Potent Activation of Human but Not Mouse TRPA1 by JT010

Transient receptor potential (TRP) ankyrin repeat 1 (TRPA1), which is involved in inflammatory pain sensation, is activated by endogenous factors, such as intracellular Zn²⁺ and hydrogen peroxide, and by irritant chemical compounds. The synthetic compound JT010 potently and selectively activates human TRPA1 (hTRPA1) among the TRPs. Therefore, JT010 is a useful tool for analyzing TRPA1 functions in biological systems. Here, we show that JT010 is a potent activator of hTRPA1, but not mouse TRPA1 (mTRPA1) in human embryonic kidney (HEK) cells expressing hTRPA1 and mTRPA1. Application of 0.3–100 nM of JT010 to HEK cells with hTRPA1 induced large Ca²⁺ responses. However, in HEK cells with mTRPA1, the response was small. In contrast, both TRPA1s were effectively activated by allyl isothiocyanate (AITC) at 10–100 μM. Similar selective activation of hTRPA1 by JT010 was observed in electrophysiological experiments. Additionally, JT010 activated TRPA1 in human fibroblast-like synoviocytes with inflammation, but not TRPA1 in mouse dorsal root ganglion cells. As cysteine at 621 (C621) of hTRPA1, a critical cysteine for interaction with JT010, is conserved in mTRPA1, we applied JT010 to HEK cells with mutations in mTRPA1, where the different residue of mTRPA1 with tyrosine at 60 (Y60), with histidine at 1023 (H1023), and with asparagine at 1027 (N1027) were substituted with cysteine in hTRPA1. However, these mutants showed low sensitivity to JT010. In contrast, the mutation of hTRPA1 at position 669 from phenylalanine to methionine (F669M), comprising methionine at 670 in mTRPA1 (M670), significantly reduced the response to JT010. Moreover, the double mutant at S669 and M670 of mTRPA1 to S669E and M670F, respectively, induced slight but substantial sensitivity to 30 and 100 nM JT010. Taken together, our findings demonstrate that JT010 potently and selectively activates hTRPA1 but not mTRPA1.     

Dehydration of d-glucose in high temperature water at pressures up to 80 MPa

Reaction of d-glucose in water to yield 5-hydroxymethylfurfural (5-HMF), 1,2,4-benzenetriol (BTO) and furfural was studied at high temperatures (up to 400 °C) and high pressures (up to 80 MPa) using a continuous flow reactor. Maximum temperature and pressure conditions gave maximum furfural yield. Increasing pressure from 40 to 70 and 80 MPa enhanced dehydration reactions to 5-HMF, but also enhanced hydrolysis of 5-HMF leading to the production of BTO and thus lead to lower yields of 5-HMF (below 10%). Remarkably, the dehydration reaction to 5-HMF and the hydrolysis of 5-HMF were both enhanced by the increase in water density at 400 °C.     

Catching a molecule at the air-water interface: Dynamic pore array for molecular recognition

As a soft and flexible porous structure, a pore array of a steroid cyclophane SC(OH), which consists of the rigid 1,6,20,25-tetraaza[6.1.6.1]paracyclophane ring connected to four steroid moieties (cholic acid) through flexible L-lysine spacers, was prepared at the air-water interface. As confirmed by surface pressure (π)-molecular area (A) isotherms, transition between open conformation and cavity conformation of the SC(OH) molecule was reversibly induced upon repeated compression and expansion of its monolayer at pH 11 where amino groups of the lysine residues are not fully deprotonated. Capture and release of an aqueous fluorescent guest (TNS) by SC(OH) was observed upon dynamic cavity formation through surface fluorescence spectroscopy. At pH 12, dynamic cavity formation of SC(OH) was sufficiently suppressed, and the capture and release of an aqueous TNS by the monolayer was not virtually observed. Lessened electrostatic repulsion between the SC(OH) molecules due to conversion of ammonium to free amine may prevent the cavity from reopening. The importance of dynamic nature of cavity formation on the guest binding was also proved by control experiments using SC(H), which cannot form cavity conformation at any surface pressures at both pH 11 and 12.     

Deoxycholic acid formation in gnotobiotic mice associated with human intestinal bacteria

In humans and animals, intestinal flora is indispensable for bile acid transformation. The goal of our study was to establish gnotobiotic mice with intestinal bacteria of human origin in order to examine the role of intestinal bacteria in the transformation of bile acids in vivo using the technique of gnotobiology. Eight strains of bile acid-deconjugating bacteria were isolated from ex-germ-free mice inoculated with a human fecal dilution of 10⁻⁶, and five strains of 7α-dehydroxylating bacteria were isolated from the intestine of limited human flora mice inoculated only with clostridia. The results of biochemical tests and 16S rDNA sequence analysis showed that seven out of eight bile acid-deconjugating strains belong to a bacteroides cluster (Bacteroides vulgatus, B. distasonis, and B. uniformis), and one strain had high similarity with Bilophila wadsworthia. All five strains that converted cholic acid to deoxycholic acid had greatest similarity with Clostridium hylemonae. A combination of 10 isolated strains converted taurocholic acid into deoxycholic acid both in vitro and in the mouse intestine. These results indicate that the predominant bacteria, mainly Bacteroides, in human feces comprise one of the main bacterial groups for the deconjugation of bile acids, and clostridia may play an important role in 7α-dehydroxylation of free-form primary bile acids in the intestine although these strains are not predominant. The gnotobiotic mouse with bacteria of human origin could be a useful model in studies of bile acid metabolism by human intestinal bacteria in vivo.     

