puffer fish and other marine organisms. It has been used as an inhibitor of voltage-gated sodium channels (VGSCs), which could selectively bind to the α-subunit on the outer vestibule of VGSCs, preventing sodium ions from entering the channel, resulting in pharmacological activities. As a typical sodium channel blocker, TTX shows a significant analgesic effect. TTX could selectively block Na+ channels without affecting other ion channels, therefore it could reduce the probability of adverse reactions caused by commonly used antiarrhythmic drugs. In addition, TTX has a significant role in detoxification and prevention of renal failure, so TTX has great potential as a medicine. The structure and physicochemical properties, mechanism of action, pharmacological activities and preparations of tetrodotoxin have been reviewed in this paper, so as to provide a general support for the evaluation of its druggability and application in the field of pharmacy.

Genomic, transcriptomic, and epigenomic analysis of a medicinal snake, Bungarus multicinctus, to provides insights into the origin of Elapidae neurotoxins

The many-banded krait, Bungarus multicinctus, has been recorded as the animal resource of JinQianBaiHuaShe in the Chinese Pharmacopoeia. Characterization of its venoms classified chief phyla of modern animal neurotoxins. However, the evolutionary origin and diversification of its neurotoxins as well as biosynthesis of its active compounds remain largely unknown due to the lack of its high-quality genome. Here, we present the 1.58 Gbp genome of B. multicinctus assembled into 18 chromosomes with contig/scaffold N50 of 7.53 Mbp/149.8 Mbp. Major bungarotoxin-coding genes were clustered within genome by family and found to be associated with ancient local duplications. The truncation of glycosylphosphatidylinositol anchor in the 3'-terminal of a LY6E paralog released modern three-finger toxins (3FTxs) from membrane tethering before the Colubroidea divergence. Subsequent expansion and mutations diversified and recruited these 3FTxs. After the cobra/krait divergence, the modern unit-B of β-bungarotoxin emerged with an extra cysteine residue. A subsequent point substitution in unit-A enabled the β-bungarotoxin covalent linkage. The B. multicinctus gene expression, chromatin topological organization, and histone modification characteristics were featured by transcriptome, proteome, chromatin conformation capture sequencing, and ChIP-seq. The results highlighted that venom production was under a sophisticated regulation. Our findings provide new insights into snake neurotoxin research, meanwhile will facilitate antivenom development, toxin-driven drug discovery and the quality control of JinQianBaiHuaShe.

The Roles of the Canonical Members of Ribonuclease A Family in Tumorigenesis / 中国生物化学与分子生物学报

In human, the ribonuclease A (RNase A) family contains 8 canonical members (RNase 1-RNase 8). Research evidence indicated that all the canonical members of this family, except RNase 8, are involved in the occurrence and development of a variety of tumors, including pancreatic cancer, colorectal cancer, bladder cancer, breast cancer, and skin cancer, etc. During tumorigenesis, the expression of specific RNase increased and glycosylation modifications changed, which are potential markers for tumor diagnosis; They also participate in tumor initiation, growth, and metastasis with a variety of mechanisms, and are potential targets for tumor therapy; Meanwhile, some members have the function of killing tumor cells and inhibiting tumor development, and it is possible to develop into tumor therapeutic drugs. Concretely, RNase 1 suppresses tumor growth by directly killing tumor cells or reducing local inflammation through its ribonuclease activity-dependent cytotoxicity and extracellular RNA degradation functions; however, its binding and activation of ephrin A4 signaling pathway promotes breast cancer initiation. RNase 2 and RNase 3 are important components of eosinophil granule proteins that play an important role in anti-tumor immune defense, and their function of killing tumor cells depends on both cationic nature and ribonuclease activity. RNase 4 and RNase 5 can promote tumorigenesis by inducing angiogenesis, promoting tumor cell proliferation, and inhibiting tumor cell apoptosis. The molecular mechanisms of RNase 5 action include promoting the transcription of 47 S precursor rRNA, activating signaling pro-tumor growth signaling pathways, and generating tRNA-derived stress-induced RNA (tiRNA). Besides, RNase 6 and RNase 7 are related to the occurrence of tumors thought their specific role and mechanism are still unclear. In this review, we summarized the relevance and mechanism of RNase A family members on promoting or inhibiting tumors and analyzed their clinical application potentials.

Significance of fecal calprotectin and eosinophil-derived neurotoxin in non-IgE-mediated food allergy / 国际儿科学杂志

Non-IgE-mediated food allergy most often presents with gastrointestinal symptoms such as diarrhoea, mucus stools, bloody stools, reflux and vomiting a few hours to days after exposure to the allergenic food.The pathogenesis may be related to the activation of intestinal T lymphocytes to secrete pro-inflammatory cytokines such as TNF-α and IL-4 by food allergens, leading to migration of neutrophils and eosinophils into the intestinal lumen, causing an intestinal inflammatory response and increased intestinal permeability.There is no rapid and specific diagnostic method.The diagnosis is mainly based on clinical manifestations combined with food avoidance and oral food challenge.In recent years, fecal biomarkers have been widely used as specific indicators for determining intestinal inflammation as an aid to diagnosis and condition assessment of intestinal infections and inflammatory bowel diseases, but their application in gastrointestinal allergic diseases is rarely reported.In this paper, we will focus on the significance of fecal calprotectin, fecal eosinophil-derived neurotoxin in non-IgE-mediated food allergy.

