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Biomedical subjects

N Kawada

Publications and source records attributed to N Kawada.

At least 91 records · Page 5Linked to original sources
PubMed1997

Electrophysiological characterization of cultured hepatic stellate cells in rats.

This study aimed to examine electrophysiological properties of cultured rat hepatic stellate cells (HSCs) using the whole cell configuration of patch-clamp technique. At least three different current components were identified. First, when the membrane was depolarized to voltages more positive than -40 mV, a transient outward K+ current was evoked. Second, membrane hyperpolarization below -60 mV evoked a sustained and inward-rectifying K+ current. The third component was a current flowing outward, which was activated when the cell was depolarized more positively than 0 mV. The channel for this current allowed Na+, K+, and Cl- to pass nonspecifically, suggesting the presence of hemi gap-junctional channel. Furthermore, a laser photobleaching technique revealed the presence of gap junctions between adjacent HSCs. A voltage-gated Ca2+ current, which is known to occur in smooth muscle cells, was searched for but was not detectable. These results suggest that membrane potential of HSCs is determined specifically by the two distinct K+ channels and by an intercellular mechanism involving gap-junctional communication.

Animals↗
PubMed1997

Immunohistochemical and ultrastructural analyses of in situ activation of hepatic stellate cells around Propionibacterium acnes-induced granulomas in the rat liver.

Stellate cells embrace the hepatic sinusoids as pericytes. To demonstrate their activation in juxta-sinusoidal location, we induced hepatic granulomatous inflammation, which did not accompany liver injury nor fibrosis, by administrating a low dose (10 mg/kg b.w.) of heat-killed Propionibacterium acnes into rats. Macrophages and lymphocytes migrated out of the sinusoid and made compact granulomas in the space of Disse. Stellate cells which faced on the granulomas were closely attached to the sinusoidal endothelial cells and extended projections into the spaces between the constituent mononuclear cells of granulomas. They did not migrate into granulomas nor displayed mitosis. Immunohistochemically, stellate cells around the granulomas expressed alpha-smooth muscle actin (alpha-SMA), whereas those in the non-granulomatous regions did not. Small deposits of type III collagen were found at the periphery of granulomas. Biochemical analysis showed an increased amount of alpha-SMA protein and type III collagen mRNA in the granuloma-bearing liver. Transforming growth factor-beta and platelet-derived growth factor were detected within the granulomas. The present study has revealed that stellate cells are activated in situ by extrasinusoidal macrophages in a paracrine manner without being detached from the sinusoidal wall.

Animals↗
PubMed1997

[Fibrin glue spray treatment on the polytetrafluoroethylene graft].

An outer surface of a 4 mm polytetrafluoroethylene (PTFE) prosthesis, Gore-Tex thin-walled stretch graft, was treated by the fibrin glue spray with a 1.0 kg/m2 compressed air. The distance between the PTFE graft and a spray-nozzle was within 0.5 cm. After the treatment, an outer surface and a cross-section of the PTFE graft were examined under a scanning electron microscope. Fibrin glue mended the gap of the PTFE fibril of an outer surface and a fibrin glue layer adhered firmly to the graft. The fibrin glue spray treatment may prevent postoperative serous fluid leakage.

Aerosols↗
PubMed1997

[Appropriate early open heart palliation of univentricular atrioventricular connection with subaortic stenosis].

Long-term conventional pulmonary artery banding deteriorates ventricular function in patients who have univentricular atrioventricular connection with subaortic obstruction. Protection of the pulmonary vascular bed and early relief of subaortic stenosis is essential to improve the outcome after Fontan operation. From January 1995 through January 1996, three infants underwent open heart palliation because of univentricular atrioventricular connection with subaortic stenosis. All infants had discordant ventriculoarterial connection. One infant underwent the Norwood procedure (patient 1). Two infants underwent palliative arterial switch operation, one with repair of aortic arch and a Blalock-Taussig shunt (patient 2) and the other with endoluminal pulmonary artery banding (patient 3). Patient 3 required a subsequent conventional pulmonary arterial banding. The postoperative recovery period were smooth in the all infants. All infants kept sufficient PO2 ranging from 34 to 37 mmHg postoperatively. On follow-up after 16 months pulmonary artery index decreased in patient 1. On the other hand, angiogram demonstrated satisfactory pulmonary arterial growth in patient 2. There were two late death, occurring in patient 1 (sudden death) and patient 3 (pneumonia). Patient 2 awaits a Fontan type procedure. It is difficult to adjust appropriate blood flow through a Blalock-Taussing shunt and a surgically isolated pulmonary artery is capable of inducing pulmonary distortion after Damus-Norwoood type operation. Whereas natural regulation of the pulmonary arterial blood flow by a restrictive ventricular outflow tract is come up after a palliative arterial switch operation. Palliative arterial switch operation is an useful alternative open heart palliation for neonates and early infants who had univentricular atrioventricular connection with subaortic stenosis.

