Block of Nav1.8 by Small Molecules

Sodium channels are key proteins in regulating neuronal excitability and accumulating data suggest that specific subtypes of voltage-dependent sodium channels are important in signaling various types of pain. Consistent with this theme, Jarvis et al (2007) recently reported the identification of a subtype-selective Nav1.8 blocker that was active in several pre-clinical models of pain. During the course of these studies compounds were also identified that showed large differences in potency when tested on Nav1.8 channels from different species. This Addendum illustrates one of these compounds along with the potency correlation between recombinant and native tetrodotoxin-resistant sodium channels for additional examples. These data show that significant differences can be observed for sodium channel blockers across species and highlight the importance of considering this possibility when searching for new compounds and research tools to probe sodium channel function.     

小分子化合物体外诱导肝上皮样干细胞-WB细胞表达PDX1

目的：近年来干细胞治疗糖尿病一直是国内外研究人员关注的焦点,而肝细胞向胰岛样细胞转变也是热点之一。本实验应用小分子化合物在体外诱导WB-F344大鼠来源肝上皮样干细胞（简写WB细胞）表达胰腺内分泌前体细胞基因PDX1,建立一种体外诱导WB细胞分化为胰腺内分泌前体细胞的实验方法。方法：选用5-AZA TSA,RA,ITS等小分子化合物,分两步法直接诱导WB细胞分化为表达PDX1的胰腺内分泌前体细胞,用含有不同浓度5-AZA分化培养基诱导WB分化,摸索诱导分化的最佳条件。观察细胞形态变化,RT-PCR及实时定量PCR检测部分基因表达情况,免疫荧光检测PDX1的表达。结果：5AZA 5 uM处理2 d,TSA 1 d,RA联合ITS诱导7天,诱导的WB细胞表达PDX1较1-4 uM 5-AZA诱导强,并表达胰腺内分泌前体细胞的相关基因,NGN3,Neurod,NKX2.2,WB表达的Nestin仍持续表达,Insulin1有少量表达。WB表达的肝干细胞基因如ALB,AFP大量下调,标志分化的基因C/EBP下调。结论：5-AZA,TSA,RA,ITS等小分子化合物能够诱导肝上皮样细胞WB表达PDX1,并且这种诱导分化的细胞具有胰腺内分泌前体细胞特征。本实验进一步证明在体外微环境中,肝干细胞能向胰腺内分泌细胞转化,而肝细胞增极强,为将来干细胞治疗糖尿病提供充足的细胞来源     

拟南芥六个新的small RNAs的克隆和功能预测

以模式植物拟南芥为例,建立了一种克隆small RNA分子的技术平台,为今后开展small RNA分子的生物学功能研究提供技术支撑.通过用抗病信号分子水杨酸(SA)处理拟南芥叶片后,进行small RNA分子群体的分离与接头连接、PCR扩增、T-载体克隆与检测、测序分析和生物信息学分析等一系列实验,成功地克隆了一些small RNAs.并对其表达和功能进行了分析.     

Induced folding in RNA recognition by Arabidopsis thaliana DCL1

DCL1 is the ribonuclease that carries out miRNA biogenesis in plants. The enzyme has two tandem double stranded RNA binding domains (dsRBDs) in its C-terminus. Here we show that the first of these domains binds precursor RNA fragments when isolated and cooperates with the second domain in the recognition of substrate RNA. Remarkably, despite showing RNA binding activity, this domain is intrinsically disordered. We found that it acquires a folded conformation when bound to its substrate, being the first report of a complete dsRBD folding upon binding. The free unfolded form shows tendency to adopt folded conformations, and goes through an unfolded bound state prior to the folding event. The significance of these results is discussed by comparison with the behavior of other dsRBDs.     

靶向小分子创新药物

寻找药物新靶点是全球创新药物研究激烈竞争的焦点."组学"、生物信息学、系统生物学、药物筛选现代检测技术等新理论、新技术的发展使新的筛选模型和评价技术不断取得突破.靶向抗肿瘤药物的开发是靶向小分子创新药物的重点任务,多靶点的抗肿瘤药物开发及新靶点的发现是抗肿瘤药物研发的新趋势.     

