Biomedical subjects

Tetsuya Ishikawa

Publications and source records attributed to Tetsuya Ishikawa.

10 recordsLinked to original sources

PubMed2003

Delivery of a growth factor fusion protein having collagen-binding activity to wound tissues.

Recently, we established a collagen-binding growth factor consisting of epidermal growth factor and the fibronectin collagen-binding domain (FNCBD-EGF). FNCBD-EGF is a biologically active fusion protein that could stably bind to collagen materials, and exert its growth factor activity even after collagen binding. In this study, we investigated the concept that FNCBD moiety with high collagen affinity may enhance the effective local concentration of EGF at the site of administration in the following tissues: skin wounds, catheter-injured arteries, and hind limb muscles. In an animal model of impaired wound healing, application of FNCBD-EGF in combination with collagen gel induced granulation tissue formation in the wounds due to its sustained retention. In the injured artery, infused FNCBD-EGF remained bound to collagen exposed on the injured tissues even after blood circulation was restored. Injection of the fusion protein into the hind limbs revealed that our delivery system was effective for direct administration to muscular tissue.

Administration, Topical↗

PubMed2003

Lack of tumor necrosis factor alpha induces impaired proliferation of hepatitis B virus-specific cytotoxic T lymphocytes.

Recent studies have shown that tumor necrosis factor alpha (TNF-alpha) plays critical roles in not only viral clearance but also lymphoid tissue development and stem cell differentiation. In this study, we attempted to induce hepatitis B virus (HBV)-specific cytotoxic T lymphocytes (CTLs) by immunization of TNF-alpha knockout (TNF-alpha(-/-)) mice with HBsAg-encoding plasmid DNA. An immunization with the HBV plasmid failed to induce CTL responses in TNF-alpha(-/-) mice, although CTLs were readily induced in wild-type mice by the same protocol. Weak CTL responses were produced in TNF-alpha(-/-) mice after two sessions of immunization with the HBV plasmid; however, TNF-alpha was required to maintain the responses of these CTL lines to in vitro stimulation and, even then, the responses were lost after 3 weeks. Interestingly, a limiting dilution of a CTL line showed that HBV-specific CTL clones with high specific cytotoxicity were present in TNF-alpha(-/-) mice, but these clones again failed to proliferate for more than 3 weeks. Furthermore, since exogenously added TNF-alpha enhanced the proliferation of a TNF-alpha(-/-) clone but suppressed that of a TNF-alpha(+/+) clone in vitro, TNF-alpha also has a direct effect on the proliferation of CTLs. In conclusion, TNF-alpha is essential rather than important for the proliferation of HBV-specific CTLs both in vivo and in vitro and this effect is not only due to the activation of dendritic cells but is also induced by the direct effect on CTLs.

Animals↗

PubMed2002

Imaging whole Escherichia coli bacteria by using single-particle x-ray diffraction.

We report the first experimental recording, to our knowledge, of the diffraction pattern from intact Escherichia coli bacteria using coherent x-rays with a wavelength of 2 A. By using the oversampling phasing method, a real space image at a resolution of 30 nm was directly reconstructed from the diffraction pattern. An R factor used for characterizing the quality of the reconstruction was in the range of 5%, which demonstrated the reliability of the reconstruction process. The distribution of proteins inside the bacteria labeled with manganese oxide has been identified and this distribution confirmed by fluorescence microscopy images. Compared with lens-based microscopy, this diffraction-based imaging approach can examine thicker samples, such as whole cultured cells, in three dimensions with resolution limited only by radiation damage. Looking forward, the successful recording and reconstruction of diffraction patterns from biological samples reported here represent an important step toward the potential of imaging single biomolecules at near-atomic resolution by combining single-particle diffraction with x-ray free electron lasers.

Escherichia coli↗

PubMed2002

Nearly diffraction-limited line focusing of a hard-X-ray beam with an elliptically figured mirror.

An elliptical mirror for X-ray microfocusing was manufactured using the new fabrication methods of elastic emission machining and plasma chemical vaporization machining. Surface profiles measured using stitching interferometry showed a maximum deviation around the ideal figure of 7 nm peak-to-valley. The mirror showed nearly diffraction-limited focusing performance, with a 200 nm line width at the focus. Wave-optical calculations, taking the measured surface profile into consideration, reproduced well the measured focusing properties both at and around the beam waist.

Journal Article↗

PubMed2002

High resolution 3D x-ray diffraction microscopy.

