Redox active or thiol reactive? Optimization of rapid screens to identify less evident nuisance compounds

Compounds that exhibit assay interference or undesirable mechanisms of bioactivity are routinely encountered in assays at various stages of drug discovery. We observed that assays for the investigation of thiol-reactive and redox-active compounds have not been collected in a comprehensive review. Here, we review these assays and subject them to experimental optimization to improve their reliability. We demonstrate the usefulness of our assay cascade by assaying a library of bioactive compounds, chemical probes, and a set of approved drugs. These high-throughput assays should complement the array of wet-lab and in silico assays during the initial stages of hit discovery campaigns to pursue only hit compounds with tractable mechanisms of action.     

中药复方肠复康抗结直肠癌血管生成作用靶基因网络药理学研究

目的运用网络药理学方法,研究中药复方肠复康(CFK)对结直肠癌(colorectal cancer,CRC)血管生成作用靶基因,为揭示其作用机制奠定理论基础。方法采用TTD、DrugBank数据库、OMIM数据库、GAD和PharmGKB等5个数据库分别检索CRC基因;采用TSMSP数据库及基于VBA工具的有效成分筛选方式检索CFK所含的5味中药,利用ADME参数进行有效成分筛选,通过TCMSP数据库检索各个有效成分的靶基因,利用cytoscape 3.2.1软件及其插件ClueGO、Bisogenet、CytoNCA对靶基因进行分析并构建CFK成分靶点网络图,并结合KEGG数据库进一步说明CFK与CRC血管生成靶基因的关系。结果 CRC靶基因339个,CFK靶基因182个;通过cytoscape构建并结合网络拓扑分析,ADRA1A、ADRA1B、ADRA1D、ADRB1、CHRM1、CHRM2、CHRM3、CHRM4、INSR、PIK3CG、RXRA、BAX、BCL2、CASP8、ICAM1、NFKBIA、CASP9、KDR、MAP2、PRKCA、PTGS2等靶点与血管生成相关;CFK治疗CRC过程中BRCA1、CDK2、CDKN1A、ITGA4、MDM2、YWHAG、CREBBP、CUL2、EP300、VHL、FLNA、SHC1、TRAF6、XPO1、EGFR、GRB2、IKBKG、NTRK1、TP53、 YWHAB、YWHAE与血管生成相关;主要通过调控PI3K-Akt、MAPK、HIF-1及VEGF信号通路抑制CRC患者新生血管生成。结论 CFK可能会通过VBRCA1、CDK2、CDKN1A、ITGA4、MDM2、YWHAG、CREBBP、CUL2、EP300、VHL、FLNA、SHC1、TRAF6、XPO1、EGFR、GRB2、IKBKG、NTRK1、TP53、 YWHAB、YWHAE等靶点调控PI3K-Akt、MAPK、HIF-1及VEGF等信号通路,进而发挥抑制CRC患者新生血管生成作用,具体机制有待进一步研究。     

祛寒逐风合剂联合西医常规疗法治疗膝骨关节炎风寒痹阻证临床研究

目的观察祛寒逐风合剂联合西医常规疗法对膝骨关节炎风寒痹阻证的临床疗效,以及对关节症状及相关实验室指标的影响。方法采用随机数字表法将94例膝骨关节炎风寒痹阻证患者分为观察组和对照组各47例。对照组采用常规药物疗法+康复训练,观察组在对照组基础上予祛寒逐风合剂,每次50m L,每日3次,口服。2组均连续治疗2周。比较2组临床疗效,观察2组治疗前后美国西大略湖和麦克马斯特大学骨关节炎指数(WOMAC)评分、视觉模拟评分法(VAS)评分、压痛指数、中医症状评分,白细胞介素(IL)-1β、IL-6、血管内皮生长因子(VEGF)、肿瘤坏死因子(TNF)-α、骨钙素、抗酒石酸盐酸性磷酸酶异构体(TRACP)-5b、骨特异性碱性磷酸酶(BALP)、纤维蛋白原、红细胞沉降率和红细胞聚集指数,对2组进行安全性评价。结果观察组总有效率为89.36%(42/47),对照组为74.47%(35/47),观察组明显优于对照组(P<0.05)。与本组治疗前比较,2组治疗后WOMAC评分、VAS评分、压痛指数、中医症状评分明显下降(P<0.05);2组治疗后比较,观察组WOMAC评分、VAS评分、压痛指数、中医症状评分明显低于对照组(P<0.05)。与本组治疗前比较,2组治疗后IL-1β、IL-6、TNF-α、VEGF、TRACP-5b水平明显下降,骨钙素、BALP水平明显升高(P<0.05);2组治疗后比较,观察组IL-1β、IL-6、TNF-α、VEGF、TRACP-5b水平明显低于对照组,骨钙素、BALP水平明显高于对照组(P<0.05)。与本组治疗前比较,观察组治疗后纤维蛋白原、红细胞沉降率、红细胞聚集指数明显下降(P<0.05);2组治疗后比较,观察组血液流变学各项指标明显低于对照组(P<0.05)。2组均未发生不良反应。结论祛寒逐风合剂联合西医常规疗法治疗膝骨关节炎风寒痹阻证患者疗效较好,可明显改善关节症状及相关实验室指标。     