Two Small Molecules Block Oral Epithelial Cell Invasion by Porphyromons gingivalis

Porphyromonas gingivalis is a keystone pathogen of periodontitis. One of its bacterial characteristics is the ability to invade various host cells, including nonphagocytic epithelial cells and fibroblasts, which is known to facilitate P. gingivalis adaptation and survival in the gingival environment. In this study, we investigated two small compounds, Alop1 and dynasore, for their role in inhibition of P. gingivalis invasion. Using confocal microscopy, we showed that these two compounds significantly reduced invasion of P. gingivalis and its outer membrane vesicles into human oral keratinocytes in a dose-dependent manner. The inhibitory effects of dynasore, a dynamin inhibitor, on the bacterial entry is consistent with the notion that P. gingivalis invasion is mediated by a clathrin-mediated endocytic machinery. We also observed that microtubule arrangement, but not actin, was altered in the host cells treated with Alop1 or dynasore, suggesting an involvement of microtubule in this inhibitory activity. This work provides an opportunity to develop compounds against P. gingivalis infection.     

Redistribution and Pinocytosis of Lymphocyte Surface Immunoglobulin Molecules Induced by Anti-Immunoglobulin Antibody

Antibody reacting with lymphocyte surface immunoglobulin molecules induces these to gather over one pole of the cell. This suggests a possible mechanism for lymphocyte triggering by antigen and raises questions about cell membrane structure.     

Effects of Small Molecules on Chaperone-Mediated Autophagy

Autophagy, including macroautophagy (MA), chaperone-mediated autophagy (CMA), crinophagy, pexophagy and microautophagy, are processes by which cells select internal components such as proteins, secretory vesicles, organelles, or foreign bodies, and deliver them to lysosomes for degradation. MA and CMA are activated during conditions of serum withdrawal in cell culture and during short-term (MA) and prolonged (CMA) starvation in organisms. Although MA and CMA are activated under similar conditions, they are regulated by different mechanisms. We used pulse/chase analysis under conditions in which most intracellular proteolysis is due to CMA to test a variety of compounds for effects on CMA. We show that inhibitors of MA such as 3-methyladenine, wortmannin, and LY294002 have no effect on CMA. Protein degradation by MA is sensitive to microtubule inhibitors such as colcemide and vinblastine, but protein degradation by CMA is not. Activators of MA such as rapamycin also have no effect on CMA. We demonstrate that CMA, like MA, is inhibited by protein synthesis inhibitors anisomycin and cycloheximide. CMA is also partially inhibited when the P38 mitogen activated protein kinase is blocked. Finally we demonstrate that the glucose-6-phophate dehydrogenase inhibitor, 6-aminonicotinamide, and heat shock protein of 90 kilodaltons inhibitor, geldanamycin, have the ability to activate CMA.     

Chapter 3: Small Molecules and Disease

“Big” molecules such as proteins and genes still continue to capture the imagination of most biologists, biochemists and bioinformaticians. “Small” molecules, on the other hand, are the molecules that most biologists, biochemists and bioinformaticians prefer to ignore. However, it is becoming increasingly apparent that small molecules such as amino acids, lipids and sugars play a far more important role in all aspects of disease etiology and disease treatment than we realized. This particular chapter focuses on an emerging field of bioinformatics called “chemical bioinformatics” – a discipline that has evolved to help address the blended chemical and molecular biological needs of toxicogenomics, pharmacogenomics, metabolomics and systems biology. In the following pages we will cover several topics related to chemical bioinformatics. First, a brief overview of some of the most important or useful chemical bioinformatic resources will be given. Second, a more detailed overview will be given on those particular resources that allow researchers to connect small molecules to diseases. This section will focus on describing a number of recently developed databases or knowledgebases that explicitly relate small molecules – either as the treatment, symptom or cause – to disease. Finally a short discussion will be provided on newly emerging software tools that exploit these databases as a means to discover new biomarkers or even new treatments for disease.

What to Learn in This Chapter

• The meaning of chemical bioinformatics
• Strengths and limitations of existing chemical bioinformatic databases
• Using databases to learn about the cause and treatment of diseases
• The Small Molecule Pathway Database (SMPDB)
• The Human Metabolome Database (HMDB)
• DrugBank
• The Toxin and Toxin-Target Database (T3DB)
• PolySearch and Metabolite Set Enrichment Analysis

This article is part of the “Translational Bioinformatics” collection for PLOS Computational Biology.

