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Biomedical subjects

M Shimizu

Publications and source records attributed to M Shimizu.

At least 559 records · Page 31Linked to original sources
PubMed1997

Sheathlin: cloning, cDNA/polypeptide sequences, and immunolocalization of porcine enamel sheath proteins.

Sheath proteins designate low-molecular-weight non-amelogenin enamel polypeptides and their parent protein, which concentrate in the sheath space separating rod and inter-rod enamel (Uchida et al., 1995). Two porcine sheath proteins, with apparent molecular weights of 13 and 15 kDa, are characterized by protein sequencing. The primary structures of these polypeptides match a portion of the derived amino acid sequences of clones isolated from a porcine enamel organ epithelia-specific cDNA library. Sheath protein RNA messages differ by the inclusion or deletion of a 45-nucleotide segment and by the use of three alternative polyadenylation/cleavage sites. The secreted proteins are 395 and 380 residues in length, with molecular masses of 42,358 and 40,279 Daltons and calculated isoelectric points of 6.3 and 6.7, respectively. Polyclonal antibodies were raised against a synthetic peptide having the sheathlin-specific sequence EHETQQYEYSGGC. Immunohistochemistry with this antibody demonstrates that the protein encoded by the sheathlin cDNA is preferentially localized in the sheath space. We propose that the porcine sheath proteins and their proteolytic cleavage products be designated "sheathlin".

Amino Acid Sequence↗
PubMed1997

Cloning and characterization of porcine enamelin mRNAs.

Dental enamel forms by matrix-mediated biomineralization. The components of the developing enamel matrix are generally specific for that matrix. The primary structures of three enamel proteins-amelogenin, tuftelin, and sheathlin (ameloblastin/amelin)-have been derived from cDNA sequences. Here we report the cloning and characterization of mRNA encoding a fourth enamel protein: enamelin. The longest porcine enamelin cDNA clone has 3907 nucleotides, exclusive of the poly(A) tail. The primary structure of the secreted protein is 1104 amino acids in length. Without post-translational modifications, the secreted protein has an isotope-averaged molecular mass of 124.3 kDa and an isoelectric point of 6.5. Polymerase chain-reaction phenotyping of enamelin cDNA suggests that porcine enamelin transcripts are not alternatively spliced and use a single polyadenylation/cleavage site. Immunohistochemical and Western blot analyses with an affinity-purified antipeptide antibody specific for the enamelin carboxyl terminus demonstrate that enamelin is synthesized and secreted by secretory-phase ameloblasts. The parent protein is a 186-kDa glycoprotein that concentrates along the secretory face of the ameloblast Tomes' process. Intact enamelin and proteolytic cleavage products containing its carboxyl terminus are limited to the most superficial layer of the developing enamel matrix, while other enamelin cleavage products are observed in deeper enamel.

Amino Acid Sequence↗
PubMed1997

Pathway and speed of calcium movement from blood to mineralizing enamel.

We studied by autoradiography the distribution of 45Ca in the enamel organ of frozen rats 4.3, 6.1, 7.8, 10.6 and 13.7 sec after an i.v. injection. The intercellular junctions of the proximal side of the smooth-ended ameloblast (SA) and the distal side of the ruffle-ended ameloblast (RA) were closed to calcium. The junctions of the distal side of SA, the proximal side of RA, and both sides of the secretory stage ameloblasts were not. The time required for calcium to pass through the ameloblast layer was less than 1.8 sec in the secretory stage and SA region. The time in the RA region was 3.5-6.3 sec. In the transitional region from RA to SA, a band of strong radioactivity appeared from the papillary layer of RA region towards the enamel of the SA region. The radioactivity in the secretory stage enamel increased almost linearly with time. The diffusion speed of calcium in the enamel was more than 50 microns for 1.8 sec in the maturation stage and less than 15 microns for 9.4 sec in the secretory stage. These results indicate that in the secretory and SA regions calcium moves to the enamel surface through the intercellular spaces of ameloblasts and in the RA region via RA cells.

Ameloblasts↗
PubMed1997

Chondrocyte migration to fibronectin, type I collagen, and type II collagen.

It is well known that cellular interactions, such as cell adhesion, migration, invasion, between cells and the extracellular matrix are mediated by the integrin family of cell surface receptors. Chondrocytes are surrounded by an abundant extracellular matrix, but there is less information on the cellular receptors which interact with this matrix. In our studies, fibronectin, type I collagen, and type II collagen promoted haptotactic and chemotactic migration of chondrocytes, as determined using a modified Boyden chamber system. Treatment of chondrocytes with tyrosine kinase inhibitor, herbimycin or genistein, resulted in a dose dependent inhibition of migration toward these matrix proteins, whereas adhesion of chondrocytes was not influenced. This indicated the existence of functional relationships between protein tyrosine phosphorylation and chondrocyte migration following the adhesion of chondrocytes to matrix proteins. Further study showed that the peptide GRGDSP inhibited chondrocyte migration to fibronectin but not to collagens. On the other hand, chondrocytes migrated toward the tetra-RGD containing peptide, but not the peptide GRGDSP, in a dose dependent fashion. These observations suggest that cross-linking or clustering of integrins is essential to induce transmembrane signaling related to tyrosine phosphorylation for chondrocyte migration toward fibronectin.

