A novel microtubule destabilizing entity from orthogonal synthesis of triazine library and zebrafish embryo screening

The first orthogonal combinatorial synthesis of a high-purity triazine library was demonstrated. Novel triazine-based microtubule inhibitors were discovered by an efficient zebrafish embryo screening and in vitro microtubule polymerization assay.     

Synthesis of novel apio carbocyclic nucleoside analogues as selective a(3) adenosine receptor agonists

On the basis of the biological activity of neplanocin A and apio-dideoxyadenosine (apio-ddA), novel apio-neplanocin A analogues 5a-d, combining the properties of two nucleosides, were stereoselectively synthesized. The apio moiety of the target nucleosides 5a-d was stereoselectively introduced by treating lactol 10 with 37% formaldehyde in the presence of potassium carbonate. The carbasugar moiety of neplanocin A was successively built by exposing diene 12 on a Grubbs catalyst in methylene chloride. The final nucleosides 5a-d were synthesized from the condensation of the glycosyl donor 14 with nucleic bases under the standard Mitsunobu conditions. Similarly, apio-aristeromycin 6 and (N)-apio-methanocarbaadenosine 7 were derived from the common intermediate 13 using catalytic hydrogenation and Simmons-Smith cyclopropanation as key steps. All of the final nucleosides 5a-d, 6, and 7 did not show significant inhibitory activity against S-adenosylhomocysteine hydrolase (SAH) up to 100 muM, maybe due to the absence of the secondary hydroxyl group at the C3'-position, which should be oxidized by cofactor-bound NAD(+). However, apio-neplanocin A (5a) showed potent and highly selective binding affinity (K(i) = 628 +/- 69 nM) at the A(3) adenosine receptor without any binding affinity at the A(1) and A(2A) adenosine receptors. In conclusion, we have first developed novel carbocyclic nucleosides with unnatural apio-carbasugars using stereoselective hydroxymethylation and RCM reaction and also discovered a new template of human A(3) adenosine receptor agonist, which play a great role in developing new A(3) adenosine receptor agonist as well as in identifying the binding site of the receptor.     

Calcium is involved in photomovement of cyanobacterium Synechocystis sp. PCC 6803

The unicellular cyanobacterium Synechocystis sp. PCC 6803 (Syn6803) exhibits photomovement through gliding motility. For a better understanding of photomovement in Syn6803, we examined the effects of Ca2+ on photoorientation and motility using a computer-assisted videomicroscope motion analysis system. When calcium ion was chelated from the basic motility medium by adding 0.5 mM ethylene glycol-bis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), the photoorientation was completely inhibited, whereas the gliding motility remained approximately 70% of the control. Photoorientation impaired by EGTA was nearly recovered within 30 min upon addition of 1 mM Ca2+. The recovery of photoorientation by Ca2+ was mimicked by either Mn2+ or Mg2+ but not by Ba2+ or Sr2+. Lanthanum ion at 10 microM completely inhibited both phototactic orientation and gliding motility of Syn6803. Furthermore, pimozide (voltage-gated L-type calcium channel inhibitor), orthovanadate (calcium efflux blocker) and A23187 (calcium ionophore) partially inhibited phototactic orientation and gliding motility. Interestingly, photoorientation was prevented with increasing concentrations of calmodulin antagonist such as trifluoperazine (TFP) and chlorpromazine, but gliding motility was inhibited in proportion to the concentration of TFP. The results we present strongly indicate that Ca2+ plays a significant role in regulating the photomovement of Syn6803.     

A critical role for Th17 cell-derived TGF-β1 in regulating the stability and pathogenicity of autoimmune Th17 cells

Pathogenic conversion of Th17 cells into multifunctional helper T cells or Th1 cells contributes to the pathogenesis of autoimmune diseases; however, the mechanism regulating the plasticity of Th17 cells remains unclear. Here, we found that Th17 cells expressed latent TGF-β1 in a manner dependent on autocrine TGF-β1. By employing IL-17-producing cell-specific Tgfb1 conditional knockout and fate-mapping systems, we demonstrated that TGF-β1-deficient Th17 cells are relatively susceptible to becoming IFN-γ producers through IL-12Rβ2 and IL-27Rα upregulation. TGF-β1-deficient Th17 cells exacerbated tissue inflammation compared to TGF-β1-sufficient Th17 cells in adoptive transfer models of experimental autoimmune encephalomyelitis and colitis. Thus, TGF-β1 production by Th17 cells provides an essential autocrine signal for maintaining the stability and regulating the pathogenicity of Th17 cells in vivo.Subject terms: Autoimmunity, Neuroimmunology     

