Search PubMed⌕ Search

Biomedical subjects

K Hirano

Publications and source records attributed to K Hirano.

At least 145 records · Page 8Linked to original sources
PubMed2000

Thrombin causes endothelium-dependent biphasic regulation of vascular tone in the porcine renal interlobar artery.

Using a method employing front-surface fura-2 fluorometry to measure the cytosolic Ca(2+) concentration, [Ca(2+)](i), the mechanism of endothelium-dependent regulation of vascular tone by thrombin was studied in porcine renal interlobar arterial strips. At concentrations lower than 3 u ml(-1), thrombin evoked only early transient relaxation, while at 3 u ml(-1) and higher concentrations, thrombin caused an early relaxation and a subsequent transient contraction. Both thrombin-induced relaxation and contraction were abolished by removing the endothelium. Similar biphasic responses were observed with a protease-activated receptor-1-activating peptide. Early relaxation was associated with a decrease in [Ca(2+)](i), while the transient contraction was not associated with a change in [Ca(2+)](i) of smooth muscle cells. A thromboxane A(2) (TXA(2))/prostaglandin H(2) (PGH(2)) receptor antagonist (10(-5) M ONO-3708) completely inhibited the thrombin-induced contraction, whereas a thromboxane A(2) synthase inhibitor (10(-5) M OKY-046) only partly inhibited it. When the thrombin-induced contraction was inhibited by ONO-3708, either pretreatment with N(omega)-nitro-L-arginine methylester (L-NAME) or an increase in the amount of external K(+) to 40 mM did not abolish thrombin-induced relaxation during phenylephrine-induced sustained contraction. However, the combination of pretreatment with L-NAME and an elevation of external K(+) to 40 mM completely abolished the relaxation. There was no significant difference in the concentration-dependent effects of thrombin on the initial early relaxation between conditions in which the contractile components either were or were not inhibited. Thrombin is thus considered to mainly activate protease-activated receptor-1 and cause a biphasic response, early relaxation and a transient contraction, in the porcine renal interlobar artery in an endothelium-dependent manner. The thrombin-induced endothelium-dependent relaxation was mediated by nitric oxide and hyperpolarizing factors, while the contraction was mediated by TXA(2) and PGH(2).

Animals↗
PubMed2000

KL-6, a mucinous glycoprotein, as an indicator of chronic lung disease of the newborn.

KL-6 is a mucinous glycoprotein that is preferentially expressed by alveolar type 2 cells. Plasma KL-6 was higher in infants with chronic lung disease (n = 12) than in infants without chronic lung disease (n = 14) on day 0-1, 10, and 30 (P =.04). KL-6 correlated with the alveolar-arterial oxygen tension difference on day 10 and day 30. Plasma KL-6 may be useful as an early marker of chronic lung disease and an indicator of severity.

Antigens↗
PubMed2000

Assessing the effect of an influenza vaccine in an encouragement design.

Many randomized experiments suffer from noncompliance. Some of these experiments, so-called encouragement designs, can be expected to have especially large amounts of noncompliance, because encouragement to take the treatment rather than the treatment itself is randomly assigned to individuals. We present an extended framework for the analysis of data from such experiments with a binary treatment, binary encouragement, and background covariates. There are two key features of this framework: we use an instrumental variables approach to link intention-to-treat effects to treatment effects and we adopt a Bayesian approach for inference and sensitivity analysis. This framework is illustrated in a medical example concerning the effects of inoculation for influenza. In this example, the analyses suggest that positive estimates of the intention-to-treat effect need not be due to the treatment itself, but rather to the encouragement to take the treatment: the intention-to-treat effect for the subpopulation who would be inoculated whether or not encouraged is estimated to be approximately as large as the intention-to-treat effect for the subpopulation whose inoculation status would agree with their (randomized) encouragement status whether or not encouraged. Thus, our methods suggest that global intention-to-treat estimates, although often regarded as conservative, can be too coarse and even misleading when taken as summarizing the evidence in the data for the effects of treatments.

