Yoshinobu Manome

Downregulation of survivin expression by induction of the effector cell protease receptor-1 reduces tumor growth potential and results in an increased sensitivity to anticancer agents in human colon cancer.

Survivin, a novel inhibitor of apoptosis, is expressed in cancer cells and not in normal adult tissues, and is recognised as a potential target in anticancer therapy. The induction of a natural antisense of survivin, effector cell protease receptor-1 (EPR-1), in a human colon cancer cell line resulted in a downregulation of survivin expression, with a similar decrease in cell proliferation, an increase in apoptosis and an increase in the sensitivity to anticancer agents. In addition, subcutaneous (s.c.) tumours from EPR-1 transfectants showed a significant reduction in size compared with parental cells, and this antitumour efficacy was further enhanced in combination with anticancer agents. These findings suggest that regulation of survivin by the induction of EPR-1 cDNA may have significant potential as a therapy for human colon cancer.

Cell-Based in Vitro Blood–Brain Barrier Model Can Rapidly Evaluate Nanoparticles’ Brain Permeability in Association with Particle Size and Surface Modification

The possibility of nanoparticle (NP) uptake to the human central nervous system is a major concern. Recent reports showed that in animal models, nanoparticles (NPs) passed through the blood–brain barrier (BBB). For the safe use of NPs, it is imperative to evaluate the permeability of NPs through the BBB. Here we used a commercially available in vitro BBB model to evaluate the permeability of NPs for a rapid, easy and reproducible assay. The model is reconstructed by culturing both primary rat brain endothelial cells and pericytes to support the tight junctions of endothelial cells. We used the permeability coefficient (Papp) to determine the permeability of NPs. The size dependency results, using fluorescent silica NPs (30, 100, and 400 nm), revealed that the Papp for the 30 nm NPs was higher than those of the larger silica. The surface charge dependency results using Qdots® (amino-, carboxyl-, and PEGylated-Qdots), showed that more amino-Qdots passed through the model than the other Qdots. Usage of serum-containing buffer in the model resulted in an overall reduction of permeability. In conclusion, although additional developments are desired to elucidate the NPs transportation, we showed that the BBB model could be useful as a tool to test the permeability of nanoparticles.

Size-Tunable Silicon/Iron Oxide Hybrid Nanoparticles with Fluorescence, Superparamagnetism, and Biocompatibility

Magnetic/fluorescent composite materials have become one of the most important tools in the imaging modality in vivo using magnetic resonance imaging (MRI) monitoring and fluorescence optical imaging. We report herein on a simplified procedure to synthesize hybrid nanoparticles (HNPs) that combine silicon and magnetic iron oxides consisting of magnetite (Fe(3)O(4)) and maghemite (γ-Fe(2)O(3)). Intriguingly, our unique synthetic approach can control magnetic and optical behaviors by reducing the particle size, demonstrating that the HNPs with the mean diameter of 3.0 nm exhibit superparamagnetic behavior and green fluorescence in an aqueous solution, ambient air, and a cellular environment, whereas the HNPs with the mean diameter more than 5.0 nm indicate ferromagnetic behavior without fluorescence. Additionally, both HNPs with different diameters possess excellent magnetic responsivity for external applied magnetic field and good biocompatibility due to the low cytotoxicity. Our biocompatible HNPs with the superparamagnetism can provide an attractive approach for diagnostic imaging system in vivo.

Deficiency of Chemokine Receptor CCR1 Causes Osteopenia Due to Impaired Functions of Osteoclasts and Osteoblasts

