Biomedical subjects

H Sugihara

Publications and source records attributed to H Sugihara.

At least 235 records · Page 13Linked to original sources

PubMed1993

Isolation and characterization of procoagulant from the venom of Vipera aspis aspis.

1. A procoagulant protein was isolated from Vipera aspis aspis (Aspic viper) venom by Sephadex G-75, DEAE-Sephacel, Q-Sepharose and Sephadex G-150 column chromatography. 2. The purified protein has a molecular weight of 125,000 and an isoelectric point of 4.3. 3. This procoagulant decreased the clotting time of plasma from humans, however, direct fibrongen clotting activity was not detected. 4. Diisopropyl fluorophosphate, a serine-protease inhibitor affected coagulant activity of purified protein significantly, while a factor Xa inhibitor (3-ABPE) possessed a slight inhibitory effect. 5. Bovine prothrombin incubated with isolated protein, phospholipid emulsion, bovine factor V and calcium ions drastically decreased the clotting time of fibrinogen and expressed hydrolytic activity against synthetic arginine esterase substrates. However, no hydrolytic activity these substrates was detected with the procoagulant alone indicating that this protein might participate in activation of prothrombin.

Amino Acid Sequence↗

PubMed1993

Effect of bilineobin, a thrombin-like proteinase from the venom of common cantil (Agkistrodon bilineatus).

A thrombin-like proteinase, named bilineobin, was isolated from Agkistrodon bilineatus venom by Sephadex G-75, DEAE-Sephacel and Heparin-Sepharose CL-6B column chromatography. The purified enzyme has a mol. wt of 57,000 and catalysed the hydrolysis of arginine esters and thrombin substrates Boc-Val-Pro-Arg-MCA and Boc-Asp(OBz)-Pro-Arg-MCA. Although bilineobin converted fibrinogen into fibrin resulting in the production of fibrinopeptides, the activity was relatively low (0.65 NIH units/mg). Fibrinopeptides released upon hydrolysis by this proteinase were identified as fibrinopeptide A (FpA) and fibrinopeptide B (FpB) by measuring fast atom bombardment (FAB) mass spectra and amino acid sequence. This indicates that bilineobin hydrolyses the Arg(19)-Gly(20) bond in the A alpha chain and the Arg(21)-Gly(22) bond in the B beta chain of the bovine fibrinogen molecule. Kinetic study of FpA and FpB release reveals that bilineobin has a preference for cleaving the B beta chain. In addition, bilineobin is resistant to thrombin inhibitors such as hirudin. These suggest that the mechanism of action of bilineobin is similar but not identical to that of thrombin. It was demonstrated that the NH2-terminal region of bilineobin has significant similarities in sequence with thrombin-like proteinases from other snake venoms; however, only three residues were common with thrombin up to residue number 24.

Amino Acid Sequence↗

PubMed1993

Barrel rotation evoked by intracerebroventricular injection of somatostatin and arginine-vasopressin is accompanied by the induction of c-fos gene expression in the granular cells of rat cerebellum.

Intracerebroventricular (i.c.v.) injection of somatostatin (SS) or arginine-vasopressin (AVP) elicits barrel rotation (BR) in rats. To identify the potential neuron structures involved in this characteristic behavior, the regional expression of the c-fos gene in rat brain after i.c.v. injection of SS (10 micrograms) or AVP (1 micrograms) was examined by hybridization histochemistry. The c-fos expression could serve as a marker of neuronal activity and/or neural transmission. Following SS-induced BR, c-fos gene expression was observed in the lingula, uvula, nodulus, simplex, centralis, and culmen of the cerebellum, while following AVP-induced BR, c-fos gene expression was observed in the first four of the above-mentioned regions of the cerebellum, but not in the centralis or culmen. In these regions, the c-fos mRNA signals were observed on the granular layer. Expression of the c-fos gene was immediately and transiently induced and was not observed in rats in which BR was not evoked after SS or AVP injection. In both control rats and SS- or AVP-injected rats, the c-fos gene expression was induced in the piriform cortex and the flocculus of the cerebellum. The findings suggest that BR is a manifestation of behavior induced by massive transsynaptic activation of the granular cells in the cerebellum.

