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The self-motion of an ester boat is investigated depending on the physicochemical properties of the surface-active substance. The results show that the ester boat moves towards the higher surface tension generating as the driving force.

The desirable properties of the sophisticated fluorescent pH probe are ratiometric detection properties and a wide detection range. In this study, three types of fluorophores with different fluorescence properties were assembled on a DNA origami nanostructure. DNA nanostructure has the advantage of being a scaffold that can assemble different types of fluorophores with control over their number and position. The defined number of three different fluorophores, i.e., pH-sensitive fluorescein (CF) and Oregon Green (OG), and pH-insensitive tetramethylrhodamine (CR), assembled on the DNA scaffold provided a ratiometric fluorescent pH probe with a wide pH detection range that could cover the variation of intracellular pH.

Photocatalysts have recently attracted attention for removing infectious-disease-causing bacteria and viruses. Among such photocatalysts, ground Rh-doped SrTiO3 (“g-STO:Rh”) has been found to have biospecificity that reduces the Qβ phage infectivity under conditions that did not decrease the E. coli survival rate. Elucidating the mechanism of selective antiphage activation is important for developing photocatalysts that act effectively against specific microorganisms. In this study, SDS-PAGE and quantitative PCR showed that a g-STO:Rh-treated Qβ phage preferentially inactivated the A2 protein involved in attachment to host cells. The analysis of the photocatalyst-treated ovalbumin using g-STO:Rh indicated that the protein’s isoelectric point significantly influenced the initial interaction with g-STO:Rh. However, once the protein is absorbed, it was decomposed without the release of intermediates. Furthermore, an inactivation assay for four different phages by photocatalyst treatment using g-STO:Rh revealed that phages with positively charged proteins are highly susceptible to inactivation, and the accessibility of critical components to g-STO:Rh influences susceptibility. We conclude that the selective antiphage activation of g-STO:Rh depends on the adsorption efficiency of the protein and g-STO:Rh.

The Acyl-activating enzyme (AAE) 3 gene encodes an oxalyl-CoA synthetase that catalyzes the conversion of oxalate to oxalyl-CoA as the first step in the CoA-dependent pathway of oxalate catabolism. Although the role of this enzyme in oxalate catabolism has been established, its biological roles in plant growth and development are less understood. As a step toward gaining a better understanding of these biological roles, we report here a characterization of the Arabidopsis thaliana aae3 (Ataae3) seed mucilage phenotype. Ruthidium red (RR) staining of Ataae3 and wild type (WT) seeds suggested that the observed reduction in Ataae3 germination may be attributable, at least in part, to a decrease in seed mucilage accumulation. Quantitative RT-PCR analysis revealed that the expression of selected mucilage regulatory transcription factors, as well as of biosynthetic and extrusion genes, was significantly down-regulated in the Ataae3 seeds. Mucilage accumulation in seeds from an engineered oxalate-accumulating Arabidopsis and Atoxc mutant, blocked in the second step of the CoA-dependent pathway of oxalate catabolism, were found to be similar to WT. These findings suggest that elevated tissue oxalate concentrations and loss of the oxalate catabolism pathway downstream of AAE3 were not responsible for the reduced Ataae3 seed germination and mucilage phenotypes. Overall, our findings unveil the presence of regulatory interplay between AAE3 and transcriptional control of mucilage gene expression.

1,5-Anhydro-D-fructose (1,5-AF) is a bioactive monosaccharide that is produced by the glycogenolysis in mammalians and is metabolized to 1,5-anhydro-D-glucitol (1,5-AG). 1,5-AG is used as a marker of glycemic control in diabetes patients. 1,5-AF has a variety of physiological activities, but its effects on energy metabolism, including feeding behavior, are unclarified. The present study examined whether 1,5-AF possesses the effect of satiety. Peroral administration of 1,5-AF, and not of 1,5-AG, suppressed daily food intake. Intracerebroventricular (ICV) administration of 1,5-AF also suppressed feeding. To investigate the neurons targeted by 1,5-AF, we investigated c-Fos expression in the hypothalamus and brain stem. ICV injection of 1,5-AF significantly increased c-Fos positive oxytocin neurons and mRNA expression of oxytocin in the paraventricular nucleus (PVN). Moreover, 1,5-AF increased cytosolic Ca2+ concentration of oxytocin neurons in the PVN. Furthermore, the satiety effect of 1,5-AF was abolished in oxytocin knockout mice. These findings reveal that 1,5-AF activates PVN oxytocin neurons to suppress feeding, indicating its potential as the energy storage monitoring messenger to the hypothalamus for integrative regulation of energy metabolism.

