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Biomedical subjects

Hiroshi Kawai

Publications and source records attributed to Hiroshi Kawai.

15 recordsLinked to original sources

Microlens arrays for integral imaging system.

When designing a system capable of capturing and displaying 3D moving images in real time by the integral imaging (II) method, one challenge is to eliminate pseudoscopic images. To overcome this problem, we propose a simple system with an array of three convex lenses. First, the lateral magnification of the elemental optics and the expansion of an elemental image is described by geometrical optics, confirming that the elemental optics satisfies the conditions under which pseudoscopic images can be avoided. In using the II method, adjacent elemental images must not overlap, a condition also satisfied by the proposed optical system. Next, an experiment carried out to acquire and display 3D images is described. The real-time system we have constructed comprises an elemental optics array with 54 H x 59 V elements, a CCD camera to capture a group of elemental images created by the lens array, and a liquid crystal panel to display these images. The results of the experiment confirm that the system produces orthoscopic images in real time, and thus is effective for real-time application of the II method.

Journal Article↗

Temperature rhythm reentrains faster than locomotor rhythm after a light phase shift.

Mammalian endogenous circadian rhythms are entrained to the environmental light-dark (LD) cycle. Although the circadian rhythms of core body temperature (Tb) and spontaneous locomotor activity (LA) are well synchronized under stable LD conditions, it is thought that these two parameters are regulated by distinct mechanisms. The purpose of the present study was to examine the adaptability of these two rhythms to an abrupt change in the environmental light phase. Tb and LA were simultaneously recorded in individual mice kept under 12:12-h LD cycle conditions before and after an 8-h photic phase advance. The onset of LA required 8 days to reentrain to the new LD cycle, whereas 6 days were required for reentrainment of the acrophase of Tb. Resting Tb, i.e., the Tb level independent of LA, was extracted from the same data source. The resting Tb level exhibited a robust daily rhythm with a difference of 1.0 degrees C between LD phases. After the photic phase advance, the resting Tb rapidly reached a stable level within 4 days, whereas the uncorrected Tb required 6 days for reentrainment. Based on these findings, we revealed that, independent of LA, the adaptability of the Tb rhythm to a new light cycle is half as rapid as that of LA. These results therefore suggest that the circadian rhythms of Tb and LA are intrinsically regulated by different pacemaker or effector mechanisms.

Adaptation, Physiological↗

Time-restricted feeding entrains daily rhythms of energy metabolism in mice.

Energy metabolism, oxygen consumption rate (VO2), and respiratory quotient (RQ) in mice were monitored continuously throughout 12:12-h light-dark cycles before, during, and after time-restricted feeding (RF). Mice fed ad libitum showed robust daily rhythms in both parameters: high during the dark phase and low during the light phase. The daily profile of energy metabolism in mice under daytime-only feeding was reversed at the beginning of the first fasting night. A few days after daytime-only feeding began, RF also reversed the circadian core body temperature rhythm. Moreover, RF for 6 consecutive days shifted the phases of circadian expression patterns of clock genes in liver significantly by 8-10 h. When mice were fed a high-fat (HF) diet ad libitum, the daily rhythm of RQ dampened day by day and disappeared on the sixth day of RF, whereas VO2 showed a robust daily rhythm. Mice fed HF only in the daytime had reversed VO2 and RQ rhythms. Similarly, mice fed HF only in the daytime significantly phase shifted the clock gene expression in liver, whereas ad libitum feeding with HF had no significant effect on the expression phases of liver clock genes. These results suggested that VO2 is a sensitive indicator of entrainment in the mouse liver. Moreover, physiologically, it can be determined without any surgery or constraint. On the basis of these results, we hypothesize that a change in the daily VO2 rhythm, independent of the energy source, might drive phase shifts of circadian oscillators in peripheral tissues, at least in the liver.

Animals↗

cDNA microarray analysis of autoimmune hepatitis, primary biliary cirrhosis and consecutive disease manifestation.

While autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) usually have distinct clinical manifestations, some patients present with features of both conditions. Using cDNA microarrays, we analyzed and compared gene expression profiles in 8 patients with AIH, 9 with PBC, 8 with chronic hepatitis C (CHC), 8 with non-alcoholic steatohepatitis (NASH) and 9 with normal livers. We subsequently applied this method to a tissue sample from a 61-year-old woman with overlapping features of both AIH and PBC. A 61-year-old woman was admitted to our hospital for evaluation of elevated serum alkaline phosphatase. A liver biopsy showed accumulation of mononuclear cells around the bile duct cells, a feature characteristic of chronic non-suppurative destructive cholangitis (CNSDC). Three years later, her serum alanine aminotransferase (ALT) level had increased, and a liver biopsy demonstrated evidence of a severe form of hepatitis. A cDNA microarray analysis of both biopsies identified the molecular events associated with her altered histology. The expression profile of this patient, which was originally different from that of the other PBC patients, changed to an AIH pattern. Our results suggest that this patient has characteristics of both AIH and PBC.

