Biomedical subjects

M Ito

Publications and source records attributed to M Ito.

At least 397 records · Page 22Linked to original sources

PubMed2000

Surgical treatment of cervical destructive spondyloarthropathy (DSA).

STUDY DESIGN: Sixteen patients with hemodialysis-associated cervical spine disorders underwent surgical treatment. After analysis of the surgical results, the optimum surgical procedures for these disorders were discussed. OBJECTIVE: To evaluate the surgical results of cervical spine disorders associated with long-term hemodialysis and to propose the optimum surgical procedures for successful outcomes. SUMMARY OF BACKGROUND DATA: There have been few reports regarding surgical results of hemodialysis-related cervical spine disorders. Surgical treatment for this disorder is still challenging. METHODS: Sixteen patients with hemodialysis-associated cervical spine disorders were treated surgically. Duration of hemodialysis ranged from 8 to 27 years (average, 17 years). Before surgery, 14 patients showed severe cervical myelopathy, and the other 2 had radiculopathy in the upper extremities. Ten patients with marked destructive changes underwent circumferential reconstructive surgery involving pedicle screw fixation, anterior strut bone grafting, and posterior and/or anterior decompression. Two patients with cervical radiculopathy underwent posterior nerve root decompression by foraminotomy and fusion by pedicle screw fixation or spinous process wiring. The remaining four patients without spinal instability underwent posterior decompression by open-door laminoplasty. RESULTS: Two patients died during follow-up. Follow-up periods in the surviving 14 patients ranged from 25 months to 92 months (average, 53 months). Marked neurologic recovery was obtained in all patients after surgery. Successful spinal fusion was obtained in all patients except one who underwent posterior fusion by spinous process wiring. Progressive destructive changes with significant instability at the adjacent mobile segments were observed in two patients who underwent circumferential fusion with a pedicle screw system more than 2 years after the initial surgery. CONCLUSIONS: The pedicle screw system achieved a high fusion rate in reconstructive surgery of cervical destructive spondyloarthropathy, even in the presence of severe bone fragility.

Adult↗

PubMed2000

Identification and characterization of a novel protein inhibitor of type 1 protein phosphatase.

We have isolated human cDNA for a novel type 1 protein phosphatase (PP1) inhibitory protein, named inhibitor-4 (I-4), from a cDNA library of germ cell tumors. I-4, composed of 202 amino acids, is 44% identical to a PP1 inhibitor, inhibitor-2 (I-2). I-4 conserves functionally important structure of I-2 and exhibited similar biochemical properties. I-4 inhibited activity of the catalytic subunit of PP1 (PP1C), specifically with an IC(50) of 0.2 nM, more potently than I-2 with an IC(50) of 2 nM. I-4 weakly inhibited the activity of myosin-associated phosphates (PP1M). However, the level of inhibition of PP1M was increased during preincubation of PP1M with I-4, suggesting that the inhibition is caused by interaction of I-4 with PP1C in such a manner that it competes with the M subunit of PP1M. Gel overlay experiments showed that I-4 binds PP1C directly. Three I-4 peptides containing the N-terminal residues 1-123, 1-131, and 1-142 all showed strong binding ability to PP1C but did not show PP1 inhibitory activity, whereas an I-2 peptide (residues 1-134), lacking the corresponding C-terminal residues, potently inhibited PP1C activity as previously reported. Removal of the 18 N-terminal amino acid residues from I-4 dramatically reduced the PP1 binding activity with a correlated loss of inhibitory activity, whereas removal of the 10 N-terminal residues had only a little effect. The two peptides GST-I-4(19-131) and GST-I-4(132-202) showed ability to bind to PP1C, albeit very weakly. These results strongly suggest a multiple-point interaction between I-4 and PP1C, which is thought to cause the inhibition of I-4 which is stronger than the inhibition of I-2.

Amino Acid Sequence↗

PubMed2000

Immunohistochemical study of Pneumocystis carinii infection in pigs: evaluation of Pneumocystis pneumonia and a retrospective investigation.