E3 ubiquitin ligases play important roles in plant immunity, but their role in soybean has not been investigated previously. Here, we used Bean pod mottle virus (BPMV)-mediated virus-induced gene silencing (VIGS) to investigate the function of GmSAUL1 (Senescence-Associated E3 Ubiquitin Ligase 1) homologs in soybean. When two closely related SAUL1 homologs were silenced simultaneously, the soybean plants displayed autoimmune phenotypes, which were significantly alleviated by high temperature, suggesting that GmSAUL1a/1b might be guarded by an R protein. Interestingly, silencing GmSAUL1a/1b resulted in the decreased activation of GmMPK6, but increased activation of GmMPK3 in response to flg22, suggesting that the activation of GmMPK3 is most likely responsible for the activated immunity observed in the GmSAUL1a/1b-silenced plants. Furthermore, we provided evidence that GmSAUL1a is a bona fide E3 ligase. Collectively, our results indicated that GmSAUL1 plays a negative role in regulating cell death and immunity in soybean.

Osteoarthritis (OA) occurs not only in the knee but also in peripheral joints throughout the whole body. Previously, we have shown that the expression of cellular communication network factor 3 (CCN3), a matricellular protein, increases with age in knee articular cartilage, and the misexpression of CCN3 in cartilage induces senescence-associated secretory phenotype (SASP) factors, indicating that CCN3 promotes cartilage senescence. Here, we investigated the correlation between CCN3 expression and OA degenerative changes, principally in human femoral head cartilage. Human femoral heads obtained from patients who received total hip arthroplasty were categorized into OA and femoral neck fracture (normal) groups without significant age differences. Gene expression analysis of RNA obtained from femoral head cartilage revealed that CCN3 and MMP-13 expression in the non-weight-bearing part was significantly higher in the OA group than in the normal group, whereas the weight-bearing OA parts and normal cartilage showed no significant differences in the expression of these genes. The expression of COL10A1, however, was significantly higher in weight-bearing OA parts compared with normal weight-bearing parts, and was also higher in weight-bearing parts compared with non-weight-bearing parts in the OA group. In contrast, OA primary chondrocytes from weight-bearing parts showed higher expression of CCN3, p16, ADAMTS4, and IL-1β than chondrocytes from the corresponding normal group, and higher ADAMTS4 and IL-1β in the non-weight-bearing part compared with the corresponding normal group. Acan expression was significantly lower in the non-weight-bearing group in OA primary chondrocytes than in the corresponding normal chondrocytes. The expression level of CCN3 did not show significant differences between the weight-bearing part and non-weight-bearing part in both OA and normal primary chondrocytes. Immunohistochemical analysis showed accumulated CCN3 and aggrecan neoepitope staining in both the weight-bearing part and non-weight-bearing part in the OA group compared with the normal group. The CCN3 expression level in cartilage had a positive correlation with the Mankin score. X-ray analysis of cartilage-specific CCN3 overexpression mice (Tg) revealed deformation of the femoral and humeral head in the early stage, and immunohistochemical analysis showed accumulated aggrecan neoepitope staining as well as CCN3 staining and the roughening of the joint surface in Tg femoral and humeral heads. Primary chondrocytes from the Tg femoral head showed enhanced expression of Ccn3, Adamts5, p16, Il-6, and Tnfα, and decreased expression of Col2a1 and -an. These findings indicate a correlation between OA degenerative changes and the expression of CCN3, irrespective of age and mechanical loading. Furthermore, the Mankin score indicates that the expression level of Ccn3 correlates with the progression of OA.

