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Biomedical subjects

T Nishizawa

Publications and source records attributed to T Nishizawa.

At least 19 recordsLinked to original sources

Nucleotide diversity of Japanese isolates of infectious hematopoietic necrosis virus (IHNV) based on the glycoprotein gene.

Infectious hematopoietic necrosis virus (IHNV), a member of the genus Novirhabdovirus, causes a highly lethal disease of salmonid fish. In the present study, G gene nucleotide sequences of 9 Japanese IHNV isolates obtained from 1971 to 1996 were analyzed to evaluate the genetic diversity and compared with IHNV isolates from North America and Europe. A radial phylogenetic tree revealed 5 major clusters including 3 genogroups (U, M and L) for North American isolates and 1 genogroup for European isolates. Five Japanese isolates from 1971 to 1982 appeared in the cluster for genogroup U, while the remaining Japanese isolates from 1980 to 1996 formed a new genogroup, JRt (Japanese rainbow trout). Maximum nucleotide diversity among the Japanese isolates was 4.5%, which was greater than that within the North American isolates (3.6%), and the degree of nucleotide diversity within Japanese isolates was increased by inclusion of the genogroup JRt isolates. It was concluded that Japanese isolates shared a common source with the genogroup U of the North American isolates and that there were large divergences between Japanese isolates before and after the 1980s.

Animals↗

Specific messenger RNA expression for signal transduction molecules by lipopolysaccharide in intestinal macrophages.

Intestinal macrophages are known to display profound inflammatory anergy in response to lipopolysacchraide (LPS). To study the mechanisms of unresponsiveness of intestinal macrophages to LPS, we compared the mRNA expression of molecules associated with signal transduction of intestinal macrophages with those of other tissue macrophages. Also cellular localization of CD14 protein was examined. Intestinal, alveolar and peritoneal macrophages were isolated from rats or mice. The expression of mRNA was assessed by real-time PCR, and cellular localization of CD14 protein was examined by flow cytometry. Cellular responses to LPS were examined by production of TNF and NO. The expression of CD14 mRNA in intestinal macrophages was lower than for peritoneal macrophages but higher than for alveolar macrophages. The mRNA expression of other molecules corresponding to intracellular signal transduction in intestinal macrophages was similar with alveolar and peritoneal macrophages. Despite the presence of CD14 mRNA, proteins of CD14 were not detected on cell surfaces of intestinal macrophages, and induction of TNF or NO responding to LPS were not detected. Flow cytometric analysis demonstrated that CD14 protein was not expressed on the cell surface but was expressed inside intestinal macrophages. The unresponsiveness of intestinal macrophages after LPS exposure is considered to be largely attributed to the lack of CD14 protein on their cell surfaces. However, CD14 protein was expressed inside of the cells, suggesting that post-transcriptional regulation rather than transcriptional suppression may play a dominant role in determining the phenotype of the intestinal macrophages.

Animals↗

A new genotype of lymphocystivirus, LCDV-RF, from lymphocystis diseased rockfish.

Lymphocystis disease virus (LCDV) is the causative agent of lymphocystis disease. In this study, nucleotide sequences of the major capsid protein (MCP) gene were analyzed among LCDV isolates from Japanese flounder and rockfish. A phylogenetic tree revealed three clusters for lymphocystiviruses. The first cluster included Japanese flounder isolates; the second cluster consisted of rockfish isolates; and the remaining one consisted of LCDV-1. Nucleotide sequence identities were > or =99.6% among Japanese flounder isolates and 100% among rockfish isolates, while between each cluster they were < or =85.2%. Experimental infections with Japanese flounder and rockfish isolates revealed that Japanese flounder and rockfish were infected by the respective homologous isolate but not by the heterologous isolate. These findings suggest that at least three genotypes exist in the genus Lymphocystivirus.

Animals↗

High prevalence of hepatitis B, C and delta virus infections among blood donors in Mongolia.