Animals↗
PubMed1997

3 beta,5 alpha-Dihydroxycholestan-6-one exists in human blood.

3 beta,5 alpha-Dihydroxycholestan-6-one was detected in fresh human plasma at the concentration of 14-44 ng/mL. The oxysterol binds specifically to phorbol ester specific binding protein in vitro, and may be an endogenous ligand of the protein.

Animals↗
PubMed1997

Establishment of a standardized assay system of fibronectin activity using fibronectin-mediated cell adhesion.

An in vitro assay of fibronectin (FN) was established based on the adhesion of baby hamster kidney (BHK) cells through the cell-binding domain of FN. Each well of a microtiter plate was coated with samples or various concentrations of standard FN. Bovine serum albumin was further coated to prevent the non-specific adhesion of the cells. Various numbers of BHK cells were plated and incubated. After washing out the non-attached cells, the number of attached cells was measured using neutral red (NR)-staining. The conditions for the assay were optimal when 1 x 10(5) cells/well were plated and incubated for 90 min. The linear relationship between the concentration of FN coated and the absorbance of NR was observed in the range of 0.1-1.0 micro/ml of FN. The inhibition of cell binding by the peptides containing an Arg-Gly-Asp (RGD) sequence demonstrated that this assay system depended on FN-mediated cell adhesion through the major cell-binding domain.

Animals↗
PubMed1997

Quantitative evaluation of the rate of myocardial interstitial fibrosis using a personal computer.

We investigated the reliability and reproducibility of an image-analyzing system run on a personal computer for measurement of myocardial interstitial fibrosis. Measurements of myocardial interstitial fibrosis in right ventricular endomyocardial biopsies obtained from patients with hypertrophic cardiomyopathy determined by this image-analyzing system were compared with measurements determined by the point-counting method. We also investigated the correlation between measurements of interstitial fibrosis obtained by image analysis and biochemical measurements of myocardial levels of hydroxyproline in normal and cardiomyopathic hamsters. The intra- and interobserver variability were significantly lower for measurements obtained by the image-analyzing system than for measurements obtained by the point-counting system. Reproducibility was superior with the image-analyzing method. The rate of myocardial interstitial fibrosis determined by the computer image-analyzing method was positively correlated with the hydroxyproline measurement (r = 0.89). Our results suggest that an image-analyzing system using a personal computer provides reproducible results with a high level of reliability.

Adult↗
PubMed1997

Toward molecular strategies for heart disease--past, present, future.

The past two decades of cardiovascular biology and medicine have been based largely upon the consideration of the heart and vasculature as an integrated physiological system, a view that has resulted in major therapeutic advances. With the advent of developments of gene transfer, mouse and human genetics, genetic engineering of intact animals, and molecular and cellular technology, cardiovascular medicine is now on the threshold of a molecular therapeutic era. Major steps have been taken toward unraveling the molecular determinants of complex, integrative, and polygenic cardiovascular disease states, including atherogenesis, hypertension, cardiac hypertrophy and failure, congenital heart disease, and coronary restenosis following balloon angioplasty. Our improved understanding of the fundamental basis of these important cardiovascular disease processes has established a scientific foundation for diagnostic, prognostic, and therapeutic advances in the mainstream of cardiovascular medicine.

Animals↗
PubMed1997

[Studies on novel, stable and potent competitive antagonists against L-DOPA].

DOPA has been proposed to be a neurotransmitter of the primary baroreceptor afferents terminating in the NTS and a neuromodulator in striata of rats. Pre- and post-synaptic recognition sites for DOPA itself may exist. DOPA methyl ester (DOPA ester), a potent competitive antagonist for DOPA in continuously superfused slices or within a several min when microinjected in the NTS has some limitation in in vivo use because of its unstableness as a prodrug for DOPA. We have explored to find stable and potent antagonists for DOPA. At first, we tried to clarify whether or not newly synthesized DOPA esters with bulky structure such as DOPA cyclohexyl ester (CHE), DOPA cyclopentyl ester (CPE) and DOPA cyclopentyldimethyl ester (CPDME) antagonize depressor responses to DOPA microinjected into the NTS, and to what degree these analogues 1 microM perfused via probes are converted to DOPA during striatal microdialysis in urethane-anesthetized rats. Then, we analyzed mode of antagonism of some candidate in the NTS system. During striatal microdialysis, DOPA ester 1 microM increased extracellular levels of DOPA to an 100 fold higher level at a peak of the 1st 20 min sample after the perfusion. Conversion ratio of CHE, CPE and CPDME, compared to DOPA ester, was less than 1/5 at the peak of the 3rd sample, 1/3.3 at the peak of the 2nd, and 1/2.7 at the peak of the 3rd, respectively. In the NTS, all analogues 1 microgram microinjected abolished depressor responses to DOPA 60 ng. Time required to recover a base was 20 min for CHE and 10 min for DOPA ester, CPE and CPDME. At 100 ng, CHE inhibited peak depressor responses to DOPA 60 ng by 53%, CPE by 40%, and CPDME by 24%, compared to DOPA ester by 22%. Then, DOPA 10-300 ng microinjected produced dose-dependent hypotension. DOPA ester 300 ng, microinjected 1 min previously, shifted a dose-response curve for DOPA 30-300 ng to the right without reduction of the maximum response, showing a competitive mode of antagonism. CHE 30 ng competitively antagonized depressor responses to DOPA 18-300 ng without reduction of the maximum response. CHE 100 ng further shifted the dose-response curve for DOPA 18-100 ng to the right with 25% reduction of the maximum response. Antagonistic activity of CHE 100 ng was equipotent with DOPA ester 300 ng. CHE is a competitive antagonist including partially some noncompetitive components. A possible component might be related to NMDA receptors, since high concentrations of CHE displaced selective binding of [3H]-MK-801, an antagonist. In conclusion, CHE is suitable for the purpose.