     

Generation of Novel Long-Acting Globular Adiponectin Molecules

Adiponectin is an adipocyte-derived hormone that has been shown to play important roles in the regulation of glucose and energy homeostasis. It exists as homotrimers or complexes containing multiple homotrimer units in plasma. The recombinant adiponectin proteins have been difficult to produce, making it challenging for both research as well as potential therapeutic development. Here, we show a novel approach for the generation of globular adiponectin that involves linking three monomer sequences together in tandem to generate one continuous polypeptide, which we have termed single-chain globular adiponectin (sc-gAd). To improve the pharmacokinetic properties of sc-gAd further, we fused it to an Fc fragment. The combined effects of single-chain and Fc fusion improved the plasma half-life from less than 2 h to close to 2 weeks. Using adeno-associated virus as a delivery method, we demonstrate that Fc-sc-gAd improved both fasting glucose levels and the tolerance to a glucose challenge in ob/ob mice without changes in body weight. Therefore, our results demonstrated the feasibility of generating globular adiponectin trimers from a single polypeptide and a long-acting globular adiponectin that could serve as a starting point for adiponectin-based therapeutics. This novel approach could also be applied to other complement factor C1q family members; in particular, this opens the possibility to study the biological functions of precisely defined heterotrimers of various family members that had not been previously possible.     

Discovery of Novel Potent Small Natural Molecules Able to Enhance Attenuation of the Pathobiology of Gastric Cancer-Associated Helicobacter pylori by Molecular Modeling

CagA is an oncoprotein that plays a significant role in the advancement of gastric cancer by interacting with phosphatidylserine of host cell plasma membrane. This protein is a potential target for the cancer therapy. In the present study, the initial and dynamic binding interaction mechanisms between CagA and herbal compounds were investigated by molecular docking of the 38 compounds followed by an analysis combining molecular dynamics simulations and molecular mechanics–Poisson Boltzmann surface area binding free energy calculations of the top four hit compounds; spinasterol, luteoxanthin, 3′-prenylrubranine and neoxanthin. Energetic analyses disclosed binding free energies pointing out that neoxanthin and luteoxanthin were potential candidates as lead compounds in drug development towards the CagA–phosphatidylserine inhibitors. Ultimately, binding free energy decomposition gave detailed information regarding the essential amino acid residues stabilizing the complexes.

     

不同视觉经验中zif268基因的表达模式

zif268基因编码一转录因子ZIF268. 在发育的视皮层中,zif268基因的表达模式受发育的调节. 在具有正常视觉经验的视皮层中,zif268基因有较高水平的表达. 视觉废置后,视皮层内该基因的表达水平显著降低. 通过视觉刺激可显著增强该基因的表达. 有关zif268基因表达模式的研究对于阐明该基因在哺乳动物视觉系统中的生理功能起到借鉴的作用.     

Excitability Changes in Intracortical Neural Circuits Induced by Differentially Controlled Walking Patterns

Our previous single-pulse transcranial magnetic stimulation (TMS) study revealed that excitability in the motor cortex can be altered by conscious control of walking relative to less conscious normal walking. However, substantial elements and underlying mechanisms for inducing walking-related cortical plasticity are still unknown. Hence, in this study we aimed to examine the characteristics of electromyographic (EMG) recordings obtained during different walking conditions, namely, symmetrical walking (SW), asymmetrical walking 1 (AW1), and asymmetrical walking 2 (AW2), with left to right stance duration ratios of 1:1, 1:2, and 2:1, respectively. Furthermore, we investigated the influence of three types of walking control on subsequent changes in the intracortical neural circuits. Prior to each type of 7-min walking task, EMG analyses of the left tibialis anterior (TA) and soleus (SOL) muscles during walking were performed following approximately 3 min of preparative walking. Paired-pulse TMS was used to measure short-interval intracortical inhibition (SICI) and intracortical facilitation (ICF) in the left TA and SOL at baseline, immediately after the 7-min walking task, and 30 min post-task. EMG activity in the TA was significantly increased during AW1 and AW2 compared to during SW, whereas a significant difference in EMG activity of the SOL was observed only between AW1 and AW2. As for intracortical excitability, there was a significant alteration in SICI in the TA between SW and AW1, but not between SW and AW2. For the same amount of walking exercise, we found that the different methods used to control walking patterns induced different excitability changes in SICI. Our research shows that activation patterns associated with controlled leg muscles can alter post-exercise excitability in intracortical circuits. Therefore, how leg muscles are activated in a clinical setting could influence the outcome of walking in patients with stroke.