Aqueous Two‐Phase System Patterning of Microbubbles: Localized Induction of Apoptosis in Sonoporated Cells

Ultrasound‐driven microbubbles produce mechanical forces that can disrupt cell membranes (sonoporation). However, it is difficult to control microbubble location with respect to cells. This lack of control leads to low sonoporation efficiencies and variable outcomes. In this study, aqueous two‐phase system (ATPS) droplets are used to localize microbubbles in select micro‐regions at the surface of living cells. This is achieved by stably partitioning microbubbles in dextran (DEX) droplets, deposited on living adherent cells in medium containing polyethylene glycol (PEG). The interfacial energy at the PEG‐DEX interface overcomes microbubble buoyancy and prevents microbubbles from floating away from the cells. Spreading of the small DEX droplets retains microbubbles at the cell surface in defined lateral positions without the need for antibody or cell‐binding ligand conjugation. The patterned microbubbles are activated on a cell monolayer exposed to a broadly applied ultrasound field (center frequency 1.25 MHz, active element diameter 0.6 cm, pulse duration 8 μs or 30 s). This system enables efficient testing of different ultrasound conditions for their effects on sonoporation‐mediated membrane disruption and cell viability. Regions of cells without patterned microbubbles show no injury or membrane disruption. In microbubble patterned regions, 8 μs ultrasound pulses (0.2‐0.6 MPa) produce cell death that is primarily apoptotic. Ultrasound‐induced apoptosis increases with higher extracellular calcium concentrations, with cells displaying all of the hallmarks of apoptosis including annexinV labeling, loss of mitochondrial membrane potential, caspase activation and changes in nuclear morphology.     

Enantiomeric distribution of odour-active epoxyaldehydes in yuzu (Citrus junos Sieb. ex Tanaka)

The odour-active epoxyaldehydes cis-4,5-epoxy-(E)-2-decenal (1), trans-4,5-epoxy-(E)-2-decenal (2), and trans-4,5-epoxy-(E,Z)-2,7-decadienal (3) are present in yuzu (Citrus junos Sieb. ex Tanaka) volatile oil. These epoxyaldehydes have two chirality centres at the C4 and C5 positions in the epoxide moiety. To date, little is known about the enantiomeric distributions of the expoxyaldehydes in natural products. In this study, we examined the enantiomeric distributions of the epoxyaldehydes in yuzu volatile oil using multidimensional gas chromatography. The standard substrates used for 1 and 3 were synthesised enantioselectively using the Katsuki-Sharpless epoxidation. Standard substrates used for 2 were made via the asymmetric organocatalytic epoxidation with a proline-derivative catalyst. The 4S,5R- and 4R,5S-enantiomers of 1 were obtained in moderate enantiomeric excess (44–66 % ee), while 4S,5S- and 4R,5R-enantiomers of 2 and 3 were obtained in high enantiomeric excess (2, 90–92 % ee; 3, 96 % ee). Enantioselective multidimensional gas chromatography-mass spectrometry (enantio-MDGC-MS) revealed that the chiral centres at C4 and C5 in the cis-form (1) have the (4R,5S) absolute configuration, while trans-forms (2 and 3) have the (4S,5S) configuration. The finding of the absolute configuration at C5 (S) suggests that the optically active epoxyaldehydes may be formed along with retention of the S configuration at C13 of the lipid hydroperoxides 13(S)-hydroperoxy-9(Z),11(E)-octadecadienoic acid (13S-HPOD) or 13(S)-hydroperoxy-9(Z),11(E),15(Z)-octadecatrienoic acid (13S-HPOT) during enzymatic lipid oxidation. Finally, we propose a pathway for the formation of the optically active epoxyaldehydes in yuzu.     

High-strength superelastic Ti–Ni microtubes fabricated by sputter deposition

Ti–Ni microtubes are attractive materials for biomedical devices, such as micro-catheters and micro-stents, but it is difficult to fabricate them with dimensions of less than 100 μm by conventional tube-drawing. In this study, Ti–Ni microtubes with 50 μm inner diameter and a tube wall thickness of 6 μm was successfully fabricated using a novel method in which Ti–Ni was sputter-deposited on a Cu wire with a diameter of 50 μm. All the microtubes exhibited shape memory behavior after crystallization at 873 K for 3.6 ks. Microtubes fabricated without rotating the Cu wire during deposition have low fracture strength due to the columnar grains and non-uniform tube wall thickness. Microtubes fabricated by depositing Ti–Ni on a rotating wire have a uniform wall thickness and the fracture strength increased with increasing rotation speed. Microtubes made by the rotating-wire method exhibited superelasticity of 3% strain at room temperature with high fracture stress of 950 MPa, suggesting that they are suitable for practical applications.     

Ex Vivo Expanded and Activated Natural Killer Cells Prolong the Overall Survival of Mice with Glioblastoma-like Cell-Derived Tumors

Glioblastoma (GBM) is the leading malignant intracranial tumor and is associated with a poor prognosis. Highly purified, activated natural killer (NK) cells, designated as genuine induced NK cells (GiNKs), represent a promising immunotherapy for GBM. We evaluated the anti-tumor effect of GiNKs in association with the programmed death 1(PD-1)/PD-ligand 1 (PD-L1) immune checkpoint pathway. We determined the level of PD-1 expression, a receptor known to down-regulate the immune response against malignancy, on GiNKs. PD-L1 expression on glioma cell lines (GBM-like cell line U87MG, and GBM cell line T98G) was also determined. To evaluate the anti-tumor activity of GiNKs in vivo, we used a xenograft model of subcutaneously implanted U87MG cells in immunocompromised NOG mice. The GiNKs expressed very low levels of PD-1. Although PD-L1 was expressed on U87MG and T98G cells, the expression levels were highly variable. Our xenograft model revealed that the retro-orbital administration of GiNKs and interleukin-2 (IL-2) prolonged the survival of NOG mice bearing subcutaneous U87MG-derived tumors. PD-1 blocking antibodies did not have an additive effect with GiNKs for prolonging survival. GiNKs may represent a promising cell-based immunotherapy for patients with GBM and are minimally affected by the PD-1/PD-L1 immune evasion axis in GBM.     

