Biomedical subjects

H Itoh

Publications and source records attributed to H Itoh.

At least 19 recordsLinked to original sources

PubMed1995

Cloning of the cDNA encoding a novel rat mast-cell proteinase, rMCP-3, and its expression in comparison with other rat mast-cell proteinases.

A cDNA encoding a novel rat mast-cell proteinase (MCP) named rMCP-3 was successfully cloned and sequenced from the peritoneal cells of Lewis rats infected with the intestinal nematode Nippostrongylus brasiliensis by using the combination of reverse transcription-PCR and rapid-amplification-of-cDNA-ends ('RACE') methods. The cDNA was 979 bp long and included a 741 bp open reading frame. When the deduced amino acid sequence was compared with those of other known mast-cell proteinases, rMCP-3 was considered to be translated as a preproenzyme with a 19-amino-acid signal peptide, a two-amino-acid activation peptide and a 226-amino-acid mature enzyme. The amino acid identity in the mature enzyme was 52.9% and 55.1% with rMCP-1 and rMCP-2 respectively. The rMCP-3 mRNA was not detected in the peritoneal cells of mast-cell-deficient Ws/Ws rats, though it was strongly detected in those of littermate +/+ and Lewis rats, indicating the mast-cell origin of rMCP-3 In addition to being present in peritoneal mast cells, the rMCP-3 mRNA was strongly detected in the skin, tongue, and RBL2H3 rat basophilic leukaemia cells and weakly in the jejunum of N. brasiliensis-infected rats by RNA blot analysis using a rMCP-3 gene-specific probe. By reverse transcription-PCR, the rMCP-3 mRNA was also detected in the lung. While the expression of rMCP-1 and rMCP-2 are clearly restricted in connective-tissue mast cells and mucosal mast cells respectively, rMCP-3 was widely expressed in both types of mast cells with a predominance in connective-tissue mast cells.

Amino Acid Sequence↗

PubMed1995

Molecular cloning of mouse intestinal trefoil factor and its expression during goblet cell changes.

A cDNA encoding mouse intestinal trefoil factor (mITF) was successfully cloned and sequenced from the small intestine of C57BL/6 mouse by using the combination of reverse transcription-PCR and rapid amplification of cDNA ends methods. The gene was, similar to rat and human ITFs, mainly expressed in the small and large intestine. The mITF expression was up-regulated during the recovery phase after depletion of goblet cells in acetic acid-induced colitis. On the other hand, the expression in the jejunum was not altered, while goblet cell hyperplasia was induced by Nippostrongylus brasiliensis infection. These results suggest that the mITF expression did not simply correlate with the number of goblet cells. The mITF may play an important role in the maintenance and repair of mucosal function of the rectum. Additionally, the mITF in the jejunum may play a role in alteration of the physicochemical nature of goblet cell mucins, thereby affecting the establishment of intestinal helminths.

Acetates↗

PubMed1995

Rapid ventricular induction of brain natriuretic peptide gene expression in experimental acute myocardial infarction.

BACKGROUND: We have demonstrated that brain natriuretic peptide (BNP) is a cardiac hormone predominantly synthesized in and secreted from the ventricle. We have also reported that, compared with atrial natriuretic peptide (ANP), the plasma concentration of BNP is increased to a greater degree in patients with congestive heart failure and more rapidly in patients with acute myocardial infarction (AMI). METHODS AND RESULTS: To investigate ventricular gene expression of BNP in AMI, we analyzed plasma and ventricular BNP concentrations along with ventricular BNP mRNA in rats with AMI produced by coronary artery ligation. The BNP concentration in the left ventricle increased about 2-fold as early as 12 hours postinfarction and 5-fold 1 day postinfarction compared with sham-operated rats, whereas left ventricular ANP concentration remained unchanged within 1 day. The tissue concentration of BNP increased in the noninfarcted region as well as in the infarcted region. The surviving myocytes in and around the necrotic tissues in the infarcted region were intensely stained with the anti-BNP antiserum, indicating augmented production in the remaining myocytes in the infarcts. The BNP concentration in the right ventricle also increased about 10-fold 12 hours postinfarction, whereas the ANP concentration remained unchanged within 12 hours. Northern blot analysis revealed that BNP mRNA expression was augmented 3-fold in the left ventricle as early as 4 hours postinfarction. In contrast, ANP mRNA expression was unchanged. Reflecting the rapid induction of ventricular BNP production, the plasma BNP concentration rose to about 100 pg/mL 12 hours postinfarction (sham-operated rats, < 70 pg/mL). CONCLUSIONS: These results demonstrate the rapid induction of ventricular BNP gene expression in rats with AMI compared with ANP and suggest that BNP gene expression in the ventricle is regulated distinctively from ANP gene expression against acute ventricular overload. They also suggest that the BNP gene can be one of the acutely responsive cardiac genes for the ventricular overload and suggest a possible pathophysiological role of BNP distinct from ANP in AMI.

