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Biomedical subjects

Y Uchiyama

Publications and source records attributed to Y Uchiyama.

At least 37 records · Page 2Linked to original sources
PubMed2001

Caspase activity is involved in, but is dispensable for, early motoneuron death in the chick embryo cervical spinal cord.

We examined the role of caspases in the early programmed cell death (PCD) of motoneurons (MNs) in the chick embryo cervical cord between embryonic day (E) 4 and E5. An increase in caspase-3-like activity in MNs was observed at E4.5. Treatment with an inhibitor of caspase-3-like activity, Ac-DEVD-CHO, for 12 h blocked this increase and revealed that caspase-3-like activity is mainly responsible for DNA fragmentation and the nuclear changes during PCD but not for degenerative changes in the cytoplasm. When a more broad-spectrum caspase inhibitor was used (bocaspartyl (OMe)-fluoromethyl ketone, BAF), the appearance of degenerative changes in the cytoplasm was delayed by at least 12 h. However, following treatment with either Ac-DEVD-CHO or BAF for 24 h, the number of surviving healthy MNs did not differ from controls, indicating a normal occurrence of PCD despite the inhibition of caspases. These results suggest that caspase cascades that occur upstream of and are independent of the activation of caspase-3-like activity are responsible for the degenerative changes in the cytoplasm of dying cervical MNs. These data also suggest that, although one function of caspases may be to facilitate the kinetics of PCD, caspases are nonetheless dispensable for at least some forms of normal neuronal PCD in vivo.

Amino Acid Chloromethyl Ketones↗
PubMed2001

Relationship between functional deficit and severity of experimental fast-strain injury of rat skeletal muscle.

We developed a rat model of fast-strain muscle injury to examine the relationship between functional deficit and the degree of muscle damage. A single fast strain was applied to the right plantaris (Plt) muscle of adult male rats. A tetanic contraction was induced by stimulating the sciatic nerve. Three types of strain injuries were produced by manipulating the timing of strain and contraction conditions, including applying the strain at: (1) the shortening phase (SP), (2) the full contraction phase (FCP) during tetanic contraction and (3) when the muscle is not contracting, i.e. non-contraction (NC). The contralateral Plt muscle was used as the control. Morphological and functional analysis were performed and the severity of strain injury was estimated by measuring [3H]thymidine labelling 48 h after the strain was applied. The results show a significant decrease in tension output in all three groups at 5 min after strain application (P < 0.01). The functional deficit lasted for 3 weeks in the NC group, while rats of the SP and FCP groups showed recovery 1 week after strain. Increased uptake of [3H]thymidine was similar in SP and FCP groups, but 3.5-fold higher in NC than in SP and FCP groups (P < 0.01). Histological analysis revealed an increase in the size of interstitial spaces of the muscle in NC compared to SP and FCP groups (P<0.05). Our results suggest that the contraction of muscle fibres limits the severity of connective tissue damage, and that muscle damage accompanied by disruption of the muscle connective tissue network requires a relatively longer recovery time.

Animals↗
PubMed2001

VCP/p97 in abnormal protein aggregates, cytoplasmic vacuoles, and cell death, phenotypes relevant to neurodegeneration.

Neuronal cell death, abnormal protein aggregates, and cytoplasmic vacuolization are major pathologies observed in many neurodegenerative disorders such as the polyglutamine (polyQ) diseases, prion disease, Alzheimer disease, and the Lewy body diseases, suggesting common mechanisms underlying neurodegeneration. Here, we have identified VCP/p97, a member of the AAA+ family of ATPase proteins, as a polyQ-interacting protein in vitro and in vivo, and report on its characterization. Endogenous VCP co-localized with expanded polyQ (ex-polyQ) aggregates in cultured cells expressing ex-polyQ, with nuclear inclusions in Huntington disease patient brains, and with Lewy bodies in patient samples. Moreover, the expression of VCP mutants with mutations in the 2nd ATP binding domain created cytoplasmic vacuoles, followed by cell death. Very similar vacuoles were also induced by ex-polyQ expression or proteasome inhibitor treatment. These results suggest that VCP functions not only as a recognition factor for abnormally folded proteins but also as a pathological effector for several neurodegenerative phenotypes. VCP may thus be an ideal molecular target for the treatment of neurodegenerative disorders.

