Biomedical subjects

T Yoshimura

Publications and source records attributed to T Yoshimura.

At least 307 records · Page 17Linked to original sources

PubMed1996

Spectral sensitivity of photoreceptors mediating phase-shifts of circadian rhythms in retinally degenerate CBA/J (rd/rd) and normal CBA/N (+/+)mice.

Light-dark cycles are the most important time cue for the circadian system to entrain the endogenous circadian clock to the environmental 24 h cycle. Although photic entrainment of circadian rhythms is mediated by the eye in mammals, photoreceptors implicated in circadian photoreception remain unknown. In our previous study, retinally degenerate CBA/J (rd/rd) mice were found to have lower circadian photosensitivity for phase-shifting the locomotor activity rhythms than normal CBA/N(+/+) mice. In the present study, the spectral sensitivity for phase-shifting the rhythms was examined in order to characterize the photopigments involved in circadian photoreception of these mice. The spectral sensitivity of CBA/J-rd/rd mice clearly fitted to the Dartnall nomogram for a retinal(1)-based pigment with a maximum at 480 nm, while the best fitted nomogram had a maximum at 500 nm in CBA/N- +/+ mice. These results suggest that circadian photopigments involved in CBA/J-rd/rd and CBA/N- +/+ mice may be different.

Animals↗

PubMed1996

The cytokines NAP-1 (IL-8), MCP-1, IL-1 beta, and GRO in rabbit inflammatory skin lesions produced by the chemical irritant sulfur mustard.

Developing and healing dermal inflammatory lesions were produced in rabbits by the topical application of dilute sulfur mustard (SM), the military vesicant. In tissue sections of such lesions, cells containing the mRNA of important cytokines were identified with in situ hybridization techniques. These cytokines were neutrophil attractant/activation protein-1 (NAP-1 (also called IL-8), monocyte chemoattractant (activating) protein 1 (MCP-1), interleukin 1 (beta) (IL-1 (beta)), and GRO (a growth factor and chemokine). Mononuclear cells (mainly macrophages and activated fibroblasts) contained the mRNA of all four of these cytokines. A higher percentage of cytokine-producing mononuclear cells (macrophages and activated fibroblasts) was present in lesions at 2 days (their peak size) than at 6 days, when they were almost healed. Granulocytes emigrated from the bloodstream, passed through the lesions, and were the major constituent of the protective crust. This sequence correlated with the distribution of cells able to produce NAP-1: At 2 days and 6 days, the mononuclears that contained messenger RNA for this granulocyte chemoattractant were found mainly in the upper part of the dermis. At 2 days and 6 days, cells containing the mRNA of IL-1, a primary cytokine, were also found predominantly in the upper dermis, i.e., nearest the site of injury. In contrast, mononuclears containing the mRNA of MCP-1 (a monocyte chemoattractant), and the mRNA of GRO (a granulocyte chemoattractant) were more equally distributed throughout the dermis. SM stimulated hair follicle epithelial cells to up-regulate GRO mRNA and, to a lesser degree, NAP-1 mRNA. Apparently, the irritation produced by SM directly or indirectly induces such epithelial cells to manufacture these growth factors. In the rabbit, hair follicles are known to be the main source of new epithelial cells after the covering epithelium has been destroyed. Therefore, GRO is probably a major autocrine-paracrine stimulus for such repair. A brief review of the role of cytokines in dermal inflammation is presented.

Animals↗

PubMed1996

Immunohistochemical study of utrophin and dystrophin at the motor end-plate in myasthenia gravis.

We studied the densities of utrophin and dystrophin at the motor end-plates of patients with myasthenia gravis (MG) using immunohistochemical analysis. The densities were compared with those found in patients with amyotrophic lateral sclerosis, Lambert-Eaton myasthenic syndrome and normal controls. Utrophin was reduced at the motor end-plates of MG patients, in association with a reduction of alpha-bungarotoxin binding sites. In contrast, the density of dystrophin at the motor end-plate of MG patients was not significantly different from that found in the controls. We conclude that, at the motor end-plate, utrophin may be more closely associated than dystrophin with the acetylcholine receptor, and that it plays a different role.