Clinical characteristics of children with atopic dermatitis and change in serum levels of Apo A1, 25(OH)D and EDN / 中国医师杂志

Objective:To investigate the clinical characteristics of children with atopic dermatitis (AD) and the changes in serum levels of apolipoprotein A1 (Apo A1), 25-hydroxyvitamin D [25(OH)D] and eosinophil derived neurotoxin (EDN).Methods:200 children with AD treated in Zhuzhou Central Hospital from January 2016 to December 2019 were selected retrospectively as AD group and 100 healthy children as control group. The clinical characteristics of children with AD were analyzed, and the differences in serum Apo A1, 25 (OH)D and EDN levels between two groups were compared. The relationships between serum Apo A1, 25(OH)D, EDN levels and severity of AD were explored.Results:The male to female composition ratio of 200 AD patients was 1.41∶1, and the age of onset <3 months was the highest (64.50%). Inhalation allergens were detected in 118 cases (59.00%) and ingestion allergens in 82 cases (41.00%). The levels of Apo A1 and EDN in AD group were significantly higher than those in control group, while the level of 25(OH)D was significantly lower than that in control group ( P<0.05). With the aggravation of the disease, the serum Apo A1 and EDN levels in AD children increased gradually, while the serum 25(OH)D level decreased significantly (all P<0.05). Severity Scoring of Atopic Dermatitis (SCORAD) was positively correlated with Apo A1 and EDN levels ( P<0.05), and was negatively correlated with 25(OH)D level (all P<0.05). Conclusions:Apo A1, 25 (OH)D and EDN are involved in the pathogenesis of AD in children, and their serum levels are closely related to the severity of AD.

Métodos de extração, estabilidade e biodisponibilidade da guanitoxina em cultivos de Sphaeropermopsis torques-reginae analisadas por espectrometria de massas / Methods of extraction, stability, and bioavailability of guanitoxin in Sphaeropermopsis torques-reginae cultures analyzed by mass spectrometry

A guanitoxina (GNT) é uma neurotoxina produzida por algumas cepas de cianobactérias dos gêneros Dolichospermum e Sphaerospermopsis>. A GNT é o único organofosforado natural, capaz de causar a morte de animais selvagens e domésticos devido à inibição irreversível da acetilcolinesterase. Apesar de sua alta toxicidade, o diagnóstico da GNT em amostras biológicas ainda é um grande desafio. A dificuldade para sua detecção está diretamente ligada à sua instabilidade em altas temperaturas e pH alcalino, tornando difícil seu monitoramento em corpos d'água. Por isso, esta pesquisa objetivou estudar a estabilidade e biodisponibilidade da GNT em amostras aquosas, com intuito de obter mais informações sobre a natureza química e biológica dessa potente neurotoxina. Para realizar este estudo, a cepa ITEP-24 (S. torques-reginae) produtora de GNT foi cultivada em laboratório sob condições controladas, para obter biomassa para os experimentos de extração, semi-isolamento, estabilidade, ensaio in vitro e identificação por LC-MS/MS. Primeiramente foram realizados testes de extração da GNT partir de células liofilizadas da cepa ITEP-24 utilizando água, metanol e etanol em pH ácido. Depois utilizou-se dois métodos de extração em fase sólida (SPE) com cartuchos preenchidos com fases estacionarias C18 em fase reversa e sílica gel em fase normal, com objetivo de avaliar qual método de SPE seria melhor para extrair e concentrar a GNT. Nós também testamos métodos para lisar as células com sondas de ultrassom, misturador e centrifugação. Além dos métodos de extração, nós avaliamos a estabilidade da toxina em diferentes temperaturas, para isso a biomassa seca contendo a GNT ficou condicionada a 4 °C, 23 °C, -20 °C, -80 °C durantes seis meses, e análises de identificação foram realizadas dentro período de 150 dias em uma sequência de 30 dias. A estabilidade da toxina foi analisada também a partir de extrações em soluções com diferentes valores de pH (1,5; 3,0; 5,0; 7,0; 8,5; 10,5) e temperatura (23 ºC e 37 ºC). Depois, analisou-se a biodisponibilidade da GNT em células frescas da linhagem ITEP-24 através de teste de dissolução in vitro. O objetivo deste teste foi avaliar a liberação da toxina intracelular em meio simulado do conteúdo gástrica e intestinal com e sem enzimas digestivas para compreender e estimar a disponibilidade da GNT in vivo. Os resultados de todos experimentos descritos neste estudo, foram obtidos a partir de análises por cromatografia líquida de interação hidrofílica (HILIC) acoplado ao espectrômetro de massas do tipo triplo quadrupolo LC-QqQ-MS/MS utilizando as transições 253>58, 253>159 e 159>58 [M+H]+ utilizando coluna com fase estacionária zwitteriônica (ZIC). A identificação da GNT foi realizada também por cromatografia líquida acoplada ao espectrômetro de massas de alta resolução (LC-HR-QTOF-MS) com coluna Luna C18, Hydro-RP C18 e ZIC-HILIC. Dos protocolos de extração testados, a combinação de metanol/água (70:30 v/v) com ácido acético (0.3%) extraiu maior quantidade relativa da GNT a partir de células frescas e liofilizadas da cepa ITEP-24 e a concentração da toxina foi maior em amostras de células frescas. Em relação aos métodos de lise celular, as extrações realizadas em sonda de ultrassom com banho-maria e centrifugação por 1h foram estatisticamente significantes para liberar a toxina intracelular. Não houve diferença significativa entre os testes de SPE, no entanto, a semipurificação da toxina foi melhor com cartucho preenchido com sílica gel em fase normal e adaptação desse método em coluna aberta permitiu obter uma fração enriquecida com GNT. A GNT mostrou ser mais estável em pH ácido, sendo o pH 3,0 o melhor para manter e extrair a toxina em amostras aquosas e a toxina intracelular presente em células secas podem degradar em temperatura de 23 °C por um período de 150 dias mesmo em solução com pH 3,0. Durante os testes de extração e purificação foi observado também a degradação da toxina em processos de secagem e ressuspensão. As análises realizadas no LC-HR-QTOF-MS com diferentes métodos cromatográficos possibilitou a identificação da GNT, porém o método realizado com coluna ZIC-HILIC mostrou melhor resolução cromatográfica dos picos relativos m/z e tempo de retenção de toxina. Os resultados obtidos nos testes de dissolução in vitro mostraram que a GNT fica mais disponível no simulado gástrico com e sem a enzima pepsina, mas também pode ser absorvida no intestino. Portanto, o teste de dissolução in vitro pode ser uma ferramenta útil para a avaliação de risco de cianotoxinas in vivo, devido ao seu potencial de monitorar qualitativa e quantitativamente substâncias dissolvidas em fluidos gastrointestinais. Os resultados apresentados neste estudo fornecem informações valiosas para uma melhor compreensão da estabilidade e biodisponibilidade do GNT. Além disso, os métodos apresentados neste estudo podem ser úteis para diversas aplicações projetadas para identificar a toxina em amostras ambientais, bem como orientações para procedimentos de purificação da GNT