Adenosine Triphosphatases↗
PubMed2001

Evaluation of the disposable Vital View laryngoscope apparatus.

The Vital View laryngoscope (Vital Signs, NJ, USA) consists of a plastic disposable blade containing a fibrelight and a non-disposable handle; there is therefore no need to sterilise the blade and no concern about disintegration of the fibrelight. In a random cross-over design, we compared the Vital View laryngoscope with a conventional metal fibrelight laryngoscope (Welch Allyn, NY, USA) in 100 patients. The Vital View laryngoscope produced a brighter field than the metal laryngoscope (p < 0.001), whereas there was no significant difference in the view of the glottis or the success rate of tracheal intubation. In no patient did any problem occur, such as damage to the laryngoscope blade or loss of light during laryngoscopy. In another 10 patients, prevention of light emission from the side of the laryngoscope blade reduced the brightness (p < 0. 01). This indicated that the brightness of the Vital View laryngoscope is produced by light emission not only from the tip of the blade but also from the side of the blade. Therefore, the disposable Vital View laryngoscope can be used as effectively as a conventional non-disposable laryngoscope.

Adolescent↗
PubMed2001

Parkin is associated with cellular vesicles.

We recently identified a novel gene, parkin, as a pathogenic gene for autosomal recessive juvenile parkinsonism. Parkin encodes a 52-kDa protein with a ubiquitin-like domain and two RING-finger motifs. To provide a insight into the function of parkin, we have examined its intracellular distribution in cultured cells. We found that parkin was localized in the trans-Golgi network and the secretory vesicles in U-373MG or SH-SY5Y cells by immunocytochemical analyses. In the subsequent subcellular fractionation studies of rat brain, we showed that parkin was copurified with the synaptic vesicles (SVs) when we used low ionic conditions throughout the procedure. An immunoelectromicroscopic analysis indicated that parkin was present on the SV membrane. Parkin was readily released from SVs into the soluble phase by increasing ionic strength at neutral pH, but not by a non-ionic detergent. To elucidate its responsible region for membrane association, we transfected with green fluorescent protein-tagged deletion mutants of parkin into COS-1 cells followed by subcellular fractionation. We demonstrated the ability of parkin to bind to the membranes through a broad region except for the ubiquitin-like domain. The significance of SV localization of parkin is discussed.

Animals↗
PubMed2001

Hemodynamic analysis of esophageal varices using color Doppler endoscopic ultrasonography to predict recurrence after endoscopic treatment.

BACKGROUND AND STUDY AIMS: The time to recurrence of esophageal varices may vary greatly between patients even after the same endoscopic therapy. To clarify the factors which contribute to recurrence after endoscopic treatment, the hemodynamics and morphology of the left gastric vein (LGV) were investigated using color Doppler endoscopic ultrasonography (EUS). PATIENTS AND METHODS: A total of 31 patients with high-risk esophageal varices underwent color Doppler-EUS before receiving endoscopic variceal ligation and endoscopic injection sclerotherapy combined therapy. Endoscopic examination was performed every 3 months after the treatment to evaluate recurrence of varices. RESULTS: A total of 18 patients responded to the therapy, while 13 patients did not respond, and had recurrence within 12 months. The hepatofugal flow velocity in the LGV trunk was significantly lower in the responders (9.9 vs. 13.9 cm/sec; P = 0.02). The branch pattern of the LGV was categorized into three groups: anterior branch dominant, posterior branch dominant, and no-dominant type. The incidence of the anterior branch dominant type was significantly less in responders (17 vs. 70 %; P = 0.01). There was no significant difference in the LGV trunk diameter and the size of the paraesophageal vein between the two groups. CONCLUSION: Risk factors for recurrence can be analyzed in detail using color Doppler-EUS. Further investigation using color Doppler-EUS may enable us to select the optimal way to treat esophageal varices to prevent recurrence.