We have imaged a 2D buried Ni nanostructure at 8 nm resolution using coherent x-ray diffraction and the oversampling phasing method. By employing a 3D imaging reconstruction algorithm, for the first time we have experimentally determined the 3D structure of a noncrystalline nanostructured material at 50 nm resolution. The 2D and 3D imaging resolution is currently limited by the exposure time and the computing power, while the ultimate resolution is limited by the x-ray wavelengths. We believe these results pave the way for the development of atomic resolution 3D x-ray diffraction microscopy.

Microscopy↗

PubMed2002

The Goos-Hänchen effect at Bragg diffraction.

The strong incident-angle dependence of the phase of complex reflectivity causes a shift of the reflected beam from the geometrically expected path. This effect, known as the Goos-Hänchen effect in the visible region, was observed for Bragg-case diffraction in the hard X-ray region. The shift was found to be in good agreement with the theory.

Journal Article↗

PubMed2002

Functional analysis of the 5'-flanking region of the human alanine:glyoxylate aminotransferase gene AGXT.

Primer extension of human liver poly(A)(+) RNA revealed that the main transcription start site of the human alanine:glyoxylate aminotransferase gene (AGXT) is situated near 45 bp upstream from the translation start site. Deletion analysis using the 1203 bp 5'-flanking region of the AGXT gene and a luciferase reporter suggested that the promoter sequence is most likely located 2-325 bp upstream from the translation start site, possibly with enhancer elements 440-700 bp upstream. It was also suggested that the region -2 to -64 is important for the expression of the AGXT gene. The region -2 to -325 has two TATA boxes and some initiator elements.

Base Sequence↗

PubMed2002

Fast multigrid fluorescent ion chamber with 0.1 ms time response.

A fast multigrid ion chamber for the detection of fluorescent X-rays has been developed. The structure of 17 grids with close separation was employed to maximize the time response as well as to give sufficient detection efficiency. The measured rise/fall response time to cyclic X-rays was shorter than that of an existing three-grid ion chamber by more than one order of magnitude. A 0.13 ms time response was obtained at the 500 V applied voltage, where the detector can stably operate without any discharge. The available frequency range is as high as 1 kHz with a practical amplitude response.

Journal Article↗

PubMed2002

Optical switching of X-rays using laser-induced lattice expansion.

The optical switching of X-rays using laser-induced crystal lattice expansion is described. Irradiation of a gallium arsenide (GaAs) crystal using picosecond laser pulses shifts the Bragg angle through the lattice expansion with a response time of a few hundred picoseconds. A single pulse was extracted from the synchrotron radiation pulse train using a double-crystal arrangement of GaAs, in which the two crystals were irradiated by way of two successive laser pulses with an appropriate time delay.

Journal Article↗

PubMed2002

A new type of antimetastatic peptide derived from fibronectin.

PURPOSE: We found previously that fibronectin (FN) has a cryptic functional site (YTIYVIAL sequence within the 14th type III repeat) opposing cell adhesion to extracellular matrix. A 22-mer FN peptide containing this site, termed FNIII14, inhibits beta1 integrin-mediated adhesion without binding to integrins. The present study shows that FNIII14 has the potential to prevent lymphoma cell metastasis. EXPERIMENTAL DESIGN: Antimetastatic effect of FNIII14 has been evaluated through in vitro or in vivo experiments. RESULTS: FNIII14 inhibited the integrin alpha4beta1-mediated B lymphoma Ramos cell adhesion to VCAM-1 on venule endothelial cells, as well as to FN. Murine T lymphoma L5178Y-ML25 cells, which are known to metastasize to liver and spleen, preferentially adhered to vitronectin (VN) and migrated toward VN concentration gradients. FNIII14 abrogated both the integrin alphavbeta3-mediated adhesion and migration of L5178Y-ML25 cells. Inhibition of the alphavbeta3mediated L5178Y-ML25 cell adhesion by FNIII14 was reversed by phenylarsine oxide, a protein tyrosine phosphatase inhibitor. In addition, FNIII14 abrogated the VN-stimulated tyrosine phosphorylation of intracellular signaling proteins, including focal adhesion kinase (p125(FAK)) and paxillin, suggesting that such a diversity of FNIII14 effects might be because of the negative regulation of p125(FAK) and paxillin tyrosine phosphorylation, which has been involved in adhesion signals transduced by different integrins. The in vivo experiment using a murine metastasis model showed that FNIII14 would inhibit liver and spleen metastases of L5178Y-ML25 cells at a dose much lower than that of RGDS. CONCLUSIONS: FNIII14 might be applicable as a new type of antimetastatic agent distinct from integrin-binding peptides.

Animals↗