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Biomedical subjects

Yutaka Kohgo

Publications and source records attributed to Yutaka Kohgo.

At least 19 recordsLinked to original sources
PubMed2004

Direct evidence that induced nitric oxide production in hepatocytes prevents liver damage during lipopolysaccharide tolerance in rats.

BACKGROUND: The role of nitric oxide (NO) in lipopolysaccharide (LPS) tolerance in the liver has been investigated in a number of previous studies, but it is still not clear whether NO is cytotoxic or cytoprotective. The aims of this study were to investigate whether low-dose LPS (LLPS)-induced hepatic production of NO is beneficial and to clarify the origins of cytoprotective NO-producing cells in the liver during LPS tolerance. MATERIALS AND METHODS: Male Wistar rats received saline or LLPS intraperitoneally (i.p.; 0.01-1000 microg/kg) followed by a high dose of LPS (HLPS, 5 mg/kg) at various time intervals (4-16 h). NG-nitro-L-arginine methyl ester (L-NAME) was used to investigate the effects of inhibition of NOS. 4,5-Diaminofluorescein (DAF-2) was used to identify NO-producing cells in isolated liver cells in vitro. At various time points (4-16 h) after saline or LLPS (1 microg/kg, i.p.) injection, hepatocytes and Kupffer cells were isolated, incubated in 7 microm DAF-2 diacetate, and perfused with Krebs solution. Illumination at 495 nm revealed DAF-fluorescence (515 nm) in isolated cells under confocal laser fluorescence microscopy. The NO production in hepatocytes and Kupffer cells was assessed by the number of labeled cells per 1000 cells or per 100 cells, respectively. RESULTS: Pretreatment with LLPS (0.1-100 microg/kg) resulted in a significant reduction (maximal at 8 h) of the HLPS-induced liver damage. L-NAME abolished the LLPS-induced protection. The NO production in hepatocytes was significantly increased and reached a maximum of 84% of all cells 8 h after LLPS administration. By contrast, the NO production in Kupffer cells remained constant at 95%, even following preinjection of LLPS. CONCLUSION: LLPS-induced NO in hepatocytes, but not in Kupffer cells, exhibits cytoprotective effects on HLPS-induced liver damage, suggesting that NO has a beneficial role in the induction of the early phase of LPS tolerance.

Animals↗
PubMed2004

Serine/threonine kinase AKT is frequently activated in human bile duct cancer and is associated with increased radioresistance.

The prognosis for patients with bile duct cancer (BDC) remains poor. Although BDC cells are essentially radioresistant, recent reports have suggested that radiation therapy, in addition to its palliative role in the management of BDC, may improve patient survival. A better understanding of the mechanisms that lead to cellular radioresistance may assist in the development of more effective BDC therapies based on radiotherapy in combination with radiosensitizing agents. The serine/threonine kinase AKT/protein kinase B, a downstream effector of phosphatidylinositol 3'-kinase, is a well-characterized kinase that is known to play a critical role in antiapoptotic signaling pathways. In this investigation, we sought to clarify the role of AKT signaling in the radioresistance in BDC cells. First, to examine whether activated AKT is expressed in BDCs, tumor specimens were obtained from 19 consecutive BDC cases. Immunohistochemical staining using an anti-phosphorylated-AKT antibody showed that phosphorylated (activated) AKT was expressed in cancer cells but not in neighboring normal mucosa in 16 cases (84.2%). Next, to evaluate the role of AKT activation in the regulation of BDC cell radiosensitivity, clonogenic assays were performed using the phosphatidylinositol 3'-kinase inhibitor LY294002 with and without irradiation. LY294002 inhibited AKT activation in BDC cells and, on irradiation, decreased clonogenic survival in a radiation dose-dependent manner. Only a small decrease in cell viability was observed in cells exposed to LY294002. Expression of constitutively active AKT in BDC cells resulted in decreased radiosensitivity, whereas a dominant-negative AKT increased radiosensitivity. Furthermore, constitutively active AKT also inhibited radiation-induced apoptosis. Collectively, these results indicate that activated AKT in BDC cells is associated with radioresistance and suggest that pharmacological or genetic modulation of AKT activity may have important therapeutic implications in BDC patients treated with radiation.

