Comparative analysis reveals selective recognition of glycans by the dendritic cell receptors DC-SIGN and Langerin

DC-SIGN (dendritic cell-specific ICAM-3 grabbing non-integrin) and Langerin are homologous C-type lectins expressed as cell-surface receptors on different populations of dendritic cells (DCs). DC-SIGN interacts with glycan structures on HIV-1, facilitating virus survival, transmission and infection, whereas Langerin, which is characteristic of Langerhans cells (LCs), promotes HIV-1 uptake and degradation. Here we describe a comprehensive comparison of the glycan specificities of both proteins by probing a synthetic carbohydrate microarray comprising 275 sugar compounds using the bacterially produced and fluorescence-labeled, monomeric carbohydrate-recognition domains (CRDs) of DC-SIGN and Langerin. In this side-by-side study DC-SIGN was found to preferentially bind internal mannose residues of high-mannose-type saccharides and the fucose-containing blood-type antigens H, A, B, Le(a), Le(b) Le(x), Le(y), sialyl-Le(a) as well as sulfatated derivatives of Le(a) and Le(x). In contrast, Langerin appeared to recognize a different spectrum of compounds, especially those containing terminal mannose, terminal N-acetylglucosamine and 6-sulfogalactose residues, but also the blood-type antigens H, A and B. Of the Lewis antigens, only Le(b), Le(y), sialyl-Le(a) and the sialyl-Le(x) derivative with 6'-sulfatation at the galactose (sialyl-6SGal Le(x)) were weakly bound by Langerin. Notably, Ca(2+)-independent glycan-binding activity of Langerin could not be detected either by probing the glycan array or by isothermal titration calorimetry of the CRD with mannose and mannobiose. The precise knowledge of carbohydrate specificity of DC-SIGN and Langerin receptors resulting from our study may aid the future design of microbicides that specifically affect the DC-SIGN/HIV-1 interaction while not compromising the protective function of Langerin.     

柯里拉京对B16细胞增殖及酪氨酸酶活性的抑制作用

采用体外培养B16细胞,在柯里拉京作用不同时间后,用MTT(四甲基偶氮唑蓝)法检测柯里拉京对B16细胞生长的影响;用多巴氧化法检测柯里拉京对B16细胞中酪氨酸酶活性的抑制作用;用形态学和Hoechst33258染色法观察柯里拉京对B16细胞生长形态的影响。结果表明,柯里拉京对B16细胞增殖的抑制作用明显,其作用12,24和48 h的IC_(50)分别是35.63,25.78和20.94 mg/L;对作用于B16细胞中酪氨酸酶12,24和48 h时的IC_(50)分别是16.38,14.00和13.85 mg/L;形态学观察表明,当ρ(柯里拉京)16 mg/L作用24 h时,B16细胞损伤明显;Hoechst33258染色显示,当ρ(柯里拉京)≤16 mg/L作用24 h时,B16细胞呈现凋亡形态。     

Sugar-Induced Chiral Orientation of Boronic-Acid-Appended Amphiphile Clusters Formed in the Dipalmitoylphosphatidylcholine (DPPC) Membrane

A boronic-acid-appended amphiphile bearing an azobenzene chromophore in the chain center ( 3 ) was synthesized. Although 3 could not form the membrane-like, ordered aggregate by itself, it formed a phase-separated aggregate in a dipalmitoylphosphatidylcholine (DPPC) matrix membrane. When saccharides were added, the boronic acid group reversibly formed the saccharide complexes, and 3 in the DPPC matrix membrane became CD-active with the appearance of exciton-coupling bands. Comparison of the saccharide absolute configuration with the CD intensity established that the saccharide possessing the OH group (as 3-OH, 4-OH, and 5-CH₂OH) in the same side as the cis-1,2-diol gives the strong CD band. Judging from the structure of 3 –saccharide complexes, these “same-side” OH groups can enjoy intermolecular hydrogen-bonding interactions, which eventually induce the chiral orientation of azobenzene chromophores. This is a novel membrane system useful to read out the information stored in the saccharide structure and to create novel membrane structures by added saccharides.     