Journal Article↗
PubMed2000

N-linked oligosaccharide processing enzyme glucosidase II produces 1,5-anhydrofructose as a side product.

alpha-1,4-Glucan lyase cleaves alpha-1,4-linkages of nonreducing termini of alpha-1,4-glucans to produce 1,5-anhydrofructose (1,5-AnFru). The enzymes isolated from fungi and algae show high homology with glycoside hydrolase family 31. Purification of alpha-1,4-glucan lyase from rat liver using DEAE Cellulose chromatography resulted in separation of two enzymatic active fractions, one was bound to the column and the other was in the flow-through. Partial amino acid sequence determined from the lyase, retained on the anion exchange column, were identical with that of the N:-linked oligosaccharide processing enzyme glucosidase II. The lyase showed similar enzymatic properties as the microsomal glucosidase such as inhibition by 1-deoxynojirimycin and castanospermine. On the other hand, glucosidase II purified from rat liver microsomes produced not only glucose but also a small amount of 1,5-AnFru using maltose as substrate. Furthermore, CHO cells overexpressing pig liver glucosidase II showed a 1.5- to 2-fold higher lyase activity compared to the nontransfected CHO cells. Conversely, no lyase activity was detectable either in PHAR2.7, the glucosidase II-deficient mutant from a mouse lymphoma cell line, or in Saccharomyces cerevisiae strain YG427 having the glucosidase II gene disrupted. These data demonstrate that glucosidase II possesses an additional enzymatic activity of releasing 1,5-AnFru from maltose.

Amino Acid Sequence↗
PubMed2000

Fluorometric assay of DNA binding proteins using class IIS endonuclease.

Fluorometric DNA-protein binding assay was developed with class IIS restriction enzyme and FITC-labeled DNA fixed on magnetic beads. We used this assay with AP1 and FokI as a case study to quantify DNA-protein binding. Fluorescent images of the treated beads showed that the transcriptional factor bound to its DNA site and inhibited the restricted digestion effectively. The affinity of AP1 to its site was estimated as a Kd value, which was approximately 10 nM.

DNA-Binding Proteins↗
PubMed2000

A novel method for DNA molecular counting.

We have developed a new method for counting DNA molecules using 'capillary-plates' consisting of a large number of small glass-capillary 'channels' fused together in parallel. PCR mixtures containing serially diluted DNA templates with the DNA indicator dye Hoechst 33258 were poured into the plates and sealed with silicone rubber-plates. Following 40 PCR cycles, fluorescence microscopy revealed that the fluorescence in some channels had increased about three-times more than in others at template concentrations of 1 fM or lower. No bright fluorescence was observed in the absence of template. The relationship between the proportion of fluorescent channels in the capillary-plates and the template concentrations was linear according to Poisson probabilities in the range of 0.1-1,000 aM. These results demonstrate the amplification of single templates in the channels, and that a small number of templates could be quantified by counting the proportion of positive channels on the capillary-plates.

Bisbenzimidazole↗
PubMed2000

Zinc induces mixed types of cell death, necrosis, and apoptosis, in molt-4 cells.

To investigate the mode of zinc-induced cell death, the associated morphological changes, and biological events were examined in zinc-treated Molt-4 cells. Fluorescence microscope observations with double staining of zinc-treated cells with Hoechst 33342 and propidium iodide (PI) indicated that the metal induced both necrosis and apoptosis. To confirm this, cells were stained with both PI and FITC-labeled annexin V, which binds phosphatidylserine, and then analyzed by flow cytometry. The results also confirmed that zinc induces mixed types of cell death, necrosis and apoptosis, and that the former induction occurs earlier and at a greater frequency. Hallmarks of apoptosis such as abnormal chromosome condensation and release of cytochrome c, as well as the appearance of annexin-positive cells, appeared along with the expression of mitochondrial membrane protein 7A6. However, zinc did not induce increases in caspase-3 like protease and caspase-8 activities, and caused slightly hypodiploid cells. Furthermore, the induction of cell death and annexin-positive cells was not blocked by the caspase inhibitors Ac-YVAD-CHO and Ac-DEVD-CHO. These results indicate that zinc induces both necrosis and apoptosis, without caspase-3 activation.

Apoptosis↗
PubMed2000

Novel mannose-binding rice lectin composed of some isolectins and its relation to a stress-inducible salT gene.