Guanitoxin (GNT) is a neurotoxin produced by some strains of cyanobacteria of the genus Dolichospermum and Sphaerospermopsis. GNT is the only natural organophosphate, capable of causing the death of animals from wild and domestic animals due to irreversible inhibition of acetylcholinesterase. Despite its high toxicity, the diagnosis of GNT in biological samples is still a significant challenge. The difficulty in its detection is directly linked to its instability at high temperatures and alkaline pH, making it difficult to monitor in bodies of water. Therefore, this research aimed to study the stability and bioavailability of GNT in aqueous samples to provide more information about the chemical and biological nature of this molecule. The strain ITEP-24 (S. torques-reginae) producing GNT was grown in the laboratory under controlled conditions to obtain biomass for the extraction, semi-isolation, stability, in vitro tests, and toxin identification by LC-MS/MS. Firstly, tests were carried out to extract GNT from lyophilized cells strain ITEP-24 using water, methanol, and ethanol at acidic pH and, two SPE methods in cartridges with stationary phases of C18 reverse phase and normal phase gel silica, to evaluate which would be better to extract and concentrate the GNT. We also tested different methods of cell lysis, such as ultrasound probes, mixers, and centrifugation. In addition to the extraction methods, the stability of the toxin was evaluated at different temperatures, for this, the dry biomass containing the toxin was conditioned at 4 °C, 23 °C, -20 °C, -80 °C for 150 days and analysis of the identification of the GNT was carried out within that period in a sequence of 30 days. The toxin stability was also analyzed from extractions in solutions with different pH values (1.5; 3.0; 5.0; 7.0; 8.5; 10.5) and temperature (23 ºC and 37 ºC). In addition, we performed dissolution tests with fresh cells of the ITEP-24 strain to evaluate the bioavailability of GNT in simulated gastric and intestinal fluids with and without digestive enzymes to understand and estimate the availability of GNT in vivo. The results of all experiments described in this study were obtained from analyzes by hydrophilic interaction liquid chromatography (HILIC) coupled to the LC-QqQ-MS/MS triple quadrupole mass spectrometer using the transitions m/z 253> 58, m/z 253> 159 and m/z 159> 58 [M + H]+ using a column with the zwitterionic stationary phase (ZIC). Liquid chromatography coupled to the high-resolution mass spectrometer (LC-HR-QTOF-MS) with Luna column C18, Hydro-RP C18, and ZIC-HILIC carried out the identification of the GNT. From the extraction protocols tested, the combination of methanol/water (70:30 v/v) with acetic acid (0.3%) extracted a greater relative amount of GNT from fresh and lyophilized ITEP-24 cells, and the concentration of the toxin is higher previously fresh. Concerning cellular methods, the ultrasound probe with a water bath and centrifugation for 1h ware statistically significant to release the intracellular toxin. There was no significant difference between the SPE tests. However, the semi-purification of the toxin was better with a cartridge filled with gel silica in the normal phase and adaptation of this method in an open column allowed to obtain a fraction enriched with GNT. GNT was more stable at acid pH, with pH 3.0 being the best to maintain and the intracellular toxin present in dry cells can degrade at a temperature at 23 °C for 150 days even in pH 3.0 solution. The toxin can also hydrolyze in the drying and resuspension processes. The analyzes carried out in LC-HR-QTOF-MS with different chromatographic methods made it possible to identify the GNT itself, however, the ZIC-HILIC column method showed excellent chromatographic resolution of the relative m/z peaks and toxin retention time. The results obtained in the in vitro dissolution tests showed that GNT is more available in the gastric simulation with and without the enzyme pepsin, but it can also be absorbed in the intestine. Thus, in vitro dissolution tests can be used as a useful tool for the risk assessment of cyanotoxins in vivo due to their potential to qualitatively and quantitatively monitor substances dissolved in gastrointestinal fluids. The results presented in this study provide valuable information for a better understanding of the stability and bioavailability of GNT. Besides, the methods presented in this study can be useful for various applications designed to identify the toxin in environmental samples, as well as guidance on procedures for purifying GNT