Effect of cell density on cytotoxicity of ether lipid analogues in variants of B16 murine melanoma

Ether lipids are defined here as analogues of naturally occurring lysophosphatidylcholines with cytotoxic activity against neoplastic cells. The activity of 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET18OMe) and 3-hexadecylmercapto-2-methoxymethyl-propyl-1-phosphocholine (Ilmofosine^®) (BM 41.440) was tested in variants of B16 murine melanoma, grown in adhesion cultures (B16F1 with low metastatic potential; B16F10 and B16BL6 with high metastatic potential). Cytotoxicity was evaluated by counting the cells that survived after 24h of drug exposure. Cholesterol, sphingomyelin, total phospholipid and phosphatidylcholine levels were determined. After 24 h of drug exposure, cultures of the B16BL6 variant contained a larger number of cells, especially when high drug concentrations (100–250 μM) were used, than cultures of the B16F1 and B16F10 variants. The sensitivity of ET18OMe of the three variants was evaluated at different cell densities (at each density the dose was equalized per number of cells/well; 0.1 μmol/10⁶ cells/well). In B16F1 and B16F10 cultures the dose-response curve was not affected by the number of cells/well, while in B16BL6 no more than 20% of the cells were killed at all cell densities measured. A linear relationship was noted between cell density and cholesterol/phospholipid and sphingomyelin/phosphatidylcholine ratios in the resistant variant B16BL6, confirming that lipid composition modulates the cytotoxic activity of ether lipids.     

Antioxidative Characteristics of Anisomeles indica Extract and Inhibitory Effect of Ovatodiolide on Melanogenesis

The purpose of the study was to investigate the antioxidant characteristics of Anisomeles indica methanol extract and the inhibitory effect of ovatodiolide on melanogenesis. In the study, the antioxidant capacities of A. indica methanol extract such as DPPH assay, ABTS radical scavenging assay, reducing capacity and metal ion chelating capacity as well as total phenolic content of the extract were investigated. In addition, the inhibitory effects of ovatodiolide on mushroom tyrosinase, B16F10 intracellular tyrosinase and melanin content were determined spectrophotometrically. Our results revealed that the antioxidant capacities of A. indica methanol extract increased in a dose-dependent pattern. The purified ovatodiolide inhibited mushroom tyrosinase activity (IC(50) = 0.253 mM), the compound also effectively suppressed intracellular tyrosinase activity (IC(50) = 0.469 mM) and decreased the amount of melanin (IC(50) = 0.435 mM) in a dose-dependent manner in B16F10 cells. Our results concluded that A. indica methanol extract displays antioxidant capacities and ovatodiolide purified from the extract inhibited melanogenesis in B16F10 cells. Hence, A. indica methanol extract and ovatodiolide could be applied as a type of dermatological whitening agent in skin care products.     

Protocatechuic Aldehyde Inhibits α-MSH-Induced Melanogenesis in B16F10 Melanoma Cells via PKA/CREB-Associated MITF Downregulation

Protocatechuic aldehyde (PA) is a naturally occurring phenolic compound that is a potent inhibitor of mushroom tyrosinase. However, the molecular mechanisms of the anti-melanogenesis activity of PA have not yet been reported. The aim of the current study was to clarify the melanogenesis inhibitory effects of PA and its molecular mechanisms in murine melanoma cells (B16F10). We first predicted the 3D structure of tyrosinase and used a molecular docking algorithm to simulate binding between tyrosinase and PA. These molecular modeling studies calculated a binding energy of −527.42 kcal/mol and indicated that PA interacts with Cu400 and 401, Val283, and His263. Furthermore, PA significantly decreased α-MSH-induced intracellular tyrosinase activity and melanin content in a dose-dependent manner. PA also inhibited key melanogenic proteins such as tyrosinase, tyrosinase-related protein 1 (TRP-1), and TRP-2 in α-MSH-stimulated B16F10 cells. In addition, PA decreased MITF expression levels by inhibiting phosphorylation of cAMP response element-binding protein (CREB) and cAMP-dependent protein kinase A (PKA). These results demonstrate that PA can effectively suppress melanin synthesis in melanoma cells. Taken together, our results show that PA could serve as a potential inhibitor of melanogenesis, and hence could be explored as a possible skin-lightening agent.     

Saccharide composition in atmospheric particulate matter in the southwest US and estimates of source contributions

Saccharide compositions were measured in ambient particulate matter (PM) samples collected at four sites in Eastern Texas and Central Arizona in an effort to assess the contribution from biomass burning, atmospheric entrainment of soil, and primary biological aerosol particles (PBAPs) in regions with different climate patterns and ecosystems. In spite of the concentration difference, samples from the four study sites showed similar saccharide composition and seasonal variation, with the only exception being trehalose, likely resulting from the influence of local climatic conditions. Comparison of samples at the Arizona site showed different saccharide enrichment patterns between PM2.5 and PM10, which is consistent with their proposed sources as determined by a correlation analysis between observations. A positive matrix factorization (PMF) model was used to resolve three saccharide-related source factors and their relative source contributions to ambient PM using saccharides as tracers. Both the correlation and PMF model results indicate a stronger influence from local biogenic sources on aerosol saccharides at the rural sites than the urban site in Texas, and a greater impact from the PBAP and other biologically derived sources for PM collected in the Arizona location. This paper will provide the first analysis of saccharides in both fine and coarse PM in two US regions with dramatically different climates and ecosystem and provide a characterization of the ambient aerosol sources.     

