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Toshio Ohtani

Publications and source records attributed to Toshio Ohtani.

14 recordsLinked to original sources
PubMed2003

Transfer-printing of highly aligned DNA nanowires.

We developed a simple method of reproducibly creating highly aligned DNA nanowires without any surface modifications or special equipment. Stretched DNA molecules initially present on the PDMS sheet were transferred onto another surface using transfer-printing (TP). Fluorescent microscopic and atomic force microscopic images revealed that many DNA molecules were highly aligned on surfaces after TP. Furthermore, it was also possible to realize the two-dimensional assembly of DNA nanowires by repeating TP.

Bacteriophage lambda↗
PubMed2003

Atomic force microscopy study of chromosome surface structure changed by protein extraction.

We applied atomic force microscopy (AFM) to investigate the surface structure of barley chromosome in combination with a chemical treatment method. As a result, we have obtained high-resolution topographic images of granular structures with a diameter of ca. 50 nm on the surface of critical-point dried metaphase chromosomes. Treatment with 2M NaCl significantly modified the chromosome surface structure: surface roughness was increased and chromosome thickness was decreased. The NaCl treatment extracted two major proteins with molecular weights of 4000 and 20,000 Da. These proteins might be belonging to non-histone protein families that do not contain any aromatic amino acid. The results demonstrate the advantage of the combined method of high-resolution AFM imaging and chemical treatments for understanding nano-scale surface structures of the chromosome.

Chromosome Structures↗
PubMed2003

Analysis by atomic force microscopy of morphological changes in barley chromosomes during FISH treatment.

We employed atomic force microscopy (AFM) to examine structural changes in barley chromosomes during the four steps of standard FISH processes. Rehydration and dehydration with alcohol accompanying RNase treatment increased chromosome arm width and decreased chromosome height about 50%. Subsequent heat denaturation reduced chromosome height further. These three-dimensional structural changes of the chromosomes were substantial, but the FISH signal produced by the hybridization of fluorescent probes was clear when observed by a fluorescence microscope. In higher-magnification images, we observed granular structures considered to represent the chromatin fiber on the surface of the chromosomes in each FISH protocol step. These our results indicate that FISH treatments result in severe damage of the three-dimensional higher-order structures of the chromosomes, although nano-structures, such as nucleosome and chromatin fibers, remain intact and relatively unaffected.

Artifacts↗
PubMed2002

Unique therapeutic effects of the Japanese-Chinese herbal medicine, Sairei-to, on Th1/Th2 cytokines balance of the autoimmunity of MRL/lpr mice.

Sairei-to, one of the Japanese-Chinese herbal medicines has been used for the treatment of various diseases, especially collagen disease and edema in nephrotic syndrome. However, the mechanism of the therapeutic effects remains uncertain. Therefore, we investigated the immunological changes of skin, kidney, spleen cells and serum in autoimmune-prone MRL/lpr, MRL/n and C57BL/6J mice treated with Sairei-to. In MRL/lpr mice treated with Sairei-to, the improvement of proteinuria, reduction in the number of hematoxylin bodies in kidney, and reduced serum levels of blood urea nitrogen were observed. These results indicate that Sairei-to can improve or inhibit the progression of lupus nephritis. The proportion of CD19 and the serum levels of IgG1, which is one of the pathogenesis of lupus dermatoses and lupus nephritis, were significantly reduced in Sairei-to-treated MRL/lpr mice. Therefore, it is suspected that the B cell function was suppressed by Sairei-to. In addition, CD4/8 ratio in spleen cells and the degree of lymphoproliferation in MRL/lpr mice also decreased. Interestingly, IL-4 producing spleen cells were increased significantly by ELISPOT assay, and IFN-gamma mRNA expressions were reduced in Sairei-to-treated MRL/lpr mice. Regarding the Th balance, an imbalance towards Th1 predominance may play a significant role in MRL/lpr mice, and the Th1 axis was suppressed and the Th2 axis became predominant in Sairei-to-treated MRL/lpr mice. On the other hand, Th2 cell type immunoglobulins (IgG1) were suppressed. These results suggested that Sairei-to is potential for impairing shifted Th1/Th2 balance and hypergammaglobulinemia resulting in therapeutic effects.

Animals↗
PubMed2002

Comparison of anti 60 and 52 kDa SS-A/Ro antibodies in the pathogenesis of cutaneous lupus erythematosus.