Preparation of neurotoxin loaded dissolving microneedles and its transdermal penetration / 中草药

Objective: To prepare Naja atra neurotoxin (NT) loaded dissolving microneedles (DMNs-NT), and investigate the physicochemical properties and in vitro transdermal properties. Methods: DMNs-NT was prepared by a two-step centrifugation method. The ratio of CS and PVP K30, the water content of the matrix solution, and the backing layer material were optimized by the indexes of formability and mechanical strength of the microneedles and flexibility of the backing layer. The drug loading content was determined by HPLC, and the morphological characteristics were observed under an optical microscope, and the stability was also examined. Franz diffusion cell was used to investigate its in vitro skin permeation characteristics. Results: Through the single-factor exploration, we confirmed that the optimal prescription technique for DMNs-NT preparation was a 1:1 ratio of CS and PVP k30, a 5:4 ratio of matrix material and water, with CMC as the backing layer material. The DMNs-NT had a pyramidal shape with a smooth surface and a length of approximately 500 μm. The drug loading content of per tablet was (15.4 ± 0.5) μg. The drug was located in the upper part of the needle. DMNs-NT had good stability within 3 months. The results of in vitro skin permeation assay showed that the cumulative penetration of NT in DMNs-NT could reach 95.8% in 4 h, while NT solution barely penetrated the skin, which proved that it had a good promoting effect on NT transdermal delivery. Conclusion: In this study, DMNs-NT had good mechanical properties and good skin penetration, which realized the transdermal drug delivery of macromolecular drugs.

Efficacy and Safety of Letibotulinum Toxin A for the Treatment of Essential Blepharospasm

PURPOSE: To evaluate the efficacy and safety of BOTULAX® in subjects with essential blepharospasm.METHODS: In this study, a total of 250 subjects with essential blepharospasm were enrolled at 15 investigational sites and a total of 220 subjects completed the study. The efficacy and safety were evaluated at weeks 4 and 16 after treatment compared with baseline. In total, 240 subjects were enrolled, treated with the investigational product, and evaluable for the primary efficacy assessment at week 4 after treatment; these subjects were included in the intention-to-treat (ITT) population. With the ITT set as the main efficacy set, efficacy assessment included Jankovic rating scale (JRS), functional disability score, investigator evaluation of global response and quality of life. Safety assessment including the incidence of adverse events was also performed.RESULTS: In terms of the primary efficacy endpoint (i.e., change in JRS total score at week 4 after treatment from baseline [ITT set]), mean change indicated a statistically significant reduction (p < 0.0001) and demonstrated the non-inferiority of the test drug to similar drugs. In terms of the secondary efficacy endpoints, mean change in JRS total score at week 16 after treatment and mean change in functional disability score at weeks 4 and 16 after treatment both exhibited a statistically significant reduction compared with baseline (p < 0.0001 for all). Among the 249 subjects treated with the investigational product in this study, 44 (17.67%) experienced 76 treatment emergent adverse events but no serious adverse events were observed.CONCLUSIONS: Based on the study results, BOTULAX® is considered to be an effective and safe treatment for essential blepharospasm.