Adult↗
PubMed2001

Low power diode laser treatment using indocyanine green for eradication of esophageal varices.

BACKGROUND AND STUDY AIMS: Endoscopic variceal ligation (EVL) is an alternative to sclerotherapy for the treatment of esophageal varices, but is associated with higher rates of recurrence and subsequent bleeding than sclerotherapy. To prevent recurrence of varices after EVL, we have developed a low-dose diode laser therapy combined with the injection of indocyanine green, which allows enhanced tissue absorption of the laser beam selectively around varices. In this study we investigated the efficacy and safety of this technique. PATIENTS AND METHODS: Eight patients with F2 or F3 esophageal varices were enrolled. At 1 week after EVL, indocyanine green solution (1 mg/ml) was injected submucosally around the remaining varices. A diode laser (power 10 watts) was applied to the surface from the esophagogastric junction to 5 cm above it. The spot size was kept to 5 mm in diameter. RESULTS: Laser irradiation was performed safely, without bleeding from the varices, or perforation. There were no major complications. Endoscopy 1 month later showed F0 forms in seven patients, F1 in one patient, and no red color sign in any patient. No recurrence of varices has been observed in any of the patients during the follow-up period of at least 12 months. CONCLUSION: This technique may provide a simple, safe and effective procedure, as an additional treatment to EVL, for the prevention of recurrence of esophageal varices.

Adult↗
PubMed2001

Early and progressive accumulation of reactive microglia in the Huntington disease brain.

Microglia may contribute to cell death in neurodegenerative diseases. We studied the activation of microglia in affected regions of Huntington disease (HD) brain by localizing thymosin beta-4 (Tbeta4), which is increased in reactive microglia. Activated microglia appeared in the neostriatum, cortex, and globus pallidus and the adjoining white matter of the HD brain, but not in control brain. In the striatum and cortex, reactive microglia occurred in all grades of pathology, accumulated with increasing grade, and grew in density in relation to degree of neuronal loss. The predominant morphology of activated microglia differed in the striatum and cortex. Processes of reactive microglia were conspicuous in low-grade HD, suggesting an early microglia response to changes in neuropil and axons and in the grade 2 and grade 3 cortex, were aligned with the apical dendrites of pyramidal neurons. Some reactive microglia contacted pyramidal neurons with huntingtin-positive nuclear inclusions. The early and proximate association of activated microglia with degenerating neurons in the HD brain implicates a role for activated microglia in HD pathogenesis.

Adult↗
PubMed2001

Expression of proteinases and inflammatory cytokines in subchondral bone regions in the destructive joint of rheumatoid arthritis.

OBJECTIVE: We previously described abnormalities in the bone marrow of patients with rheumatoid arthritis (RA), but were able to shed little light on the pathogenic roles of inflammatory cytokines and proteinases in joint destruction in the subchondral region in RA. This is the first report to describe the co-localization of cytokines and proteinases in this area. METHODS: Decalcified paraffin-embedded sections from 10 patients with RA and five patients with osteoarthritis (OA) were examined for the immunolocalization of cathepsins B, K and L and the localization of messenger RNAs for interleukin 1beta (IL-1beta), tumour necrosis factor alpha (TNF-alpha) and matrix metalloproteinase 9 (MMP-9). The cells were double-stained with anti-CD68 or anti-prolyl 4-hydroxylase (PH) antibody. RESULTS: An immunohistochemical study confirmed the expression of cathepsins B and L by CD68-positive mononuclear cells at the sites of significant cartilage and bone erosion from the subchondral region in all RA specimens. Osteoclast-like cells showed intense staining for cathepsin K and MMP-9. Osteoblast-like cells strongly expressed MMP-9. Analysis of serial sections revealed that expression of the IL-1beta and TNF-alpha genes occurred near that of the cathepsins and MMP-9 in the subchondral region. CONCLUSION: We conclude that inflammatory cytokines and tissue-damaging proteinases play important roles in joint destruction in the subchondral region in RA.