Infection with Pneumocystis carinii was demonstrated immunohistochemically in the lungs of pigs 15 to 75 days of age from a herd with epidemic pneumonia due to the organism. The distribution of the organism was centered on the airways, and extended progressively with age from the alveolar ducts to the alveoli. In a retrospective immunohistochemical study of 245 newborn to adult pigs which were necropsied between 1988 and 1995, P carinii infection was found in 87 pigs (35.5 per cent) aged between 17 days to seven months. In the pigs aged between one and three months the infection rate was 63.1 per cent. Pigs from herds in which suckler and weaner pigs shared the same air space were more heavily infected than those from the herds in which they were reared separately. There were no regional or seasonal variations in the level of infection, and the infection was not associated with any single disease.

Animals↗

PubMed2000

Expression of human alpha 2-adrenergic receptors in adipose tissue of beta 3-adrenergic receptor-deficient mice promotes diet-induced obesity.

Catecholamines play an important role in controlling white adipose tissue function and development. beta- and alpha 2-adrenergic receptors (ARs) couple positively and negatively, respectively, to adenylyl cyclase and are co-expressed in human adipocytes. Previous studies have demonstrated increased adipocyte alpha 2/beta-AR balance in obesity, and it has been proposed that increased alpha 2-ARs in adipose tissue with or without decreased beta-ARs may contribute mechanistically to the development of increased fat mass. To critically test this hypothesis, adipocyte alpha 2/beta-AR balance was genetically manipulated in mice. Human alpha 2A-ARs were transgenically expressed in the adipose tissue of mice that were either homozygous (-/-) or heterozygous (+/-) for a disrupted beta 3-AR allele. Mice expressing alpha 2-ARs in fat, in the absence of beta 3-ARs (beta 3-AR -/- background), developed high fat diet-induced obesity. Strikingly, this effect was due entirely to adipocyte hyperplasia and required the presence of alpha2-ARs, the absence of beta 3-ARs, and a high fat diet. Of note, obese alpha 2-transgenic beta 3 -/- mice failed to develop insulin resistance, which may reflect the fact that expanded fat mass was due to adipocyte hyperplasia and not adipocyte hypertrophy. In summary, we have demonstrated that increased alpha 2/beta-AR balance in adipocytes promotes obesity by stimulating adipocyte hyperplasia. This study also demonstrates one way in which two genes (alpha 2 and beta 3-AR) and diet interact to influence fat mass.

Adipose Tissue↗

PubMed2000

A naturally occurring steroidogenic factor-1 mutation exhibits differential binding and activation of target genes.

Steroidogenic factor-1 (SF-1) is an orphan nuclear receptor that binds DNA as a monomer and regulates the transcription of multiple target genes. A mutation in the proximal (P)-box of the first zinc finger of SF-1 (G35E) has been reported to cause complete XY sex reversal and adrenal insufficiency. Because this P-box region dictates DNA binding specificity, we investigated the effect of this mutation on DNA binding and regulation of target genes. Binding of the P-box mutant was markedly impaired for most native SF-1 response elements. However, mutant SF-1 bound to a subset of response elements containing a CCA AGGTCA motif. Mutagenesis studies of response elements revealed that the first nucleotide position in the 5'-flanking sequence triplet and the central part of the half-site dictate DNA binding specificity by the mutant SF-1. Further, introduction of a mutation into the SF-1 A-box, which has been proposed to bind to the 5'-flanking sequence triplet, eliminated binding by mutant SF-1 to all response elements tested. These data support the idea that the A-box stabilizes monomeric binding by nuclear receptors. This action may be particularly important when P-box binding affinity is compromised either by mutations in SF-1 or by sequence alterations in its binding site.

Amino Acid Sequence↗

PubMed2000

Purification, characterization, and cDNA cloning of a novel acidic endoglycoceramidase from the jellyfish, Cyanea nozakii.