Bile Duct Neoplasms↗
PubMed2004

Involvement of p38 mitogen-activated protein kinase in gemcitabine-induced apoptosis in human pancreatic cancer cells.

In this study, we investigated the involvement of Akt and members of the mitogen-activated protein kinase (MAPK) superfamily, including ERK, JNK, and p38 MAPK, in gemcitabine-induced cytotoxicity in human pancreatic cancer cells. We found that gemcitabine induces apoptosis in PK-1 and PCI-43 human pancreatic cancer cell lines. Gemcitabine specifically activated p38 MAPK in a dose- and time-dependent manner. A selective p38 MAPK inhibitor, SB203580, significantly inhibited gemcitabine-induced apoptosis in both cell lines, suggesting that phosphorylation of p38 MAPK may play a key role in gemcitabine-induced apoptosis in pancreatic cancer cells. A selective JNK inhibitor, SP600125, failed to inhibit gemcitabine-induced apoptosis in both cell lines. MKK3/6, an upstream activator of p38 MAPK, was phosphorylated by gemcitabine, indicating that the MKK3/6-p38 MAPK signaling pathway is indeed involved in gemcitabine-induced apoptosis. Furthermore, gemcitabine-induced cleavage of the caspase substrate poly(ADP-ribose) polymerase was inhibited by pretreatment with SB203580, suggesting that activation of p38 MAPK by gemcitabine induces apoptosis through caspase signaling. These results together suggest that gemcitabine-induced apoptosis in human pancreatic cancer cells is mediated by the MKK3/6-p38 MAPK-caspase signaling pathway. Further, these results lead us to suggest that p38 MAPK should be investigated as a novel molecular target for human pancreatic cancer therapies.

Antimetabolites, Antineoplastic↗
PubMed2004

Growth arrest by troglitazone is mediated by p27Kip1 accumulation, which results from dual inhibition of proteasome activity and Skp2 expression in human hepatocellular carcinoma cells.

In our study, we examined whether human hepatocellular carcinoma (HCC) expresses peroxisome proliferator-activated receptor gamma (PPARgamma) and the effects of PPAR gamma activation by its selective ligands on cell growth and cell invasion in HCC cells. RT-PCR and Western blot analysis revealed that HCC-derived cell lines, HepG2 and HLF, express PPARgamma mRNA and protein. Luciferase assay in HLF cells showed that troglitazone, a selective ligand for PPAR gamma, transactivated the transcription of a peroxisome proliferator response element-driven promoter in a dose-dependent manner, suggesting that the expressed PPARgamma functions as a transcriptional factor. Not only troglitazone but pioglitazone dose-dependently inhibited cell growth in HepG2 and HLF cells. Invasion assay using a transwell chamber demonstrated that troglitazone also inhibited cell invasion in HCC cells. To examine the mechanism of the troglitazone-induced growth inhibition, we determined p27(Kip1), a cyclin dependent kinase inhibitor, expression by Western blot analysis in troglitazone-treated HLF cells. Troglitazone increased p27(Kip1) in time- and dose-dependent manners, suggesting that p27(Kip1) may be involved in the growth inhibition by troglitazone in HLF cells. To further examine the mechanism of the troglitazone-induced p27(Kip1) protein accumulation, 2 major systems for regulation of p27(Kip1) protein, proteasome activity and Skp2, an F-box protein that targets p27(Kip1) for degradation, were evaluated. Troglitazone potently inhibited proteasome activity and decreased Skp2 protein levels. All these results suggest that human HCC cells express functional PPAR gamma and PPARgamma activation resulted in growth inhibition. The growth inhibition was mediated by p27(Kip1) accumulation, which is induced by both inhibition of ubiquitylation of p27(Kip1) and reduction of degradation activity of p27Kip1 by proteasome.

Adenosine Triphosphatases↗
PubMed2004

Reduction of syndecan-1 expression in differentiated type early gastric cancer and background mucosa with gastric cellular phenotype.