Animals↗

PubMed1995

Carboxyl terminal of G protein beta subunit is required for association with gamma subunit.

To investigate a complex formation of heterotrimeric G protein beta and gamma subunits, we constructed a deletion mutant which lacked the C-terminal WD40 repeating unit of beta 1 and cotransfected into HEK293 cells with gamma 2 which was tagged with FLAG-epitope. Co-immunoprecipitation assay indicated that the mutant beta 1 failed to associate with FLAG gamma 2. Furthermore, from the analysis of additional deletion mutants, the last 10 amino acids of C-terminal region of beta 1 were found to be necessary to associate with gamma 2. These results suggest that the C-terminal region, in addition to the N-terminal alpha-helical structure, of beta subunit may be involved in the beta gamma complex formation.

Amino Acid Sequence↗

PubMed1995

The complete amino acid sequence of chum salmon stanniocalcin, a calcium-regulating hormone in teleosts.

Stanniocalcin (STC) is a hypocalcemic hormone secreted from the corpuscles of Stannius of bony fish. Chum salmon STC was isolated in pure form by ion-exchange chromatography and reversed phase high performance liquid chromatography following ethanol-ammonium extraction of the tissues. A cDNA was cloned from cDNAs of the tissue by the PCR method using two primers corresponding to the N- and C-terminal amino acid sequence of the hormone. Sequence analysis of the protein and the cDNA revealed that chum salmon STC is a homodimer, and that the monomer consists of 179 amino acids including 11 half-Cys residues and one N-linked glycosylation site, which is 44 residues smaller at the C-terminal region than the sequence deduced from coho salmon STC cDNA.

Amino Acid Sequence↗

PubMed1995

Young borderline hypertensives are hyperreactive to mental arithmetic stress: spectral analysis of R-R intervals.

To investigate whether sympathetic tone and its reactivity to stress are increased in borderline hypertension, we compared pressor and autonomic nervous responses to mental arithmetic stress in male borderline hypertensives (BH) and normotensive volunteers (NT). Three age groups, 30, 40 and 50-year-old groups, which included 30 to 39, 40 to 49 and 50 to 59-year-old subjects, were studied. Fractional LF (%LF), fractional HF (%HF) and L/H, obtained from the power spectrum of R-R intervals, were used as indices of autonomic nervous function. Baseline autonomic nervous indices did not differ between NT and BH of any age group. Blood pressure rose higher during mental arithmetic stress in 50 than in 30-year-old NT but not in comparable age groups of BH. Pressor responses were augmented in BH compared to NT only in the 30-year-old group. However, the differences were not significant when pressor responses were expressed as percent increases in blood pressure. Both %L and L/H increased during arithmetic stress. The increase in %L did not differ between NT and BH but that in L/H (% delta L/H) was larger in 50 than in 30-year-old NT. % delta L/H was larger in BH than in NT only in the 30-year-old group. These findings suggest that both pressor and autonomic nervous responses to metal arithmetic stress were altered by aging and augmented in BH compared to NT in the 30-year-old group.

Adult↗

PubMed1995

Cloning of the cDNA encoding mast cell tryptase of Mongolian gerbil, Meriones unguiculatus, and its preferential expression in the intestinal mucosa.