Aortic Valve Stenosis↗
PubMed1997

The hepatic perisinusoidal stellate cell.

Hepatic stellate cell (also referred to as Ito cell, fat-storing cell, perisinusoidal cell, lipocyte) is one of the sinusoid-constituent cells that play multiple roles in liver pathophysiology. Although identification of the stellate cell had taken about 100 years because of the misconception caused by the discoverer von Kupffer, Wake made a great contribution to the "re" discovery of the cell in 1971. Establishment of the isolation of hepatic non-parenchymal cells from rats by Knook has made it possible to uncover the metabolic function of individual cells. Now, the stellate cell function is expanding from a retinol (fat)-storing site to a center of extracellular matrix metabolism and mediator production in the liver. Function as a liver specific pericyte has also been elucidated. Transition of the stellate cells from the vitamin A-storing phenotype to "activated" or "myofibroblastic" cells that produce a large amount of type I collagen and transforming growth factor beta triggers the progress of liver fibrosis in the course of hepatic inflammation. Communication of the stellate cells with the other hepatic constituent cells and invading inflammatory cells is also an important factor that regulates the local pathological reaction. Analysis of cellular and molecular aspects of the stellate cell activation would lead to the establishment of a novel therapeutic strategy against the progress of liver fibrosis in human liver disease.

Animals↗
PubMed1997

TCV-116 inhibits interstitial fibrosis and HSP47 mRNA in rat obstructive nephropathy.

Unilateral ureteral obstruction (UUO) is a well established disease model leading to fibrosis of the obstructed kidney. In this model, involvement of enhanced renin-angiotensin system in the pathogenesis of interstitial fibrosis has been demonstrated. A 47-kDa heat-shock protein (HSP47) was originally identified as a collagen-binding stress protein, and is currently considered to be a collagen-specific molecular chaperone that plays a pivotal role during the biosynthesis and secretion of procollagen from endoplasmic reticulum. To test if HSP47 is involved in interstitial fibrosis in UUO, we examined the expression of HSP47 mRNA in rat UUO kidneys after 12 hours. 1, 4, 7 days of obstruction. HSP47 mRNA expression was significantly increased as early as 12 hours after obstruction and was sustained at the increased level until seven days. Type I collagen mRNA significantly increased after four days of UUO. Fibrotic changes of interstitium appeared in Masson's trichrome stained section after four days. To explore the possible involvement of angiotensin II (Ang II) in HSP47 induction, the effect of Ang II receptor antagonist (TCV-116) and angiotensin converting enzyme inhibitor (lisinopril) was tested. TCV-116 or lisinopril was given to the animals orally once a day at the dose of 10 mg/kg. TCV-116 or lisinopril significantly ameliorated the fibrotic change of interstitium seven days after obstruction. HSP47 and type I collagen mRNA levels in the TCV-116- or lisinopril-treated groups were reduced to about 60% of untreated UUO. A possible involvement of HSP47 in the pathogenesis of interstitial fibrosis in UUO is suggested; however, further investigation is required to identify the signals involved in the induction of HSP47 in UUO.

Angiotensin II↗
PubMed1996

Smooth muscle alpha-actin expression in rat hepatic stellate cell is regulated by nitric oxide and cGMP production.