Adult↗
PubMed2001

Anabolic steroids increase exercise tolerance.

The influence of an anabolic androgenic steroid (AAS) on thymidine and amino acid uptake in rat hindlimb skeletal muscles during 14 days after a single exhaustive bout of weight lifting was determined. Adult male rats were divided randomly into Control or Steroid groups. Nandrolone decanoate was administered to the Steroid group 1 wk before the exercise bout. [3H]thymidine and [14C]leucine labeling were used to determine the serial changes in cellular mitotic activity, amino acid uptake, and myosin synthesis. Serum creatine kinase (CK) activity, used as a measure of muscle damage, increased 30 and 60 min after exercise in both groups. The total amount of weight lifted was higher, whereas CK levels were lower in Steroid than in Control rats. [3H]thymidine uptake peaked 2 days after exercise in both groups and was 90% higher in Control than in Steroid rats, reflecting a higher level of muscle damage. [14C]leucine uptake was approximately 80% higher at rest and recovered 33% faster postexercise in Steroid than in Control rats. In a separate group of rats, the in situ isometric mechanical properties of the plantaris muscle were determined. The only significant difference was a higher fatigue resistance in the Steroid compared with the Control group. Combined, these results indicate that AAS treatment 1) ameliorates CK efflux and the uptake of [3H]thymidine and enhances the rate of protein synthesis during recovery after a bout of weight lifting, all being consistent with there being less muscle damage, and 2) enhances in vivo work capacity and the in situ fatigue resistance of a primary plantarflexor muscle.

Anabolic Agents↗
PubMed2001

Angiotensin II type 2 receptor inhibits epidermal growth factor receptor transactivation by increasing association of SHP-1 tyrosine phosphatase.

Angiotensin (Ang) II has 2 major receptor isoforms, Ang type 1 (AT(1)) and Ang type (AT(2)). AT(1) transphosphorylates epidermal growth factor receptor (EGFR) to activate extracellular signal-regulated kinase (ERK). Although AT(2) was shown to inactivate ERK, the action of AT(2) on EGFR activation remains undefined. Using AT(2)-overexpressing vascular smooth muscle cells from AT(2) transgenic mice, we studied these undefined actions of AT(2). Maximal ERK activity induced by Ang II was increased 1.9- and 2.2-fold by AT(2) inhibition, which was abolished by orthovanadate but not okadaic acid or pertussis toxin. AT(2) inhibited AT(1)-mediated EGFR tyrosine phosphorylation by 63%. The activity of SHP-1 tyrosine phosphatase was significantly upregulated 1 minute after AT(2) stimulation and association of SHP-1 with EGFR was increased, whereas AT(2) failed to tyrosine phosphorylate SHP-1. Stable overexpression of SHP-1-dominant negative mutant completely abolished AT(2)-mediated inhibition of EGFR and ERK activation. AT(1)-mediated c-fos mRNA accumulation was attenuated by 48% by AT(2) stimulation. Induction of fibronectin gene containing an AP-1 responsive element in its 5'-flanking region was decreased by 37% after AT(2) stimulation, corresponding to the results of gel mobility assay with the AP-1 sequence of fibronectin as a probe. These findings suggested that AT(2) inhibits ERK activity by inducing SHP-1 activity, leading to decreases in AP-1 activity and AP-1-regulated gene expression, in which EGFR dephosphorylation plays an important role via association of SHP-1.

Angiotensin II↗
PubMed2001

Different distribution patterns of the two mannose 6-phosphate receptors in rat liver.