Epidemiologia e fatores de riscos relacionados à intoxicação alimentar causada por Clostridium Botulinum: uma revisão narrativa / Epidemiology and risk factors related to food intoxication caused by Clostridium Botulinum: a narrative review

O botulismo é uma doença resultante da ação de uma toxina produzida pelo Clostridium botulinum. Devido à sua gravidade e alta mortalidade é considerado um problema de saúde pública. Nesta revisão apresentamos os principais fatores de riscos associados à intoxicação alimentar provocada pelo Clostridium botulinum, bem como realizamos um levantamento epidemiológico sobre o botulismo alimentar e infantil. A busca bibliográfica considerou as bases de dados Scielo, Medline, Lilacs e PubMed. Foram selecionados artigos originais e relatos de caso publicados em inglês, espanhol e português, incluindo publicações dos últimos dez anos. A partir das análises dos títulos, resumos e artigos, um total de 26 artigos foram incluídos nesta revisão. Verificou-se predomínio de 54% dos casos referentes ao botulismo alimentar, dos quais aproximadamente 58% confirmaram a ocorrência da toxina tipo A; e 35% referente ao botulismo infantil. Na literatura consultada os principais sintomas, relacionados ao botulismo alimentar, identificados foram: visão turva, vômito, paralisia flácida, náuseas, tontura, diplopia, dificuldade respiratória, disatria, disfagia, fraqueza muscular, boca seca, ptose e cefaleia. Dentre as principais fontes de contaminação, 65% das publicações selecionadas identificaram as conservas como principal causa do botulismo alimentar. Embora o mel (42%) seja a única fonte registrada de alimento veiculador do agente causador do botulismo infantil, alguns relatos na literatura (25%) associaram à doença com a inalação de poeira contendo esporos do Clostridium botulinum, bem como o uso de plantas medicinais (25%). Os sintomas mais comuns observados na literatura foram: constipação dificuldade respiratória e dificuldade de sucção. Apesar de vários relatos na literatura acerca das duas doenças, o botulismo ainda é muito subnotificado dado ao diagnóstico muitas vezes equivocado, ressaltando-se a importância do diagnóstico precoce no tratamento da doença pelos profissionais de saúde, bem como a disponibilidade de informações relevantes para a investigação epidemiológica de doenças de notificação compulsória. Os dados apresentados também demonstram a importância de sensibilizar a população dos principais riscos e medidas de prevenção, já que a maioria dos casos relatados está relacionada a práticas inadequadas de preparo dos alimentos. (AU)

Botulism is a disease resulting from the action of a toxin produced by Clostridium botulinum. Because of its severity and high mortality, it is considered a public health problem. In this review, we present the main risk factors associated with food poisoning caused by Clostridium botulinum, as well as an epidemiological survey on foodborne and infant botulism. A bibliographic search was conducted in SciELO, MEDLINE, LILACS and PubMed databases. Original articles and case reports published in English, Spanish and Portuguese in the past ten years were selected. After analyzing titles, abstracts and articles, 26 articles were used in this review. In total, 54% of the cases were related to foodborne botulism, of which approximately 58% had confirmed type A botulism, and 35% were related to infant botulism. In the literature consulted, the main symptoms related to foodborne botulism were blurred vision, vomiting, flaccid paralysis, nausea, dizziness, diplopia, respiratory distress, dysarthria, dysphagia, muscle weakness, dry mouth, ptosis and headache. Among the sources of contamination, 65% of the published studies reported home-canned foods as the main cause of foodborne botulism. Although honey (42%) is the only reported food source for the agent causing infant botulism, some reports in the literature (25%) associated the disease with inhalation of dust containing Clostridium botulinum spores, as well as use of medicinal plants (25%). The most common symptoms observed in the literature were constipation, difficulty breathing and difficulty suckling. Although several reports on the two forms of the disease exist, botulism remains under-reported because of often incorrect diagnosis. Thus, early diagnosis is important for an adequate treatment provided by health professionals, as well as availability of relevant information for the epidemiological investigation of notifiable diseases. The data presented in this study also demonstrate the importance of raising people's awareness to main risks and prevention measures, as most reported cases were related to inadequate food preparation practices. (AU)

Serum Levels of Eosinophil-Derived Neurotoxin: A Biomarker for Asthma Severity in Adult Asthmatics

PURPOSE: Eosinophilic inflammation is a key component of severe asthma (SA). However, there has been no reliable serum biomarker for the eosinophilic inflammation of SA. We hypothesized that serum eosinophil-derived neurotoxin (EDN) could predict the eosinophilic inflammation of SA in adult asthmatics. METHODS: Severe asthmatics (n = 235), nonsevere asthmatics (n = 898), and healthy controls (n = 125) were enrolled from Ajou University Hospital, South Korea. The serum levels of EDN and periostin were measured by enzyme-linked immunosorbent assay and compared between severe and nonsevere asthmatics. Their associations with total eosinophil count (TEC) and clinical parameters were evaluated; clinical validation of the K-EDN kit for the measurement of serum EDN was evaluated. RESULTS: Severe asthmatics were older and had longer disease duration with significantly lower levels of forced expiratory volume in 1 second and methacholine PC20 than nonsevere asthmatics. Significant differences were found in TEC or sputum eosinophil count (%) between the groups. The serum levels of EDN and periostin were significantly higher in severe asthmatics than in nonsevere asthmatics and in healthy controls (all P < 0.05). Although significant correlations were found between serum EDN levels measured by the 2 kits (ρ = 0.545, P < 0.0001), higher correlation coefficients between serum EDN levels measured by the K-EDN kit and TEC were higher (ρ = 0.358, P < 0.0001) than those between serum EDN levels measured by the MBL kit and TEC (ρ = 0.319, P < 0.0001) or serum periostin level (ρ = 0.222, P < 0.0001). Multivariate regression analysis demonstrated that serum EDN levels measured by the K-EDN kit predicted the phenotype of SA (P = 0.003), while 2 other biomarkers did not. CONCLUSIONS: The serum EDN level may be a useful biomarker for assessing asthma severity in adult asthmatics.