The molecular chaperones HdeA and HdeB of the Escherichia coli (E. coli) periplasm protect client proteins from acid denaturation through a unique mechanism that utilizes their acid denatured states to bind clients. We previously demonstrated that the active, acid-denatured form of HdeA is also prone to forming inactive, amyloid fibril-like aggregates in a pH-dependent, reversible manner. In this study, we report that HdeB also displays a similar tendency to form fibrils at low pH. HdeB fibrils were observed at pH < 3 in the presence of NaCl. Similar to HdeA, HdeB fibrils could be resolubilized by a simple shift to neutral pH. In the case of HdeB, however, we found that after extended incubation at low pH, HdeB fibrils were converted into a form that could not resolubilize at pH 7. Fresh fibrils seeded from these “transformed” fibrils were also incapable of resolubilizing at pH 7, suggesting that the transition from reversible to irreversible fibrils involved a specific conformational change that was transmissible through fibril seeds. Analyses of fibril secondary structure indicated that HdeB fibrils retained significant alpha helical content regardless of the conditions under which fibrils were formed. Fibrils that were formed from HdeB that had been treated to remove its intrinsic disulfide bond also were incapable of resolubilizing at pH 7, suggesting that certain residual structures that are retained in acid-denatured HdeB are important for this protein to recover its soluble state from the fibril form.

This study targets on-site/real-time taxonomic identification and metabolic profiling of seven different Candida auris clades/subclades by means of Raman spectroscopy and imaging. Representative Raman spectra from different Candida auris samples were systematically deconvoluted by means of a customized machine-learning algorithm linked to a Raman database in order to decode structural differences at the molecular scale. Raman analyses of metabolites revealed clear differences in cell walls and membrane structure among clades/subclades. Such differences are key in maintaining the integrity and physical strength of the cell walls in the dynamic response to external stress and drugs. It was found that Candida cells use the glucan structure of the extracellular matrix, the degree of α-chitin crystallinity, and the concentration of hydrogen bonds between its antiparallel chains to tailor cell walls’ flexibility. Besides being an effective ploy in survivorship by providing stiff shields in the α–1,3–glucan polymorph, the α–1,3–glycosidic linkages are also water-insoluble, thus forming a rigid and hydrophobic scaffold surrounded by a matrix of pliable and hydrated β–glucans. Raman analysis revealed a variety of strategies by different clades to balance stiffness, hydrophobicity, and impermeability in their cell walls. The selected strategies lead to differences in resistance toward specific environmental stresses of cationic/osmotic, oxidative, and nitrosative origins. A statistical validation based on principal component analysis was found only partially capable of distinguishing among Raman spectra of clades and subclades. Raman barcoding based on an algorithm converting spectrally deconvoluted Raman sub-bands into barcodes allowed for circumventing any speciation deficiency. Empowered by barcoding bioinformatics, Raman analyses, which are fast and require no sample preparation, allow on-site speciation and real-time selection of appropriate treatments.

Plant cell deformation is a mechanical process that is driven by differences in the osmotic pressure inside and outside of the cell and is influenced by cell wall properties. Legume leaf movements result from reversible deformation of pulvinar motor cells. Reversible cell deformation is an elastic process distinct from the irreversible cell growth of developing organs. Here, we begin with a review of the basic mathematics of cell volume changes, cell wall function, and the mechanics of bending deformation at a macro scale. Next, we summarize the findings of recent molecular genetic studies of pulvinar development. We then review the mechanisms of the adaxial/abaxial patterning because pulvinar bending deformation depends on the differences in mechanical properties and physiological responses of motor cells on the adaxial versus abaxial sides of the pulvinus. Intriguingly, pulvini simultaneously encompass morphological symmetry and functional asymmetry along the adaxial/abaxial axis. This review provides an introduction to leaf movement and reversible deformation from the perspective of mechanics and molecular genetics.