Mechanism by which lipopolysaccharide (LPS) and interferon gamma (IFN gamma) downregulate smooth muscle (SM) alpha-actin expression in rat hepatic stellate cell was studied. Culture-induced SM alpha-actin expression was suppressed in the presence of LPS (> 5 ng/ml) and/or IFN gamma (> 1 U/ml) dose-dependently. Inhibition of nitric oxide (NO) production by N omega-nitro-L-arginine (LNA) canceled such an inhibitory effect of LPS and/or IFN gamma. LPS and/or IFN gamma increased cellular level of cGMP in an LNA inhibitable manner, and cGMP analogues attenuated SM alpha-actin expression of the cells. These results indicate that NO-dependent increase of cellular cGMP level mediates the inhibitory effect of LPS and IFN gamma on SM alpha-actin expression in stellate cells.

Actins↗
PubMed1996

Retinoic acid-stimulated liver stellate cells suppress the production of albumin from parenchymal cells via TGF-beta.

We studied a cell-cell interaction via transforming growth factor-beta (TGF-beta ) between liver stellate cells (SCs) and parenchymal cells (PCs) using co-cultures of rat primary SCs and PCs. Both TGF-beta added exogenously to the culture medium, and TGF-beta produced endogenously from SCs after stimulation with retinoic acid (RA), suppressed the production and secretion of albumin from PCs. This effect occurred at the translational level, but not at the transcriptional level; TGF-beta, as well as SC culture medium conditioned by RA, did not affect the albumin mRNA levels, but decreased the biosynthesis of [35-S]methionine-labeled albumin without altering its post-translational degradation rate. These results suggest that TGF-beta generated from SCs facilitates the development of liver cirrhosis not only by inducing the production of fibrotic components from SCs, but also by impairing the function of the surrounding PCs.

Albumins↗
PubMed1996

Inhibition of myofibroblastic transformation of cultured rat hepatic stellate cells by methylxanthines and dibutyryl cAMP.

Stellate cells isolated from rat liver and cultured on uncoated plastic plates in serum-containing medium started proliferating and transforming to myofibroblastic cells. However, stellate cells did not proliferate when cultured in the presence of 3-isobutyl-1-methylxanthine or dibutyryl cAMP (dBcAMP). These substances significantly reduced [3h] thymidine incorporation of the proliferating cells. Morphologically, stellate cells cultured in the presence of 3-isobutyl-1-methylxanthine or dibutyryl cAMP kept well-developed processes and lipid droplets while untreated cells exhibited myofibroblastic characteristics. Western blot analysis and immunocytochemical studies revealed that 3-isobutyl-1-methylxanthine and dBcAMP suppressed the expression of alpha-smooth muscle actin in stellate cells. 3-isobutyl-1-methylxanthine increased the cellular levels of cAMP from a basal value of 0.7 +/- 0.1 to 8.5 +/- 1.7 pmol/well in stellate cells. Thus, 3-isobutyl-1-methylxanthine and dBcAMP inhibit the myofibroblastic transformation of stellate cells in vitro in some cAMP-related mechanism.

Animals↗
PubMed1996

Expression of neural cell adhesion molecule (N-CAM) in perisinusoidal stellate cells of the human liver.

Neural cell adhesion molecule (N-CAM) is distributed in most nerve cells and some non-neural tissues. The present immunohistochemical study has revealed, for the first time, the expression of N-CAM in perisinusoidal stellate cells of the human liver. Liver specimens were stained with monoclonal antibody against human Leu19 (N-CAM) by a streptoavidin-biotin-peroxidase-complex method. Light- and electron-microscopic analyses have shown that N-CAM-positive nerve fibers are distributed in the periportal and intermediate zones of the liver lobule. Perisinusoidal stellate cells in these zones are also positive for N-CAM. N-CAM is expressed on the surface of the cell, including cytoplasmic projections. Close contact of N-CAM-positive nerve endings with N-CAM-positive stellate cells has been observed. On the other hand, stellate cells in the centrilobular zone exhibit weak or no reaction for N-CAM. Perivascular smooth muscle cells and fibroblasts in the portal area and myofibroblasts around the central veins are negative for N-CAM. The present results indicate that the perisinusoidal stellate cells in the periportal and intermediate zones of the liver lobule characteristically express N-CAM, unlike other related mesenchymal cells, and suggest that the intralobular heterogeneity of N-CAM expression by stellate cells is related to the different maturational stages of these cells.

Antibodies, Monoclonal↗
PubMed1996

Expression of heat-shock protein 47 in mouse liver.