BACKGROUND: Syndecan-1 is known to play a role as a cell adhesion molecule, similar to E-cadherin, and is associated with the maintenance of epithelial morphology. The purpose of this study was to elucidate the role and alterations of syndecan-1 expression in comparison with those of E-cadherin in different cellular phenotypes of differentiated-type gastric cancers (DGCs). METHODS: A total of 80 DGCs at an early stage, and their adjacent mucosa, were evaluated by both immunohistochemistry and in situ hybridization. Syndecan-1 and E-cadherin were assessed by immunohistochemical staining with an anti-syndecan-1 and an anti-E-cadherin antibody, respectively. Based on immunohistochemistry, DGCs and their surrounding mucosa were divided into four types: gastric type (G-type), ordinary type (O-type), complete-intestinal type (CI-type), and null type. RESULTS: The expression sites of syndecan-1 mRNA mostly coincided with those of syndecan-1 protein. Syndecan-1 expression was significantly lower in G-type cancers (30%) than in O- (81%) and CI-type cancers (92%) ( P = 0.0001 and P = 0.004, respectively), but E-cadherin did not show this result. In addition, syndecan-1 expression was significantly reduced in DGCs comprised partly of poorly differentiated adenocarcinoma or signet-ring cell carcinoma, compared to DGCs demonstrating papillary and/or tubular adenocarcinoma ( P = 0.02). G-type intestinal metaplasia (IM) surrounding the tumors was observed in 21% of G-type cancers, in 0% of O-, and in 10% of CI-type cancers ( P = 0.01; G-type vs O-type). Reduction of syndecan-1 expression was significant in G-type IM (25%) compared to non-G-type IM (75%; P = 0.02). CONCLUSIONS: Syndecan-1 plays a role in the growth of G-type cancers at an early stage compared with E-cadherin changes, and the reduction of syndecan-1 expression in IM surrounding the tumors may influence the growth of G-type cancer.

Adenocarcinoma↗
PubMed2004

Inhibition of cell invasion and morphological change by troglitazone in human pancreatic cancer cells.

BACKGROUND: We have recently demonstrated that peroxisome proliferator activated receptor (PPAR) gamma activation by its selective ligand, troglitazone, potently inhibited cell proliferation in human pancreatic cancer cells. The present study was performed to clarify the role of PPARgamma in cell invasion/motility in human pancreatic cancer cells. METHODS: Cell invasive activity was assessed by an in vitro invasion assay, using a Transwell chamber, and by a wound-healing assay, in the human pancreatic cancer cell lines, PK-1 and PK-9. Cell morphology and actin structure were evaluated by phase-contrast and fluorescence microscopy. RESULTS: PPARgamma activation by troglitazone inhibited cell invasion and cell migration in PK-1 and PK-9 cells. We also examined the effect of troglitazone on cell morphology and actin structure because of its effect on cell motility. The size of PK-1 and PK-9 cells that had been incubated with troglitazone became smaller, and the in shape changed from flat to spindle, followed by round. The troglitazone-induced cell rounding was reversible by replacement with troglitazone-free medium. Rhodamine-phalloidin staining revealed a decreased number of actin filaments in PK-1 cells treated with troglitazone. In cells treated with mycalolide B, an actin depolymerizing agent, troglitazone failed to induce cell rounding. CONCLUSIONS: These results suggest that PPARgamma activation by troglitazone inhibited cell motility and changed cell morphology through modulating actin organization.

Actins↗
PubMed2004

A significant reduction in serum alanine aminotransferase levels after 3-month iron reduction therapy for chronic hepatitis C: a multicenter, prospective, randomized, controlled trial in Japan.

BACKGROUND: Increasing evidence indicates that iron cytotoxicity plays an important role in the pathogenesis of chronic hepatitis C (CHC). However, the biochemical effects of iron reduction therapy on CHC remain to be confirmed in a controlled study. This study aimed to test whether iron removal by repeated phlebotomy improves serum alanine aminotransferase (ALT) levels in patients with CHC. METHODS: Patients were randomly assigned to an iron reduction therapy or control group. The patients in the treatment group received 3-month iron reduction therapy by biweekly phlebotomy, while the patients in the control group were followed up for 3 months with regular blood tests alone. RESULTS: Thirty-three patients completed the 3-month treatment, while 29 patients received the complete follow-up. The serum ALT levels were reduced from 118 +/- 79 to 73 +/- 39 IU/L in the treatment group, but did not change in the control group (106 +/- 45 versus 107 +/- 48 IU/L). Posttreatment enzyme activity was decreased significantly from the baseline. Furthermore, it was significantly lower than the 3-month control level. Although 5 patients withdrew from the study, none was affected by any side effects of repeated phlebotomy that required them to discontinue the treatment. CONCLUSIONS: This short-term controlled trial demonstrated the biochemical efficacy and safety of iron reduction therapy for patients with CHC.