Size reduction of PtRu catalyst particle deposited on carbon support by addition of non-metallic elements

Addition of non-metallic elements such as N, P and S was examined to reduce the catalyst particle size of PtRu deposited on a carbon support and to improve the catalytic performance. It was found that the addition of N, P and S reduced the size of PtRu catalyst particle and that P was the most effective additive on the size reduction. A well dispersed PtRu catalyst particle 2 nm in size was obtained by the addition of P. The maximum power density was observed to be 64 mW/cm² in a passive state of direct methanol fuel cell (DMFC) at room temperature by using the PtRuP anode catalyst. The PtRuP catalyst retained the size independently with surface area of the carbon support. This made it possible to use a carbon support with low surface area (lower porosity), resulting in better accessibility by molecules on the noble metal particles which were deposited prior to the external surface in compared to the micropore walls. The maximum power density was improved from 44 to 64 mW/cm² by using a less porous carbon support (140 m²/g).     

Higher-order structures and mechanical properties of stereocomplex-type poly(lactic acid) melt spun fibers

Equimolar blend of poly(l-lactic acid) (PLLA) and poly(d-lactic acid) (PDLA) was melt spun into fibers and the relations among the processing conditions, crystalline structures, thermal properties, and mechanical properties were investigated. Drawing and annealing were performed in order to obtain fiber mainly consisting of the stereocomplex crystal phase. Fibers drawn at various temperatures exhibited either amorphous, highly oriented homo crystal, or the mixture of homo and stereocomplex with a fairly low orientation depending on the drawing temperature. Annealing of the drawn fibers at an elevated temperature higher than the melting temperature of homo crystal increased the stereocomplex content significantly. The fractions of the homo and the stereocomplex crystals strongly depended on the higher-order structure of the drawn fibers and the annealing temperature.     

Sol–Gel‐Derived High‐Performance Stacked Transparent Conductive Oxide Thin Films

Single‐ and multi‐layer transparent conductive oxide (TCO) thin films exhibiting high performance, good packing density and low surface/interface roughness are deposited on silica glass substrates by the sol–gel method. The crystal and microstructural properties of the TCO thin films are evaluated as an alternate to films prepared by ultra‐high vacuum deposition. Tin‐doped indium oxide (ITO) thin films produced using a two‐step drying process showed low surface roughness because of dense packing structure not only horizontal but also vertical directions. As a result, electrical conductivity, carrier concentration, carrier mobility, and optical transmittance of 2.3 × 10³ S/cm, 8 × 10²⁰ cm^?3, 18 cm²/Vs, and over 98% at 500 nm, respectively, were achieved. A multilayer ZnO/ITO stacked structure was also fabricated using the sol–gel process. Our findings suggest that solution‐based methods show promise as an alternative to existing ultra‐high vacuum methods to fabricate TCO thin films.     

Mixed Lubrication with C18 Fatty Acids: Effect of Unsaturation

In this paper, the lubrication mechanism of fatty acids is revisited with a new approach combining experimental and computational chemistry studies. The lubricating properties of single and mixtures of stearic, oleic and linoleic acids in a synthetic Poly-Alpha-Olefin base oil (PAO4) on iron oxide surface are investigated under mixed boundary regime with temperatures from 50 °C up to 150 °C. Low friction coefficient (about 0.055) with no visible wear is reported in the presence of single stearic acid at high temperature. This lubricating behavior is inhibited in the presence of unsaturated fatty acids highlighting an anti-synergic effect of a saturated/unsaturated mixture, especially at 150 °C. To understand the anti-synergic effect and the adsorption mechanism of these molecules, molecular dynamic (MD) and quantum chemistry simulations are performed to evaluate their diffusion coefficient in PAO4 and their adsorption mechanism on iron oxide at different temperatures. MD simulation results show a faster diffusion toward the surface for unsaturated fatty acids than for saturated fatty acid at all the studied temperatures. This means that unsaturated molecules arrive and mainly adsorb before stearic acid on the surface leading to a tribological behavior of the mixture characteristic of the unsaturated molecule. Computational chemistry suggests that all fatty acids (saturated and unsaturated) adsorption mechanism is due to the chemisorption of the carboxylic group on iron oxide surface with no desorption up to 150 °C.     

Accumulation of triosephosphate isomerase, with sequence homology to Beta amyloid peptides, in vessel walls of the newborn piglet hippocampus

We investigated whether beta-amyloid (Abeta)-like immunoreactivity was seen in the brains of newborn piglets. The immunoreactivity for Abeta(1-42) and Abeta(1-40) proteins, but not Abeta precursor protein, was present in CD68-positive perivascular cells of the hippocampus and in parts of the meninges. It was colocalized with immunoreactivity for receptor for advanced glycation end product and tumor necrosis factor-alpha. The protein with a molecular mass of 27 kDa, which was recognized by the Abeta antibodies, was identified as triosephosphate isomerase (TPI) with sequence homology to Abeta peptides by N-terminal amino acid sequencing, mass fingerprint analysis using matrix-associated laser desorption/ionization mass spectrometry, and Western blotting. Western blotting assay also revealed that detectable expression of Abeta proteins were not seen in the piglet brains. These findings indicate that TPI with sequence homology to Abeta peptides accumulates in perivascular cells of the microglia/macrophage lineage located around arterial vessels of the newborn piglet hippocampus.