Endoglycoceramidase (EC ) is an enzyme capable of cleaving the glycosidic linkage between oligosaccharides and ceramides in various glycosphingolipids. We report here the purification, characterization, and cDNA cloning of a novel endoglycoceramidase from the jellyfish, Cyanea nozakii. The purified enzyme showed a single protein band estimated to be 51 kDa on SDS-polyacrylamide gel electrophoresis. The enzyme showed a pH optimum of 3.0 and was activated by Triton X-100 and Lubrol PX but not by sodium taurodeoxycholate. This enzyme preferentially hydrolyzed gangliosides, especially GT1b and GQ1b, whereas neutral glycosphingolipids were somewhat resistant to hydrolysis by the enzyme. A full-length cDNA encoding the enzyme was cloned by 5'- and 3'-rapid amplification of cDNA ends using a partial amino acid sequence of the purified enzyme. The open reading frame of 1509 nucleotides encoded a polypeptide of 503 amino acids including a signal sequence of 25 residues and six potential N-glycosylation sites. Interestingly, the Asn-Glu-Pro sequence, which is the putative active site of Rhodococcus endoglycoceramidase, was conserved in the deduced amino acid sequences. This is the first report of the cloning of an endoglycoceramidase from a eukaryote.

Amidohydrolases↗

PubMed2000

Bacillus subtilis YqkI is a novel malic/Na+-lactate antiporter that enhances growth on malate at low protonmotive force.

Bacillus subtilis yheL encodes a Na(+)/H(+) antiporter, whereas its paralogue, yqkI, encodes a novel antiporter that achieves a simultaneous Na(+)/H(+) and malolactate antiport. B. subtilis yufR, a control in some experiments, encodes a Na(+)/malate symporter. YqkI complemented a malate transport mutant of Escherichia coli if Na(+) and lactate were present. YheL conferred Na(+) uptake capacity on everted membrane vesicles from an antiporter-deficient E. coli mutant that was consistent with a secondary Na(+)/H(+) antiport, but YqkI-dependent Na(+) uptake depended on intravesicular malate and extravesicular lactate. YqkI-dependent lactate uptake depended on intravesicular malate and extravesicular Na(+). YqkI mediated an electroneutral exchange, which is proposed to be a malic(-2)-2H(+) (or fully protonated malate)/Na(+)-lactate(-1) antiport. Because the composite YqkI-mediated exchanges could be driven by gradients of the malate-lactate pair, this transporter could play a role in growth of B. subtilis on malate at low protonmotive force. A mutant with a disruption of yqkI exhibited an abrupt arrest in the mid-logarithmic phase of growth on malate when low concentrations of protonophore were present. Thus growth of B. subtilis to high density on a putatively nonfermentative dicarboxylic acid substrate depends on a malolactate exchange at suboptimal protonmotive force.

Antiporters↗

PubMed2000

Increased intestinal phospholipase A(2) activity catalyzed by phospholipase B/lipase in WBN/Kob rats with pancreatic insufficiency.

Male WBN/Kob rats derived from the Wistar strain spontaneously develop chronic pancreatitis as late as 3 months old. To assess the degree of disease severity, we compared the lipolytic enzyme levels in pancreas of 2-, 4-, and 6-month-old WBN/Kob rats fed isocaloric no fat (NF) and high fat (HF, 57% of total calories) diets and its pathology. Diet treatment did not significantly affect lipase and group Ib phospholipase A(2) (PLA(2)) levels in the pancreas at all ages. Development of chronic pancreatitis at the age of 4 and 6 months was consistent with the tendency of decreasing group Ib PLA(2) specific content determined by enzyme immunoassay and lipase activity, and the decreased number of group Ib PLA(2)-positive acinar cells. Pancreatic lipase and group Ib PLA(2) levels of 4-month-old WBN/Kob rats were significantly lower than those of control Wistar rats at age 4 months irrespective of diet. This allowed us to adopt 4-month-old WBN/Kob rats as a model of pancreatic insufficiency, which could be a useful tool to examine the role of gastrointestinal enzymes in lipid digestion. Ca(2+)-independent PLA(2) activity of brush border membrane-associated phospholipase B/lipase (PLB/LIP) in ileal mucosa increased significantly in 4-month-old WBN/Kob rats while its content and transcript levels remained constant, suggesting its activation at the enzyme level. In WBN/Kob rats fed the HF diet at age 4 months, PLA(2) activity catalyzed by PLB/LIP in the proximal ileal mucosa was four times the total PLA(2) activity in the intestinal lumen. These results indicate that PLB/LIP compensates for the depletion of pancreatic lipolytic enzymes in WBN/Kob rats with pancreas insufficiency.