Serum samples obtained from 289 first-time and 114 repeat donors at the Blood Center of Mongolia (MBC) were tested for serological and molecular markers of hepatitis B virus (HBV), hepatitis C virus (HCV), and hepatitis delta virus (HDV) infections. Among the 403 blood donors, 33 (8.2%), 21 (5.2%), and 27 (6.7%) tested positive for hepatitis B surface antigen (HBsAg) and/or HBV DNA, HCV RNA, and HDV RNA, respectively. Collectively, 55 donors were viremic for one or more of these viruses, and included 54 first-time donors (18.7%) and 1 repeat donor (0.9%) (P < 0.0001). One discrepant case with HBsAg detectable only at MBC was negative for HBsAg, HBV DNA and anti-HBc in this study. Four donors who were HCV-viremic in this study were negative for anti-HCV by the MBC method. Further efforts to increase the sensitivity and specificity of the currently-used tests are urgently required in Mongolia. Three donors who were positive for anti-HBc and anti-HDV but negative for HBsAg, had both HBV DNA and HDV RNA. This suggests that introduction of a new anti-HDV serological test is useful for not only HDV screening but also HBV screening of anti-HBc-positive, HBsAg negative donors, considering a possibility of viral interference by coexisting HDV.

Adolescent↗

Raman microspectroscopic study on polymerization and degradation processes of a diacetylene derivative at surface enhanced Raman scattering active substrates. 1. Reaction kinetics.

Raman microspectroscopy was applied to study the polymerization kinetics of the Langmuir-Blodgett (LB) films of 10,12-pentacosadiynoic acid (DA) adsorbed on surface enhanced Raman scattering (SERS) active Ag island films. A two-dimensional (2D) Raman microscopic image measured at 1520 cm(-1) exhibits bright and dim spots with the diameter of several hundred nanometers. Raman microscopic spectra, measured by defocusing the excitation laser light (532 nm, diameter of ca. 10 mum) on the samples at room temperature, proved the occurrence of the surface processes consisting of the formation of polydiacetylene (PDA) in the blue phase, its conversion to the red phase, and subsequent bleaching. These reactions were negligible under the same condition for the DA-LB films prepared on a smooth (i.e., SERS-inactive) Ag film, indicating that the 532-nm-induced polymerization and the bleaching process are enhanced by the SERS-active substrates. At low temperatures below -50 degrees C, the Raman microscopic measurements proved the formation of the blue phase and its conversion to the red phase with much lower reaction rates compared to the corresponding rates at room temperature. The bleaching, however, was much suppressed at the low temperatures. The kinetic analyses of the formation of the blue phase and its conversion to the red phase were performed by using the intensity changes of the Raman bands due to the blue (1477 cm(-1)) and red (1517 cm(-1)) phases as a function of the irradiation time. The results strongly suggested the existence of at least two processes taking place simultaneously on the SERS-active substrates; that is, one of the processes is a sequential reaction, DA-monomers --> PDA in the blue phase --> PDA in the red phase, and the other is another sequential reaction, DA-monomers --> PDA in the red phase --> degradation species (probably amorphous carbon). Thus, even at the low temperatures, there occurs the surface reaction consisting of the formation of PDA and its degradation. The reaction can be ascribed to a process taking place at the highly SERS-active site, which gives the bright spot (so-called "hot spot") on the 2D Raman image, as proved by the confocal Raman microscopic measurement in the following paper.

Adsorption↗

Raman microspectroscopic study on polymerization and degradation processes of a diacetylene derivative at surface enhanced Raman scattering active substrates. 2. Confocal Raman microscopic observation of polydiacetylene adsorbed on active sites.