Chemokines are characterized by the homing activity of leukocytes to targeted inflammation sites. Recent research indicates that chemokines play more divergent roles in various phases of pathogenesis as well as immune reactions. The chemokine receptor, CCR1, and its ligands are thought to be involved in inflammatory bone destruction, but their physiological roles in the bone metabolism in vivo have not yet been elucidated. In the present study, we investigated the roles of CCR1 in bone metabolism using CCR1-deficient mice. Ccr1−/− mice have fewer and thinner trabecular bones and low mineral bone density in cancellous bones. The lack of CCR1 affects the differentiation and function of osteoblasts. Runx2, Atf4, Osteopontin, and Osteonectin were significantly up-regulated in Ccr1−/− mice despite sustained expression of Osterix and reduced expression of Osteocalcin, suggesting a lower potential for differentiation into mature osteoblasts. In addition, mineralized nodule formation was markedly disrupted in cultured osteoblastic cells isolated from Ccr1−/− mice. Osteoclastogenesis induced from cultured Ccr1−/− bone marrow cells yielded fewer and smaller osteoclasts due to the abrogated cell-fusion. Ccr1−/− osteoclasts exerted no osteolytic activity concomitant with reduced expressions of Rank and its downstream targets, implying that the defective osteoclastogenesis is involved in the bone phenotype in Ccr1−/− mice. The co-culture of wild-type osteoclast precursors with Ccr1−/− osteoblasts failed to facilitate osteoclastogenesis. This finding is most likely due to a reduction in Rankl expression. These observations suggest that the axis of CCR1 and its ligands are likely to be involved in cross-talk between osteoclasts and osteoblasts by modulating the RANK-RANKL-mediated interaction.

Chemical Identity of a Rotting Animal-Like Odor Emitted from the Inflorescence of the Titan Arum (Amorphophallus titanum)

The titan arum, Amorphophallus titanum, is a flowering plant with the largest inflorescence in the world. The flower emits a unique rotting animal-like odor that attracts insects for pollination. To determine the chemical identity of this characteristic odor, we performed gas chromatography-mass spectrometry-olfactometry analysis of volatiles derived from the inflorescence. The main odorant causing the smell during the flower-opening phase was identified as dimethyl trisulfide, a compound with a sulfury odor that has been found to be emitted from some vegetables, microorganisms, and cancerous wounds.

Tetrahydrouridine inhibits cell proliferation through cell cycle regulation regardless of cytidine deaminase expression levels.

Tetrahydrouridine (THU) is a well characterized and potent inhibitor of cytidine deaminase (CDA). Highly expressed CDA catalyzes and inactivates cytidine analogues, ultimately contributing to increased gemcitabine resistance. Therefore, a combination therapy of THU and gemcitabine is considered to be a potential and promising treatment for tumors with highly expressed CDA. In this study, we found that THU has an alternative mechanism for inhibiting cell growth which is independent of CDA expression. Three different carcinoma cell lines (MIAPaCa-2, H441, and H1299) exhibited decreased cell proliferation after sole administration of THU, while being unaffected by knocking down CDA. To investigate the mechanism of THU-induced cell growth inhibition, cell cycle analysis using flow cytometry was performed. This analysis revealed that THU caused an increased rate of G1-phase occurrence while S-phase occurrence was diminished. Similarly, Ki-67 staining further supported that THU reduces cell proliferation. We also found that THU regulates cell cycle progression at the G1/S checkpoint by suppressing E2F1. As a result, a combination regimen of THU and gemcitabine might be a more effective therapy than previously believed for pancreatic carcinoma since THU works as a CDA inhibitor, as well as an inhibitor of cell growth in some types of pancreatic carcinoma cells.

Effects of silica and titanium oxide particles on a human neural stem cell line: morphology, mitochondrial activity, and gene expression of differentiation markers.

Several in vivo studies suggest that nanoparticles (smaller than 100 nm) have the ability to reach the brain tissue. Moreover, some nanoparticles can penetrate into the brains of murine fetuses through the placenta by intravenous administration to pregnant mice. However, it is not clear whether the penetrated nanoparticles affect neurogenesis or brain function. To evaluate its effects on neural stem cells, we assayed a human neural stem cell (hNSCs) line exposed in vitro to three types of silica particles (30 nm, 70 nm, and <44 μm) and two types of titanium oxide particles (80 nm and < 44 μm). Our results show that hNSCs aggregated and exhibited abnormal morphology when exposed to the particles at concentrations ≥ 0.1 mg/mL for 7 days. Moreover, all the particles affected the gene expression of Nestin (stem cell marker) and neurofilament heavy polypeptide (NF-H, neuron marker) at 0.1 mg/mL. In contrast, only 30-nm silica particles at 1.0 mg/mL significantly reduced mitochondrial activity. Notably, 30-nm silica particles exhibited acute membrane permeability at concentrations ≥62.5 μg/mL in 24 h. Although these concentrations are higher than the expected concentrations of nanoparticles in the brain from in vivo experiments in a short period, these thresholds may indicate the potential toxicity of accumulated particles for long-term usage or continuous exposure.