Anti SS-A/Ro antibodies are involved in the pathogenesis of cutaneous lupus erythematosus (CLE) in a part through antibody dependent cellular cytotoxicity (ADCC). However, it is still obscure which and how anti 60 and anti 52 kDa Ro antibodies are involved. To address the issue, we examined both types of anti Ro antibodies of sera from patients with systemic LE (SLE) or discoid LE (LE). The titer of anti 60 kDa antibody in SLE was significantly much higher than that of DLE or control. The positive ratio of DLE showed more higher tendency than control, but it was not statistically significant. The similar tendency was observed in the titer of anti 52 kDa antibody. An association between the anti 60 kDa antibody and the anti 52 kDa antibody was statistically significant in SLE patients. Although the relative index (RI) was statistically significant in DLE, they included many negative sera, which biased the statistics. Both anti 60 kDa antibody and anti 52 kDa antibody could induce significant ADCC of ultraviolet B (UVB) light-irradiated keratinocytes from neonatal foreskins and/or normal adults, in which anti 60 kDa antibody showed higher cytotoxicity than anti 52 kDa antibody. The autologous combination studies (keratinocytes and monospecific anti 60 or anti 52 kDa antibody from patients) suggested anti 60 kDa antibody was more potent to induce ADCC activity than anti 52 kDa antibody. Taken together, it is likely that anti 60 kDa antibody dependent keratinocyte damage plays more significant role in the pathogenesis of SLE skin lesions than anti 52 kDa dependent damage. However, both type antibodies seemed to have little contribution to the pathogenesis of DLE skin lesion.

Adult↗
PubMed2002

Simultaneous collection of topographic and fluorescent images of barley chromosomes by scanning near-field optical/atomic force microscopy.

The present study investigated the applicability of scanning near-field optical/atomic force microscopy (SNOM/AFM) to observations of YOYO-1 stained barley chromosomes. Using this technique, topographic and fluorescent images in the same portion were obtained simultaneously, thus enabling the precise analysis of fluorescent structures in relation to the morphology of chromosomes. In YOYO-1 stained chromosomes, the fluorescent intensity roughly reflected the local amount and/or density of DNA in chromosomes. R-banding of chromosomes stained with YOYO-1 and methyl green was also observed clearly as bright spots of various sizes by this microscope. Thus, the SNOM/AFM is expected to become a useful tool for analysing the structure and function of chromosomes more precisely than before.

Benzoxazoles↗
PubMed2002

Useful technique for DNA-stretching and fixation.

We report on a useful technique for reproducibly straightening and fixing DNA molecules. When a droplet of DNA solution on surfaces is sucked up by pipet, surface tension at the moving air-water interface is sufficient to stretch the molecules. The point of this technique is the interaction control between surfaces and DNA molecules by polymer coating. With polymers containing pi-conjugation units (polyphenazasiline, PPhenaz and poly(vinylcarbazole), PVCz), many DNA were nicely stretched and fixed on surfaces. Furthermore, structural changes of pi-units in polymers affected DNA-stretching and fixation. By using atomic force microscopy (AFM) and scanning near-field optical/atomic force microscopy (SNOM/AFM), the feasibility of this technique for DNA observations was demonstrated.

DNA↗
PubMed2002

Pseudoxanthoma elasticum.

Pseudoxanthoma elasticum (PXE) is a heritable disease characterized by dermal, ocular, and vascular lesions that result from degeneration of the elastic fibers. Recently, the ATP-binding cassette subfamily C member 6 (ABCC6) gene has been demonstrated to be responsible for PXE, and 43 mutations have been identified to date. However, it is still unknown now mutations in the ABCCC6 gene can lead to manifestations of PXE.

Humans↗
PubMed2002

Direct visualization of abasic sites on a single DNA molecule using fluorescence microscopy.

A new method was developed to allow direct visualization of damaged sites on individual DNA molecules. Fluorescence in situ hybridization on extended DNA molecules was modified to detect a single abasic site. Abasic sites were specifically labeled with a biotinylated aldehyde-reactive probe and fluorochrome-conjugated streptavidin. The light emitted by a single fluorochrome-DNA complex was calibrated. The number of abasic sites on the DNA molecule was estimated by counting each fluorochrome-DNA complex. The present study directly visualized and characterized the abasic sites of single DNA molecules.