The novel mannose-binding rice lectin (MRL) purified by Sephadex G-50 or maltamyl Sepharose 4B affinity chromatography was not homogeneous, but the components were separated clearly by two dimensional polyacrylamide gel electrophoresis (1st; isoelectric focusing with Immobiline, 2nd; SDS-PAGE). The major spots were located at pI 4.85 and 4.74, and minor spots at pI 4.66, 4.56, and 4.44; all spots were distributed at about MW 45,000. Other faint spots were sometimes detected just below the major spots. In the western blot analysis, all the spots reacted with the monoclonal antibodies specific to MRL, which bound to MRL and inhibited the lectin activity to agglutinate rabbit erythrocytes. The proteins of the spots at pI 4.85, 4.77, 4.66, and 4.56 had lectin activity. The major proteins at pI 4.85 and 4.77 also had the common amino acid sequence at N-terminus, TLVKIGPWGGNGGSAQDISV, which is almost identical to salt and drought stress-inducible salT gene products in rice plants. High homology was also conserved in both the cDNA and the genomic clones encoding the MRL component at pI 4.85, which were selected with MRL-specific antibodies and an oligonucleotide designed from the partial amino acid sequence. All results suggest that MRL is composed of several isolectins, if not, related proteins having a common epitope and may belong to a family of stress-inducible proteins.

Amino Acid Sequence↗
PubMed2000

Small amount of low-residue diet with parenteral nutrition can prevent decreases in intestinal mucosal integrity.

OBJECTIVE: To investigate the suitable combination ratio of low-residue diet (LRD) and parenteral nutrition (PN) for nutritional support of surgical patients. SUMMARY BACKGROUND DATA: Bacterial translocation (BT) is a severe complication of total parenteral nutrition (TPN). However, it is sometimes impossible to supply sufficient amounts of nutrients to surgical patients by the enteral route. The authors reported previously that concomitant use of LRD with PN provided preferable nutritional support for patients undergoing surgery for colorectal cancer. METHODS: Ninety male Donryu rats were used for three experiments. In experiment 1, rats were divided into two groups to receive TPN or total enteral nutrition with LRD. In experiment 2, rats were divided into six groups, receiving variable amounts of LRD. In experiment 3, rats were divided into five groups to receive isocaloric nutritional support with variable proportions of PN and LRD. Intestinal permeability was assessed by monitoring urinary excretion of phenolsulfonphthalein. BT was assessed in tissue cultures of mesenteric lymph nodes and spleen. RESULTS: In experiment 1, increases in intestinal permeability and BT were observed in rats maintained on 7-day TPN, but not in those maintained on total enteral nutrition for up to 14 days. In experiment 2, the changes in body weight of rats were correlated with the dose of LRD. However, the intestinal permeability was increased only in rats receiving LRD at 15 kcal/kg per day. In experiment 3, additive LRD corresponding to 15% of total caloric intake prevented increases in intestinal permeability and BT. CONCLUSION: Combined nutritional therapy consisting of PN and small amounts of LRD can provide better nutritional support than TPN for surgical patients.

Animals↗
PubMed2000

Pros and cons of inhibiting cholesteryl ester transfer protein.

Whether or not it is desirable to inhibit cholesteryl ester transfer protein (CETP) has been an important question for over fifteen years since genetic CETP deficiency was found. Recently, some epidemiological studies which have been reported in Japan as well as Western countries help to clarify the atherogenicity of human subjects with mutations or polymorphisms in the CETP gene. In addition, some experimental atherosclerosis studies, in which CETP was inhibited in rabbits with different approaches, have been reported. There was a considerable difference in the atherogenicity of human CETP deficiency and CETP-inhibited rabbits. In this review, we summarize the recent advances in this field as well as discussing the significance of CETP in reverse cholesterol transport, a major protective system against atherosclerosis.

Animals↗
PubMed2000

Increased expression after X-irradiation of MUC1 in cultured human colon carcinoma HT-29 cells.

The effect of X-irradiation on production of MUC1 was studied with human colon carcinoma HT-29 cells. As evaluated by immunocytochemical staining, the percentages of MUC1-positive cells in cells at 4 days after 6 Gy irradiation and in unirradiated control cells were 52 +/- 3.5% (n = 6) and 26 +/- 2.8% (n = 6), respectively. Flow-cytometric analysis of living cells showed that MUC1 began to rise from day 1, reaching a plateau by day 4 after 6 Gy irradiation. Western blot analysis with monoclonal antibody MY.1E12 against glycosylated MUC1 (mature form) showed dose-dependent increases of two bands (500 and 390 kDa) corresponding to two polymorphic MUC1 alleles. Premature forms of MUC1 (350 and 240 kDa) were detectable with monoclonal antibody HMFG-2 only in irradiated cells, suggesting that new core protein synthesis had been induced. The transcriptional activity of the MUC1 gene was analyzed in terms of transient expression of MUC1-CAT reporter plasmids containing 5'-flanking sequences of the MUC1 gene fused to the bacterial chloramphenicol acetyltransferase (CAT) gene. The results of CAT assay indicate that enhanced expression of MUC1 in irradiated HT-29 cells was due to upregulation of MUC1 transcription, and required the upstream promoter.