Two mannose 6-phosphate receptors, cation-dependent and -independent receptors (CDMPR and CIMPR), play an important role in the intracellular transport of lysosomal enzymes. To investigate functional differences between the two in vivo, their distribution was examined in the rat liver using immunohistochemical techniques. Positive signals corresponding to CIMPR were detected intensely in hepatocytes and weakly in sinusoidal Kupffer cells and interstitial cells in Glisson's capsule. In the liver acinus, hepatocytes in the perivenous region showed a more intense immunoreactivity than those in the periportal region. On the other hand, positive staining of CDMPR was detected at a high level in Kupffer cells, epithelial cells of interlobular bile ducts, and fibroblast-like cells, but the corresponding signal was rather weak in hepatocytes. In situ hybridization analysis also revealed a high level of expression of CIMPR mRNAs in hepatocytes and of CDMPR mRNA in Kupffer cells. By double immunostaining, OX6-positive antigen-presenting cells in Glisson's capsule were co-labeled with the CDMPR signal but were only faintly stained with anti-CIMPR. These different distribution patterns of the two MPRs suggest distinct functional properties of each receptor in liver tissue.

Animals↗
PubMed2001

Relation of fibrillatory wave amplitude with hemostatic abnormality and left atrial appendage dysfunction in patients with chronic nonrheumatic atrial fibrillation.

Large left atrium (LA) and LA appendage (LAA) dysfunction are known to relate to cardiogenic thromboembolism, so the present study investigated the relation of the atrial fibrillatory wave (F wave) amplitude to hemostatic markers and LAA function. Transthoracic and transesophageal echocardiographic studies were performed in 82 consecutive patients with chronic, nonrheumatic atrial fibrillation (AF). Patients were divided into 2 groups according to F wave amplitude in lead V1 on the 12-lead ECG: coarse AF (the greatest amplitude of F wave > or =1 mm, n=44) and fine AF (<1 mm, n=38). Plasma levels of thrombin-antithrombin III complex, D-dimer, platelet factor 4 and beta-thromboglobulin were determined. Compared with patients with coarse AF, those with fine AF had lower LAA peak flow velocity (p<0.05) and higher prevalence of embolic cerebral infarction (50% vs 27%, p<0.05). Platelet activity did not differ between the 2 groups; however, plasma levels of thrombin-antithrombin III complex and D-dimer were significantly higher in patients with fine AF than in those with coarse AF (p<0.05). Multiple logistic regression analysis showed that fine AF was independently associated with cerebral embolism. Therefore, the presence of fine F wave in V1 would be a useful marker of LAA dysfunction and hypercoagulability, and indicate a risk for cerebral embolism in patients with chronic, nonrheumatic AF.

Aged↗
PubMed2001

Prevention of apoptosis of mammalian cells by the CED-3-cleaved form of CED-9.

CED-9 prevents apoptosis in embryonic cells of Caenorhabditis elegans but not in mammalian cells. We show here that the prevention of apoptosis in mammalian cells requires a CED-3-cleaved form (68-280) of CED-9 which is localized in the inner mitochondrial membrane. The viability of PC12 and HeLa cells was significantly increased after death stimuli when truncated CED-9 was expressed in these cells but full-length CED-9 did not. The truncated CED-9 expressed in these cells was largely localized to the inner mitochondrial and the endoplasmic reticulum membranes, while full-length CED-9 was detected mainly in endoplasmic reticulum fractions. Moreover, truncated CED-9 in purified mitochondria was resistant to trypsin digestion, but full-length CED-9 was not. These results suggest that the CED-3-cleaved form of CED-9 prevents apoptosis in mammalian cells by localizing to the inner mitochondrial membrane.

Animals↗
PubMed2001

Autophagic cell death and its execution by lysosomal cathepsins.