Adult↗
PubMed2004

Impaired nitric oxide production of the myenteric plexus in colitis detected by a new bioimaging system.

Direct measurement of the release of nitric oxide (NO) from the myenteric plexus has been extremely difficult to date, due to the lack of suitable methodologies. We have developed a new bioimaging system to visualize the nitrergic neurons of the myenteric plexus and investigated whether NO production is impaired in dextran sulfate sodium (DSS)-induced colitis. Longitudinal muscle layers with the myenteric plexus intact were obtained from the rat colon and were incubated with 4,5-diaminofluorescein-2-diacetate (DAF-2DA) (7 microm) for 30 min. Illumination at 450-490 nm revealed the fluorescence in the myenteric plexus. Confocal laser microscopy and three-dimensional reconstruction techniques were used to quantify the changes in the amount of NO production by the myenteric plexus. Fluorescent double-labeled immunostaining for nNOS was performed to confirm the colocalization of nNOS in 4,5-diaminofluorescein (DAF-2)-positive cells. DAF-2 fluorescence was abolished by pretreatment with N(G)-nitro-l-arginine methyl ester (l-NAME; a nonselective NOS inhibitor), 1-(2-trifluoromethylphenyl) imidazole (TRIM; a selective neuronal NOS inhibitor), and omega-conotoxin GVIA (an N-type Ca(2+) channel blocker), but not by nifedipine (an l-type Ca(2+) channel blocker). Fluorescent double-labeled immunostaining showed that DAF-2-positive cells colocalized with nNOS-positive cells. Oral administration of 5% DSS for 7 days induced distal colitis and the number of DAF-2-positive neurons were significantly reduced to 55 +/- 17% of control. DAF-2 offers a sensitive indicator for visualizing production of NO with high spatial resolution. This new system may contribute to the study of the pathophysiological role of the nitrergic pathway in the gastrointestinal tract.

Animals↗
PubMed2004

The role of Paneth cells and their antimicrobial peptides in innate host defense.

The intestinal epithelium is the largest surface area that is exposed to various pathogens in the environment, however, in contrast to the colon the number of bacteria that colonize the small intestine is extremely low. Paneth cells, one of four major epithelial cell lineages in the small intestine, reside at the base of the crypts and have apically oriented secretory granules. These granules contain high levels of antimicrobial peptides that belong to the alpha-defensin family. Paneth cells secrete these microbicidal granules that contain alpha-defensins when exposed ex vivo to bacteria or their antigens, and recent evidence reveals that antimicrobial peptides, particularly alpha-defensins, that are present in Paneth cells contribute to intestinal innate host defense.

Animals↗
PubMed2004

Induction of transferrin receptor by ethanol in rat primary hepatocyte culture.

BACKGROUND: It is not uncommon for alcoholics to have iron accumulation in the liver, a condition that may contribute to the development of alcoholic liver disease. Recently, we reported that the expression of transferrin receptor, which mediates cellular iron uptake, was increased in hepatocytes in patients with alcoholic liver disease. To elucidate the mechanism of the iron accumulation in hepatocytes in such disease, we examined whether ethanol exposure induced the transferrin receptor expression and increased the cellular iron uptake. METHODS: Rat primary hepatocytes were isolated and cultured in the presence of 20 micromol/liter of iron and 25 mmol/liter of ethanol. RESULTS: Ethanol exposure to the hepatocytes demonstrated an ~2-fold increase in transferrin receptor expression for 24 hr, shown by Western blot analysis and S-methionine metabolic labeling, 19% increase in Fe-transferrin uptake by hepatocytes, and 20% increase in activity of iron regulatory protein examined by band shift assay. CONCLUSION: Ethanol exposure induced the transferrin receptor expression, partially through the activation of iron regulatory protein, and increased the transferrin-bound iron uptake in rat hepatocyte cultures. The induction of transferrin receptor by ethanol might be one of the mechanisms of iron accumulation in the hepatocytes in alcoholic liver disease.