We have prepared two tris(2,2’-bipyridine)ruthenium(II) complex-based new chymotrypsin (CHT) inhibitors, [Ru(bpy)2(bsfbpy)]Cl2 and [Ru(bsfbpy)3]Cl2 {bsfbpy = 4-methyl-2,2′-bipyridine-4′-carboxylic acid [2-(4-benzenesulfonylfluoride)-ethyl]-amide}. They were covalently modified with Ser195 in the active site of CHT. Photoinduced electron-transfer (ET) systems comprising a Ru-CHT complex and an electron acceptor such as methylviologen (MV2+) were studied. After photoirradiation, the photo-excited triplet state of 2CHT-3{[Ru(bsfbpy)3]2+}* and CHT-3{[Ru(bpy)2(bsfbpy)]}* were both quenched by MV2+ through an intermolecular photoinduced ET mechanism, forming oxidized 2CHT-[Ru(bsfbpy)3]3+ and CHT-[Ru(bpy)2(bsfbpy)]3+, respectively. The intramolecular ET from the amino acid residue of Tyr94 near the active site of CHT proceeded. Transient absorption spectra display the formation of a Tyr●+ radical in the 2CHT-[Ru(bsfbpy)3]2+-MV2+ and the CHT-[Ru(bpy)2(bsfbpy)]2+-MV2+ systems, having a lifetime of 2.24 ms for 2CHT-[Ru(bsfbpy)3]2+-MV2+. The present model system could be one of the available models for the further photochemical reaction of Tyr•+ formed by intramolecular photoinduced ET reactions within a protein scaffold. To clarify the photophysical properties of these Ru inhibitors and their multi step photoinduced ET reactions in the protein matrix, the detailed ET reaction kinetics are discussed.

Structural programmability and accurate addressability of DNA nanostructures are ideal characteristics for the platform of arranging enzymes with the nanoscale precision. In this study, a three-dimensional DNA scaffold was designed to enable a dynamic shape transition from an open plate-like structure to its closed state of a hexagonal prism structure. The two domains in the open state were folded together to transform into the closed state by hybridization of complementary short DNA closing keys at both of the facing edges in over 90% yield. The shape transformation of the DNA scaffold was extensively studied by means of the fluorescence energy transfer measurement, atomic force microscope images, and agarose gel electrophoretic analyses. A dimeric enzyme xylitol dehydrogenase was assembled on the DNA scaffold in its open state in a high-loading yield. The enzyme loaded on the scaffold was subsequently transformed to its closed state by the addition of short DNA closing keys. The enzyme encapsulated in the closed state displayed comparable activity to that in the open state, ensuring that the catalytic activity of the enzyme was well maintained in the DNA nanocarrier. The nanocarrier with efficient encapsulation ability is potentially applicable for drug delivery, biosensing, biocatalytic, and diagnostic tools.

Oral candidiasis, a common opportunistic infection of the oral cavity, is mainly caused by the following four Candida species (in decreasing incidence rate): Candida albicans, Candida glabrata, Candida tropicalis, and Candida krusei. This study offers in-depth Raman spectroscopy analyses of these species and proposes procedures for an accurate and rapid identification of oral yeast species. We first obtained average spectra for different Candida species and systematically analyzed them in order to decode structural differences among species at the molecular scale. Then, we searched for a statistical validation through a chemometric method based on principal component analysis (PCA). This method was found only partially capable to mechanistically distinguish among Candida species. We thus proposed a new Raman barcoding approach based on an algorithm that converts spectrally deconvoluted Raman sub-bands into barcodes. Barcode-assisted Raman analyses could enable on-site identification in nearly real-time, thus implementing preventive oral control, enabling prompt selection of the most effective drug, and increasing the probability to interrupt disease transmission.