Antibodies, Monoclonal↗
PubMed2000

Corneal amyloidosis caused by Leu518Pro mutation of betaig-h3 gene.

AIM: To report a Japanese family diagnosed clinically as having lattice corneal dystrophy type I (LCDI) in which a Leu518Pro mutation in the betaig-h3 gene and not the R124C mutation reported previously was found. METHODS: Molecular genetic analysis was performed on DNA extracted from peripheral leucocytes from four members (three affected and one unaffected) of a family. Exon 4 of the betaig-h3 gene was amplified by PCR and directly sequenced. Histopathological study was performed on the corneal tissue from the proband obtained during deep lamellar keratoplasty. RESULTS: All the affected members were clinically diagnosed as having LCDI, and the pedigree indicated an autosomal dominant inheritance. A heterozygous single base pair transition (CTG to CCG, leucine to proline) was detected in codon 518 of the betaig-h3 gene in the three affected members, and not in the unaffected member. No mutation was found in codon 124. Amyloid deposits were observed between the collagen bundles of the corneal stroma and were seen to extend deep into the stroma. CONCLUSION: The Leu518Pro mutated betaig-h3 forms amyloidogeneic intermediates which precipitate in the cornea and gives rise to a clinical appearance of LCDI.

Adolescent↗
PubMed2000

Human carcinoma: early experience with phase-contrast X-ray CT with synchrotron radiation--comparative specimen study with optical microscopy.

Phase-contrast x-ray computed tomography (CT) indicates the distribution of the refractive index and has potential to reveal the structures inside soft tissues without a contrast agent. With a synchrotron x-ray source, phase-contrast x-ray CT with a triple Laue-case x-ray interferometer clearly differentiated various human pathologic tissues in the cases of hepatocellular carcinoma with cirrhosis and metastatic colon carcinoma to the liver, and the images closely corresponded to those obtained with low-magnification optical microscopy.

Carcinoma, Hepatocellular↗
PubMed2000

Expression of macrophage (Mphi) scavenger receptor, CD36, in cultured human aortic smooth muscle cells in association with expression of peroxisome proliferator activated receptor-gamma, which regulates gain of Mphi-like phenotype in vitro, and its implication in atherogenesis.

CD36 is one of the major receptors for oxidized low density lipoproteins belonging to macrophage (Mphi) scavenger receptor (SR) class B and is thought to play an important role in the foam cell formation from monocyte-Mphi in the atherosclerotic lesions. Although it has been hypothesized that smooth muscle cells (SMCs) may be the other origin of foam cells in vivo, supporting data are still very limited. In the present study, we have tested the expression of a variety of SRs, including CD36, in 8 lots of primary human aortic SMCs (HASMCs) explanted from 8 different donors. Functional CD36 was expressed in cultured HASMCs, and the levels of expression were widely ranged between the lots. SR class A (SR-A) was expressed abundantly in CD36-negative lots. Other Mphi markers, such as CD32 and CD68, were expressed in all lots tested. These data suggest that the cultured HASMCs gained an Mphi-like phenotype. To determine the mechanism for the above-described phenotypic change, we have tested the expression of a nuclear receptor, peroxisome proliferator activated receptor-gamma, in those cells. This nuclear receptor was abundantly expressed in CD36-positive lots, whereas c-fms was expressed abundantly in CD36-negative/SR-A-positive lots. The synthetic ligand of peroxisome proliferator activated receptor-gamma, troglitazone, upregulated the expression of CD36 only in CD36-positive lots. These observations demonstrate that cultured HASMCs can gain an Mphi-like phenotype, possibly classified by the expression of CD36 or SR-A. The present study may support the possibilities of transformation of HASMCs into foam cells in vivo.

Adult↗
PubMed2000

Oxidized LDL-induced NF-kappa B activation and subsequent expression of proinflammatory genes are defective in monocyte-derived macrophages from CD36-deficient patients.