Cytoskeletal Proteins↗

PubMed1996

Effects of nimesulide, a preferential cyclooxygenase-2 inhibitor, on carrageenan-induced pleurisy and stress-induced gastric lesions in rats.

Intrapleural injection of carrageenan in rats increased prostaglandin E2 (PGE2) production and induced newly synthesized cyclooxygenase-2 (COX-2) in pleural exudate cells without affecting COX-1 levels. Nimesulide, a preferential inhibitor of COX-2, reduced pleural PGE2 production and was almost as active as indomethacin and 10 times more active than ibuprofen. Only COX-1, and nc COX-2, was detected in gastric mucosal cells, and PGE2 concentration of gastric mucosa was significantly decreased by indomethacin and ibuprofen. The decrease in gastric PGE2 production induced by indomethacin and ibuprofen was enhanced in stressed rats, resulting in aggravation of stress-induced gastric lesions at anti-inflammatory doses. However, nimesulide did not produce stress-induced gastric lesions even at 30 times the anti-inflammatory dose. This supports the hypothesis that inhibition of COX-1 causes unwanted side effects and inhibition of COX-2 produces anti-inflammatory effects.

Animals↗

PubMed1996

Differential expression of macrophage inflammatory protein-2 and monocyte chemoattractant protein-1 in experimental glomerulonephritis.

We examined the relation between glomerular expression of chemokines from alpha-subfamily (macrophage inflammatory protein-2, MIP-2) and beta-subfamily (monocyte chemoattractant protein-1, MCP-1) and infiltration of neutrophils and monocytes in antibody mediated glomerulonephritis in rats. In the accelerated model of nephrotoxic nephritis (NTN), glomerular expression of MIP-2 and MCP-1 genes correlated with the sequential migration of neutrophil and monocyte influx, respectively. These relationships were investigated further in the heterologous phase of NTN by applying various treatments known to modulate the severity of injury. Pretreatment with bacterial lipopolysaccharide resulted in greater injury, MIP-2 expression increased 25- to 50-fold, and the glomerular neutrophil count increased two- to fourfold. Both MIP-2 mRNA levels and neutrophil infiltration were reduced by additional pretreatment with IL-6, IL-1 receptor antagonist, soluble IL-1 receptor or soluble TNF receptor (Spearman correlation coefficient r = 0.897, P < 0.005). In the heterologous phase of NTN, different pre-treatments only resulted in trivial changes in MCP-1 expression and monocyte infiltration. In conclusion, glomerular MIP-2 gene expression correlates with neutrophil infiltration both temporally during the evolution of nephritis, and when glomerular injury is modified by treatment. Glomerular MCP-1 gene expression correlates with monocyte influx. The data show chemokines of alpha- and beta-subfamilies co-operative to cause selective and sequential migration of different leukocyte subsets during development of antibody mediated glomerulonephritis.

Animals↗

PubMed1996

In vivo EPR detection and imaging of endogenous nitric oxide in lipopolysaccharide-treated mice.

Nitric oxide (NO), a simple diatomic free radical, is known to play a critical physiological role in diverse organisms. An iron complex, with N-(dithiocarboxy)sarcosine (Fe-DTCS), has a high affinity for endogenous NO and can trap, stabilize, and accumulate it. The stable NO adduct thus formed is detectable at room temperature with electron paramagnetic resonance (EPR) spectrometry. We report in vivo EPR imaging of endogenous NO, trapped by an Fe-DTCS complex, in the abdomen of a live mouse. To our knowledge, this is the first report on EPR imaging of endogenous free radicals produced in vivo. This EPR imaging method will be useful for the noninvasive investigation of the spatial distribution of NO in pathologic organs or tissues.

Animals↗

PubMed1996

Connexin43 is another gap junction protein in the peripheral nervous system.