DNA↗
PubMed2002

Imaging of chromosomes at nano-meter scale resolution using scanning near-field optical/atomic force microscopy.

Topographic and fluorescent images of whole barley chromosomes stained with YOYO-1 were observed simultaneously by scanning near-field optical/ atomic force microscopy (SNOM/AFM). The chromosome was relatively smooth and flat in the topographic images and no significant difference in height was present between regions of high fluorescent and low fluorescent intensity in the chromosomes. The telomeric region, labeled by fluorescence in situ hybridization (FISH) method, was also observed by SNOM/AFM at high resolution, and fluorescent signals of the telomeric region were clearly defined on the topographic image of chromatin fibers on the chromosome at the nano-meter scale level. Although the telomeric signals were usually visualized as a single fluorescent region at the end of sister chromatids by conventional light microscopy, they were observed separately as two fluorescent regions, less than 100-200 nm distance, using the SNOM/AFM. The SNOM/AFM offers great potential in identifying particular single gene location on chromosomes in the near future.

Benzoxazoles↗
PubMed2002

Mechanical elongation of the centromere in the barley metaphase chromosome.

The present study investigated the mechanical elongation of the centromere in the barley chromosomes by a microneedle manipulation method for the structural analysis of the chromosomes. Chromosomes were extracted from barley root cells, affixed on a cover slip by a standard preparation method, and elongated in either distilled water, phosphate buffered saline (PBS), or 2 x sodium saline citrate (SSC). The mechanical property of the chromosome elongation was assessed by the measurement of the force required for the elongation of chromosomes. This assessment has shown that the chromosomes in distilled water were much firmer than those in the PBS or 2 x SSC. To confirm the elongation of the centromere, the elongated chromosomes were investigated by fluorescence in situ hybridization with a centromere probe. The fluorescence information indicated that the extent of the loosening of the centromere during elongation differed depending on the buffers used; the centromere elongated in 2 x SSC was more loosened than that in the PBS. Atomic force microscopy also revealed the structure of the unpacked centromere after the mechanical elongation, when rows of fibrous structures about 30 to 50 nm thick were clearly observed in the centromere elongated in 2 x SSC. The investigation of elongated chromosomes should prove useful for an understanding of the structural analysis of chromosomes.

Centromere↗
PubMedDate not supplied

Nano-scale imaging of chromosomes and DNA by scanning near-field optical/atomic force microscopy.

Nano-scale structures of the YOYO-1-stained barley chromosomes and lambda-phage DNA were investigated by scanning near-field optical/atomic force microscopy (SNOM/AFM). This technique enabled precise analysis of fluorescence structural images in relation to the morphology of the biomaterials. The results suggested that the fluorescence intensity does not always correspond to topographic height of the chromosomes, but roughly reflects the local amount and/or density of DNA. Various sizes of the bright fluorescence spots were clearly observed in fluorescence banding-treated chromosomes. Furthermore, fluorescence-stained lambda-phage DNA analysis by SNOM/AFM demonstrated the possibility of nanometer-scale imaging for a novel technique termed nano-fluorescence in situ hybridization (nano-FISH). Thus, SNOM/AFM is a powerful tool for analyzing the structure and the function of biomaterials with higher resolution than conventional optical microscopes.

Bacteriophage lambda↗
PubMedDate not supplied

Topical FK506 (tacrolimus) therapy for facial erythematous lesions of cutaneous lupus erythematosus and dermatomyositis.

Tacrolimus is a prototype of a class of topical immunosuppressive agents with great potential for the treatment of inflammatory skin diseases. Topical tacrolimus therapy was applied to facial skin lesions in 11 cases of cutaneous lupus erythematosus (LE) and dermatomyositis. Of the 11 patients, 6 (3 systemic LE, one discoid LE and 2 dermatomyositis) showed a marked regression of their skin lesions after tacrolimus therapy, but 4 patients (3 discoid LE and one dermatomyositis) were resistant to the therapy. A good response was observed for facial erythematous lesions with edematous or telangiectatic changes in systemic LE and dermatomyositis. In discoid LE with typical discoid lesions, tacrolimus brought no improvement. Topical tacrolimus will become a new tool for managing the skin lesions of collagen diseases.

Administration, Topical↗