Animals↗
PubMed2003

Usefulness of contrast-enhanced wide-band Doppler ultrasonograpy to diagnose alveolar echinococcosis of the liver and evaluate the effect of the treatment.

Alveolar echinococcosis is a rare parasitic disease caused by Echinococcus multicularis and most commonly involves the liver. Early diagnosis and accurate evaluation of the effect of the treatment are essential to improve the prognosis of patients with alveolar echinococcosis of the liver (AEL). The aim of this study was to demonstrate the characteristic imaging of AEL by contrast-enhanced Dynamic Flow (CE-DF) employing a wide-band Doppler technique for the diagnosis and the accurate evaluation of the effect of the treatment. Four patients with five AEL lesions before treatment or during medication were examined by CE-DF. All of the five AEL lesions examined by CE-DF revealed a defect with an irregular and distinct margin like a worm-eaten defect appearance, which was never observed on other hepatic lesions, in liver perfusion image during post-vascular phase. In addition, CE-DF made it possible to measure the size of AEL lesions accurately because the margin was detected distinctly. These data suggest that CE-DF is clinically useful for the diagnosis of AEL and the evaluation of the effect of the treatment.

Aged↗
PubMed2003

Clinical utility of sequential imaging of hepatocellular carcinoma by contrast-enhanced power Doppler ultrasonograpy.

The aim of this study was to investigate the clinical utility of sequential imaging of hepatocellular carcinoma (HCC) by contrast-enhanced power Doppler ultrasonograpy (CE-PDUS) to differentiate hepatocellular carcinoma from adenomatous hyperplasia (AH) and regenerated nodule (RN) and to predict the degree of differentiation of HCC. Fifty-one patients with 62 hepatic lesions including 33 moderately and poorly differentiated HCCs, 19 well-differentiated HCCs, seven AHs and three large RNs were examined by CE-PDUS. The imaging patterns during early arterial phase (tumor vessel image), late vascular phase (tumor perfusion image) and post-vascular phase (liver perfusion image) were classified as diffuse, basket, peripheral, central and no enhancement; as whole tumor, partial tumor and no enhancement; as whole tumor, partial tumor and no defect, respectively. The diffuse pattern in the tumor vessel image, the whole enhancement pattern in the tumor perfusion image and the whole defect pattern in the liver perfusion image were observed in moderately and poorly differentiated HCCs only. The basket pattern in the tumor vessel image and the partial defect pattern in the tumor perfusion image were observed in HCCs only. All AH/RNs showed no defect pattern in the liver perfusion image. The sequential imaging of HCC during early arterial, late vascular and post-vascular phases by CE-PDUS is clinically useful to differentiate HCC from AH/RN and to predict the degree of differentiation of HCC.

Adult↗
PubMed2003

Cell kinetics and genetic instabilities in differentiated type early gastric cancers with different mucin phenotype.

To clarify the biological impact and molecular pathogenesis of cellular phenotype in differentiated-type gastric cancers (DGCs), we investigated cell kinetics and genetic instabilities in early stage of DGCs. A total of 43 early gastric cancers (EGCs) were studied. EGCs were divided into 3 phenotypic categories: gastric (G type, n = 11), ordinary (O type, n = 20), and complete intestinal (CI type, n = 12) based on the combination of HGM, ConA, MUC2, and CD10. Proliferative index (PI), apoptotic index (AI), and p53 overexpression were investigated by immunohistochemical staining with anti-Ki-67, the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling method, and p53 antibody, respectively. Using a high-resolution fluorescent microsatellite analysis system, microsatellite instability (MSI) and loss of heterozygosity (LOH) were examined. Frameshift mutation analysis of transforming growth factor-beta type II receptor (TGF-betaRII) and bcl-2-associated X (BAX) in cancers with MSI was also performed. The mean AI/PI ratio values were 0.04 for G-type, 0.10 for O-type, and 0.13 for CI-type cancers--significantly lower in G type than in O and CI types (P = 0.02 and P = 0.001, respectively). No difference in the incidence of MSI and LOH was seen among the 3 cellular phenotypes. However, the major pattern of MSI, which showed drastic and widely dispersed changes and is related to an increased risk for cancer, was significantly higher in G and O types than in CI type (P <0.005). No frame shift mutations of TGF-betaRII or BAX were found in CI-type cancers. These results indicate that G-type cancers are likely to show more aggressive behaviors than CI-type cancers, and that O-type cancers show the intermediate characteristics of both types. However, the molecular pathogenesis of each phenotypic cancer is not associated with microsatellite alterations.