Expression of heat-shock protein 47 in intact and fibrotic liver and in hepatic constituent cells was investigated in mice. Immunohistochemical study of intact liver and Western blot analysis of the protein from isolated liver cells revealed that stellate cells and smooth muscle cells of interlobular vessels, but not hepatocytes, Kupffer cells, or endothelial cells, expressed heat-shock protein 47. The protein was found in both vitamin-A-storing stellate cells and myofibroblast-like cells. The amount of the protein in cultured stellate cells was reduced by dexamethasone but was not regulated by quercetin, transforming growth factor beta, interferon gamma, or retinoic acid. In CCl4-treated or bile-duct-ligated mouse liver, the number of cells positive for heat-shock protein 47 markedly increased in the centrilobular area or around the periportal area, respectively, and the level of heat-shock protein 47 also increased.

Animals↗
PubMed1996

Morphological study of endothelin-1-induced contraction of cultured hepatic stellate cells on hydrated collagen gels.

Hepatic stellate cells become activated and aquire contractility on being cultured. In order to characterize the morphology of contracted and relaxed stellate cells, we performed light- and electron-microscopic analyses of cultured stellate cells on collagen gels. Incubation of stellate cells with medium alone, 10 nM endothelin (ET)-1, or 1 mM N6,2' dibutyryladenosine 3':5'-cyclic monophosphate (dBcAMP) for 48 h induced contraction of the underlying collagen gels to 83%, 57%, and 97%, respectively, of their original size. Stellate cells relaxed by dBcAMP exhibited a round cell body and extended several long thin cytoplasmic processes with several varicosities. Culture with ET-1 accelerated spreading of the stellate cells on collagen gels and decreased the number of processes. Each such flattened stellate cell attached itself to the underlying collagen matrix by bending its cell body. Collagen fibers around the cell were pulled toward the cell and stretched. Thus, the present study has revealed that ET-1-stimulated cultured stellate cells adduct associated collagen fibers by the retraction of cytoplasmic processes and the bending of their spread cell bodies.

Animals↗
PubMed1996

Interleukin 1 beta markedly stimulates nitric oxide formation in the absence of other cytokines or lipopolysaccharide in primary cultured rat hepatocytes but not in Kupffer cells.

To investigate whether a single inflammatory cytokine could stimulate nitric oxide formation in the absence of other cytokines or lipopolysaccharide (LPS), NO was measured by the redox chemiluminescence method in primary cultured rat hepatocytes and in rat Kupffer cells. Interleukin (IL) 1 beta, but neither IL-6, tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), nor LPS stimulated NO formation in a dose-dependent manner and induced half-maximal effects at 30 pmol/L. Maximal stimulation was achieved at 12 to 16 hours after the addition of 1I nmol/L of IL-1 beta, and was 50- to 60-fold above basal levels in rat hepatocytes. The combined effect of these cytokines with LPS or IFN-gamma on NO formation was also examined. Neither LPS nor IFN-gamma affected the IL-1 beta-induced NO formation. TNF-alpha, however, stimulated IL-1 beta-induced NO formation, while IL-6 inhibited it, although independently these cytokines had no effect on NO formation. None of the cytokines tested stimulated NO formation in cultured rat Kupffer cells. In hepatocytes, the NO formation induced by IL-l beta was blocked by both the NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA) and by IL-1 receptor antagonist (IL-1ra). Furthermore, IL-1 beta markedly increased NOS activity, and this increase in activity was accompanied by the expression of inducible NOS (iNOS) messenger RNA (mRNA). This study clearly demonstrated that IL-1 beta markedly stimulates NO formation in hepatocytes, in the absence of other cytokines or LPS.

Animals↗
PubMed1996

Mechanism that regulates nitric oxide production by lipopolysaccharide-stimulated rat Kupffer cells.