Whole body vibration combined with botulinum neurotoxin A injection in the treatment of spastic diplegic cerebral palsy / 中华物理医学与康复杂志

Objective@#To observe the effect of combining whole body vibration with botulinum neurotoxin A injections on tiptoe and the gross motor function of children with spastic diplegic cerebral palsy.@*Methods@#Sixty spastic diplegic children with tipped foot aged between 2 to 5 were equally divided into a control group and an experimental group randomly. The control group received 3 IU/kg botulinum neurotoxin A injections to the medial and lateral heads of the gastrocnemius muscle. Then 5 daily courses of conventional training were administered 5 days a week for 3 weeks beginning 24 hours after the injections. The experimental group additionally received 2min of whole body vibration 3 or 4 times per day with one-minute rests, 5 days per week for 5 weeks. All of the children were assessed before the experiment and 1, 3 and 6 months later using the modified Tardieu scale (MTS) and the R1 and R2 ankle and dimensions D and E of the gross motor function measurement scale (GMFM-88).@*Results@#There were no significant differences between the two groups before the treatment. Afterward, the average MTS, R1, R2 and GMFM-88 scores of both groups were significantly improved. The average MTS, R1 and R2 scores of the experimental group after treatment were significantly better than the control group′s averages. The average GMFM-88 score of the experimental group was not significantly different from that of the control group after 1 month, but after 3 and 6 months significant differences emerged.@*Conclusion@#Whole body vibration improves the effectiveness of botulinum neurotoxin A injections in relieving tiptoe and improving the gross motor function of children with spastic diplegic cerebral palsy.

Whole body vibration combined with botulinum neurotoxin A injection in the treatment of spastic diplegic cerebral palsy / 中华物理医学与康复杂志

Objective To observe the effect of combining whole body vibration with botulinum neurotoxin A injections on tiptoe and the gross motor function of children with spastic diplegic cerebral palsy. Methods Sixty spastic diplegic children with tipped foot aged between 2 to 5 were equally divided into a control group and an ex-perimental group randomly. The control group received 3 IU/kg botulinum neurotoxin A injections to the medial and lateral heads of the gastrocnemius muscle. Then 5 daily courses of conventional training were administered 5 days a week for 3 weeks beginning 24 hours after the injections. The experimental group additionally received 2min of whole body vibration 3 or 4 times per day with one-minute rests, 5 days per week for 5 weeks. All of the children were assessed before the experiment and 1, 3 and 6 months later using the modified Tardieu scale ( MTS) and the R1 and R2 ankle and dimensions D and E of the gross motor function measurement scale ( GMFM-88) . Results There were no significant differences between the two groups before the treatment. Afterward, the average MTS, R1, R2 and GMFM-88 scores of both groups were significantly improved. The average MTS, R1 and R2 scores of the experimental group after treatment were significantly better than the control group' s averages. The average GMFM-88 score of the experimental group was not significantly different from that of the control group after 1 month, but after 3 and 6 months significant differences emerged. Conclusion Whole body vibration improves the effectiveness of botulinum neurotoxin A injections in relieving tiptoe and improving the gross motor function of chil-dren with spastic diplegic cerebral palsy.

Biomarkers for Recurrent Wheezing and Asthma in Preschool Children

Wheezing is one of the characteristic symptoms of asthma, but all preschool children with wheezing are not diagnosed with asthma. Preschool children are not cooperative enough to participate in spirometry and invasive tests. Thus, there is no conventional method to diagnose asthma in preschool children. We reviewed studies on non-invasive biomarkers for assessing asthma in preschool children. Specimens that can be easily obtained by non-invasive methods are blood, exhaled breath and urine. Eosinophils, eosinophil cationic protein and eosinophil-derived neurotoxin (EDN) in blood are helpful in evaluating eosinophilic inflammation of the airways. Exhaled breath contains nitric oxide, volatile organic compounds, various cytokines and mediators as analytical components. Fraction of exhaled nitric oxide has been used to assess the degree of eosinophil inflammation and has been standardized in school-age children and adults, but not yet in preschool children. Exhaled breath condensate (EBC) pH and various cytokines/mediators that are detected in EBC seem to be promising biomarkers for assessing asthma, but need more standardization and validation. There are several biomarkers useful for assessing asthma, but none are ideal. Some biomarkers need standardized methods of obtaining samples from uncooperative preschool children for clinical use and require sufficient validation. Recently, another activated eosinophil marker, serum EDN, has shown promising results as a biomarker for recurrent wheezing and asthma in preschool children.