Despite numerous treatment methods, there is no gold standard for the treatment of peri-implantitis—an infectious peri-implant disease. Here, we examined selenium nanoparticles (SeNPs) at a wide range of concentrations to investigate their cytotoxicity, regulation of osteoblastic differentiation, and assessed the antibacterial effect against Porphyromonas gingivalis. SeNPs (mean size: 70 nm; shape: near-spherical; concentration: 0–2048 ppm) were tested against the MC3T3-E1 osteoblast precursor cell line and P. gingivalis red complex pathogen. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) analysis was used to evaluate the bone morphogenetic protein 2 (BMP-2) signaling pathway. SeNPs at concentrations of 2–16 ppm showed no obvious cytotoxicity and promoted good mineralization and calcification. SeNPs at concentrations 64 ppm and below influenced gene expression promoting osteoblastic differentiation, whereas at high concentrations inhibited the expression of Runt-related transcription factor 2 (Runx2). The growth of P. gingivalis was significantly inhibited at SeNP concentrations of more than 4 ppm. SeNPs at low concentrations promoted osteoblastic differentiation while strongly inhibiting peri-implantitis pathogen growth. This study represents one of the few in vitro assessments of SeNPs against a red complex pathogen and the regulatory effect on osteoblastic differentiation. The findings demonstrate SeNPs could potentially be used for future application on implant coating.

A contribution of the cholinergic system to immune cell function has been suggested, though the role of nicotine and its receptors in T cells, especially regulatory T (Treg) cells, is unclear. We herein investigated the expression and function of nicotinic acetylcholine receptors (nAChRs) in murine-induced Treg (iTreg) cells. Upon differentiation of naive BALB/c T cells into iTreg cells and other T-cell subsets, the effect of nicotine on cytokine production and proliferation of iTreg cells was examined. The expression of nAChRs and its regulatory mechanisms were comparatively analyzed among T-cell subsets. Stimulation-induced transforming growth factor-β1 (TGF-β1) production of iTreg cells was suppressed by nicotine, whereas interleukin (IL)-10 production and proliferation was not affected. α2-, α5-, α9-, and β2-nAChRs were differentially expressed in naive, Th1, Th2, Th9, Th17, and iTreg cells. Among these cell types, the α9-nAChR was particularly upregulated in iTreg cells via its gene promoter, but not through tri-methylation at the 4th lysine residue of the histone H3-dependent mechanisms. We conclude that the immunoregulatory role of Treg cells is modified by the cholinergic system, probably through the characteristic expression of nAChRs.

Stem cell-based therapies and experimental methods rely on efficient induction of human pluripotent stem cells (hPSCs). During limb development, the lateral plate mesoderm (LPM) produces limb-bud mesenchymal (LBM) cells that differentiate into osteochondroprogenitor cells and form cartilage tissues in the appendicular skeleton. Previously, we generated PRRX1-tdTomato reporter hPSCs to establish the protocol for inducing the hPSC-derived PRRX1+ LBM-like cells. However, surface antigens that assess the induction efficiency of hPSC-derived PRRX1+ LBM-like cells from LPM have not been identified. Here, we used PRRX1-tdTomato reporter hPSCs and found that high pluripotent cell density suppressed the expression of PRRX1 mRNA and tdTomato after LBM-like induction. RNA sequencing and flow cytometry suggested that PRRX1-tdTomato+ LBM-like cells are defined as CD44high CD140Bhigh CD49f−. Importantly, other hPSC lines, including four human induced pluripotent stem cell lines (414C2, 1383D2, HPS1042, HPS1043) and two human embryonic stem cell lines (SEES4, SEES7), showed the same results. Thus, an appropriate cell density of hPSCs before differentiation is a prerequisite for inducing the CD44high CD140Bhigh CD49f− PRRX1+ LBM-like cells.

Over the past 24 years, the AErosol RObotic NETwork (AERONET) program has provided highly accurate remote-sensing characterization of aerosol optical and physical properties for an increasingly extensive geographic distribution including all continents and many oceanic island and coastal sites. The measurements and retrievals from the AERONET global network have addressed satellite and model validation needs very well, but there have been challenges in making comparisons to similar parameters from in situ surface and airborne measurements. Additionally, with improved spatial and temporal satellite remote sensing of aerosols, there is a need for higher spatial-resolution ground-based remote-sensing networks. An effort to address these needs resulted in a number of field campaign networks called Distributed Regional Aerosol Gridded Observation Networks (DRAGONs) that were designed to provide a database for in situ and remote-sensing comparison and analysis of local to mesoscale variability in aerosol properties. This paper describes the DRAGON deployments that will continue to contribute to the growing body of research related to meso- and microscale aerosol features and processes. The research presented in this special issue illustrates the diversity of topics that has resulted from the application of data from these networks.