Focal hole versus screw stimulation to prevent false negative results in detecting pedicle breaches during spinal instrumentation

ObjectiveWe describe a stimulus-evoked EMG approach to minimize false negative results in detecting pedicle breaches during lumbosacral spinal instrumentation.MethodsIn 36 patients receiving 176 lumbosacral pedicle screws, EMG threshold to nerve root activation was determined using a focal probe inserted into the pilot hole at a depth, customized to the individual patients, suitable to position the stimulating tip at the point closest to the tested nerve root. Threshold to screw stimulation was also determined.ResultsMean EMG thresholds in 161 correctly fashioned pedicle instrumentations were 7.5 mA ± 2.46 after focal hole stimulation and 21.8 mA ± 6.8 after screw stimulation. Direct comparison between both thresholds in individual pedicles showed that screw stimulation was always biased by an unpredictable leakage of the stimulating current ranging from 10 to 90%. False negative results were never observed with hole stimulation but this was not true with screw stimulation.ConclusionsFocal hole stimulation, unlike screw stimulation, approaches absolute EMG threshold as shown by the lower normal limit (2.6 mA; p < 0.05) that borders the upper limit of threshold to direct activation of the exposed root.SignificanceThe technique provides an early warning of a possible pedicle breakthrough before insertion of the more harmful, larger and threaded screw.     

Cholinergic hyperactivity in patients with myasthenia gravis with MuSK antibodies: A neurophysiological study

ObjectivePatients with myasthenia gravis associated with muscle-specific tyrosine kinase antibodies (MuSK-MG) often manifest signs of cholinergic hyperactivity with standard doses of acetylcholinesterase inhibitors (AChE-Is). Aim of the study was to investigate whether repetitive compound muscle action potential (R-CMAP), the neurophysiological correlate of cholinergic hyperactivity, was present in MuSK-MG irrespective of AChE-I treatment.MethodsPatients with confirmed diagnosis of MuSK-MG were consecutively enrolled during follow-up visits, from January 2019 to April 2020. All these subjects underwent the same neurophysiological protocol, including motor nerve conduction studies and repetitive nerve stimulation. In patients taking pyridostigmine, neurophysiological testing was performed at least 12 hours after the last dose. For comparison, the presence of R-CMAP was investigated in 20 consecutive acetylcholine receptor antibody positive myasthenia gravis (AChR-MG) patients.ResultsWe enrolled 25 MuSK-MG patients (20 females), aged 16–79 years at the study time, with disease duration ranging 0.6–48.8 years (median: 17.7 years). R-CMAP was detected in 12/25 (48%) MuSK-MG cases and in none of the AChR-MG controls (p = 0.0003). In the MuSK-MG population, a history of muscle cramps and fasciculations, during low-dose pyridostigmine therapy, was significantly more frequent in R-CMAP positive than in R-CMAP negative patients (100% vs 31%, p = 0.001). At the time of the study, the proportion of patients still symptomatic for MG was higher among R-CMAP positive cases (92% vs 23%, p = 0.0005).ConclusionsCholinergic hyperactivity is a relatively common finding in MuSK-MG patients, independent of AChE-I treatment, and may constitute an intrinsic feature of the disease.SignificanceR-CMAP detection can represent a useful diagnostic clue for MuSK-MG and predicts poor tolerance to AChE-Is.     