Aged↗

Simultaneous real-time detection of initiator- and effector-caspase activation by double fluorescence resonance energy transfer analysis.

Fluorescence resonance energy transfer (FRET) with green fluorescent protein (GFP) variants has become widely used for biochemical research. In order to expand the choice of fluorescent range in FRET analysis, we designed various color versions of the FRET-based probes for caspase activity, in which the substrate sequence of the caspase was sandwiched by donor and acceptor fluorescent proteins, and studied the potential of these color versions as fluorescent indicators. Six color versions were constructed by a combination of cyan fluorescent protein (CFP), GFP, yellow fluorescent protein (YFP), and DsRed. Real-time monitoring in single cells revealed that all probes could detect caspase activation during tumor necrosis factor (TNF)-alpha-induced cell death as a fluorescent change. GFP-DsRed and YFP-DsRed were as sensitive as CFP-YFP, and CFP-DsRed also showed a large fluorescent change. By using two probes, CFP-DsRed and YFP-DsRed, we carried out simultaneous multi-FRET analysis and revealed that the initiator- and effector-caspases were activated almost simultaneously in TNF-alpha-induced cell death. These findings may give experimental bases for the development of novel techniques to analyze multi-events simultaneously in single cells by using FRET probes in combination.

Apoptosis↗

Differential gene expression profiles in stage I primary biliary cirrhosis.

OBJECTIVES: Primary biliary cirrhosis (PBC) is a progressive disease. However, little is understood about the molecular mechanisms underlying its features. METHODS: We analyzed gene expression profiles of liver biopsy samples from 16 patients with PBC, seven with autoimmune hepatitis, eight with chronic hepatitis C, and eight normal control livers. In addition to whole liver samples, we selectively analyzed chronic nonsuppurative destructive cholangitis (CNSDC) lesions by laser capture microdissection. RESULTS: Hierarchical clustering analysis using only early-stage liver disease demonstrated 85 genes were upregulated in stage I PBC specifically. Surprisingly, the expression of these genes was not maintained in advanced-stage PBC, while other gene clusters were upregulated. Expression analysis of CNSDC lesions in stage I PBC showed the presence of active inflammatory changes, characterized by the significant elevation of interferon-gamma and the development and maturation of lymphocytes. Expression of these genes was diminished in lymphoid cells aggregation in stage III PBC, and genes reflecting hepatocyte damage were upregulated with disease progression. CONCLUSION: Gene expression patterns in stage I PBC are different from others. There are distinct changes in molecular pathology from early- to late-stage PBC, which might be a clue to reveal the etiology and progression of PBC.

Aged↗

Promoter methylation downregulates CDX2 expression in colorectal carcinomas.

CDX2 (caudal type homeobox transcription factor 2) is a homeobox protein, which is expressed in intestinal epithelium. CDX2 has been considered to play a role as a tumor suppressor gene in colorectal cancer because its expression is lacking in colorectal carcinomas, but preserved in adenomas. The point mutations have been found to account for loss of CDX2 expression, but the main mechanism responsible for CDX2 gene inactivation is less understood. We analyzed methylation and expression status of CDX2 in colorectal cancer cell lines and primary tumors. There are two CpG-rich sites in promoter region of CDX2 gene, -1570 to -1200 and -220 to +880. By COBRA (combined bisulfite restriction analysis) assay, the upper CpG-rich site was heavily methylated in all cell lines, but the lower CpG-rich site was methylated in limited cell lines. Bisulfite sequencing analysis and RT-PCR revealed that methylation of the lower CpG site was associated with down-regulation of CDX2. In addition, gene expression was restored in COLO201, a methylated cell line with 5-aza-2'-deoxycytidine, confirming that methylation caused gene down-regulation. We also examined CDX2 promoter methylation of primary tumors by MSP (methylated-allele specific PCR) assay and found that nearly 40% of cases have a methylated CDX2 gene. Our results demonstrate that CDX2 methylation is frequently present in colorectal cancers and may play a key role in inactivating CDX2 expression.

CDX2 Transcription Factor↗

Simultaneous imaging of initiator/effector caspase activity and mitochondrial membrane potential during cell death in living HeLa cells.