Polyphosphate (polyP), a biopolymer of inorganic phosphate, is widely distributed in living organisms. In platelets, polyP is released upon activation and plays important roles in coagulation and tissue regeneration. However, the lack of a specific quantification method has delayed the in-depth study of polyP. The fluorescent dye 4′,6-diamidine-2-phenylindole dihydrochloride (DAPI) has recently received attention as a promising probe for the visualization and quantification of cellular polyP levels. In this study, we further optimized quantification conditions and applied this protocol in quantification of platelet polyP levels in a Japanese population. Blood samples were collected from non-smoking, healthy Japanese subjects (23 males, 23 females). Washed platelets were fixed and probed with DAPI for fluorometric determination. PolyP levels per platelet count were significantly higher in women than that in men. A moderate negative correlation between age and polyP levels was found in women. Responsiveness to CaCl2 stimulation was also significantly higher in women than that in men. Overall, our optimized protocol requires neither purification nor degradation steps, reducing both the time and bias for reproducible quantification. Thus, we suggest that despite its low specificity, this DAPI-based protocol would be useful in routine laboratory testing to quantify platelet polyP levels efficiently and economically.

To overcome the limitations of the Limulus amebocyte lysate (LAL) assay method for the diagnosis of invasive fungal infection, we applied a reaction system combining recombinant β-glucan binding proteins and a scanning single-molecule counting (SSMC) method. A novel (1→3)-β-D-glucan recognition protein (S-BGRP) and a (1→6)-β-glucanase mutant protein were prepared and tested for the binding of (1→6)-branched (1→3)-β-D-glucan from fungi. S-BGRP and (1→6)-β-glucanase mutant proteins reacted with β-glucan from Candida and Aspergillus spp. Although LAL cross-reacted with plant-derived β-glucans, the new detection system using the SSMC method showed low sensitivity to plant (1→3)-β-D-glucan, which significantly improved the appearance of false positives, a recognized problem with the LAL method. Measurement of β-glucan levels by the SSMC method using recombinant β-glucan-binding proteins may be useful for the diagnosis of fungal infections. This study shows that this detection system could be a new alternative diagnostic method to the LAL method.

Aerosol mass concentrations are affected by local emissions as well as long-range transboundary (LRT) aerosols. This work investigates regional and local variations of aerosols based on Distributed Regional Aerosol Gridded Observation Networks (DRAGON). We constructed DRAGON-Japan and DRAGON-Osaka in spring of 2012. The former network covers almost all of Japan in order to obtain aerosol information in regional scale over Japanese islands. It was determined from the DRAGON-Japan campaign that the values of aerosol optical thickness (AOT) decrease from west to east during an aerosol episode. In fact, the highest AOT was recorded at Fukue Island at the western end of the network, and the value was much higher than that of urban areas. The latter network (DRAGON-Osaka) was set as a dense instrument network in the megalopolis of Osaka, with a population of 12 million, to better understand local aerosol dynamics in urban areas. AOT was further measured with a mobile sun photometer attached to a car. This transect information showed that aerosol concentrations rapidly changed in time and space together when most of the Osaka area was covered with moderate LRT aerosols. The combined use of the dense instrument network (DRAGON-Osaka) and high-frequency measurements provides the motion of aerosol advection, which coincides with the wind vector around the layer between 700 and 850 hPa as provided by the reanalysis data of the National Centers for Environmental Prediction (NCEP).