Mechanism that regulates nitric oxide (NO) production by stimulated Kupffer cells was studied. Lipopolysaccharide (LPS) stimulated NO production by primary-cultured Kupffer cells in a time- and dose-dependent manner. Twenty-four h after incubation with 1 microgram/ml of LPS, the nitrite concentration in the culture medium increased from 2.87 +/- 1.4 to 73.6 +/- 6.9 nmol/ml. NO production started to increase around 6 h after adding LPS and reached a maximum within 24 h. NO production was inhibited by 0.3 mM of NG-monomethyl-L-arginine and was reversed by adding 3 mM of L-arginine. When incubated with 1 microgram/ml of LPS for 24 h, NO synthase activity in Kupffer cells increased from 0.164 +/- 0.035 to 3.16 +/- 0.11 nmol/min/mg protein. Dexamethasone and prednisolone significantly inhibited the induction of NO synthase and NO production in Kupffer cells. Expression of mRNA for inducible type of NO synthase (iNOS) started to increase around 4 h after adding LPS (1 microgram/ml) and reached a maximum by about 20 h. The enhanced expression of iNOS mRNA was also suppressed by 1 microM dexamethasone. On the other hand, calcium ionophore A23187 significantly enhanced the induction of NO synthase and iNOS mRNA expression in LPS-stimulated Kupffer cells. In addition, a tyrosine kinase inhibitor, genistein (100 microM), significantly inhibited NO production by LPS-stimulated Kupffer cells. Neither phorbol 12-myristate 13-acetate, dibutyryl cAMP, indomethacin nor a protein kinase C inhibitor H-7 affected NO production by Kupffer cells. These results suggested that iNOS mRNA expression, NO synthase activity and NO production increased in LPS-stimulated Kupffer cells by a glucocorticoid-inhibitable mechanism and that Ca2+ and tyrosine phosphorylation might play important roles in LPS-dependent induction of NO synthase.

Animals↗
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

Action of endothelins on hepatic stellate cells.

To elucidate the role played by hepatic sinusoidal cells in the regulation of the circulatory status in the liver, the effect of endothelins (ETs) on primary-cultured stellate cells was examined. Kinetic analysis with 125I-labeled ET-1 revealed that stellate cells have ET receptors with a Kd value of 141 pM and a Bmax of 12.3 fmol/10(5) cells. ET-1, -2, and -3 dose-dependently increased inositol monophosphate (InsP) levels in stellate cells with an EC50 of 0.53, 1.63, and 1.88nM, respectively. Binding of 125I-labeled ET-1 to stellate cells and the ET-enhanced InsP formation were suppressed by preincubating the cells with 10 nM of unlabeled ET-1 or ET-3 for more than 3 h, indicating down-regulation and desensitization of ET receptors by homologous ligands. Binding of ETs to surface receptors induced a marked contraction of stellate cells. Stellate cells rapidly reacted to ETs, as detected by the flexible silicone-rubber-membrane method; 78%, 73%, and 58% of the stellate cells contracted 2.5 min after the addition of 10 nM of ET-1, ET-2, or ET-3, respectively. On the other hand, ETs also triggered a long-lasting contraction of the cells, as revealed with hydrated collagen gels. The ET-induced contraction of stellate cells decreased the diameter of the collagen lattice by about 60%, and this action was inhibited either by cytochalasin B or by H-7, a protein kinase C inhibitor. These and other results suggest that ETs induced cell contraction by some mechanism that involved protein kinase C.

Animals↗
PubMed1995

Characterization of arachidonate 12-lipoxygenase found in the liver of mongrel dog and its immunohistochemical localization in neutrophils.

The cytosol fraction of non-parenchymal cells isolated from the liver of adult mongrel dogs converted arachidonic acid to 12S-hydroxy-5,8,10,14-eicosatetraenoic acid. The arachidonate 12-lipoxygenase enzyme reacted with linoleic and alpha- and gamma-linolenic acids as well as arachidonic acid. The enzyme was immunoprecipitable with an antibody against the leukocyte 12-lipoxygenase, but not with an antibody against the platelet 12-lipoxygenase. By immunohistochemical observation with anti-leukocyte 12-lipoxygenase antiserum, hepatocytes, Kupffer cells and sinusoidal endothelial cells were not immunostained, but a large number of neutrophils located in sinusoidal cavities were positively stained. The 12-lipoxygenase activity thus detected may be attributed predominantly to the neutrophils appearing in sinusoidal cavities rather than the non-parenchymal cells in the liver of mongrel dog upon infection. In agreement with this finding there was essentially no neutrophil accumulation in the liver of Beagle dog which had no bacterial and parasitic infection.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