That many cells express more than one connexin (Cx) led us to examine whether Cxs other than Cx32 are expressed in the PNS. In addition to Cx32 mRNA, Cx43 and Cx26 mRNAs were detected in rat sciatic nerve by northern blot analysis. Cx43 mRNA, but not Cx26 mRNA, was expressed in both the primary Schwann cell culture and immortalized Schwann cell line (T93). The steady-state levels of the Cx43 mRNA in the primary Schwann cell culture increased 2.0-fold with 100 microM forskolin, whereas that of Po increased 7.0-fold. Immunoreactivity to Cx43 was detected on western blots of cultured Schwann cells, T93 cells, and sciatic nerves but not on blots of PNS myelin. Immunohistochemical study using human peripheral nerves revealed that anti-Cx43 antibody stained cytoplasm around nucleus of Schwann cells but not myelin, confirming western blot results. Although Po expression was markedly decreased by crush injury of the sciatic nerves, Cx43 expression showed no apparent change. Developmental profiles showed that Cx43 expression in the sciatic nerve increased rapidly after birth, peaked at about postnatal day 6, and then decreased gradually to a low level. In adult rats, the Cx43 mRNA value was much lower than that of Cx32. These findings suggest that Cx43 is localized in Schwann cell bodies and that, compared with Po, its expression is less influenced by axonal contact and cyclic AMP levels. The high expression on postnatal day 6 indicates that Cx43 may be related to PNS myelination. Cx43 is another gap junction, but its function appears to differ from that of Cx32, as judged by the differences in their localization and developmental profiles.

Animals↗

PubMed1996

Enzyme immunoassay for conjugated cholic and 1 beta-hydroxycholic acids in urine of early infancy.

A direct competitive heterologous enzyme immunoassay (EIA) for conjugated cholic acid (CA) was developed using horseradish peroxidase labeled antigen having a shorter bridge length than that of the immunogen. An appropriate dose-response curve for conjugated CA was obtained in the range of 0.05-50 ng/well. Specificity of the EIA proved satisfactory in terms of cross-reactivities to 23 kinds of related bile acids. The proposed method was evaluated to be useful for the determination of conjugated CA in urine with acceptable accuracy and inter- and intra-assay precision. The results of analysis showed a reverse relationship between age and urinary excretion ratio of conjugated 1 beta-hydroxy-CA to conjugated CA in the first 9 months after birth.

Bile Acids and Salts↗

PubMed1996

Activities of an occupational health organization in Japan, in special reference to services for small- and medium-scale enterprises.

This paper reports the activities of occupational health organizations in Japan, taking Nishinihon Occupational Health Service Center, one of the largest scale occupational health service providers, as an example. The organizations, at the request of enterprises, provide occupational health services on a profit basis especially for small- and medium-scale enterprises which lack the human resources and/or facilities to meet legal requirements on occupational health. The main services include: (1) providing various types of medical examinations for workers, (2) measuring work environment, (3) occupational physicians and occupational health nurses providing a comprehensive service. Among these services, legally required health examinations and work environment measurements are well served even for small-scale enterprises. Less effective are the health promotion and occupational health service by the occupational health staffs. In the future, these service will be better provided by a newly developed network system to support occupational heath activities in small-scale enterprises.

Industry↗

PubMed1996

Differential expression of three genes for different beta-tubulin isotypes during the initial culture of Zinnia mesophyll cells that divide and differentiate into tracheary elements.

Complementary DNA clones for three different beta-tubulin isotypes (ZeTubB1, ZeTubB2 and ZeTubB3) were isolated from a cDNA library generated from RNA of cultured mesophyll cells of Zinnia elegans that were differentiating into tracheary elements and/or dividing. Sequence analysis revealed that the proteins encoded by ZeTubB1 and ZeTubB3 and that encoded by ZeTubB2 were each homologous to two of three groups of beta-tubulin isotypes in Arabidopsis. RNA gel blot analysis of the expression of the ZeTubB transcripts indicated that transcripts that corresponded to each clone were differently expressed during culture of Zinnia mesophyll cells. In particular, the level of expression of ZeTubB1 and ZeTubB3 transcripts increased rapidly prior to cell division and secondary wall formation, and such expression was promoted by combinations of auxin and cytokinin that induced tracheary element differentiation as well as cell division. Results of an in situ hybridization experiment with an antisense RNA probe derived from ZeTubB1 cDNA suggested the preferential expression of ZeTubB transcripts in differentiating xylem cells, as well as in the ground meristem and the procambium, of Zinnia seedlings.