The Effects of Intradetrusor BoNT-A Injections on Vesicoureteral Reflux in Children With Myelodysplasia / 대한배뇨장애요실금학회지

PURPOSE: We retrospectively evaluated the efficacy of botulinum neurotoxin A (BoNT-A) on vesicoureteral reflux (VUR), continence status, and urodynamic parameters in children with myelodysplasia who were not responsive to standard conservative therapy.METHODS: The study included 31 children (13 boys, 18 girls) with a mean age of 9.2±2.3 years (range, 5–14 years) with myelodysplasia, retrospectively. All children were fully compatible with clean intermittent catheterization (CIC) and did not respond to the maximum tolerable anticholinergic dose. All children received an intradetrusor injection of 10 U/kg (maximum, 300 U) of BoNT-A into an infection-free bladder. All patients had VUR (22 unilateral, 9 bilateral) preoperatively. The grade of reflux was mild (grades 1, 2), intermediate (grade 3), and severe (grades 4, 5) in 25, 7, and 8 ureters, respectively.RESULTS: The mean maximum bladder capacity increased from 152.9±76.9 mL to 243.7±103 mL (P<0.001), and the maximum detrusor pressure decreased from 57±29.4 cm H₂O to 29.6±13.9 cm H₂O (P<0.001). After BoNT-A treatment, 16 refluxing ureters (40%) completely resolved, 17 (42.5%) improved, 5 (12.5%) remained unchanged, and 2 (5%) became worse. Of the 31 children with urinary leakage between CICs, 22 (71%) became completely dry, 6 (19%) improved, and 3 (10%) experienced partial improvement.CONCLUSIONS: In children with myelodysplasia, we were able to increase bladder capacity, enhance continence, and prevent VUR by using intradetrusor BoNT-A injections. Although our results are promising, a larger group of long-term prospective studies are warranted to investigate this method of treatment.

Antidepressant-Like Action of Single Facial Injection of Botulinum Neurotoxin A is Associated with Augmented 5-HT Levels and BDNF/ERK/CREB Pathways in Mouse Brain / 神经科学通报·英文版

The present study was designed to examine the therapeutic effects of Botulinum neurotoxin A (BoNT/A) on depression-like behaviors in mice and to explore the potential mechanisms. These results revealed that a single facial injection of BoNT/A induced a rapid and prolonged improvement of depression-like behaviors in naïve and space-restriction-stressed (SRS) mice, reflected by a decreased duration of immobility in behavioral despair tests. BoNT/A significantly increased the 5-hydroxytryptamine (5-HT) levels in several brain regions, including the hippocampus and hypothalamus, in SRS mice. BoNT/A increased the expression of the N-methyl-D-aspartate receptor subunits NR1 and NR2B in the hippocampus, which were significantly decreased in SRS mice. Furthermore, BoNT/A significantly increased the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus, hypothalamus, prefrontal cortex, and amygdala, which were decreased in SRS mice. Finally, BoNT/A transiently increased the levels of phosphorylated extracellular signal-regulated kinase (p-ERK) and cAMP-response element binding protein (p-CREB), which were suppressed in the hippocampus of SRS mice. Collectively, these results demonstrated that BoNT/A treatment has anti-depressant-like activity in mice, and this is associated with increased 5-HT levels and the activation of BDNF/ERK/CREB pathways in the hippocampus, supporting further investigation of BoNT/A therapy in depression.

Preparation of angiopep-2-modified FITC-labeled neurotoxin nanoparticles and in vitro release characteristics / 中国中药杂志

In order to prepare angiopep-2 modified fluorescein isothiocyanate-labeled neurotoxin nanoparticles( ANG-NPs/FITCNT),emulsion/solvent evaporation method was used with m PEG-PLA and ANG-PEG-PLA( in proper proportions) as carriers and with FITC-NT as drug. With particle size and encapsulation efficiency as comprehensive indexes,the effects of different ultrasound power and ultrasound time combinations on the process were investigated. The in vitro release characteristics of nanoparticles in PBS buffer at p H 7. 4 and p H 6. 5 were investigated by dialysis method. The results indicated that the optimum process for preparing ANG-NPs/FITC-NT was as follows: ultrasonic power 90 W,ultrasonic time 30 s. In such optimal process,ANG-NPs/FITC-NT were well-shaped under the transmission electron microscope,with an average particle size of( 123. 9±0. 5) nm,Zeta potential of(-10. 5±0. 5) m V,encapsulation efficiency of( 68. 1±0. 4) %,and the drug loading of( 0. 82±0. 01) %. The in vitro drug release profiles of the nanoparticles in PBS buffer at p H 7. 4 and p H 6. 5 were both consistent with Ritger-Peppas equation,ln Q = 0. 508 8 lnt-2. 285 0,r = 0. 961 5( p H 7. 4) and ln Q= 0. 449 9 lnt-1. 855 3,r = 0. 970 3( p H 6. 5),respectively. The experiment results proved that the nanoparticles prepared by emulsion/solvent evaporation method had uniform particle size,high encapsulation efficiency and in vitro sustained release characteristic,which might be a potential carrier for NT intracerebral drug delivery.