Animals↗

PubMed1993

Differentiation of brown adipose cells in three-dimensional collagen gel culture.

Brown adipose cells are heat-producing cells through non-shivering thermogenesis by intramitochondrial "thermogenin". This specific protein is a marker for their cellular differentiation. It has long been known that cultured brown adipose cells in monolayer rapidly lose the thermogenin bioactivity. In this study, we cultured brown adipose cells in three-dimensional type I collagen gel matrix. Under this culture condition, they were able to survive, and differentiated morphologically and functionally for a long period of time, especially exhibited the characteristic immunohistochemical activity of thermogenin. These findings suggest that brown adipose cells differentiate in type I collagen gel. In this condition, cholera toxin or BRL 37344, one of beta 3-adrenoceptor agonists, specifically stimulated the brown adipose cells.

Adipose Tissue, Brown↗

PubMed1993

Influence of extracellular matrix on the proliferation and differentiation of adrenocortical cells in culture.

Extracellular matrix is indispensable for cell differentiation in vivo. The aim of this study is to examine effects of extracellular matrix on adrenocortical cells embedded in collagen gel in terms of cellular proliferation and differentiation. Dissociated human or bovine adrenocortical cells were embedded in collagen gel; the gel was then immediately thin-coated on the bottom of culture dishes. The cortical cells were thus trapped in collagen and cultured in monolayer, a suitable means for observation of culture cells. To the authors' best knowledge, this "collagen gel-embedded" monolayer culture is described here for the first time. Cortical cells were spherical in shape and had the activities of proliferation and steroid production in collagen gel matrix of this culture method. Aldosterone production in particular tended to be maintained much longer than in conventional monolayer culture. This method seems to provide a physiological environment in which to study the differentiation of cortical cells.

Adrenal Cortex↗

PubMed1993

Primary structures of platelet aggregation inhibitors (disintegrins) autoproteolytically released from snake venom hemorrhagic metalloproteinases and new fluorogenic peptide substrates for these enzymes.

A hemorrhagic protein (60 kDa), HR1B, present in the venom of Trimeresurus flavoviridis is a mosaic protein consisting of an NH2-terminal metalloproteinase-domain, a disintegrin (platelet aggregation inhibitor)-like domain, and a unique COOH-terminal Cys-rich domain. Since the gross structures of HR1B and protein precursors of disintegrins, trigramin, and rhodostomin, all of which contain the metalloproteinase domain, are similar, many disintegrins so far detected in snake venoms are assumed to be autoproteolytic fragments released from precursors. In ongoing related experiments, the newly purified hemorrhagic metalloproteinases, HR1A from T. flavoviridis venom and HT-1 from Crotalus ruber ruber venom, in addition to HR1B, were autoproteolyzed, in the absence of Ca2+, at 37 degrees C for 3-12 h. Under these conditions, HR1A, HR1B, and HT-1 each released a single major fragment of 32, 34, and 31 kDa, respectively. The entire amino acid sequences of the isolated fragments indicated the presence of disintegrin-like and Cys-rich domains in the COOH-terminal regions of HR1A, HR1B, and HT-1, respectively. It seems likely that so-called disintegrins probably originate from various metalloproteinases present in venom. On the bases of peptide sequences close to the autoproteolytic cleavage sites of these metalloproteinases and the sites of fibrinogen cleaved by these enzymes, we synthesized new intramolecularly quenched fluorogenic peptide substrates. Among the 10 peptides tested, 2-aminobenzoyl (Abz)-Ser-Pro-Met-Leu-2,4-dinitroanilinoethylamide (Dna) proved to be the best substrate for venom metalloproteinase, as deduced from kinetic analyses.