By using the combination of reverse-transcription PCR and rapid amplification of cDNA ends methods, a cDNA encoding mast cell tryptase was successfully cloned from the small intestine of Mongolian gerbil, Meriones unguiculatus, infected with Nippostrongylus brasiliensis. The cDNA was 1219 bp long including 810 bp of an open reading frame. Based on the deduced amino acid sequences of known mast cell tryptases of other species, the gerbil mast cell tryptase (gMCT) was highly similar to mouse mast cell protease (mMCP)-7, and seems to be translated as a prepro-enzyme with 25 amino acids of signal and activation peptides and 245 amino acids of mature enzyme. The gMCT mRNA was preferentially transcribed in the intestinal mucosa and to a far lesser extent in the connective tissue such as skin and tongue. Moreover, kinetic study after infection revealed that the amount of gMCT mRNA in the small intestine correlated well with the degree of intestinal mastocytosis. Throughout the course of infection, enzyme-histochemically detectable tryptase activity was limited to mucosal mast cells. Since mucosal mast cells of other rodents, including mice and rats, do not express tryptases, this is the first report of rodent mast cell tryptase expressed in the intestinal mucosa.

Amino Acid Sequence↗

PubMed1995

G protein beta gamma subunit activates Ras, Raf, and MAP kinase in HEK 293 cells.

Using transient transfection of HEK 293 cells, we have studied the activation of Ras, c-Raf, and MAP kinase by G protein-coupled receptors, activated G protein alpha subunit (G alpha), and beta gamma subunits (G beta gamma). The expression of constitutively activated Gs alpha, Gi alpha, and G11 alpha did not have any effect on MAP kinase phosphorylation. In contrast, overexpression of G beta gamma could stimulate the phosphorylation of MAP kinase and enhance the MEK kinase activity of c-Raf. Coexpression of dominant negative Ras inhibited G beta gamma-induced phosphorylation of MAP kinase. Furthermore, the GTP-bound form of Ras was increased by overexpression of G beta gamma. These results strongly suggest that the G beta gamma may play an important role in signaling from G protein-coupled receptors to the MAP kinase pathway, and the activation of Ras and c-Raf may be involved in this signaling cascade in HEK 293 cells.

Cell Line↗

PubMed1995

Mammalian 60-kDa stress protein (chaperonin homolog). Identification, biochemical properties, and localization.

Mammalian chaperonin homolog (HSP60) was purified from porcine livers cytosol using a tandem ATP-Sepharose column and Mono Q column chromatography. A partial amino acid sequence (96 amino acid residues) of this protein was determined and coincided with those of human HSP60 with 96.9% homology, which was deduced from the nucleotide sequence of the cDNA. The sequence of the NH2 termini of the purified protein (5 amino acid residues) coincided with the signal sequence of HSP60. These facts led to the identification of the 60-kDa liver protein with the chaperonin homolog. Dihydrofolate reductase was able to form a stable complex with the liver chaperonin homolog. The liver chaperonin homolog was detected by at least five spots around pI = 5.6 on two-dimensional gel electrophoresis. Immunoblotting studies using an antibody against chaperonin homolog showed that the chaperonin homolog was localized in the cytosol, mitochondrial, and nuclear fractions of porcine liver. The chaperonin homolog was localized both in the mitochondria and cytoplasm of rat kidneys at the electron microscopic level. The chaperonin homolog in the cytosol, but not in the other subcellular fractions, was cross-reacted with an antibody against the synthetic peptide corresponding to the signal peptide of HSP60 as well as the purified chaperonin homolog on immunoblotting. These results suggested that the functional chaperonin homolog in the cytosol may be transported into the mitochondria and the protein may be processed to mitochondrial HSP60 in the organella.

Amino Acid Sequence↗

PubMed1995

Structural organization and chromosomal assignment of the human prostacyclin receptor gene.