Thermal and Mechanical Properties of Polyurethanes Derived from Mono- and Disaccharides

Thermal and mechanical properties of polyurethane (PU) sheets pre-pared from the glucose/fructose/sucrose–polyethylene glycol (PEG)–diphenylmethane diisocyanate (MDI) system were examined by differential scanning calorimetry, thermogravimetry, dynamic mechanical analysis and tensile tests. The saccharide content was varied at a constant NCO: OH ratio of 1·0. The glass transition temperature (T_g) increased with increasing saccharide content. The incorporation of saccharides into the PU structure results in a higher crosslinking density and a higher content of hard segments. The thermal decomposition was dependent on the saccharide content, an increase leading to a lower thermal decomposition temperature (T_d). The dissociation of saccharide OH groups and NCO groups is a major part of the thermal decomposition of these PUs. Dynamic mechanical analysis revealed two kinds of relaxation: the high temperature relaxation corresponds to main chain motion and the other is a local mode relaxation due to non-reacted isocyanate groups. The tensile stress and Young’s modulus increased with the saccharide content. © of SCI.     

重组腺病毒Ad-CMV-TK的构建及其对Lewis细胞的抑制作用

目的构建重组腺病毒Ad-CMV-TK,并检测Ad-CMV-TK/GCV系统对小鼠Lewis肺癌细胞的抑制作用。方法PCR扩增HSV-TK基因,以复制缺陷型腺病毒为载体构建重组腺病毒Ad-CMV-TK,在HEK293细胞中包装,CsCl2梯度离心法纯化后,感染Lewis肺癌细胞,Western blot检测HSV-TK蛋白的表达;MTT法检测Ad-CMV-TK/GCV系统对Lewis细胞的体外抑制作用;同时检测Ad-CMV-TK/GCV系统对C57荷瘤小鼠的抑瘤作用。结果重组腺病毒Ad-CMV-TK感染的Lewis细胞可特异地表达HSV-TK蛋白,Ad-CMV-TK/GCV系统在体外能明显抑制Lewis细胞的生长,当GCV浓度为100μmol/L时,Lewis细胞的存活率仅为16%;在体内能明显抑制荷瘤小鼠肿瘤的生长,与Ad-Blank/GCV对照组和PBS对照组相比,差异具有统计学意义。结论已成功构建了重组腺病毒Ad-CMV-TK,Ad-CMV-TK/GCV系统在体内及体外对小鼠Lewis肺癌细胞均有明显的抑制作用,为进一步研究其抗肿瘤机制及应用奠定了基础。     

Nanofiltration for separation and purification of saccharides from biomass

Saccharide production is critical to the development of biotechnology in the field of food and biofuel. The extraction of saccharide from biomass-based hydrolysate mixtures has become a trend due to low cost and abundant biomass reserves. Compared to conventional methods of fractionation and recovery of saccharides, nanofiltration (NF) has received considerable attention in recent decades because of its high selectivity and low energy consumption and environmental impact. In this review the advantages and challenges of NF based technology in the separation of saccharides are critically evaluated. Hybrid membrane processes, i.e., combining NF with ultrafiltration, can complement each other to provide an efficient approach for removal of unwanted solutes to obtain higher purity saccharides. However, use of NF membrane separation technology is limited due to irreversible membrane fouling that results in high capital and operating costs. Future development of NF membrane technology should therefore focus on improving material stability, antifouling ability and saccharide targeting selectivity, as well as on engineering aspects such as process optimisation and membrane module design.     

Hydration of Saccharides: Estimation of Reaction Properties and Equilibrium Conversion

Biomass holds great promise as a renewable source of hydrogen and thus as a zero‐emission, carbon‐neutral, and nearly inexhaustible energy resource. Thermodynamic analysis of biomass hydration is carried out to study the reaction properties considering a series of saccharides as a model. Equilibrium constants and composition are estimated in dependence on the saccharide chain length and temperature. The latter is also studied as a function of the reactant ratios and in the presence of nonreacting additives. Being highly endothermic, the reaction is thermodynamically favorable due to a high entropic contribution. Increase in the saccharide chain length affects the conversion only slightly, so the results for low‐molecular‐weight saccharides can be transferred to long‐chain ones.     