Animals↗
PubMed1997

Homozygous Q258X mutation in the steroidogenic acute regulatory gene in a Japanese patient with congenital lipoid adrenal hyperplasia.

Congenital lipoid adrenal hyperplasia (CLAH) is an autosomal recessive disorder characterized by impaired synthesis of all adrenal and gonadal steroid hormones. It has recently been reported that mutations in the steroidogenic acute regulatory protein (StAR) gene cause CLAH. We analyzed the nucleotide sequences of exon 7 of the StAR gene in a Japanese CLAH patient with a karyotype of 47,XYY, and her parents. The patient was homozygous for a nonsense mutation Q258X, which changed codon 258 (CAG) encoding Gln to the stop codon TAG, and the her parents were heterozygous for the Q258X mutation. Since the Q258X mutation destroys a MvaI site normally present in the StAR gene sequence, we confirmed the Q258X mutation by means of the restriction endonuclease MvaI digestion of the PCR products. Endocrinological examinations of the parents revealed normal responses of adrenal steroid hormones to exogenous adrenocorticotropin administration, confirming the failure to detect the heterozygous carriers of CLAH by hormonal evaluation.

Adrenal Hyperplasia, Congenital↗
PubMed1997

Stimulus alignments in the selection of bigger rectangles by four- and five-year-old children.

63 four- and five-yr.-olds were presented pairs consisting of one square plus one rectangle with the bases, centers, or tops aligned and asked to select the bigger ones. The larger stimuli were selected more often under all aligned conditions, whereas the taller stimuli were also selected more often under the base- and the center-aligned conditions. In the pairs with the larger square and the smaller rectangle, children selected the taller stimuli most often under the base-aligned and least often under the top-aligned conditions and selected the larger ones more often under the center-aligned than under the base-aligned conditions. These effects of alignments did not appear in the pairs with the smaller square and the larger rectangle. Under the base-aligned condition, children selected the taller stimuli more often and the larger ones less often in the pairs with the larger square and the smaller rectangle than in the pairs with the smaller square and the larger rectangle. These results confirmed that 4- and 5-yr.-olds tend to attend to height dimension, and that, when height is salient, they tend to perceive taller objects as larger.

Attention↗
PubMed1997

[Frequency of hepatitis GB virus C in patients on maintenance hemodialysis].

Hepatitis GB virus C(HGBV-C) RNA was detected in 10(4.5%), among the 223 patients on maintenance hemodialysis. Of the 10 patients with HGBV-C RNA, 4(40%) were also infected with HCV. All patients with HGBV-C RNA had a history of blood transfusion. HGBV-C is suspected to be transmitted by blood transfusion. And also, none of the 10 HGBV-C RNA positive-patients had evidence of liver dysfunction. All patients are asymptomatic HGBV-C carrier. The patients on maintenance hemodialysis are high risk group for HGBV-C infection.

Adult↗
PubMed1997

[Surgical treatment for primary cardiac malignant lymphoma].

A 60-year-old woman who complained of palpitation was diagnosed as ventricular tachycardia on electrocardiography and admitted to our hospital. The ultrasonic cardiography showed cardiac tumor in right ventricle and right atrium. Due to the obstruction of the right ventricle inflow by the tumor, we immediately performed resection of cardiac tumor and repair of right ventricle wall under cardiopulmonary bypass. The tumor was diagnosed as malignant lymphoma by pathological examination for surgical specimen of tumor. After operation her general condition was good, but residual cardiac lymphoma developed large size. We performed radiation therapy for cardiac lymphoma. Therefore the lymphoma was reduced to minimum size. Six months after operation metastatic malignant lymphoma appeared at whole body. So we performed chemotherapy for reduction of systemic malignant lymphoma. At first the chemotherapy was very effective. But metastasis spread rapidly and effectiveness of chemotherapy reduced. Thirteen months after operation she died for respiratory distress, probably due to metastatic brain tumor.

Female↗