New signaling procedures in ATM networks for advance resource reservation

Currently, communication with resource reservation on ATM networks has been required by popularization of multimedia applications. Types of resource reservation are classified into two types according to reservation timing. One type invokes resource reservation just before communication between users. Another type invokes it in advance. In this type, resources can be reserved with communication time periods invoked by a user, well before actual communication. For the former type, several standardization committees, such as the ATM Forum and ITU‐T, have provided some signaling protocols. However, the latter type has no signaling protocol although necessity of this type has increased. This paper presents approaches of this type at first. Next, it proposes new signaling protocols to support this type keeping consistency with the conventional protocols for the former type, and compares proposed protocols. Moreover, it presents mechanisms to reserve resources cooperating with proposed signaling protocols. This revised version was published online in June 2006 with corrections to the Cover Date.     

AlxGa1-xAs三元化合物共沉积研究

在简单盐溶液中添加络合剂－柠檬酸,电共沉积ＡｌｘＧａ１－ｘＡｓ三元化合物,用能谱分析仪进行成分分析,获得化学计量比接近Ａｌ０．３Ｇａ０．７Ａｓ的三元化合物半导体材料。     

Path of ordering under confinement in alloy films

The effects of surface confinement on the kinetics of ordering in fcc alloy films are studied by the Monte Carlo method. Under weak surface confinement, there are two kinds of kinetic path for alloy films with even layer. One is the films directly relaxing to the equilibrium state, the other is the films relaxing to a metastable state first. Even for the same initial atomic configuration, the kinetic path of ordering is randomized, and the possibility that the films relax to a metastable state is independent with the initial atomic configurations. We attribute this randomicity to the competition between surface confinement and chemical ordering. The evolution of the metastable state to the equilibrium state is also simulated. The results indicate that an alloy film with weak surface segregation can easily be trapped into metastable state.     

Formation of molecular doping patterns in organic–inorganic hybrid films by a capillary electrophoresis doping technique

A new technique is proposed for the fabrication of fine patterns of molecular doping in organic–inorganic hybrid materials by the combination of capillary electrophoresis doping (CED) and photolithography. The UV-induced polymerization of bonds in organic groups yields a fine contrast of structures with the desired pattern in organic–inorganic hybrid films, and CED treatment introduces functional molecules only into unirradiated regions to form the doping patterns of molecules inside the films. The fine patterning of rhodamine-6G doping with from 2 to 4 μm resolution is demonstrated in hybrid films of 10 μm thickness.     

Elevated Alpha 1(I) to Alpha 2(I) Collagen Ratio in Dermal Fibroblasts Possibly Contributes to Fibrosis in Systemic Sclerosis

Systemic sclerosis (SSc) is characterized by excessive collagen deposition in the skin and internal organs. Activated fibroblasts are the key effector cells for the overproduction of type I collagen, which comprises the α1(I) and α2(I) chains encoded by COL1A1 and COL1A2, respectively. In this study, we examined the expression patterns of α1(I) and α2(I) collagen in SSc fibroblasts, as well as their co-regulation with each other. The relative expression ratio of COL1A1 to COL1A2 in SSc fibroblasts was significantly higher than that in control fibroblasts. The same result was observed for type I collagen protein levels, indicating that α2(I) collagen is more elevated than α2(I) collagen. Inhibition or overexpression of α1(I) collagen in control fibroblasts affected the α2(I) collagen levels, suggesting that α1(I) collagen might act as an upstream regulator of α2(I) collagen. The local injection of COL1A1 small interfering RNA in a bleomycin-induced SSc mouse model was found to attenuate skin fibrosis. Overall, our data indicate that α2(I) collagen is a potent regulator of type I collagen in SSc; further investigations of the overall regulatory mechanisms of type I collagen may help understand the aberrant collagen metabolism in SSc.     

A pseudo-mastication sound presentation device to improve the texture of nursing care foods

The texture of foods is affected by concurrent auditory sensations. To improve the texture of nursing care foods, we developed a pseudo-mastication sound presentation device based on electromyogram (EMG) signals from the muscles of mastication. EMG signals have enabled us to promptly present care recipients with pseudo-mastication sounds. However, actual mastication sounds vary in intensity and duration more than EMG signals. Here, we investigated changes in EMG signals and actual mastication sounds during the mastication of two food types (rice crackers and Japanese pickles) to improve our device. We found that the intensity and duration of mastication sounds decreased as the number of mastication strokes increased. Furthermore, the intensity and duration of mastication sounds and the latency between the onset of EMG signals and the onset of mastication sounds also varied by food type. For EMG signals, only the intensity varied by food type. Based on our findings, we modified our pseudo-mastication sound presentation device to enable control of the intensity and duration of pseudo-mastication sounds based on the number of mastication strokes and food type. Reproducing more natural pseudo-mastication sounds can improve care recipients’ motivation for ingesting nursing care foods, thus preventing malnutrition and frailty.