Diffusion Mechanism of Carbon Dioxide in Zeolite 4A and CaX Pellets

The adsorption kinetics and equilibria of CO₂ in commercial zeolite 4A and CaX pellets were theoretically and experimentally studied by a gravimetric method in the range of 273–313 K and 0.0–0.8 atm. The diffusion mechanism of an adsorbate into a pellet is composed of micropore and macropore diffusion due to the bidisperse structure of the pellet. When one diffusion mechanism played a more important role than the other in determining the overall diffusion rate, the diffusion rate was estimated by the nonisothermal monodisperse diffusion model (NMDM). However, when the combined effects of both mechanisms controlled the overall adsorption kinetics, the experimental uptake was analyzed by the nonisothermal bidisperse diffusion model (NBDM). The CO₂ diffusion in zeolite 4A pellets was controlled by micropore diffusion within the experimental pressure and temperature ranges. However, both macropore and micropore diffusion contributed to CO₂ diffusion in the zeolite CaX pellet. The overall CO₂ diffusion rate in zeolite CaX became faster as pressure increased mainly due to its highly favorable isotherm in the zeolite CaX. The micropore diffusion time constant of CO₂ in the zeolite CaX pellet was approximately one hundred times greater than that in the zeolite 4A pellet. In addition, the activation energy of micropore diffusion of CO₂ diffusion in the zeolite CaX pellet was smaller than that in the zeolite 4A pellet. In this study, the dimensionless parameter, , indicating the relative importance of macropore and micropore diffusion, was modified to consider non-zero coverage as an initial condition for each step in the gravimetric method. When is greater than 100, the overall adsorption rate is controlled by macropore diffusion. However, in cases where is less than 0.1, micropore diffusion is the dominant mechanism in the overall adsorption rate. In the case of a system with between these values, both macropore and micropore diffusion contributed to the overall diffusion rate.     

An internal doping method for the preparation of transparent hybrid xerogels containing nano-sized iron particles

Sol-gel copolymerization of iron tricarbonyl-2-(triisopropoxysilyl)-1,3-butadiene with 1,6-bistriethoxysilylhexane and 1,4-bistriethoxysilylbenzene followed by drying produced bridged polysilsesquioxane xerogels. These porous, transparent hybrid materials containing the iron metal precursor were irradiated (UV) and heated under vacuum resulting in the deposition of nano-sized iron particles doped in the xerogels. EDAX and electron diffraction techniques were used to characterize the iron phases. The TEM images of these doped xerogels provided additional information regarding the domain size of the iron phase.Using a combination of external doping of Cd²⁺ and S^2– ions and internal doping of Fe°, mixed Fe/CdS phases were prepared within the porous bridged polysilsesquioxane xerogels. The resulting doped xerogels were found to have retained their porous morphology.     

Size control of self-assembled nanoparticles by an emulsion/solvent evaporation method

A novel and simple method for size control of self-assembled nanoparticles is suggested in this paper. Polymeric nanoparticles were prepared from amphiphilic chitosan derivatives fluorescein isothiocyanate (FITC)-conjugated glycol chitosans (FGCs). The attachment of hydrophobic FITC onto hydrophilic glycol chitosan induced the amphiphilic conjugate to form self-assembled nanoparticles in aqueous media, depending on degree of substitution. The size of self-assembled nanoparticles was controlled by a novel emulsion/solvent evaporation method. Adding a small amount of an immiscible solvent with water (chloroform) to FGC nanoparticle suspensions in aqueous media followed by ultrasonification and solvent evaporation led to partial dissociation and subsequent reformation of nanoparticles. The evaporation of chloroform facilitated the hydrophobic association, which resulted in more dense and hardened hydrophobic cores. The size of nanoparticles was closely related with the FGC concentration in the emulsion. The mean diameters of self-assembled nanoparticles were 150–500 nm at the FGC concentrations of 0.3–2.5 mg/ml. Higher FGC concentration resulted in larger particles. The polydispersity factors (μ ₂/Γ ²) of the reformed nanoparticles were fairly low (0.001–0.094), indicating narrow size distribution. The FGC nanoparticles were stable in phosphate-buffered saline at 37°C up to 20 days. Lactose was a good excipient for maintaining the structural integrity of nanoparticles during freeze-drying. Without lactose, the freeze-dried nanoparticles were not homogeneously redispersed in aqueous media. However, the freeze-dried nanoparticles with lactose were spontaneously redispersed in aqueous milieu with their own sizes.     

Two-dimensional infrared spectroscopy of antiparallel beta-sheet secondary structure

We investigate the sensitivity of femtosecond Fourier transform two-dimensional infrared spectroscopy to protein secondary structure with a study of antiparallel beta-sheets. The results show that 2D IR spectroscopy is more sensitive to structural differences between proteins than traditional infrared spectroscopy, providing an observable that allows comparison to quantitative models of protein vibrational spectroscopy. 2D IR correlation spectra of the amide I region of poly-l-lysine, concanavalin A, ribonuclease A, and lysozyme show cross-peaks between the IR-active transitions that are characteristic of amide I couplings for polypeptides in antiparallel hydrogen-bonding registry. For poly-l-lysine, the 2D IR spectrum contains the eight-peak structure expected for two dominant vibrations of an extended, ordered antiparallel beta-sheet. In the proteins with antiparallel beta-sheets, interference effects between the diagonal and cross-peaks arising from the sheets, combined with diagonally elongated resonances from additional amide transitions, lead to a characteristic "Z"-shaped pattern for the amide I region in the 2D IR spectrum. We discuss in detail how the number of strands in the sheet, the local configurational disorder in the sheet, the delocalization of the vibrational excitation, and the angle between transition dipole moments affect the position, splitting, amplitude, and line shape of the cross-peaks and diagonal peaks.