Objective Display and Discrimination of Floral Odors from Amorphophallus titanum, Bloomed on Different Dates and at Different Locations, Using an Electronic Nose

As olfactory perceptions vary from person to person, it is difficult to describe smells objectively. In contrast, electronic noses also detect smells with their sensors, but in addition describe those using electronic signals. Here we showed a virtual connection method between a human nose perceptions and electronic nose responses with the smell of standard gases. In this method, Amorphophallus titanum flowers, which emit a strong carrion smell, could objectively be described using an electronic nose, in a way resembling the skill of sommeliers. We could describe the flower smell to be close to that of a mixture of methyl mercaptan and propionic acid, by calculation of the dilution index from electronic resistances. In other words, the smell resembled that of “decayed cabbage, garlic and pungent sour” with possible descriptors. Additionally, we compared the smells of flowers which bloomed on different dates and at different locations and showed the similarity of odor intensities visually, in standard gas categories. We anticipate our assay to be a starting point for a perceptive connection between our noses and electronic noses.

A Culture Device Demonstrates that Hydrostatic Pressure Increases mRNA of RGS5 in Neuroblastoma and CHC1-L in Lymphocytic Cells

Previous studies demonstrated that mechanical forces affect a wide range of cellular behaviors. These forces regulate important cellular responses in the human body and consist of gravity, hydrostatic pressure, stretch, and shear stress, which is exerted on the vascular system by the passage of blood flow. We reasoned that these forces might be significant and dynamic regulators of cellular functions within the human body. While cellular effects of stretch and shear stress have been studied particularly with endothelial cells, little is known about the effects of gravity and hydrostatic pressure to cells. To examine the direct effect of hydrostatic pressure, we developed a culture device to confer hydrostatic pressures to cells ranging from 0 to 1,000 psi. We subjected human neuroblastoma cells and rIL-2-activated lymphocytes to a constant pressure of 20 or 100 psi for 48 h and attempted to identify genes regulated by hydrostatic pressure. Genes of regulator of G-protein signaling 5 in neuroblastoma cells and CHC1-L in lymphocytes increased after exposure to hydrostatic pressure. The results demonstrated that hydrostatic pressure directly regulates the expression of specific genes in mammalian cells. Moreover, there may be some underlying mechanisms that have common effects in altered physical environments. Our in vitro culture system may provide some insight into the mechanisms through the intracellular processes affected by mechanical forces.

Discrimination Method of the Volatiles from Fresh Mushrooms by an Electronic Nose Using a Trapping System and Statistical Standardization to Reduce Sensor Value Variation

Electronic noses have the benefit of obtaining smell information in a simple and objective manner, therefore, many applications have been developed for broad analysis areas such as food, drinks, cosmetics, medicine, and agriculture. However, measurement values from electronic noses have a tendency to vary under humidity or alcohol exposure conditions, since several types of sensors in the devices are affected by such variables. Consequently, we show three techniques for reducing the variation of sensor values: (1) using a trapping system to reduce the infering components; (2) performing statistical standardization (calculation of z-score); and (3) selecting suitable sensors. With these techniques, we discriminated the volatiles of four types of fresh mushrooms: golden needle (Flammulina velutipes), white mushroom (Agaricus bisporus), shiitake (Lentinus edodes), and eryngii (Pleurotus eryngii) among six fresh mushrooms (hen of the woods (Grifola frondosa), shimeji (Hypsizygus marmoreus) plus the above mushrooms). Additionally, we succeeded in discrimination of white mushroom, only comparing with artificial mushroom flavors, such as champignon flavor and truffle flavor. In conclusion, our techniques will expand the options to reduce variations in sensor values.