Amino Acid Sequence↗

PubMed1996

Detection of K-ras point mutations at codon 12 in pancreatic juice for the diagnosis of pancreatic cancer by hybridization protection assay: a simple method for the determination of the types of point mutation.

The present study was undertaken to detect K-ras oncogene point mutations at codon 12 in pure pancreatic juice (PPJ) by the hybridization protection assay (HPA) method for the diagnosis of pancreatic cancer (PC). This assay can be carried out within 30 min and can determine not only the presence of a mutation, but also the mutational type of K-ras at codon 12. The minimal ratio of mutant DNA detectable by the HPA was 5-10% of the total DNA. PPJ was collected through a cannula under duodenal fiberscope control from 20 patients with PC and 20 patients with chronic pancreatitis (CP). Analysis of PPJ by the HPA revealed that the incidence of K-ras point mutations at codon 12 was 55% (11/20) in patients with PC and 0% (0/20) in those with CP. Mutational types of K-ras at codon 12 in PC were aspartic acid (Asp) in nine cases, both Asp and cysteine in one case, and arginine in one case. Analysis of K-ras point mutations at codon 12 in PPJ using the HPA method seems promising as a new genetic test for the diagnosis of PC, because the HPA method is simple, and can easily determine the mutational type.

Base Sequence↗

PubMed1996

Production of monocyte chemotactic protein-1 by rat brain macrophages.

In the present study, we show that cultured rat brain macrophages release a soluble factor that stimulates the migration of bone marrow-derived macrophages, as determined by an in vitro chemotaxis assay. A checkerboard analysis indicated that most of this effect resulted from a polarized migration of the cells (chemotactic phenomenon), rather than in an increase in cell motility (chemokinesis). This activity was significantly decreased by an immune serum directed against the rat monocyte chemoattractant protein-1 (chemokine MCP-1). Northern blot analysis demonstrated expression of the MCP-1 gene in cultured brain macrophages, but its absence in unstimulated bone marrow-derived macrophages. Up-regulation of MCP-1 expression was observed when lipopolysaccharide was added to cultured brain macrophages, a peak occurring after a 6 h period of stimulation. Also, inflammatory cytokines such as interleukin (IL)-1 beta, colony stimulating factor-1, tumour necrosis factor-alpha and IL-6 individually increased the basal level of MCP-1 mRNA. Subsequently, we demonstrated the in vivo production of MCP-1 in the adult rat brain following injury induced by a local injection of kainic acid. MCP-1 synthesis was localized in both astrocytes and brain macrophages. These results suggest that the activation of resting microglial cells into brain macrophages and their subsequent secretion of chemokines could contribute to the mechanism(s), leading to the infiltration of the CNS by blood-derived monocytes, as observed in several pathologies.

Animals↗

PubMed1996

Differential effects of protein kinase C inhibitors on chemokine production in human synovial fibroblasts.