Age Factors↗

PubMed2000

Isoforms of JSAP1 scaffold protein generated through alternative splicing.

We have identified four isoforms of c-Jun NH(2)-terminal kinase (JNK)/stress-activated protein kinase-associated protein 1 (JSAP1), a scaffold protein that participates in JNK mitogen-activated protein kinase cascades, termed JSAP1a, JSAP1b, JSAP1c, and JSAP1d. The previously identified JSAP1 was renamed JSAP1a to avoid confusion. Analyses of the exon-intron structure of the jsap1 gene indicated that the isoforms are generated through alternative splicing involving exons 5 and 6. The mRNA expression levels of the JSAP1 isoforms differed among the mouse tissues examined. We also investigated the region of JSAP1 responsible for its interaction with JNK, and found that the JNK-binding domain is located between aa residues 201 and 217 in JSAP1a, which is encoded by part of exon 6. As all the JSAP1 isoforms contain this binding domain, we examined the binding affinity of the JSAP1 isoforms for JNK1, JNK2, and JNK3. JSAP1c and JSAP1d, which contain a 31-aa sequence not present in JSAP1a or JSAP1b, had a lower binding affinity for the JNKs, especially JNK3. These results suggest that JSAP1c and JSAP1d may attenuate the scaffolding activity of JSAP1a and/or JSAP1b in JNK cascades, especially the JNK3 cascades.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Induction of c-Src in human blood monocytes by anti-CD98/FRP-1 mAb in an Sp1-dependent fashion.

Freshly isolated human blood monocytes expressed neither c-src mRNA nor c-Src. However, when monocytes were incubated with anti-CD98 heavy chain (HC) mAb, expression of c-src mRNA, c-Src, and activated c-Src was induced. Many binding sites for the ubiquitous transcription factor Sp1 were identified in the promoter region of the c-src gene. Surprisingly, Sp1 and Sp1 mRNA were not found in monocytes that were freshly isolated or incubated with control antibody. Stimulation with anti-CD98HC mAb also resulted in the expression of Sp1 and its translocation to the nucleus. Herbimycin A, genistein, manumycin A, PD-98059, SB203580, and HBJ127 suppressed CD98HC-mediated c-src and Sp1 mRNA induction. On the contrary, H-7, Wortmannin, HA1077, and Y-27632 showed no effect on c-Src and Sp1 induction. Furthermore, anti-CD98HC mAb induced activation of tyrosine kinases and ERK kinases. These findings suggest that the tyrosine kinase(s)-Ras-MAPK-Sp1 pathway(s) is involved in CD98HC-mediated induction of c-Src in human blood monocytes.

Antibodies, Monoclonal↗

PubMed2000

Characterization of Solt, a novel SoxLZ/Sox6 binding protein expressed in adult mouse testis.

SoxLZ/Sox6, a member of the Sox protein family, contains a leucine zipper motif in addition to an HMG box, which is its DNA binding domain. Here we have identified a novel SoxLZ/Sox6 binding protein, termed Solt, which we obtained independently using both a far-Western blot and a yeast two-hybrid screen. Like SoxLZ/Sox6 mRNA, Solt mRNA was exclusively expressed in the testis in mouse. Solt contains an unusual leucine zipper, which bound to the leucine zipper region of SoxLZ/Sox6 in vitro. In transient transfection assays in CHO cells with SoxLZ/Sox6 containing the transactivational region of herpes simplex virus VP16, expression of a reporter gene that carries a cis binding region for Sox proteins was significantly enhanced by the co-expression of Solt and Ca(2+)/calmodulin-dependent protein kinase IV.

Amino Acid Sequence↗

PubMed2000

Rapidly turned over protein maintains metabotropic synaptic transmission in Purkinje cells.