Long-term Efficacy of Botulinum Neurotoxin-A Treatment for Essential Blepharospasm

PURPOSE: In the present study, we investigated the treatment efficacy and clinical outcomes of botulinum neurotoxin-A (BoNT-A) administered for longer than 5 years to patients with essential blepharospasm. METHODS: We retrospectively reviewed 19 patients (male : female = 8 : 11) diagnosed with essential blepharospasm between March 2006 and July 2016 who underwent BoNT-A injections for over 5 years and were followed. Efficacy of 297 injections of Botox (n = 162), Meditoxin (n = 75), Hugel-tox (n = 40), or Dysport (n = 20) was based on the symptom improvement score at the final injection (−1, worse; 0, same; 1, better). Injection dose (botox unit), duration of efficacy (months), and adverse events were also investigated. RESULTS: Based on product type, significant differences in patient age (59.3 ± 9.8 years), disease period (5.0 ± 5.4 years), number of botulinum neurotoxin injections before visiting our clinic (1.6 ± 2.6), and follow-up period (7.2 ± 1.6 years) were not observed. Treatment efficacy score and injection dose of repetitive injections were 0.1 ± 0.5 and 39.1 ± 4.0 units, respectively, and did not show significant differences with repeated injections. Duration of response was 5.9 ± 5.4 months, but this significantly decreased as the injections were repeated (p < 0.01). Among the 297 injections, adverse events occurred 12 times (4.0%) with no severe sequelae. CONCLUSIONS: In this study, we showed that repetitive, long-term BoNT-A injections are considered a stable and effective treatment for essential blepharospasm in terms of consistent injection dose and maintenance of treatment efficacy. However, the duration of long-term efficacy could be decreased in patients injected repetitively.

Evaluation of activity of botulinum neurotoxin A based on primary cerebellar granu-losa cells / 军事医学

Objective To isolate and culture the primary cerebellar granulosa cells(CGNs)of SD rats and evaluate the activity of botulinum neurotoxin A(BoNT/A)based on CGNs.Methods CGNs of 6-to 8-days-old SD rats were isolated and cultured.After 5-7 d,the cells were treated with BoNT/A.The activity of the toxin was evaluated with immunofluo-rescence,and the relationships between the activity and dose of the toxin were analyzed with Western blotting.Results and Conclusion CGNs Of SD rats were successfully cultured,a method for evaluating the activity of BoNT/A was established at the level of primary nerve cells,and the relationships between the activity and dose of toxin were analyzed.This study provides a tool for further detailing the biochemical mechanism of BoNT /A.

Progresses in the serum eosinophil-derived neurotoxin in pediatric asthma / 国际儿科学杂志

Asthma is the most common chronic airway disease in children.The main manifestations are recurrent cough,wheezing,and dyspnea.In recent years,the morbidity and mortality rate of childhood asthma shows a clear rising trend.However,the exact mechanism of this disease still remains uncertain.There is still no specific laboratory test to diagnose pediatric asthma,especially for children under five years who cannot finish pulmonary function test.Serum eosinophil-derived neurotoxin (EDN),as a degranulation of eosinophil,the most important participants in childhood asthma,is drawing an increasing attention of investigators in the assistance of diagnosis,classification and prognosis of childhood asthma.This review concentrates on the relationship between EDN and pediatric asthma.

Evaluation of the effect of botulinum neurotoxin type A (BoNT/A) on daily activity performance in chronic migraine patients using VAS, MIDAS AND HIT-6 tests

@#Background & Objectives: According to ICHD-III beta 2013 criteria, chronic migraine is defined as having headaches more than 15 times a month, for a period of more than 3 months, at least 8 must have migrainous features or good response to migraine-specific treatment; there must also be a history of 5 or more migraine attacks. The aim of the present study was to evaluate the effect of Botulinum Neurotoxin A (BONT/A) on headache and daily activities in chronic migraine patients using VAS, MIDAS and HIT-6 tests. Methods: Twenty five patients admitted to Hospital Department of Neurology were reviewed retrospectively. In order to evaluate the severity of headache and effects on daily performance, MIDAS (Migraine Disability Assessment Test), VAS (Visual Analogue Scale for Pain) and HIT-6 results after the baseline assessment, first and second administration of (BONT/A) were examined retrospectively from patients’ records. Results: VAS, MIDAS and HIT-6 scores were compared after baseline assessment and the first and second administrations. Results showed that VAS, MIDAS and HIT-6 scores decreased. This difference was statistically significant (p<0.05). Correlation analysis was conducted and significant correlations between scores on these three tests were found.Conclusions: The results showed that BoNT/A is an important and effective treatment option for chronic migraine patients not responding to migraine-specific prophylactic treatment and having alterations in daily life due to frequency and severity of pain.