Lewis (y) Antigen Overexpression Increases the Expression of MMP-2 and MMP-9 and Invasion of Human Ovarian Cancer Cells

Lewis (y) antigen is a difucosylated oligosaccharide present on the plasma membrane, and its overexpression is frequently found in human cancers and has been shown to be associated with poor prognosis. Our previous studies have shown that Lewis (y) antigen plays a positive role in the process of invasion and metastasis of ovarian cancer cells. However, the mechanisms by which Lewis (y) antigen enhances the invasion and tumor metastasis are still unknown. In this study, we established a stable cell line constitutively expressing Lewis (y) antigen (RMG-1-hFUT) by transfecting the cDNA encoding part of the human α1,2-fucosyltransferase (α1,2-FUT) gene into the ovarian cancer cell line RMG-1, and investigated whether Lewis (y) antigen regulates the expression of matrix metalloproteinase-2 (MMP-2) and MMP-9, and tissue inhibitors of metalloproteinases (TIMP-1) and TIMP-2. We found that RMG-1-hFUT cells exhibited higher invasive capacities than their control cells. In addition, expression of TIMP-1 and TIMP-2 was down-regulated and expression of MMP-2 and MMP-9 was up-regulated. Anti-Lewis (y) antigen antibody treatment significantly reversed the expression of TIMP-1, TIMP-2, MMP-2 and MMP-9. Taken together, we provide the first evidence that down-regulation of TIMP-1 and TIMP-2 and up-regulation of MMP-2 and MMP-9 represents one of the mechanisms by which Lewis (y) antigen promotes cell invasion.     

The Anticancer Activity of Organotelluranes: Potential Role in Integrin Inactivation

Organic Te^IV compounds (organotelluranes) differing in their labile ligands exhibited anti‐integrin activities in vitro and anti‐metastatic properties in vivo. They underwent ligand substitution with l ‐cysteine, as a thiol model compound. Unlike inorganic Te^IV compounds, the organotelluranes did not form a stable complex with cysteine, but rather immediately oxidized it. The organotelluranes inhibited integrin functions, such as adhesion, migration, and metalloproteinase secretion mediation in B16F10 murine melanoma cells. In comparison, a reduced derivative with no labile ligand inhibited adhesion of B16F10 cells to a significantly lower extent, thus pointing to the importance of the labile ligands of the Te^IV atom. One of the organotelluranes inhibited circulating cancer cells in vivo, possibly by integrin inhibition. Our results extend the current knowledge on the reactivity and mechanism of organotelluranes with different labile ligands and highlight their clinical potential.     

Assembly of naturally occurring glycosides, evolved tactics, and glycosylation methods

Glycosylation of proteins and lipids is critical to many life processes. Secondary metabolites (or natural products), such as flavonoids, steroids, triterpenes, and antibiotics, are also frequently modified with saccharides. The resulting glycosides include diverse structures and functions, and some of them have pharmacological significance. The saccharide portions of the glycosides often have specific structural characteristics that depend on the aglycones. These molecules also form heterogeneous "glycoform" mixtures where molecules have similar glycosidic linkages but the saccharides vary in the length and type of monosaccharide unit. Thus, it is difficult to purify homogeneous glycosides in appreciable amounts from natural sources. Chemical synthesis provides a feasible access to the homogeneous glycosides and their congeners. Synthesis of a glycoside involves the synthesis of the aglycone, the saccharide, the connection of these two parts, and the overall manipulation of protecting groups. However, most synthetic efforts to date have focused on the aglycones, treating the attachment of saccharides onto the aglycones as a dispensable topic. The synthesis of the aglycone and the synthesis of the saccharide belong to two independent categories of chemistry, and different types of the aglycones and saccharides pose as specific synthetic subjects in their own disciplines. The only reaction that integrates the broad chemistry of glycoside synthesis is the glycosidic bond formation between the saccharide and the aglycone. Focusing on this glycosylation reaction in this Account, we string together our experience with the synthesis of the naturally occurring glycosides. We briefly describe the synthesis of 18 glycosides, including glycolipids, phenolic glycosides, steroid glycosides, and triterpene glycosides. Each molecule represents a prototypical structure of a family of the natural glycosides with interesting biological activities, and we emphasize the general tactics for the synthesis of these diverse structures. We provide a rationale for four tactics for the synthesis of glycosides, based on the stage at which the glycosidic bond is formed between the saccharide and the aglycone. This choice of tactic determines the success or failure of a synthesis, and the flexibility and the overall efficiency of the synthesis as well. Toward the synthesis of heterogeneous glycoform mixtures, we discuss successive and random glycosylation reactions. Finally, we have developed two new glycosylation protocols that address the challenges in the glycosylation of aglycones that are poorly nucleophilic, extremely acid labile, or extremely electrophilic. One of these new protocols takes advantage of glycosyl trifluoroacetimidate donors, and a second protocol uses gold(I)-catalyzed glycosylation with glycosyl ortho-alkynylbenzoate donors.     

