Biomedical subjects

Katsuhiko Enomoto

Publications and source records attributed to Katsuhiko Enomoto.

17 recordsLinked to original sources

PubMed2006

Hyperplasia and carcinomas in Pten-deficient mice and reduced PTEN protein in human bladder cancer patients.

PTEN is a tumor suppressor gene mutated in many human cancers. We used the Cre-loxP system to generate an urothelium-specific null mutation of Pten in mice [FabpCrePten(flox/flox) (FPten(flox/flox)) mice]. Histologic examination revealed that all FPten(flox/flox) mice exhibited urothelial hyperplasia in which component cells showed enlarged nuclei and increased cell size. With time, 10% of FPten(flox/flox) mice spontaneously developed pedicellate papillary transitional cell carcinomas (TCC). This type of tumor also arose in FPten(flox/flox) mice treated with the chemical carcinogen N-butyl-N-(4-hydroxybutyl) nitrosamine. FPten(flox/flox) urothelial cells were hyperproliferative and showed increased activation of the survival signaling molecules Akt and extracellular signal-regulated kinase. In humans, 53% of primary bladder cancer patients exhibited decreased or absent expression of PTEN protein in either the cytoplasm or nucleus of tumor cells. In early bladder cancers, PTEN expression was repressed in 42% of superficial papillary TCC but in only 8% of cases of carcinoma in situ (CIS). In advanced bladder cancers, PTEN protein was significantly reduced (particularly in the nucleus) in 94% of cases, and this decrease in PTEN correlated with disease stage and grade. Thus, PTEN deficiency may contribute to bladder cancer both by initiating superficial papillary TCC and by promoting the progression of CIS to advanced invasive and metastatic forms.

Animals↗

PubMed2006

Maintenance of Bad phosphorylation prevents apoptosis of rat hepatic sinusoidal endothelial cells in vitro and in vivo.

To elucidate the mechanism of apoptosis of liver sinusoidal endothelial cells (SECs), we examined the phosphorylation status of Bad and its upstream signaling molecules during apoptosis in culture and after ischemia-reperfusion injury. Rat SECs were isolated by the immunomagnetic method, and 2 days after culture, most SECs underwent apoptosis, which was associated with decreased tyrosine phosphorylation of cellular proteins. Addition of orthovanadate (OV), a protein tyrosine phosphatase inhibitor, sustained cellular protein phosphorylation and strongly inhibited apoptosis. Bad was dephosphorylated at Ser-112 and Ser-136 during apoptosis, but the phosphorylation status of Bad was maintained in the presence of OV. OV activated the Akt, extracellular signal-regulated protein kinase, and p38 mitogen-activated protein kinase pathways, which are involved in Bad phosphorylation. In the absence of OV, depletion of Bad by RNA interference conferred resistance to apoptosis. Hepatic injury after ischemia-reperfusion was alleviated by OV treatment, with significant inhibition of SEC apoptosis. SEC apoptosis in vivo was associated with dephosphorylation of Bad, Akt, and extracellular signal-regulated protein kinase, which was blocked by OV treatment. Our data suggest that maintenance of Bad phosphorylation is important in the prevention of SEC apoptosis and that the anti-apoptotic property of OV might have therapeutic utility.

14-3-3 Proteins↗

PubMed2005

Campylobacter jejuni isolated from retail poultry meat, bovine feces and bile, and human diarrheal samples in Japan: comparison of serotypes and genotypes.

To determine the significance of poultry and bovine as infectious sources of Campylobacter jejuni in Japan, the serotype distribution and pulsed-field gel electrophoresis (PFGE) patterns of poultry and bovine isolates were compared with those of isolates from patients with diarrhea in Akita (Japan). Serotypes O:2 and O:4-complex were common in human, poultry, and bovine isolates, and serotype O:23,36,53 was common in human and bovine isolates. SmaI PFGE patterns of isolates belonging to these serotypes were generated. Eight PFGE patterns were shared by poultry and human isolates and three patterns were shared by human and bovine isolates. Further analysis of the isolates having the same SmaI PFGE pattern by KpnI PFGE confirmed that four patterns and two patterns were still shared by poultry and human isolates, and bovine and human isolates, respectively. Thus, serotypic and genotypic data indicated a possible link between sporadic human campylobacteriosis and C. jejuni from retail poultry and bovine bile and feces, suggesting that bovine serves as an infectious source of C. jejuni in Japan, as is observed in other countries.

Animals↗

PubMed2005

Transdifferentiation of mature rat hepatocytes into bile duct-like cells in vitro.