Amino Acid Sequence↗

PubMed1993

Expression of growth hormone receptor gene in rat hypothalamus.

Growth hormone receptor (GHR) mRNA-expressing cells in the hypothalamus were observed using hybridization histochemistry in adult male rats. Digoxigenin-labeled cRNA corresponding to the extracellular part of rat GHR was used as a probe. Northern blotting analysis of hypothalamic total RNA from adult male rats revealed that the 4.5 kilobase (kb) transcript of the GHR gene corresponding to the GHR messenger RNA (mRNA) predominated over the 1.2 kb transcript corresponding to GH-binding protein mRNA. GHR mRNA-containing cells were observed in the arcuate nucleus (ARC), the periventricular nucleus (PeV), ventrolateral region of the ventromedial nucleus, the paraventricular nucleus and the supraoptic nucleus. To further understand the significance of the GHR gene expression in the hypothalamus, the effect of in vivo manipulation of GH on the somatostatin (SS) gene expression in the ARC and PeV, and the GRF gene expression in the ARC was observed among adult male rats using in situ hybridization histochemistry. Ten days after hypophysectomy, the SS mRNA level in the ARC as well as PeV was significantly lower than that in the respective nuclei of sham-operated control rats, while the GRF mRNA level in the ARC was significantly higher than that in the ARC of control animals. Subcutaneous injection of recombinant human GH (0.33 mg) to hypophysectomized rats every 12 h for 5 days restored the SS mRNA level in the ARC and PeV, and reduced the GRF mRNA level in the ARC to that of control animals. The data suggest that GH directly acts on the hypothalamic PeV and ARC, and alters the gene expression of SS and GRF.

Animals↗

PubMed1993

Effect of parenteral administration of selected nutrients and central injection of gamma-globulin from antiserum to neuropeptide Y on growth hormone secretory pattern in food-deprived rats.

Serial changes in growth hormone (GH) secretory pattern during 72 h food deprivation and in response to refeeding after 72 h food deprivation were observed in adult male Wistar rats. In addition, the effects of intravenous administration of glucose, fatty acids or amino acids and central injection of gamma-globulin from antiserum to human neuropeptide Y (NPY) on the GH secretory pattern were examined in 72 h food-deprived rats. Rats were provided with chronic indwelling right atrial cannula and serial blood specimens were withdrawn via the cannula every 10 min using an automatic blood-sampling device. The GH secretory pattern was analyzed using a Pulsar computer program. In fed rats, episodic GH secretion with an amplitude exceeding 350 ng/ml occurred at 170-min intervals and intervening trough GH levels were less than 10 ng/ml. During the 72 h food deprivation, the amplitude of pulsatile GH secretion decreased progressively without altering the pulse frequency. When 72 h food-deprived rats were refed lab chow, the pulse frequency and the pulse amplitude increased rapidly and lasted for 6-8 h. Normal pulsatile GH secretion resumed by 10-12 h after refeeding following the 72 h food deprivation. To identify the specific nutrient (i.e. glucose, fatty acids or amino acids) serving as the signal causing restoration of the central drive to GH secretion, glucose solution (2 or 6 kcal), lipid emulsion (2 or 6 kcal) or amino acid solution (2 kcal) was administered intravenously to the 72 h food-deprived rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

PubMed1993

Somatostatin reduces transcription of the growth hormone gene in rats.