Confocal Raman microscopic measurements were performed at room temperature on the Langmuir-Blodgett (LB) monolayer of 10,12-pentacosadiynoic acid (DA) prepared on surface enhanced Raman scattering (SERS) active Ag island films, two-dimensional (2D) Raman images of which exhibit bright and dim spots on a dark background. The measurements performed by focusing the excitation laser light (488 nm) on the dark background indicate the prompt appearance of the Raman bands (1515 and 2115 cm(-1)) due to polydiacetylene (PDA) in the red phase and subsequent diminution of the Raman bands. On the other hand, the spectra observed by focusing the excitation laser spot on the dim and bright spots exhibit almost random fluctuations, giving rather narrow Raman bands in the 1620-1000 cm(-1) region, which appear and disappear temporarily with varying intensities under the continuous irradiation at 488 nm. Broad Raman bands appear around 1580 and 1360 cm(-1), which are ascribable to amorphous carbon, at a later stage of the observation, the intensities from the bright spot being more than 100 times stronger than those from the dim spot. The narrow bands are ascribed to a series of carbonaceous intermediates such as polyenes, graphite sheets with various sizes, and folded or reorganized forms of the sheets including carbon nanotubes and fullerenes, which are formed during the conversion of PDA to amorphous carbon. The random spectral fluctuation was interpreted by considering that the intermediates undergo thermally activated diffusion and get temporarily in contact with the SERS-active site, resulting in the enhancement of their Raman bands and the fluctuation.

Adsorption↗

Effect of tacrolimus in a patient with pure red-cell aplasia.

A 78-year-old woman has suffered from pure red-cell aplasia (PRCA) associated with generalized myasthenia gravis and thymoma. Cyclosporin A (CyA) with corticosteroid increased numbers of erythroid cells in her bone marrow cells but she required monthly blood transfusions. Administration of tacrolimus as a substitution for CyA inhibited progression of anemia without the need for further blood transfusion. No serious side effects were observed. This case demonstrates that tacrolimus is another option of treatment for PRCA in patients who fail to respond to CyA.

Aged↗

Experimental horizontal transmission of viral hemorrhagic septicemia virus (VHSV) in Japanese flounder Paralichthys olivaceus.

Infection by viral hemorrhagic septicemia virus (VHSV) has recently occurred among wild and farmed Japanese flounder Paralichthys olivaceus in Japan. In the present study, horizontal transmission of VHSV among Japanese flounder was experimentally demonstrated by immersion challenge. Exposure to a flounder isolate (Obama25) of VHSV revealed a dose-response, with higher mortality (81 and 70%) at the 2 higher exposure levels (6.0 and 4.0 log10 TCID50 ml(-1)). In a second experiment, high titers of VHSV were expressed from moribund and dead flounder based on virus detection in holding-tank waters 2 to 3 d prior to death of the fish and 1 d after death. The virus could not be detected in tank waters 2 d after death. Finally, a third cohabitation experiment in small tanks demonstrated horizontal transmission of VHSV from experimentally infected to uninfected fish.

Animals↗

Genetic relatedness among Japanese, American and European isolates of viral hemorrhagic septicemia virus (VHSV) based on partial G and P genes.

Molecular virological analyses of 8 Japanese VHSV (viral hemorrhagic septicemia virus) isolates from wild and farmed Japanese flounder Paralichthys olivaceus were performed to investigate their genetic relatedness to American and European isolates of VHSV. Phylogenetic analyses based on the partial nucleotide sequences of G and P genes revealed that there are 2 genogroups of VHSV in Japan. The first one represented by the Obama25 isolate is closely related to the American isolates (Genogroup I) while the other, the KRRV9601 isolate, is closely related to the traditional European isolates (Genogroup III). The 2 types of Japanese VHSV showed differences in the relative mobility of the G protein and intensity of the antibody reaction on the P and M proteins. The Obama25 type of VHSV is widely distributed as a native virus in the coastal areas of western Japan and has been responsible for the occurrence of VHSV infection in farmed Japanese flounder while the KRRV9601 isolate is considered to have been introduced from a foreign country.

Animals↗

A time-course study on the resistance of Penaeus japonicus induced by artificial infection with white spot syndrome virus.