We investigated the mechanism of phenotypic plasticity of hepatocytes in a three-dimensional organoid culture system, in which hepatocytic spheroids were embedded within a collagen gel matrix. Hepatocytes expressed several bile duct markers including cytokeratin (CK) 19 soon after culture and underwent branching morphogenesis within the matrix in the presence of insulin and epidermal growth factor. Cultured hepatocytes did not express Delta-like, a specific marker for oval cells and hepatoblasts. Furthermore, hepatocytes isolated from c-kit mutant rats (Ws/Ws), which are defective in proliferation of oval cells, showed essentially the same phenotypic changes as those isolated from control rats. The bile duct-like differentiation of hepatocytes was associated with increased expression of Jagged1, Jagged2, Notch1, and several Notch target genes. CK19 expression and branching morphogenesis were inhibited by dexamethasone, a mitogen-activated protein kinase kinase 1 (MEK1) inhibitor (PD98059), and a phosphatidyl inositol 3-kinase inhibitor (LY294002). After being cultured for more than 3 weeks within the gels, hepatocytes transformed into ductular structures surrounded by basement membranes. Our results suggest that hepatocytes might have the potential to transdifferentiate into bile duct-like cells without acquiring a stem-like phenotype and that this is mediated through specific protein tyrosine phosphorylation pathways.

Animals↗

PubMed2004

Restoration of copper metabolism and rescue of hepatic abnormalities in LEC rats, an animal model of Wilson disease, by expression of human ATP7B gene.

Hepatic abnormalities in Long-Evans Cinnamon (LEC) rats, an animal model of Wilson disease (WD), were restored by the expression of the human ATP7B cDNA under the control of CAG promoter. Expression of ATP7B transcript and protein in the liver of the transgenic rats resulted in the restoration of biosynthesis of holoceruloplasmin and biliary copper excretion. Meanwhile, transgenic rats showed striking improvements in their hepatic abnormalities, i.e., rescue from fulminant hepatitis, late onset of hepatic cholangiofibrosis, suppression of hepatocellular carcinoma and much improved survival rates. Moreover, dramatic decreases were noted both in the levels of hepatic copper and iron in transgenic rats before the occurrence of hepatitis. These results indicated that the human ATP7B product compensated for the deficiency of the endogenous rattus protein and did function in intrahepatic copper transport by secreting copper into the plasma via incorporation into ceruloplasmin and by the excretion of copper into the bile, and that ATP7B is critical to hepatic dysfunctions in WD. This first successful transgenic rescue has important implications for the gene therapy of WD.

Adenosine Triphosphatases↗

PubMed2004

Cell biology and pathology of liver sinusoidal endothelial cells.

Growing evidence revealed that liver sinusoidal endothelial cells (SEC) play several important roles in physiology and pathology of the liver. It has been well understood that their structural characteristics, such as the membrane sieve and lack of basement membrane, facilitate direct contact of soluble and insoluble serum substances with hepatic parenchymal cells, resulting in enhancement of hepatic metabolic activity. In addition, SEC is now regarded as a member of the scavenger endothelial cells, which have potential to eliminate a variety of macromolecules from the blood circulation by receptor-mediated endocytosis. It is reported that molecules preferentially eliminated by SEC are denatured or modified proteins such as advanced glycation end products, extracellular matrix components including hyaluronic acid, and some lipoproteins. The nature of the scavenger receptors corresponding to these molecules remains to be clarified. Recently, it was noted that SEC has an antigen-presenting function similar to dendritic cells. Taken together, it is suggested that SEC, cooperating with Kupffer cells and hepatic dendritic cells, may partake of immunoregulatory functions in the liver. SEC also plays a pivotal role in the pathological process of ischemia-reperfusion injury following liver surgery and liver transplantation. Thus, it is of importance to elucidate the mechanisms of apoptosis and proliferation of SEC. Recent results on the regulation of growth and apoptotic signaling of SEC are discussed.

Animals↗

PubMed2004

Reduced expression and aberrant localization of p120catenin in human squamous cell carcinoma of the skin.

BACKGROUND: p120catenin (p120ctn), a member of the Armadillo protein family, is ubiquitously expressed and binds to classical cadherins together with other catenins to participate in the adherens junction. p120ctn has numerous isoforms generated through extensive splicing and alternative usage of translation initiation codons. Although the involvement of p120ctn in cell growth control has been postulated, little has so far been known about its expression patterns in the skin and epidermal tumors. OBJECTIVE: To identify the isoforms expressed in benign and malignant keratinocytes and to analyze the expression patterns of p120ctn in epidermal tumor specimens. METHODS: HaCaT, DJM-1, BSCC-93, HSC-1, HSC-5 and A431 cells along with normal skin tissue were subjected to RT-PCR to identify the expressed isoforms. For immunofluorescence study, surgical specimens including 29 squamous cell carcinomas (SSCs), 4 basal cell carcinomas (BCCs) and 4 seborrheic keratoses as well as 3 normal skin samples were stained with specific antibodies to detect p120ctn and E-cadherin. RESULTS: RT-PCR revealed that the isoform 3A was predominantly expressed with faint expression of the isoform 4 and that there was no difference in expressed isoforms between the examined cell lines. Immunofluorescent staining indicated that the expression of p120ctn was significantly reduced in all of the examined squamous cell carcinomas and that p120ctn was frequently localized in cytoplasm. In benign tumors and normal samples, p120ctn was properly expressed in cell-cell boundaries. Furthermore, there were several SCCs where E-cadherin continued to be expressed with no expression of p120ctn. CONCLUSION: Reduced expression and aberrant localization of p120ctn are among the most common events during the development and/or progression of SCCs.