To examine whether somatostatin (SS) exerts influences on the steady state levels of GH-releasing factor (GRF), the effect of SS on GH gene transcription was examined in rats. This approach was used because it has been shown that GRF stimulates GH gene transcription independent of GH release, and SS does not inhibit basal or GRF-stimulated GH gene transcription. Therefore, it is assumed that an effect of SS on GH gene transcription would be mediated by a change in GRF levels. Adult female Sprague-Dawley rats were provided with right atrial cannulae. Studies were performed using unanesthetized rats. Pituitary GH gene transcription was measured by transcription assay. An iv administration of antiserum to rat GRF 150 min previously significantly decreased GH gene transcription compared with that in control rats given normal goat serum. A continuous infusion of SS (300 micrograms/kg.h) via the cannula for 150 min significantly decreased GH gene transcription compared with that in control rats receiving 0.9% NaCl. When GRF (3 micrograms/kg.h) was given simultaneously with SS (300 micrograms/kg.h), GH gene transcription increased significantly compared with that in rats receiving SS infusion alone. After the withdrawal of SS infusion, GH gene transcription rapidly and significantly increased. The data suggest that SS reduces the steady state levels of GRF.

Animals↗

PubMed1993

Plural cells organize thyroid follicles through aggregation and linkage in collagen gel culture of porcine follicle cells.

Some follicle cells organize thyroid follicles through proliferation of single cells in collagen gel culture. The aim of this study was to clarify whether two or more (plural) cells can form follicles through aggregation and linkage in this culture system. To address this question, we performed collagen gel culture of porcine follicle cells, using cell labeling with PKH 2 dye. A mixture of dye-labeled and nonlabeled cells was cultured in collagen gel. Organized follicles consisted of both dyed and nondyed cells. This suggested that plural cells reconstructed follicles through aggregation and linkage. To further confirm this finding, cells embedded in collagen gel were cultured in inhibition of cell proliferation with 2 micrograms/ml aphidicolin. Forty to 60 percent of the cells formed follicles, which did not grow larger. Electron microscopy showed that intracytoplasmic cavities appeared in the cells. In contrast, 20-30% of the embedded cells developed into cavity-embracing single cells, which remained signet rings. These results indicate that follicle cells organize follicles in collagen gel culture by two means: through cell division of cavity-embracing single cells and through aggregation and linkage of plural cells.

Animals↗

PubMed1993

Occurrence of paroxysmal synchronous EEG discharges in subcortical arteriosclerotic encephalopathy (Binswanger's disease).

A 72-year-old patient with von Recklinghausen's disease showed akinetic mutism within 6 months of the onset of dementia. The findings of diffuse cerebral atrophy on CT and periodic synchronous discharges (PSDs) in EEG suggested Creutzfeldt-Jakob disease. However, autopsy findings of diffuse softening of the subcortical white matter and marked arteriosclerotic changes of the subcortical arterioles with sparing of the cortex in the cerebrum confirmed a diagnosis of Binswanger's disease. Binswanger's disease should be included in the differential diagnosis of dementia showing PSDs in EEG.

Aged↗

PubMed1993

Reconstruction of cornea in three-dimensional collagen gel matrix culture.

PURPOSE: To report on an in vitro reconstruction of cornea composed of three main layers. METHODS: These layers were reconstructed from isolated bovine corneal cells in three-dimensional collagen gel culture conditions. RESULTS: The topmost layer, corneal epithelial cells on collagen gel, was treated by what was designated as the "air-liquid interface method," a method developed specifically for this study. The treatment induced proliferation and differentiation of this layer. Those epithelial cells were positively immunostained with anti-epithelial keratin (55-kD/64-kD) antibody, AE5, which was specific for in vivo corneal epithelial cells. The middle layer consisted of keratocytes in the three-dimensional collagen gel matrix. This layer was comparable to the stroma in vivo. The underlayer, the counterpart of in vivo endothelial layer, was composed of corneal endothelial cells. CONCLUSIONS: This culture system probably provides a physiologic environment for detailed investigations of corneal cells.

Animals↗

PubMed1993

[A case of acute coronary syndrome followed by 201Tl, 123I-BMIPP and 123I-MIBG myocardial imagings before and after PTCA].