The onset and duration of resistance in experimental survivors of Penaeus japonicus produced by an intramuscular injection with white spot syndrome virus (WSSV) were surveyed by re-challenge tests with the virus conducted at weeks 1-4 and months 1-3 post initial exposure (PIE) to the virus. Virus neutralising activity in the survivors' plasma was also examined. Plasma-treated WSSV was separated from the plasma by centrifugation and then injected into naïve shrimp, in parallel with each re-challenge test. Re-challenge tests of the survivors conducted at weeks 1-4 PIE revealed that the resistance commenced at week 3 (relative percent survival, RPS: 39%) and almost fully developed at week 4 (RPS: 58%), because statistically significant differences in survival rates were observed between the test (previously virus exposed) and control groups at weeks 3 and 4. Re-challenge at months 1-3 PIE resulted in RPS values of 67, 54 and 6%, respectively, indicating the resistance persisted until month 2. RPS values in neutralisation tests performed at weeks 1-4 and months 1-3 PIE were -5, 14, 36, 50, 100, 38 and 6%, respectively, which coincided with the RPS values in each re-challenge test conducted in parallel. The present results demonstrated that resistance of P. japonicus against the viral pathogen developed 3 or 4 weeks after an exposure to the virus, and it persisted for another month at 24 degrees C. The resistance was paralleled by a humoral neutralising factor(s) in the plasma of shrimp.

Animals↗

Analysis of the entire genomes of thirteen TT virus variants classifiable into the fourth and fifth genetic groups, isolated from viremic infants.

TT virus (TTV) DNA in serum samples obtained from 24 TTV-infected infants was amplified by polymerase chain reaction (PCR) with inverse primers derived from the untranslated region. The amplified PCR products were molecularly cloned; six clones each were analyzed. Seventy-six (53%) of the 144 TTV clones were classified into group 4 (YONBAN isolates), and 22 (15%) into a novel genetic group (group 5). The TTV clones in group 4 were classified into 9 types, and those in group 5 into 4 types. The entire nucleotide sequence of one representative clone each from the 13 types were determined; they comprised 3570-3770 nucleotides, and had poor homology to TTVs of groups 1-3 (TA278, PMV and SANBAN isolates). A phylogenetic tree based on the entire nucleotide sequence of open reading frame 1 confirmed the presence of five distinct clusters separated by a bootstrap value of 100%. Analysis of 13 TTV variants demonstrated preservation of the genomic organization and transcription profile in all TTV groups. TTV group 4 was detected in 54% or 72% of 7-to-12-month-old infants in Japan and China, respectively, which is comparable with that among adults in the respective country, indicating early and frequent acquisition of this TTV group in infancy.

Adult↗

Analysis of the complete genome of indigenous swine hepatitis E virus isolated in Japan.

Using reverse transcription-polymerase chain reaction with primers derived from well-conserved genomic areas among all four hepatitis E virus (HEV) genotypes (I-IV), the HEV sequence was identified in serum samples obtained from 3 (3%) out of 95 60- to 90-day-old pigs in Japan and characterized molecularly. In the partial sequence of open reading frame (ORF) 2 of 421 nucleotides, the three swine isolates (swJ570, swJ681, and swJ791) showed the highest similarity of 83-87% to genotype III HEV representing human and swine strains (US1, US2, and swUS1) in the United States. The full-length nucleotide sequence of swJ570 consisted of 7225 nucleotides excluding the poly(A) tail and contained ORF 1 encoding 1703 amino acids (aa), ORF2 encoding 660 aa, and ORF3 encoding 122 aa. The swJ570 strain was most closely related to a Japanese strain (JRA1), which had been obtained from a hepatitis patient who had not traveled outside Japan. The overall nucleotide sequence identity between them was 89% and the deduced amino acid sequence identities of ORF1, ORF2, and ORF3 were 96, 99, and 98%, respectively. These results indicate that a certain proportion of pigs in Japan are HEV-viremic and may act as reservoirs of HEV infection, and that the presence of an indigenous strain(s) of HEV should be taken into consideration for the diagnosis of acute hepatitis in Japan.

3' Untranslated Regions↗

An outbreak of VHSV (viral hemorrhagic septicemia virus) infection in farmed Japanese flounder Paralichthys olivaceus in Japan.