Pharmacological Classification and Activity Evaluation of Furan and Thiophene Amide Derivatives Applying Semi-Empirical ab initio Molecular Modeling Methods

Pharmacological and physicochemical classification of the furan and thiophene amide derivatives by multiple regression analysis and partial least square (PLS) based on semi-empirical ab initio molecular modeling studies and high-performance liquid chromatography (HPLC) retention data is proposed. Structural parameters obtained from the PCM (Polarizable Continuum Model) method and the literature values of biological activity (antiproliferative for the A431 cells) expressed as LD(50) of the examined furan and thiophene derivatives was used to search for relationships. It was tested how variable molecular modeling conditions considered together, with or without HPLC retention data, allow evaluation of the structural recognition of furan and thiophene derivatives with respect to their pharmacological properties.     

Pyrrolo[1,2-a]quinoxalines: Insulin Mimetics that Exhibit Potent and Selective Inhibition against Protein Tyrosine Phosphatase 1B

PTP1B dephosphorylates insulin receptor and substrates to modulate glucose metabolism. This enzyme is a validated therapeutic target for type 2 diabetes, but no current drug candidates have completed clinical trials. Pyrrolo[1,2-a]quinoxalines substituted at positions C1–C4 and/or C7–C8 were found to be nontoxic to cells and good inhibitors in the low- to sub-micromolar range, with the 4-benzyl derivative being the most potent inhibitor (0.24 μm ). Some analogues bearing chlorine atoms at C7 and/or C8 kept potency and showed good selectivity compared to TCPTP (selectivity index >40). The most potent inhibitors behaved as insulin mimetics by increasing glucose uptake. The 4-benzyl derivative inhibited insulin receptor substrate 1 and AKT phosphorylation. Molecular docking and molecular dynamics simulations supported a putative binding mode for these compounds to the allosteric α3/α6/α7 pocket, but inconsistent results in enzyme inhibition kinetics were obtained due to the high tendency of these inhibitors to form stable aggregates. Computational calculations supported the druggability of inhibitors.     

Small‐Molecule Proteomimetic Inhibitors of the HIF‐1α–p300 Protein–Protein Interaction

The therapeutically relevant hypoxia inducible factor HIF‐1α–p300 protein–protein interaction can be orthosterically inhibited with α‐helix mimetics based on an oligoamide scaffold that recapitulates essential features of the C‐terminal helix of the HIF‐1α C‐TAD (C‐terminal transactivation domain). Preliminary SAR studies demonstrated the important role of side‐chain size and hydrophobicity/hydrophilicity in determining potency. These small molecules represent the first biophysically characterised HIF‐1α–p300 PPI inhibitors and the first examples of small‐molecule aromatic oligoamide helix mimetics to be shown to have a selective binding profile. Although the compounds were less potent than HIF‐1α, the result is still remarkable in that the mimetic reproduces only three residues from the 42‐residue HIF‐1α C‐TAD from which it is derived.     

Chemistry of Intermolecular Frustrated Lewis Pairs in Motion: Emerging Perspectives and Prospects

This feature article describes the chemistry in motion of frustrated Lewis pairs (FLPs). With state-of-the-art ab initio molecular dynamics (AIMD) simulations supplemented by minimum energy path (MEP) and potential energy surface (PES) calculations, we examine the binding of CO₂ and the heterolytic cleavage of H₂ by a Lewis base (LB), tBu₃P, and a Lewis acid (LA), B(C₆F₅)₃. We strive to uncover and understand mechanistic implications of the physical laws that govern the behavior of a LB and a LA when they react with a third species (e.g., CO₂ or H₂) at finite temperature. The approximation that we necessarily must make at present is to forgo the quantization of the movement of atoms in favor of the Born-Oppenheimer molecular dynamics (BOMD), which unfold according to the classical (Newton’s) laws of motion. However, strict quantum chemical theory is used to compute all of the forces that govern the dynamics of the macromolecular FLP system. Using physical reasoning and innovative computer simulations, we show that multi-scale motion is the predominant mechanistic aspect in reactions of the tBu₃P/B(C₆F₅)₃ FLP, as well as, conceivably, those of other similar intermolecular FLPs. Insight achieved thus far leads to a novel activity model for intermolecular FLPs and specific predictions, which could be useful for future experimental and theoretical studies of FLP and other chemistries.