Microhaplotypes in forensic genetics

Microhaplotype loci (microhaps, MHs) are a novel type of molecular marker of less than 300 nucleotides, defined by two or more closely linked SNPs associated in multiple allelic combinations. The value of these markers is enhanced by massively parallel sequencing (MPS), which allows the sequencing of both parental haplotypes at each of the many multiplexed loci. This review describes the features of these multi-SNP markers and documents their value in forensic genetics, focusing on individualization, biogeographic ancestry inference, and mixture deconvolution. Foreseeable applications also include missing person identification, relationship testing, and medical diagnostic applications. The technique is not restricted to humans.     

表没食子儿茶素没食子酸酯通过下调Toll样受体4对高盐诱导大鼠高血压的降压作用

目的研究表没食子儿茶素没食子酸酯(EGCG)对高盐引起的高血压模型大鼠的降压作用及作用机制。方法大鼠随机分为对照组、模型组和EGCG 50、100 mg/kg组,测定大鼠的大鼠平均动脉压,并采用Western blotting印迹法检测大鼠下丘脑室旁核(PVN)中Toll样受体4(TLR4)、核因子κB(NF-κB)p65、白细胞介素-1β(IL-1β)的水平;采用酶联免疫吸附试验(ELISA)法检测大鼠PVN中单核细胞趋化蛋白1(MCP-1)、肿瘤坏死因子α(TNF-α)和IL-6水平。结果高盐诱导的高血压模型大鼠中TLR4表达量明显增加(P<0.05)。EGCG 50、100 mg/kg显著降低大鼠的平均动脉压(P<0.05),并明显降低PVN中TLR4、NF-κBp65、IL-1β和TNF-α、IL-6、IL-1β水平(P<0.05)。结论 EGCG可降低高盐诱导大鼠的高血压,其机制可能与阻断TLR4信号有关。     

Early axonal loss predicts long-term disability in chronic inflammatory demyelinating polyneuropathy

ObjectiveTo investigate early pre-treatment nerve fiber loss as a predictor of long-term clinical outcome in chronic inflammatory demyelinating polyneuropathy (CIDP).MethodsIn 14 patients, motor and sensory conduction studies of the median, fibular, and sural nerves were performed at pre-treatment and follow-up 11–28 years later. Z-scores of amplitudes were combined as biomarkers of axonal loss and Z-scores of conduction properties as demyelination scores. The axonal loss was further examined by electromyography (EMG) and motor unit number estimation. Axonal and demyelination scores were compared to clinical outcomes in the Inflammatory Rasch-built Overall Disability Scale, the Neuropathy Impairment Score, and dynamometry.ResultsAt follow-up 12 patients walked independently, one needed support and one could not walk. The initial and follow-up axonal and demyelination scores were markedly abnormal. The initial axonal loss but not demyelination was strongly associated with both the follow-up axonal loss and the clinical measures. Moreover, delay of treatment initiation negatively influenced the axonal scores and clinical outcomes.ConclusionIn this hypothesis generating limited study, we found that axonal loss at early CIDP was highly predictive for long-term nerve fiber loss and disability.SignificanceThe study indicates that prompt initiation of treatment to prevent nerve fiber loss is necessary for outcome in CIDP.     

BOPPPS教学模式在医科院校医用化学教学中的应用与思考

医用化学是医学院校一门重要的基础必修课程,内容多为知识理解型,这导致了学生兴趣薄弱,学习效果欠佳。因此,需要通过改革传统"灌输式"课堂授课方式,让学生真正参与到教学活动中。BOPPPS教学模式分为导言、学习目标、前测、参与式学习、后测和总结六个环节。突出"以学生为中心"的教学理念,遵循教学活动的客观规律。在医用化学课程的教学过程中引入BOPPPS教学,能够有效提高学生的课堂参与度,增强学生的学习动力,培养学生独立思考、自主学习的能力。