-CD36 is 1 of the class B scavenger receptor expressed on monocytes, monocyte-derived macrophages (Mphi), platelets, and adipocytes. In our previous studies, we reported that the uptake of oxidized low density lipoproteins (OxLDLs) is reduced by approximately 50% in Mphi from CD36-deficient patients compared with that in control subjects. Recently, we have shown that CD36 is highly expressed in human atherosclerotic aorta. Possibilities have been raised that besides the wide distribution and multifunctional characteristics of CD36, this molecule may also be involved in the mediation of intracellular signaling. The aim of the present study was to elucidate the role of CD36 in cytokine secretion and to investigate the CD36-mediated intracellular signaling stimulated by OxLDL. On addition of OxLDL or thrombospondin-1, the Mphi from CD36-deficient patients secreted significantly less amounts of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) compared with those from controls. RNase protection assay with multiprobe template sets demonstrated that after incubation with OxLDL, the mRNAs of a variety of cytokines, including genes encoding IL-1Ra, IL-1beta, IL-6, TNF-alpha and -beta, and interferon (IFN)-gamma and -beta, were significantly lower in the Mphi of patients. The addition of antibody against CD36 attenuated this OxLDL-induced response in controls. We also observed a reduced response in nuclear factor-kappa B (NF-kappa B) activity in OxLDL-stimulated Mphi from CD36-deficient patients. Unlike OxLDL, stimulation by lipopolysaccharide induced an increase in NF-kappa B activity in Mphi from CD36-deficient patients, suggesting that lipopolysaccharide-mediated signaling was conserved. These results demonstrate that in addition to the reduced OxLDL uptake that we reported previously, CD36-deficient patients may also have an impaired response of OxLDL-induced NF-kappa B activation and subsequent cytokine expression.

Arteriosclerosis↗
PubMed2000

Hydrogen peroxide is an endothelium-derived hyperpolarizing factor in mice.

The endothelium plays an important role in maintaining vascular homeostasis by synthesizing and releasing several endothelium-derived relaxing factors, such as prostacyclin, nitric oxide (NO), and the previously unidentified endothelium-derived hyperpolarizing factor (EDHF). In this study, we examined our hypothesis that hydrogen peroxide (H(2)O(2)) derived from endothelial NO synthase (eNOS) is an EDHF. EDHF-mediated relaxation and hyperpolarization in response to acetylcholine (ACh) were markedly attenuated in small mesenteric arteries from eNOS knockout (eNOS-KO) mice. In the eNOS-KO mice, vasodilating and hyperpolarizing responses of vascular smooth muscle per se were fairly well preserved, as was the increase in intracellular calcium in endothelial cells in response to ACh. Antihypertensive treatment with hydralazine failed to improve the EDHF-mediated relaxation. Catalase, which dismutates H(2)O(2) to form water and oxygen, inhibited EDHF-mediated relaxation and hyperpolarization, but it did not affect endothelium-independent relaxation following treatment with the K(+) channel opener levcromakalim. Exogenous H(2)O(2) elicited similar relaxation and hyperpolarization in endothelium-stripped arteries. Finally, laser confocal microscopic examination with peroxide-sensitive fluorescence dye demonstrated that the endothelium produced H(2)O(2) upon stimulation by ACh and that the H(2)O(2) production was markedly reduced in eNOS-KO mice. These results indicate that H(2)O(2) is an EDHF in mouse small mesenteric arteries and that eNOS is a major source of the reactive oxygen species.

Acetylcholine↗
PubMed2000

Improvement in the histopathology of hearts from cardiomyopathic BIO TO-2 hamsters following long-term administration of amlodipine and cilnidipine.

The effect of long-term administration of amlodipine and cilnidipine was examined on the histopathology and 1,4-dihydropyridine (DHP) calcium channel antagonist receptors in the left ventricle of BIO TO-2 hamsters, a model of dilated cardiomyopathy (DCM). Oral administration of amlodipine (3 and 10 mg/kg/d, 19 weeks) in 7 week-old BIO TO-2 hamsters produced a significant reduction in calcium deposition and necrosis with little change in the cavity area and fibrosis. A reduction of calcium deposition and necrosis in the myocardium of BIO TO-2 hamsters was also seen following similar administration of cilnidipine (10 mg/kg/d). The long-term administration of amlodipine (3 and 10 mg/kg/d) caused a significant increase (36.6 and 21.7%, respectively) in the Bmax for specific (+)-[3H]PN 200-110 binding in the myocardium from BIO TO-2 hamsters, compared with that in control hamsters. In conclusion, the present study has shown that long-term administration of amlodipine and cilnidipine improves calcium deposition and necrosis in the myocardium from BIO TO-2 hamsters. Thus, these data suggest that both agents may be effective pharmacological treatments of DCM.

Amlodipine↗