Prostacyclin receptor is a member of the prostanoid receptor family in the G protein-coupled receptor superfamily with seven transmembrane domains. We report here the isolation and structural organization of the human prostacyclin receptor gene. Southern blot analysis demonstrated a single copy of the human prostacyclin receptor gene in the human genome. The human prostacyclin receptor gene spanned approximately 7.0 kb and was composed of three exons separated by two introns. The first intron occurred in the 5'-untranslated region, 13 bp upstream to the ATG start codon. The second intron was located at the end of the sixth transmembrane domain, thereby separating it from the downstream coding region and the 3'-untranslated region. By primer extension analysis, the transcription initiation sites were mapped 870-872 bp upstream to the ATG start codon. The 1.2-kb human prostacyclin receptor 5'-flanking region lacked conventional TATA and CCAAT boxes, but it contained several cis-acting regulatory elements including an inverted CCAAT box (Y box) and two copies of SP-1 binding sites. Using human-rodent somatic hybrid cell DNA, the human prostacyclin receptor gene was assigned to human chromosome 19. The present study helps establish the genetic basis for prostacyclin receptor research and provides further insight into the molecular mechanisms underlying the prostanoid receptor family.

Amino Acid Sequence↗

PubMed1995

Activity in the human primary motor cortex related to arm and finger movements.

With the purpose of mapping representations of the finger and proximal arm in the human primary motor cortex (MI), we measured regional cerebral blood flow (rCBF) in eight right-handed normal volunteers during self-paced finger and proximal arm movements. Each movement activated two different fields in MI, one located deep in the anterior lip of the central sulcus and the other relatively close to the surface of the precentral gyrus. Arm fields were located higher than finger fields. A superficial arm field and a deep finger field partially, overlapped. The results indicate that there are two spatially separate motor representations of the finger and proximal arm in the human MI.

Adult↗

PubMed1995

Interaction stabilizing tertiary structure of bacteriorhodopsin studied by denaturation experiments.

The structural stability of bacteriorhodopsin was studied by denaturation experiments, using aliphatic alcohol as denaturants. The disappearance of a positive peak at 285 nm of the circular dichroism spectra, the change in the intrinsic fluorescence decay time, and the decrease of the regeneration activity bacteriorhodopsin indicated the denaturation of the tertiary structure of this protein at a methanol concentration of about 3 M. The circular dichroism band at 222 nm was unchanged by the denaturation. It was concluded that the alcohol-denatured state in water was similar to the molten globule state of soluble proteins, in which only the tertiary structure was destroyed. Solvent substitution from water to hexane did not cause denaturation of bacteriorhodopsin. However, further addition of alcohol destroyed the secondary as well as the tertiary structures. Comparing the alcohol effects of bacteriorhodopsin in water to that in hexane, the dominant interactions for the structure formation of this protein could be revealed: the hydrophobic interaction that arose from the structure of water is essential for the stability of membrane spanning helices, while the interaction which binds the helices is polar in nature.

Alcohols↗

PubMed1995

Thymoma: tumour type related to expression of epidermal growth factor (EGF), EGF-receptor, p53, v-erb B and ras p21.

As clinicopathological features may not be sufficient to predict the progression of thymoma, we have carried out what we believe to be the first immunohistochemical study describing the relationship between the different types of thymoma and the tumour stage, on the one hand, and the expression of epidermal growth factor (EGF), EGF-receptor (EGFR), p53, v-erb B and ras p21, on the other. The positive rates versus histological types and Masaoka's clinical stages in the 47 cases were as follows: p53 (non-invasive thymoma: 41.7%; malignant thymoma category I: 82.4%; malignant thymoma category II: 83.3%), EGF (non-invasive thymoma: 4.2%; malignant thymoma category I: 11.8%; malignant thymoma category II: 33.3%) and EGFR (non-invasive thymoma: 8.3%; malignant thymoma category I: 35.3%; malignant thymoma category II: 66.7%); p53 (stages I and II: 51.7%; stages III and IV: 77.8%), EGF (stages I and II: 3.4%; stages III and IV: 22.2%) and EGFR (stages I and II: 13.8%; stages III and IV: 44.4%). These data suggest that p53 may be implicated in the initial stages of tumorigenesis and that increased expression of EGF and EGFR may play a role in thymoma progression.

Adult↗

PubMed1995

Late graft failure of autologous vein grafts for arterial occlusive disease: clinical and experimental studies.