It has generally been thought that protein synthesis is required for relatively slow cellular processes such as synaptogenesis and synaptic plasticity. In this study on rat cerebellar slices, we found that metabotropic glutamate receptor-mediated synaptic transmission to cerebellar Purkinje cells was quickly and persistently depressed by brief (5 min) applications of translational inhibitors, which were confirmed to induce quick and persistent depression of protein synthesis in cerebellar tissues. Brief applications of transcriptional inhibitors also depressed metabotropic synaptic transmission, but progressively over 1 h, presumably due to depletion of mRNAs in the dendrites. Results of this study indicate the presence of a unique protein(s) that is dynamically involved in metabotropic synaptic transmission.

Animals↗

PubMed2000

Mutant presenilin 2 transgenic mice. A large increase in the levels of Abeta 42 is presumably associated with the low density membrane domain that contains decreased levels of glycerophospholipids and sphingomyelin.

The N141I mutation in presenilin (PS) 2 is tightly linked with a form of autosomal dominant familial Alzheimer's disease in the Volga German families. We previously reported that mouse brains harboring mutant PS2 contained increased levels of amyloid beta protein (Abeta) 42 in the Tris-saline-soluble fraction (Oyama, F., Sawamura, N., Kobayashi, K., Morishima-Kawashima, M., Kuramochi, T., Ito, M., Tomita, T., Maruyama, K., Saido, T. C., Iwatsubo, T., Capell, A., Walter, J., Grünberg, J., Ueyama, Y., Haass, C. and Ihara, Y. (1998) J. Neurochem. 71, 313-322). Here, using a new extraction protocol, we quantitated the Abeta40 and Abeta42 levels in the Tris-saline-insoluble fraction. The insoluble Abeta levels were found to be higher than the soluble Abeta levels, and the insoluble Abeta42 levels were markedly increased in mutant PS2 transgenic mice. To investigate the origin of the insoluble Abeta42, we prepared the detergent-insoluble, low density membrane fraction. This fraction from two independent lines of mutant PS2 transgenic mice contained remarkably increased levels of Abeta42 and significantly low levels of glycerophospholipids and sphingomyelin. This unexpected finding suggests that a large increase in the levels of Abeta42 in mutant PS2 mice is presumably induced through alterations of the lipid composition in the low density membrane domain in the brain.

Amino Acid Substitution↗

PubMed2000

Neutral/alkaline and acid ceramidase activities are actively released by murine endothelial cells.

Ceramidases (CDase(s)) play a key role in sphingolipid metabolism by hydrolyzing ceramide into sphingosine. Here we report that murine endothelial cells, macrophages, and human fibroblasts are all able to release acid as well as neutral/alkaline CDase activities in the culture medium. Endothelial cells were characterized by the highest specific activity of cellular as well as secreted CDases. The release of both enzymatic activities was reduced by protein synthesis inhibitor cycloheximide but was unaffected by the blocking of RNA transcription with actinomycin D. The discharge of acid and neutral/alkaline CDases was also diminished by brefeldin A, a fungal metabolite which disrupts Golgi apparatus. Remarkably, treatment of endothelial cells with bradykinin resulted in a significant increase of neutral/alkaline but not acid CDase release. This report represents the first evidence for the existence of constitutive and regulated release of CDase activities by endothelial cells. In view of the known ability of these cells to secrete sphingomyelinase, this finding suggests that CDase may participate in extracellular sphingomyelin metabolism which is presently known to have a role in atherogenesis and could be involved in other physiological or pathological events.

Acid Ceramidase↗

PubMed2000

Endogenous and ectopic gland induction by FGF-10.

FGF-10, a member of the fibroblast growth factor family, is expressed in mesodermally derived cell populations during embryogenesis. During normal ocular development, FGF-10 is expressed in the perioptic mesenchyme adjacent to the Harderian and lacrimal gland primordia. In this report, we provide evidence that FGF-10 is both necessary and sufficient to initiate glandular morphogenesis. Lens-specific expression of FGF-10 was sufficient to induce ectopic ocular glands within the cornea. In addition, lacrimal and Harderian glands were not seen in FGF-10 null fetuses. Based on these results we propose that FGF-10 is an inductive signal that initiates ocular gland morphogenesis.