201TlCl (Tl), 123I-beta-methyl-p-iodophenyl pentadecanoic acid (BMIPP) and 123I-metaiodobenzylguanidine (MIBG) images were applied before and after PTCA to a patient with acute coronary syndrome who had repeated ischemic attacks on light effort. A decreased tracer uptake was noted at the mid and apical anteroseptal regions in each image before PTCA. MIBG uptake was most decreased and Tl uptake was least decreased, but Tl uptake at exercise decreased as severely as MIBG uptake at rest. After PTCA an uptake was improved in each image. Both before and after PTCA, the area with decreased BMIPP uptake corresponded to the region with deteriorated wall motion obtained by echocardiography. It was suspected that the decreased MIBG uptake correlates with "area at risk" and that the decreased BMIPP uptake correlates with abnormal wall motion. Taken together those findings, it was indicated that an application of these cardiac scintigraphy is useful for the evaluation of comprehending metabolic abnormality and recovering process of ischemic myocardium.

3-Iodobenzylguanidine↗

PubMed1993

Isolation and characterization of fibrinogenase from Candida albicans NH-1.

1. Fibrinogenase was isolated from Candida albicans NH-1 by DEAE-Cellulose, Sephadex G-75 and Sephadex G-100 column chromatographies. 2. The purified fibrinogenase gave a single band on disc polyacrylamide gel electrophoresis, isoelectric focusing and sodium dodecyl sulfate polyacrylamide gel electrophoresis. 3. The enzyme preparation had a molecular weight of 13,000, isoelectric point of pH 4.2 and possessed 117 amino acid residues. 4. The purified fibrinogenase possessed capillary permeability-increasing activity. 5. The enzyme hydrolyzed fibrinogen, casein, hide powder azure, azocoll hydrolytic activities and also hydrolyzed the oxidized B chain of insulin. The cleavage sites in the oxidized B chain of insulin were identified as Asp(3)-Glu(4), Glu(13)-Ala(14), Ala(14)-Leu(15), Tyr(16)-Leu(17), Arg(22)-Gly(23), Phe(25)-Tyr(26) and Tyr(26)-Thr(27). 6. Fibrinogenase activity of this preparation was inhibited by alpha 2-macroglobulin antithrombin-III, o-phenanthroline, disodium ethylenediaminetetra acetic acid and dithiothreitol.

Amino Acid Sequence↗

PubMed1993

[Assessment of myocardial perfusion abnormalities in patients with apical hypertrophic cardiomyopathy using exercise 201Tl scintigraphy].

Regional myocardial perfusion abnormalities commonly occur during exercise in patients with hypertrophic cardiomyopathy (HCM). Exercise 201Tl myocardial scintigraphy has provided a noninvasive means of identifying myocardial perfusion abnormalities in patients with HCM. On the other hand, apical hypertrophic cardiomyopathy (APH) is reported as a subtype of HCM. Whether APH is essentially equal to HCM or not is controversial. To assess myocardial ischemia in patients with APH, we studied 28 patients with APH, with exercise 201Tl SPECT. Myocardial perfusion images were obtained immediately after submaximal exercise and again after a 3-hour delay. Regional perfusion defects during exercise were identified in 19 of the 28 patients (68%) with APH. Complete reversible defects were observed in 15 (79%) patients with APH. Although perfusion defects were present in all regions of the left ventricle in patients with HCM, they were present only in the apical region in patients with APH. Thus, reversible 201Tl perfusion abnormalities commonly occur during exercise in patients with APH as well as in patients with HCM.

Adult↗

PubMed1993

[The distribution and kinetics of 123I-MIBG in normal human hearts].