1. Rheumatoid arthritis is associated with the accumulation and activation of selected populations of inflammatory cells within the arthritic joint. One putative signal for this process is the production, by resident cells, of a group of inflammatory mediators known as the chemokines. 2. The chemokines interleukin-8 (IL-8), monocyte chemotactic protein-1 (MCP-1) and RANTES (regulated on activation normal T-cell expressed and presumably secreted) are target-cell specific chemoattractants produced by synovial fibroblasts in response to stimulation with interleukin-1 alpha (IL-1 alpha) or tumour necrosis factor alpha (TNF alpha). The signalling pathways involved in their production are not well defined. We therefore used four different protein kinase C inhibitors to investigate the role of this kinase in the regulation of chemokine mRNA and protein expression in human cultured synovial fibroblasts. 3. The non-selective PKC inhibitor, staurosporine (1-300 nM) significantly increased the production of IL-1 alpha-induced IL-8 mRNA and protein. A specific PKC inhibitor, chelerythrine chloride (0.1-3 microM), also caused a small concentration-dependent increase in IL-8 mRNA and protein production. In contrast, 3-[1-[3-(amidinothio)propyl]-3-indoly]-4-(1-methyl-3-indolyl )- 1H-pyrrole-2,5-dione methanesulphonate (Ro 31-8220) and 2[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3- yl)-maleimide (GF 109203X), two selective PKC inhibitors of the substituted bisindolylmaleimide family had a concentration-dependent biphasic effect on IL-1 alpha or TNF alpha-induced chemokine expression. At low concentrations they caused a stimulation in chemokine production, which was especially evident at the mRNA level. At higher concentrations both inhibited IL-1 alpha or TNF alpha-induced chemokine mRNA and protein production. Ro 31-8220 was 10 fold more potent than GF 109203X, with an IC50 of 1.6 +/- 0.08 microM (mean +/- s.e.mean, n = 4) for IL-1 alpha induced IL-8 production. Ro 31-8220 also inhibited the expression of IL-1 alpha or TNF alpha-induced MCP-1 and RANTES mRNA with a similar potency. 4. The stimulatory effect of staurosporine is discussed in relation to the known poor selectivity of this inhibitor for PKC. It is proposed that activation of an isoform of PKC, possibly PKC epsilon or zeta, which is inhibited by higher concentrations of the bisinodolylmaleimides, plays a role in the regulation of chemokine expression induced by IL-1 alpha or TNF alpha in synovial cells. 5. The inhibition of chemokine production by bisindolylmaleimide compounds heralds a novel approach for future anti-inflammatory therapies.

Alkaloids↗

PubMed1996

Congenital plantar melanocytic nevus with satellite lesions.

We report a case of a congenital melanocytic nevus accompanied by a mucous cyst and satellite pigmented macules on the undersurface of the left great toe in a 17-year-old Japanese man. Myxoid change was confirmed by the discharge of viscous liquid when conducting biopsy of the tumor. Besides such an unusual clinical feature of the tumor, accompanying satellite lesions led us initially to consider the possibility of malignant melanoma. However, a biopsy specimen obtained from the tumor revealed the typical features of intradermal type melanocytic nevus. In a specimen obtained from the satellite pigmented lesion, there was a small mass of nevus cells underneath the basement membrane zone, suggestive of having originated from the spread of nevus cells through lymphatic vessels.

Adolescent↗

PubMed1996

Intra-alveolar macrophage-inflammatory peptide 2 induces rapid neutrophil localization in the lung.

Endotoxin-induced lung injury is characterized by neutrophil infiltration of the lungs. The various mechanisms which mediate movement of neutrophils from vascular space to lung interstitium and alveoli remain unclear. Macrophage-inflammatory protein 2 (MIP-2) is a potent chemoattractant for neutrophils and may play a significant role in recruiting neutrophils in acute lung injury in rats. Experiments were performed in male Sprague Dawley rats to: (1) evaluate the kinetics of neutrophil influx in the lung following intraperitoneal administration of Salmonella enteritidis lipopolysaccharide (LPS); (2) determine the expression of transcripts for chemokines and adhesion molecules in the lung following intraperitoneal LPS; and (3) elucidate the effects of intra-alveolar instillation of recombinant rat MIP-2 on neutrophil influx into the lung. Intraperitoneal LPS resulted in an increase in neutrophil sequestration in the lung capillaries of rats as early as 45 min following administration, and there was a parallel increase in lung myeloperoxidase activity. There were also major increases in mRNA in whole-lung homogenates of LPS-treated rats for chemokines MIP-2 and KC (cytokine-induced neutrophil chemoattractant) and adhesion molecules P- and E-selectin at 1 and 2 h following LPS. When recombinant rat MIP-2 was instilled into the alveolar space of rats through a catheter wedged into a bronchus, there was profound neutrophil localization both in the vascular and alveolar space which significantly differed (P < 0.05) from the contralateral lungs of the same animals, and lungs of control animals instilled with control buffer. These observations reveal that MIP-2 is a potent chemoattractant in rat lungs, and suggest that chemoattractants locally released in alveoli can recruit neutrophils to those alveoli. This suggests that alveolar macrophages may play an important role in neutrophil sequestration in sepsis and other inflammatory lung diseases which produce a neutrophilic alveolitis.