Animals↗

PubMed2000

Distinction of differentiated type early gastric carcinoma with gastric type mucin expression.

BACKGROUND: Intestinal and diffuse gastric carcinomas differ in morphology and growth behavior. Differentiated type gastric carcinoma (DGC), which corresponds roughly with the intestinal type of Lauren, can demonstrate phenotypic properties associated with mucin expression and brush border. However, their clinical significance is controversial. A classification based on mucin phenotype and brush border was performed to determine the clinicopathologic diversity of DGCs in their early stage. METHODS: A total of 120 specimens from 116 DGC patients with definite submucosal invasion were evaluated both macroscopically and histologically. All sections were examined immunohistochemically with human gastric mucin, Muc-2, and CD10 and with mucin histochemically with paradoxical concanavalin A staining and high iron Diamine-Alcian Blue. They were classified into gastric type (G-type), intestinal type (I-type), mixed gastric and intestinal type (M-type), or null type (N-type) phenotypes. The immunoreactivity of E-cadherin and beta-catenin also was investigated to determine the correlation between mucin phenotype and clinicopathologic factors. RESULTS: The G-type phenotype was found to be in contrast to I-type: G-type was an independent factor associated with lymph node metastasis. Significant correlations were observed between the G-type phenotype and the complex type carcinoma found that was histologically: lymphatic invasion, lymph node metastasis, and the abnormal expression of E-cadherin. A significant difference in the proportion of mucin phenotypes between papillary type and tubular type carcinoma was observed. G-type was found to be the predominant phenotype in papillary carcinoma in contrast to tubular carcinoma. CONCLUSIONS: The G-type mucin phenotype and papillary adenocarcinoma should be distinguished from other types of DGCs because of their increased malignant potential in the incipient phase of invasion and metastasis. The significance of G-type and papillary adenocarcinoma should be reflected in the treatment of patients with early stage DGCs, including endoscopic mucosal resection.

Adenocarcinoma↗

PubMed2000

Phosphorylation of CPI-17, an inhibitor of myosin phosphatase, by protein kinase N.

CPI-17 is a phosphorylation-dependent inhibitory protein for smooth muscle myosin phosphate. Phosphorylation at Thr(38), in vitro, by protein kinase C or Rho-kinase enhances the inhibitory potency toward myosin phosphatase. Phosphorylation of CPI-17 by protein kinase N (PKN), a fatty acid- and Rho-activated serine/threonine kinase, and its effect on smooth muscle myosin phosphatase activity were investigated. CPI-17 was phosphorylated by GST-PKN-CAT, a constitutively active GST-fusion fragment of PKN, to 1.46 mol of P/mol of CPI-17, in vitro. The K(m) value of CPI-17 for PKN was 0.96 microM. Phosphorylation of PKN dramatically increased the inhibitory effect of CPI-17 on myosin phosphatase activity. The major and inhibitory phosphorylation site was identified as Thr(38) using a point mutant of CPI-17 and a phosphorylation-state specific antibody. Thus, CPI-17 is a substrate of PKN and might be involved in the Ca(2+) sensitization of smooth muscle contraction as a downstream effector of Rho and/or arachidonic acid.

Animals↗

PubMed2000

Mapping of domains on the human parainfluenza type 2 virus P and NP proteins that are involved in the interaction with the L protein.

Eleven monoclonal antibodies (MAbs) directed against the large (L) protein of human parainfluenza type 2 virus (hPIV-2) were prepared to examine the interactions of the L protein with other viral proteins. Coimmunoprecipitation assays using these MAbs revealed that the L protein directly interacted with the phospho- (P) and nucleocapsid (NP) proteins in vivo and in vitro. Mutational analysis of the P or NP protein was performed to identify the region(s) on these proteins interacting with L protein, indicating that amino acids 278-353 on the P protein and amino acids 403-494 on the NP protein are essential for the binding to the L protein.

Antibodies, Monoclonal↗