A rhabdoviral disease occurred in farmed populations of market sized Japanese flounder (hirame) Paralichthys olivaceus in the Seto Inland Sea of Japan in 1996. The causative agent was identified as viral hemorrhagic septicemia virus (VHSV) based on morphological, immunological, and genetic analyses. Diseased fish that were artificially injected with a representative virus isolate showed the same pathological signs and high mortality as observed in the natural outbreak. This is the first report of an outbreak of VHSV infection in cultured fish in Japan. Clinical signs of diseased fish included dark body coloration, an expanded abdomen due to ascites, congested liver, splenomegaly, and a swollen kidney. Myocardial necrosis was most prominent and accompanied by inflammatory reactions. Necrotic lesions also occurred in the liver, spleen and hematopoietic tissue, and were accompanied by circulatory disturbances due to cardiac failure. Hemorrhagic lesions did not always appear in the lateral musculature. Transmission electron microscopy revealed many rhabdovirus particles and associated inclusion bodies containing nucleocapsids in the necrotized myocardium. The histopathological findings indicated that the necrotizing myocarditis could be considered a pathognomonic sign of VHSV infection in Japanese flounder.

Animals↗

Effects of shrimp density on transmission of penaeid acute viremia in Penaeus japonicus by cannibalism and the waterborne route.

To investigate the effects of shrimp density on mortalities of Penaeus japonicus in experimental penaeid acute viremia (= white spot syndrome), shrimp injected intramuscularly with penaeid rod-shaped DNA virus (PRDV) were reared at different densities. In Expt 1, challenged (10(-6) dilution of a PRDV preparation) shrimp were reared collectively in a tank or individually in separate chamber units. A significant difference in cumulative mortalities was found between collectively (75.6%) and individually (1.2%) reared groups after 30 d. In Expt 2, effects of density on mortality were clearly shown when challenged (10(-5) dilution) shrimp were reared collectively in tanks at high (260 shrimp m(-2)), middle (135 shrimp m(-2)) and low densities (73 shrimp m(-2)). The cumulative mortalities for 14 d in the high, middle and low density groups were 72, 46 and 18%, respectively. In Expt 3, challenged (10(-5) dilution) shrimp were reared collectively in 3 tanks (Groups A, B and C) at the same high density (260 shrimp m(-2)): Group A, dead shrimp were immediately removed to avoid transmission of the pathogen through cannibalism and the waterborne route; Group B, dead shrimp were removed at scheduled times but were separated from living shrimp by a net partition to avoid cannibalism; and Group C, dead shrimp were removed twice a day at scheduled times. Resulting cumulative mortalities for 20 d in Groups A, B and C were 4, 24 and 64 %, respectively. These results show that the higher mortalities occur in P. japonicus reared at the higher densities in experimental PRDV infection, and this phenomenon is caused mainly by a higher opportunity of horizontal transmission of the virus through cannibalism and the waterborne route.

Acute Disease↗

Size-related and size-unrelated functional heterogeneity among pancreatic islets.

Functional heterogeneity of pancreatic islets was systematically analyzed for the first time using freshly isolated single rat pancreatic islets. First, 60 islets were sequentially exposed to 3, 9.4, 15.6, and 24.1 mM glucose for 30 min each in incubation experiments: 36 (60%) responded in a concentration-dependent and 19 (32%) in an all-or-none manner, and 5 (8%) islets did not respond to high glucose. As a group, the larger the islet, the higher the beta cell glucose sensitivity. However, glucose-stimulated elevation of [Ca2+]i in the beta cell. insulin/glucagon ratio in the islet, and expression of glucose transporter 2, glucokinase, and pancreatic duodenal homeobox factor-1 in the beta cell were not significantly related to islet size. Second, 50 islets were stimulated with 16.7 mM glucose in perifusion. A biphasic insulin release was found in 39 (78%), and no or little first phase response in 11 (22%) islets, irrespective of the islet size. Nevertheless, when the response was plotted as a group, it was clearly biphasic. Islet size, insulin content and the amount of insulin release were positively correlated with each other. In conclusion, there are size-related and size-unrelated functional diversity among pancreatic islets. The reason for such heterogeneity remained to be determined.