123I-metaiodobenzylguanidine (MIBG) myocardial scintigraphy was performed in twelve normal human subjects, and the distribution and clearance of MIBG were estimated. In PLANAR studies, heart to mediastinum (H/M) and lung to mediastinum activity ratio (L/M) were 2.68 +/- 0.45 and 1.82 +/- 0.23 in initial images, and 2.75 +/- 0.44 and 1.55 +/- 0.13 in delayed images. The clearance of MIBG in the lung was more rapid than that in the heart, 41.6 +/- 4.1 vs. 29.8 +/- 3.2 (%), therefore MIBG cardiac profiles were more clarified in delayed images. In SPECT studies, the regional relative uptake (RRU) in anterior, septal, inferior and lateral wall were 100.0 +/- 0, 93.9 +/- 6.9, 85.0 +/- 10.0 and 104.9 +/- 8.3 (%) in initial images, and 100.0 +/- 0, 96.8 +/- 9.6, 79.4 +/- 8.3 and 99.1 +/- 7.9 (%) in delayed images, respectively. The RRU in inferior wall was significantly lower than those in the other walls. The regional clearance rate (RCR) in these four walls were 25.1 +/- 4.6, 22.5 +/- 8.2, 29.7 +/- 8.3, 29.3 +/- 3.4 (%), respectively. The RCR in inferior and lateral wall were significantly larger than those in anterior and septal walls. The RRU in basal, mid and apical portion were 100.0 +/- 0, 104.5 +/- 3.3, 98.9 +/- 12.1 (%) in initial images, and 100.0 +/- 0, 103.9 +/- 4.5, 96.8 +/- 15.2 (%) in delayed images, respectively. The RRU in mid portion was significantly higher than that in basal portion.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Iodobenzylguanidine↗

PubMed1993

[Quantitative analysis of right ventricular overloading by 201Tl myocardial SPECT].

Clinical usefulness of quantitative analysis of right ventricular overloading was evaluated by 201Tl myocardial SPECT in comparison with cardiac catheterization and MRI. Seventy-four MBq of 201TlCl was intravenous injected and 201Tl myocardial SPECT was performed on 40 patients (mean age: 61.0 +/- 11.8) with right ventricular overloading. Regions of interest (ROI) were selected on right and left ventricular walls in a midventricular short axis image of SPECT and uptake of each ROI were counted. The right ventricle (RV)/left ventricle (LV) 201Tl uptake ratio (R/L-Tl) was calculated. Wall thickness of RV and LV were measured on MRI and the RV/LV wall thickness ratio (R/L-WT) was calculated. RV and LV pressure were recorded in cardiac catheterization, and the RV/LV systolic pressure ratio (R/L-P) was calculated. There was significant positive correlation (Y = 0.73X + 0.19, r = 0.71, p < 0.001) between R/L-Tl and R/L-WT. R/L-Tl was positively correlated with R/L-P in patients with pressure overload (Y = 1.14X - 0.049, r = 0.85, p < 0.001) and in patients with volume overload (Y = 0.51X + 0.023, r = 0.88, p < 0.001) and the slope of the regression line in patients with pressure overload was significantly steeper than that in patients with volume overload (p < 0.001). In conclusion, quantitative analysis of right ventricular overload by 201Tl myocardial SPECT is useful to estimate RV/LV wall thickness ratio and pressure ratio.

Adult↗

PubMed1993

[Assessment of reverse redistribution on exercise 201Tl myocardial SPECT in patients with hypertrophic cardiomyopathy].

Reverse redistribution is revealed on exercise 201Tl myocardial SPECT in some cases with hypertrophic cardiomyopathy. Ten patients with hypertrophic cardiomyopathy without coronary artery disease who showed reverse redistribution visually were studied. Reverse redistribution was evaluated by the early and delayed images of exercise 201Tl myocardial SPECT. Relation between reverse redistribution and distribution of left ventricular hypertrophy estimated by echocardiography and magnetic resonance imaging was investigated, and the mechanism of reverse redistribution was analyzed by referring to Bull's eye display of washout rate. Reverse redistribution was displayed in non-hypertrophic region and washout rate was normal in that region and low in hypertrophic region. In conclusion, in order to interpret reverse redistribution it is necessary to recognize hypertrophic region and to refer to washout rate.

Adult↗