A family of cystein proteases, the caspases, plays a central role in mediating cell death. In this study, we measured the activation of the initiator and effector caspase in real time, and studied the relationship between caspase activity and mitochondrial membrane potential in living cells by means of bioimaging. We also designed and developed a fluorescence resonance energy transfer (FRET)-based genetically encoded fluorescent indicator, which consisted of yellow fluorescent protein (YFP), a peptide sequence which can be cleaved by specific caspases, and cyan fluorescent protein (CFP). Two peptide sequences which could be cleaved by initiator caspases and effector caspases, respectively, were used. Simultaneous real-time measurements of the caspase activity and mitochondrial membrane potential in the cells treated with TNF-alpha and staurosporine revealed that dying cells showed caspase activation and mitochondrial depolarization, and that these events, however, were not firmly linked. Although it takes anywhere from 1 to over 10 h after the addition of the cell death inducer for the caspases to begin to be activated, initiator caspases and effector caspases are activated within a short period of time at the last stage in the entire process leading to cell death.

Amino Acid Sequence↗

Improved sensitivity for insulin in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry by premixing alpha-cyano-4-hydroxycinnamic acid matrix with transferrin.

This report describes an enhancement of the signal intensities of proteins and peptides in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). When alpha-cyano-4-hydroxycinnamic acid (CHCA) premixed with human transferrin (Tf) was used as a matrix, the signal intensity of insulin was amplified to more than ten times that of the respective control in CHCA without Tf. The detection limit of insulin was 0.39 fmol on-probe in the presence of Tf, while it was 6.3 fmol in the absence of Tf. The signal intensity of insulin was also enhanced when the CHCA matrix was premixed with proteins other than Tf (80 kDa), such as horse ferritin (20 kDa), bovine serum albumin (BSA, 66 kDa), or human immunoglobulin G (150 kDa). The optimum spectrum of insulin was obtained when the added amount of protein was in the range 0.26-0.62 pmol, regardless of the molecular weight of the added protein. Tf and BSA outperformed the other tested proteins, as determined by improvements in the resulting spectra. When the mass spectra of several peptides and proteins were recorded in the presence of Tf or BSA, the signal intensities of large peptides such as glucagon were enhanced, though those of smaller peptides were not enhanced. In addition, the signal enhancement achieved with Tf and BSA was more pronounced for the proteins, including cytochrome C, than for the large peptides. This enhancement effect could be applied to improve the sensitivity of MALDI-TOFMS to large peptides and proteins.

Animals↗

[Rice pollen asthma and pollinosis in childhood: seasonal asthma and allergic rhinoconjunctivitis during the period of rice pollen emission in the surrouding area of rice field].

Although in 1969, rice pollen was first reported as a cause of asthma, rice pollen allergy has not been studied after the first report and thus the allergic significance of rice pollen is not well recognized at present. We investigated the sensitization to various allergens and the residential areas in children with the symptoms of asthma or rhinoconjunctivitis during the first decade of August, and measured rice pollen-specific IgE antibodies. Eighty-eight children (57 boys and 31 girls, mean age 8.5+/-2.9 years) with bronchial asthma or allergic rhinitis/conjunctivitis were included in this study and divided into two groups: children with (n=21) or without (n=67) symptoms during this period. The positivity rate to orchard grass pollen and the rate of residence in the surrounding area of rice field were high (81%, P=0.008 and 86%, P<0.001, respectively) in children with allergic symptoms, as compared to the values (48% and 27%, respectively) in children without symptoms. As the rice pollen season in Nagano occurs in the first decade of August, we measured rice pollen-specific IgE antibodies in 8 patients with symptoms during this period; all of these patients showed positive IgE antibodies to rice pollen. The RAST-inhibition assay using orchard grass and rice pollen indicated cross-allergenecity between these two pollen and also the existence of rice pollen-specific allergens. These results suggest that rice pollen induces seasonal asthma and allergic rhinoconjunctivitis during the first decade of August, which is the rice pollen season, in the surrounding area of rice fields.

Adolescent↗

Impacts by heavy-oil spill from the Russian tanker Nakhodka on intertidal ecosystems: recovery of animal community.

The impact of a heavy-oil spill from the Nakhodka on an intertidal animal community, and the recovery process of animals from the damage were surveyed from the autumn of 1997 to the spring of 2001. The field study was carried out in the rocky coast of Imago-Ura Cove, located along the Sea of Japan, where clean-up operations for oil pollution had been conducted less intensely than in other polluted areas. We have examined individual number of each animal taxon by continuously placing a quadrat of 5 m width along the entire intertidal zone of the cove. A total of 76 invertebrate taxa including 57 species of mollusks, 10 species of crustaceans were observed during the survey. The number of taxa increased from 1998 to 1999 in areas where the initial oil pollution was intense. Total individual number of benthic animals continued to increase from 1998 to 2000 in the polluted areas. The impact of oil on benthic animals was different from species to species. Some species such as Cellana toreuma and Monodonta labio confusa increased rapidly after the oil spill, whereas other species such as Patelloida saccharina lanx and Septifer virgatus did not show any apparent temporal tendencies. Population size structure of P. saccharina lanx varied greatly among years, however that of M. labio confusa did not. For P. saccharina lanx, recruitment was unsuccessful in 1997, possibly due to the effect of oil pollution. These differences in responses to oil pollution among benthic animals are considered to be caused by the differences in habitat use, susceptibility to heavy-oil, life history and migration ability. The findings suggest that it took at least 2-3 years for the intertidal animal community to recover to its original level after the oil spill.