Central nervous system tumor with BCL6-corepressor internal tandem duplication (CNS-BCOR ITD) is a malignant entity characterized by recurrent alterations in exon 15 encoding the essential binding domain for the polycomb repressive complex (PRC). In contrast to deletion or truncating mutations seen in other tumors, BCOR expression is upregulated in CNS-BCOR ITD, and a distinct oncogenic mechanism has been suggested. However, the effects of this change on the biology of neuroepithelial cells is poorly understood. In this study, we introduced either wildtype BCOR or BCOR-ITD into human and murine neural stem cells and analyzed them with quantitative RT-PCR and RNA-sequencing, as well as growth, clonogenicity, and invasion assays. In human cells, BCOR-ITD promoted derepression of PRC2-target genes compared to wildtype BCOR. A similar effect was found in clinical specimens from previous studies. However, no growth advantage was seen in the human neural stem cells expressing BCOR-ITD, and long-term models could not be established. In the murine cells, both wildtype BCOR and BCOR-ITD overexpression affected cellular differentiation and histone methylation, but only BCOR-ITD increased cellular growth, invasion, and migration. BCOR-ITD overexpression drives transcriptional changes, possibly due to altered PRC function, and contributes to the oncogenic transformation of neural precursors.

The non-adiabatic dynamics of furan excited in the ππ* state (S2 in the Franck–Condon geometry) was studied using non-adiabatic molecular dynamics simulations in connection with an ensemble density functional method. The time-resolved photoelectron spectra were theoretically simulated in a wide range of electron binding energies that covered the valence as well as the core electrons. The dynamics of the decay (rise) of the photoelectron signal were compared with the excited-state population dynamics. It was observed that the photoelectron signal decay parameters at certain electron binding energies displayed a good correlation with the events occurring during the excited-state dynamics. Thus, the time profile of the photoelectron intensity of the K-shell electrons of oxygen (decay constant of 34 ± 3 fs) showed a reasonable correlation with the time of passage through conical intersections with the ground state (47 ± 2 fs). The ground-state recovery constant of the photoelectron signal (121 ± 30 fs) was in good agreement with the theoretically obtained excited-state lifetime (93 ± 9 fs), as well as with the experimentally estimated recovery time constant (ca. 110 fs). Hence, it is proposed to complement the traditional TRPES observations with the trXPS (or trNEXAFS) measurements to obtain more reliable estimates of the most mechanistically important events during the excited-state dynamics.

Considering the widespread occurrence of oxalate in nature and its broad impact on a host of organisms, it is surprising that so little is known about the turnover of this important acid. In plants, oxalate oxidase is the most well-studied enzyme capable of degrading oxalate, but not all plants possess this activity. Recently, acyl-activating enzyme 3 (AAE3), encoding an oxalyl-CoA synthetase, was identified in Arabidopsis. This enzyme has been proposed to catalyze the first step in an alternative pathway of oxalate degradation. Since this initial discovery, this enzyme and proposed pathway have been found to be important to other plants and yeast as well. In this study, we identify, in Arabidopsis, an oxalyl-CoA decarboxylase (AtOXC) that is capable of catalyzing the second step in this proposed pathway of oxalate catabolism. This enzyme breaks down oxalyl-CoA, the product of AtAAE3, into formyl-CoA and CO2. AtOXC:GFP localization suggested that this enzyme functions within the cytosol of the cell. An Atoxc knock-down mutant showed a reduction in the ability to degrade oxalate into CO2. This reduction in AtOXC activity resulted in an increase in the accumulation of oxalate and the enzyme substrate, oxalyl-CoA. Size exclusion studies suggest that the enzyme functions as a dimer. Computer modeling of the AtOXC enzyme structure identified amino acids of predicted importance in co-factor binding and catalysis. Overall, these results suggest that AtOXC catalyzes the second step in this alternative pathway of oxalate catabolism.

Noble metal nanodots have been applied to plasmonic devices, catalysts, and highly sensitive detection in bioinstruments. We have been studying the fabrications of them through a laser-induced dot transfer (LIDT) technique, a type of laser-induced forward transfer (LIFT), in which nanodots several hundred nm in diameter are produced via a solid–liquid–solid (SLS) mechanism. In the previous study, an interference laser processing technique was applied to LIDT, and aligned Au nanodots were successfully deposited onto an acceptor substrate in a single shot of femtosecond laser irradiation. In the present experiment, Pt thin film was applied to this technique, and the deposited nanodots were measured by scanning electron microscopy (SEM) and compared with the Au nanodots. A typical nanodot had a roundness fr=0.98 and circularity fcirc=0.90. Compared to the previous experiment using Au thin film, the size distribution was more diffuse, and it was difficult to see the periodic alignment of the nanodots in the parameter range of this experiment. This method is promising as a method for producing large quantities of Pt particles with diameters of several hundred nm.