Adenocarcinoma↗
PubMed2003

Ethanol induces transforming growth factor-alpha expression in hepatocytes, leading to stimulation of collagen synthesis by hepatic stellate cells.

BACKGROUND: Liver fibrosis often develops in alcoholic liver diseases without accompanying inflammation; however, the underlying mechanism is unclear. Using ethanol-exposed human HepG2 hepatoblastoma cells as a model for alcoholic liver diseases, we previously found that ethanol exposure causes HepG2 cells to secrete an approximately 6,000 Da nonheparin-binding polypeptide that stimulates collagen synthesis in human IMR-90 fibroblasts. The aim of the current study was to characterize and identify this factor. METHODS: Concentration of type I procollagen peptide and transforming growth factor (TGF)-alpha was assessed by enzyme-linked immunosorbent assay. TGF-alpha protein expression was examined by Western blot. Type I collagen messenger RNA expression in rat hepatic stellate cells was assessed by reverse transcription-polymerase chain reaction. RESULTS: The collagen-stimulating activity in conditioned media from ethanol-exposed HepG2 cells to stimulate type I procollagen peptide synthesis of IMR-90 cells was specifically inhibited by addition of anti-TGF-alpha antibodies. Western blot analysis showed increased TGF-alpha protein expression in ethanol-treated HepG2 cells. TGF-alpha in conditioned medium from ethanol-exposed HepG2 cells stimulated type-I collagen messenger RNA expression in rat hepatic stellate cells. CONCLUSIONS: These results suggest that TGF-alpha derived from ethanol-exposed hepatocytes may contribute to the development of hepatic fibrosis in alcoholic liver diseases.

Blotting, Western↗
PubMed2003

PPAR gamma ligand-induced apoptosis through a p53-dependent mechanism in human gastric cancer cells.

We have recently demonstrated that the PPAR gamma ligand troglitazone induced cell growth arrest and evoked apoptosis in a gastric cancer cell line, MKN-45. Since in general, p53 plays an important role in the induction of apoptosis and growth inhibition, we tried to clarify whether or not p53 mediates troglitazone-induced apoptosis and growth arrest in gastric cancer cells. Troglitazone increased the number of apoptoic cells in MKN-28, MKN-45 and MKN-74, but not in KATO-III cells. The troglitazone-induced apoptotic change was significantly reduced by coincubation with bisphenol A digycidyl ether (BADGE), a synthetic PPAR gamma antagonist, in MKN-74 cells, suggesting that PPAR gamma mediates the apoptotic effect of troglitazone. Since KATO-III lacks the p53 gene, we speculated that p53 might be implicated in the PPAR gamma ligand-induced apoptosis. Western blot analysis revealed that p53 expression was increased by troglitazone in a time-dependent manner in MKN-74 cells, further suggesting that p53 may mediate the apoptotic process induced by troglitazone. We next established a dominant-negative p53 mutant by stable transfection of p53 mutant into MKN-74 cells. In the dominant-negative p53 mutant cells, troglitazone failed to induce apoptosis, strongly supporting the hypothesis that p53 indeed mediates the process of the troglitazone-induced apoptosis. In the dominant-negative p53 mutant cells, troglitazone significantly induced cell growth arrest and increased expression of p27(Kip1) protein, which is thought to be the key molecule to evoke growth arrest, suggesting that p53 is not involved in the growth inhibition by troglitazone. All these results suggest that p53 mediates the PPAR gamma ligand-induced apoptosis, but not the cell growth inhibition.

Antineoplastic Agents↗