Accidents↗

Schizocladia ischiensis: a new filamentous marine chromophyte belonging to a new class, Schizocladiophyceae.

A new marine filamentous chromophyte Schizocladia ischiensis sp. nov. is described from Naples, Italy, and a new class, Schizocladiophyceae, is proposed to accommodate the species based on morphology, photosynthetic pigment analysis, and rbcL and 18S rRNA gene sequences. The vegetative thallus is composed of branched filaments, 3-7 microm in diameter, containing one to two light brown parietal plastids. Cell walls are composed of layered fibers containing alginates, but lacking cellulose. Plastids are of the typical chromophyte type, containing chlorophylls a and c, and abundant fucoxanthin. Zoospores are formed by direct transformation of vegetative cells or through a process including a multinucleated cell stage. Zoospores are teardrop-shaped with a longer anterior flagellum with tubular mastigonemes and a shorter smooth posterior flagellum with a basal swelling. Flagella have a single basal plate and multi-gyred transitional helix distal to the basal plate. Each zoospore has an eyespot. Phylogenetic analyses using rbcL and 18S rDNA sequences suggest the closest phylogenetic relationship with Phaeophyceae, and then with Xanthophyceae and Phaeothamniophyceae. Nevertheless, Schizocladia differs from Phaeophyceae in some essential features (i.e. cell wall lacking cellulose and plasmodesmata, presence of flagellar transitional helix). Therefore, an independent class Schizocladiophyceae is proposed to accommodate this new taxon.

Animals↗

[Multi-center analysis of calcinosis in children with juvenile dermatomyositis].

OBJECTIVES: To reveal the frequency and the clinical characteristics of dystrophic calcification that occurs in children with juvenile dermatomyositis, multi-center analysis was constructed. METHOD: Fifty children with JDM were enrolled, and 14 of them (28.0%) were complicated with calcinosis. Clinical symptoms and laboratory tests at onset, initial therapy and disease course were compared in children with and without calcinosis. RESULTS: The mean age of the onset of calcinosis was 4.78 +/- 3.33 years, and it was younger than those of children without calcinosis (8.66 +/- 3.85 years) (P = 0.0017). No differences of clinical manifestation except Gower's sign were observed. The frequency of positive anti-nuclear antibody was 7.1% in children with calcinosis and 52.9% without calcinosis (P = 0.0112). The initial therapy of methylprednisolon pulses gave no effects on prognosis of calcium deposition. The calcinosis appeared in 1.56 +/- 1.91 year after the onset of the disease. The various types of calcium deposition including large tumorous clumps, subcutaneous plaques or nodules, sheet-type calcification were deserved. They appeared over knee joints (64.3%), elbow joint (64.3%), and hip processes (50.0%). Calcinosis affecting the subcutaneous tissues frequently resulted in painful superficial ulceration of the overlying skin (42.9%), local infection (50.0%), and limitation of joint movement (14.3%). Although aluminum phosphate was effective in 2 children among 7, no other effective treatment was recommended. In 5 cases, surgical removal of tumorous clumps was operated. Thus, juvenile dermatomyositis is frequently complicated with calcinosis. This type of calcinosis was found to be unlikely to resolve completely, and resulted in severe disability in children.

Age of Onset↗

Photomovement of the swarmers of the brown algae Scytosiphon lomentaria and Petalonia fascia: effect of photon irradiance, spectral composition and UV dose.

Photomovement measurements were carried out with swarmers of the brown algae Scytosiphon lomentaria (Lyngb.) Link and Petalonia fascia (O. F. Müll.) as a function of irradiance direction, photon irradiance, spectral composition and ultraviolet radiation (UVR, lambda=280-400 nm) dose. Swarmers from both species showed similar photomovement patterns: negative phototaxis occurred under photon irradiances of 10-90 micromol photons m(-2) s(-1), and no movement was observed at 190 micromol photons m(-2) s(-1). The translocational velocity measured between 10 and 90 micromol m(-2) s(-1) ranged from 100 to 200 microm s(-1). The accumulation of swarmers presented a peak at 450 nm (waveband of 50 nm), and smaller peaks at 400 and 500 nm; no effect was observed at wavelengths of 550 nm and above. The decline in phototactic index (an estimator of photomovement response) of swarmers was linearly correlated with the logarithm of UVR doses. These data were correlated with levels of natural solar radiation in the field. It is hypothesized that motility of swarmers could be a critical factor in the survival of these species under a scenario of increased UVR.

Phaeophyceae↗