Animals↗

Intraspinal cyst communicating with the intervertebral disc in the lumbar spine: discal cyst.

STUDY DESIGN: A retrospective case study of patients with intraspinal cyst having a distinct connection with the corresponding intervertebral disc. OBJECTIVES: To propose a new clinical entity, "discal cyst," by clarifying the clinical, radiographic, and histologic aspects of the disease. SUMMARY OF BACKGROUND DATA: Several types of intraspinal cysts with different pathogenesis, causing symptoms indistinguishable from those of lumbar disc herniation, have been reported, such as perineural cysts, synovial cysts, and ganglion cysts. However, to the authors' knowledge, no detailed analysis has been made of cysts that have a distinct connection with the corresponding intervertebral disc. METHODS: Clinical pictures, radiographic findings, and surgical and histologic findings in eight surgically treated patients with intraspinal cyst having a distinct connection with the intervertebral disc were reviewed. Possible pathogenesis and a proposal for nomenclature were also discussed. RESULTS: This disease can be characterized by (1) clinical symptoms indistinguishable from those of typical disc herniation, manifesting as a unilateral single nerve root lesion; (2) incidence at slightly younger age and at upper intervertebral levels than with typical disc herniation; (3) T1 low signal and T2 high signal intensity, round to oval mass lesion on magnetic resonance imaging, compatible with a liquid-containing cyst; (4) minimal degeneration of the involved disc, either on discography/computed tomographic discography or magnetic resonance imaging; (5) a connection between the cyst and the corresponding intervertebral discs on discograms with severe radiating pain in the affected leg at the time of injection; (6) immediate relief of symptoms after simple removal of the cyst; (7) cyst wall consisting of dense fibrous connective tissue containing bloody to clear serous discharge; and (8) absence of disc materials and a specific lining cell layer on histologic examination. Although the exact cause is unknown, underlying minor disc injury may serve as a basis for cyst formation. CONCLUSION: Eight cases of intraspinal cysts communicating with the intervertebral disc presenting symptoms identical to those of disc herniation are presented. Because all cysts were connected to the corresponding disc and the development of the cyst was assumed to be related to underlying disc injury, it is proposed to name this clinical entity discal cyst.

Adult↗

Heterogeneous distribution of TT virus of distinct genotypes in multiple tissues from infected humans.

TT virus (TTV) DNA was quantitated in the serum and nine autopsy tissues (bone marrow, lymph node, muscle, thyroid gland, lung, liver, spleen, pancreas, and kidney) obtained from each of three TTV-infected subjects by real-time polymerase chain reaction (PCR), which can detect all TTV genotypes. TTV DNA was detected in all examined tissues, with the viral load being equal to or up to 300 times higher than that in the corresponding serum (2.1 x 10(5) to 5.3 x 10(7) copies/g vs 1.2-3.9 x 10(5) copies/ml). Generally, the TTV viral load was higher in the bone marrow, lung, spleen, and liver than in the other tissues, although it varied by individual. Restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified TTV DNA of 3.3 kilobases (kb) revealed considerable differences among the TTVs in the serum and tissue specimens from each subject. Further, the 3.3-kb amplicons from the serum and tissue specimens from one subject were molecularly cloned, and 30 clones each from the serum and each tissue specimen were subjected to RFLP and sequence analysis (total, 300 clones): the TTV clones were classified into six genotypes including four novel genotypes. The genotypic variability was remarkable: each specimen had one to five TTV genotypes at different frequencies. TTV DNA in replicative intermediate forms and TTV mRNA were detectable in all tissues tested. These results indicate the broad, uneven distribution of TTV genotypes in tissues and suggest that viral replication takes place in multiple tissues at distinct levels in infected individuals.

Base Sequence↗