Synovial mesenchymal stem cell (SMSC) is the promising cell source of cartilage regeneration but has several issues to overcome such as limited cell proliferation and heterogeneity of cartilage regeneration ability. Previous reports demonstrated that basic fibroblast growth factor (bFGF) can promote proliferation and cartilage differentiation potential of MSCs in vitro, although no reports show its beneficial effect in vivo. The purpose of this study is to investigate the promoting effect of bFGF on cartilage regeneration using human SMSC in vivo. SMSCs were cultured with or without bFGF in a growth medium, and 2 × 105 cells were aggregated to form a synovial pellet. Synovial pellets were implanted into osteochondral defects induced in the femoral trochlea of severe combined immunodeficient mice, and histological evaluation was performed after eight weeks. The presence of implanted SMSCs was confirmed by the observation of human vimentin immunostaining-positive cells. Interestingly, broad lacunae structures and cartilage substrate stained by Safranin-O were observed only in the bFGF (+) group. The bFGF (+) group had significantly higher O’Driscoll scores in the cartilage repair than the bFGF (−) group. The addition of bFGF to SMSC growth culture may be a useful treatment option to promote cartilage regeneration in vivo.

Aging is a major risk factor of osteoarthritis, which is characterized by the degeneration of articular cartilage. CCN3, a member of the CCN family, is expressed in cartilage and has various physiological functions during chondrocyte development, differentiation, and regeneration. Here, we examine the role of CCN3 in cartilage maintenance. During aging, the expression of Ccn3 mRNA in mouse primary chondrocytes from knee cartilage increased and showed a positive correlation with p21 and p53 mRNA. Increased accumulation of CCN3 protein was confirmed. To analyze the effects of CCN3 in vitro, either primary cultured human articular chondrocytes or rat chondrosarcoma cell line (RCS) were used. Artificial senescence induced by H2O2 caused a dose-dependent increase in Ccn3 gene and CCN3 protein expression, along with enhanced expression of p21 and p53 mRNA and proteins, as well as SA-β gal activity. Overexpression of CCN3 also enhanced p21 promoter activity via p53. Accordingly, the addition of recombinant CCN3 protein to the culture increased the expression of p21 and p53 mRNAs. We have produced cartilage-specific CCN3-overexpressing transgenic mice, and found degradative changes in knee joints within two months. Inflammatory gene expression was found even in the rib chondrocytes of three-month-old transgenic mice. Similar results were observed in human knee articular chondrocytes from patients at both mRNA and protein levels. These results indicate that CCN3 is a new senescence marker of chondrocytes, and the overexpression of CCN3 in cartilage may in part promote chondrocyte senescence, leading to the degeneration of articular cartilage through the induction of p53 and p21.

The skin loses its moisture with advancing age, causing cosmetic issues such as wrinkles. In addition, the loss of moisture leads to hypersensitivity to external stimuli such as UV light. Transcutaneous supplementation with hyaluronic acid (HA) is an effective and safe method of recovering the moisturizing function and elasticity of the skin. However, the transcutaneous delivery of HA remains challenging owing to the barrier function of the stratum corneum (SC) layer. To penetrate the SC barrier, we used a reverse micelle formulation that does not require high energy consumption processes for preparation. We aimed to enhance the skin permeability of HA by incorporating glyceryl monooleate—a skin permeation enhancer—into the formulation. A fluorescently-labeled HA-loaded reverse micelle formulation showed significantly enhanced permeation across Yucatan micro pig skin. Fourier transform infra-red spectroscopy of the surface of the skin treated with the reverse micelle formulation showed blue shifts of the CH2 symmetric/asymmetric stretching peaks, indicating a reduction in the barrier function of the SC. Further study revealed that HA was released from the reverse micelles at the hydrophobic/hydrophilic interface between the SC and the living epidermis. The results demonstrated that our reverse micellar system is an easy-to-prepare formulation for the effective transcutaneous delivery of HA.