Late graft failure following arterial reconstructive surgery, especially after infrainguinal reconstruction, remains a major concern for vascular surgeons. To more effectively predict the outcome of reconstructed arteries, we herein propose an intraoperative flow waveform analysis which correlates well with the long-term patency rate of grafts. According to this flow waveform analysis, late graft failure was occasionally seen in grafts with type II waveforms when poor distal runoff vessels had been shown by the preoperative arteriogram. Next, to investigate which events occurring in autologous vein grafts under abnormal hemodynamics may contribute to late graft failure, a distal poor-runoff model was made in the canine femoral artery. In this review, we present the results of our investigation on autologous vein grafts using this poor-runoff model. We also relate our recent findings on the function of regenerated endothelium in autologous vein grafts.

Animals↗

PubMed1995

A 10-year survivor with unresectable hepatic metastases from sigmoid colon carcinoma treated with regional chemotherapy.

We treated a man with unresectable hepatic metastases from sigmoid colon carcinoma who has since survived for more than 10 years. A sigmoidectomy with lymph node dissection was performed and a continuous hepatic arterial infusion of 5-fluorouracil (5-FU) with intermittent infusion of mitomycin C (MMC) was administered for about 3 months after this operation. The total doses of 5-FU and MMC were 16 g and 84 mg, respectively. Tegafur also was administered orally at a dose of 600 mg/day for about 8 months. The carcinoembryonic antigen (CEA) level (which had reached 4,409 ng/ml preoperatively) normalized 4 months after surgery, and still remains normal. Very few patients with unresectable hepatic metastases survive for 5 or more years. However, regional chemotherapy can be effective in some patients.

Adenocarcinoma↗

PubMed1995

Serum antibody against Helicobacter pylori assayed by a new capture ELISA.

We developed a highly specific detection technique for serum antibody, using a monoclonal antibody to a specific antigen of Helicobacter pylori. A monoclonal antibody preparation that reacted with the 54-kDa molecule of H. pylori antigens was obtained. Using this preparation, an antigen-capture enzyme-linked immunosorbent assay (ELISA) was established by fixation of the monoclonal antibody, followed by reaction with sonicated whole cell antigens. The serum antibody titers of patients with gastritis and peptic ulcers were significantly higher than control titers, and the antibody titer correlated with the histological severity of gastritis. Patients positive for H. pylori by bacterial culture had higher titers than H. pylori-negative patients. Our new ELISA may be useful for the diagnosis of H. pylori infections and for evaluation of the severity of gastric inflammation.

Antibodies, Bacterial↗

PubMed1995

Significance of aminopyrine breath test as a parameter of hepatic functional reserve in 40% partial hepatectomy of rats with CCl4-induced liver injury.

Rats with CCl4-induced liver injury underwent partial (40%) hepatectomy. The [14C]aminopyrine breath test (ABT) values in rats with CCl4-induced liver injury were reduced by 34% compared with those in rats with normal liver. Preoperative ABT values clearly discriminated between survivors and those that died following 40% partial hepatectomy in rats CCl4-induced liver injury (P < 0.05). Hepatic protein synthesis was remarkably enhanced in CCl4-induced liver injury compared with normal liver (P < 0.001), and this was inversely correlated with ABT values (P < 0.001). These data show that the enhanced hepatic protein synthesis could induce a decrease of hepatic functional reserve. ABT seems to be a useful preoperative test for predicting surgical mortality following hepatectomy.

Alanine Transaminase↗

PubMed1995

Goblet cell hyperplasia induced by Strongyloides venezuelensis-infection in Syrian golden hamster, Mesocricetus auratus.

The goblet cell response was examined in the jejunum of the Syrian golden hamster, Mesocricetus auratus, following infection with an intestinal helminth, Strongyloides venezuelensis. After subcutaneous inoculation with 3000 infective larvae (L3), the number of adult worms in the small intestine remained stable up to day 20 and then decreased. Before infection, the number of goblet cells in the jejunum was about 70/10 villi. The number of goblet cells slowly increased, though not significantly, until day 20 post-infection, and then rapidly increased with an inverse relationship to the worm burden. By day 43, when adult worms were almost completely expelled, the number of goblet cells was about 3-fold higher than that of control animals. Similar vigorous goblet cell hyperplasia has not been reported in Strongyloides spp. infection in other experimental animals. Histochemical study revealed that goblet cells in the jejunum of Syrian golden hamsters, unlike those of other experimental animals, produced and released highly sulfated mucins.

Animals↗