Animals↗

PubMed1996

Comparison of thermal and guanidine hydrochloride denaturation behaviors of glucoamylase from the STA1 gene of Saccharomyces cerevisiae var. diastaticus.

To investigate the relationships between enzyme inactivation and conformational change, the effects of heat and guanidine hydrochloride (GnHC1) on the STA1 gene glucoamylase (STA1GA) of Saccharomyces cerevisiae var. diastaticus were examined by circular dichroism and fluorescence spectroscopies. A conformational change was observed in the thermal denaturation of STA1GA, while extensive enzyme inactivation occurred in GnHC1 denaturation before noticeable conformational change.

Circular Dichroism↗

PubMed1996

Stereospecificity for the hydrogen transfer and molecular evolution of pyridoxal enzymes.

We here describe the stereochemical aspects of the reactions of pyridoxal 5'-phosphate (PLP)-dependent enzymes, and the relationship between the stereochemistry of the enzyme reaction and molecular evolution of the enzyme. The reactions of PLP-dependent enzymes proceed through the formation of an anionic Schiff base intermediate between the substrate and the coenzyme. Three stereochemical possibilities exist for the formation and cleavage of bonds in the intermediate: the reaction occurs stereospecifically on either the si- or the re-face of the planar intermediate, or alternatively, non-stereospecifically on both faces. The stereospecificities for hydrogen transfer between C-4' of the cofactor and substrate in the transamination catalyzed by various PLP-dependent enzymes have been studied. The stereospecificities reflect the active-site structures of the enzymes, especially the topographical situation of a coenzyme-substrate Schiff base and a catalytic base for the hydrogen transfer. The aminotransferases and other PLP-enzymes catalyzing the transamination as a side-reaction so far studied catalyze only the si-face specific hydrogen transfer. This suggests that these PLP enzymes have similar active-site structures and are evolved divergently from a common ancestral protein. We recently established a new method for the identification of stereospecificity for the hydrogen transfer, and found that D-amino acid aminotransferase and branched chain L-amino acid aminotransferase, which have significant sequence similarity to each other, catalyze the re-face hydrogen transfer on the intermediate. The X-ray crystallographic studies of D-amino acid aminotransferase showed that the relative arrangement of the catalytic base of the enzyme active center to the C4' of the bound cofactor is opposite to that of other aminotransferases catalyzing the si-face hydrogen transfer. The folding of D-amino acid aminotransferase is also different from those of the other aminotransferase so far studied. Therefore, the classifications of the aminotransferases based on their primary structures, three dimensional structures, and stereochemistry of their hydrogen transfer coincide with one another. We also found that PLP-dependent amino acid racemases, the primary structures of which are similar to none of the other PLP-enzymes, catalyze the non-stereospecific hydrogen transfer on both faces of the planar intermediate. Stereospecificities for the hydrogen transfer suggest convergent evolution of the PLP-dependent enzymes. The stereochemical aspects of the enzyme reactions give a clue to the molecular evolution of the enzymes as well as the primary structures and three-dimensional structures of the enzymes.

Amino Acids↗

PubMed1996

Hypercholesterolemia and the progression of the renal dysfunction in chronic renal failure patients.

The effect of hyperlipidemia on the progression of chronic renal failure was investigated in 104 chronic renal failure patients, aged 39.3 +/- 2.9 years. The follow up period was 4.1 +/- 2.9 years. The serum creatinine level was 2.1 +/- 1.1 (mean +/- SD) mg/dl at the beginning of study and increased to 8.7 +/- 4.4 mg/dl at the end of the study. The reciprocal serum creatinine concentration (1/Cr) was plotted against the observation time, and the slope was calculated. The absolute value of the slope was used as the progression rate of renal impairment. The progression rate was positively related to total cholesterol level or urinary protein score, while it was negatively related to total protein level. Without the influence of urinary protein score, the progression rate correlated with total cholesterol level. The result suggests that hypercholesterolemia may be an independent aggravating factor in the progression of renal dysfunction in chronic renal failure patients.

Adolescent↗