In the last decade, the molecular mechanisms of apoptosis, a major type of active cell death (type I cell death) have largely been clarified in mammalian cells. Particularly, the caspase family of proteinases has been shown to play crucial roles in the execution of apoptosis. Differing from apoptosis, type II cell death is known to be associated with autophagosomes/autolysosomes and appear in the developing nervous system (CLARKE, 1990). We have previously shown that delayed neuronal death occurring in the CA1 pyramidal layer of the gerbil hippocampus after brief forebrain ischemia is apoptotic in nature and autophagosomes/autolysosomes abundantly appear in the neurons before DNA fragmentation. To further understand the roles of autophagosomes/autolysosomes in active cell death, we examined the apoptosis of PC12 cells using morphological and biochemical techniques. PC12 cells are known to undergo apoptosis when cultured in the absence of serum. In such an environment, the mitochondrial pathway of apoptosis is activated; cytochrome c is released from mitochondria, and caspase-9/caspase-3 are activated. We have first examined morphological features of PC12 cells during the apoptotic process following serum deprivation, and found that autophagy is induced from the early stage of the process in the cells before typical nuclear changes. When autophagy is inhibited in the cells by 3-methyladenine, an autophagy inhibitor, they are largely protected from apoptosis. In relation to the induction of autophagy in PC12 cells following serum deprivation, immunoreactivity, protein amounts, and the proteolytic activity of lysosomal proteinases, particularly cathepsins B and D, are all greatly altered; those of cathepsin B drastically decrease in the cells from the early stage of serum-deprived cultures, whereas those of cathepsin D increase. Moreover, PC12 cells overexpressing cathepsin D undergo apoptosis more rapidly in serum-deprived cultures than wild-type cells, whereas those overexpressing cathepsin B increase the viability. These lines of evidence suggest that autophagy is involved in PC12 cell death following serum deprivation, this type of cell death being regulated by lysosomal proteinases, cathepsins B and D, downstream autophagy.

Animals↗
PubMed2001

Specific localization of lysosomal aminopeptidases in type II alveolar epithelial cells of the rat lung.

We previously demonstrated that lysosomal cysteine proteinases, cathepsins B, H, and L were localized in lysosomes of alveolar macrophages and bronchial epithelial cells in the rat lung, while cathepsin H, a typical aminopeptidase, was additionally distributed in lamellar bodies containing surfactant in type II alveolar epithelial cells (ISHII et al., 1991). The present immunohistochemical study further examined the localization of lysosomal aminopeptidases, cathepsin C, and tripeptidyl peptidase I (TPP-I) in the rat lung. Western blotting confirmed the presence of cathepsin C and TPP-I as active forms in the pulmonary tissue, showing 25 kD and 47 kD, respectively. Immunohisto/cytochemical observations demonstrated that positive staining for cathepsin C and TPP-I was more intensely localized in alveolar epithelial regions than in bronchial or bronchiolar epithelial cells. By double immunostaining using confocal laser microscopy, immunoreactivity for cathepsin H was found to be co-localized with that for cathepsin C or TPP-I in both type II cells and macrophages. Moreover, when doubly stained with anti-cathepsin C and ED2, single-positive type II cells could be clearly distinguished from double-positive macrophages in the alveolar region. Immunoelectron microscopy revealed the gold labeling of cathepsin C or TPP-I in multivesicular and composite bodies, and lamellar bodies of Type II cells. These results showing that lysosomal aminopeptidases such as cathepsin H, cathepsin C and TPP-I are localized in lamellar bodies of type II alveolar epithelial cells strongly argue for the participation of lysosomal aminopeptidases in the formation process of surfactant containing specific proteins.

Aminopeptidases↗
PubMed2001

Peripheral ophthalmic artery aneurysm. Report of two cases.

The authors report on two patients with peripheral ophthalmic artery (OphA) aneurysms and demonstrate the mechanism by which these lesions are formed by using OphA color Doppler flow velocimetry imaging. During formation of these two aneurysms, the hemodynamic stress on the OphA was quite significant.

Adult↗