Carcinoma, Squamous Cell↗

PubMed2004

Increased expression of H19 non-coding mRNA follows hepatocyte proliferation in the rat and mouse.

BACKGROUND/AIMS: H19 is a paternally imprinted gene that is believed to function as non-coding mRNA. While H19 is only faintly expressed in the normal adult liver, it is abundantly expressed during the fetal period. We explored the possibility that H19 might participate in the regulation of hepatocyte proliferation. METHODS: Adult male rats and mice were subjected to a two-thirds partial hepatectomy, and after various time periods, hepatocytes were isolated and analyzed for H19 gene expression. The expression was also examined in cultured rat hepatocytes. RESULTS: The expression of H19 was dramatically increased after 2 days (rat) and 4 days (mouse), peaked at 3 days (rat) and 6 days (mouse), and then gradually declined. In both species, the increase in H19 gene expression was preceded by the induction of proliferating cell nuclear antigen and DNA synthesis. An allele-specific RT-PCR analysis in the mouse showed that the paternally imprinted status of the gene was maintained after a partial hepatectomy. H19 was strongly induced in spheroid cultures after transient hepatocyte proliferation, but not in conventional monolayer cultures, in which persistent proliferation occurred. CONCLUSIONS: Our results demonstrated that H19 gene expression was dynamically regulated in adult hepatocytes in close association with their proliferation.

Animals↗

PubMed2004

Class 1 integron containing metallo-beta-lactamase gene blaVIM-2 in Pseudomonas aeruginosa clinical strains isolated in Japan.

Four bla(VIM-2) gene-harboring Pseudomonas aeruginosa strains were identified. These strains possessed a class 1 integron harboring ORF1, bla(VIM-2), and aacA4 gene cassettes. The transposon-mediated horizontal spread of the bla(VIM-2) gene among these strains was suggested, which increases the threat that the bla(VIM-2) gene will disseminate among diverse genera of bacteria.

Blotting, Southern↗

PubMed2004

Hepatocyte-specific Pten deficiency results in steatohepatitis and hepatocellular carcinomas.

PTEN is a tumor suppressor gene mutated in many human cancers, and its expression is reduced or absent in almost half of hepatoma patients. We used the Cre-loxP system to generate a hepatocyte-specific null mutation of Pten in mice (AlbCrePten(flox/flox) mice). AlbCrePten(flox/flox) mice showed massive hepatomegaly and steatohepatitis with triglyceride accumulation, a phenotype similar to human nonalcoholic steatohepatitis. Adipocyte-specific genes were induced in mutant hepatocytes, implying adipogenic-like transformation of these cells. Genes involved in lipogenesis and beta-oxidation were also induced, possibly as a result of elevated levels of the transactivating factors PPARgamma and SREBP1c. Importantly, the loss of Pten function in the liver led to tumorigenesis, with 47% of AlbCrePten(flox/flox) livers developing liver cell adenomas by 44 weeks of age. By 74-78 weeks of age, 100% of AlbCrePten(flox/flox) livers showed adenomas and 66% had hepatocellular carcinomas. AlbCrePten(flox/flox) mice also showed insulin hypersensitivity. In vitro, AlbCrePten(flox/flox) hepatocytes were hyperproliferative and showed increased hyperoxidation with abnormal activation of protein kinase B and MAPK. Pten is thus an important regulator of lipogenesis, glucose metabolism, hepatocyte homeostasis, and tumorigenesis in the liver.

Adipocytes↗

PubMed2003

A novel method for sentinel lymph node mapping using magnetite in patients with non-small cell lung cancer.

OBJECTIVE: The detection rate of sentinel lymph nodes in patients with non-small cell lung cancer using isosulfan blue dye is too low for clinical use. Although exposure to radioactivity is reportedly minimal, special procedures are nonetheless required when a radioactive isotope is used as a tracer. Therefore, to eliminate the need for a radioactive tracer and to obtain a better detection rate than is obtained with isosulfan blue dye, we have developed a novel method that employs magnetite as the tracer. The aim of the present study was to test the feasibility of this technique. METHODS: The tracer employed was ferumoxides, a colloidal superparamagnetic iron oxide of nonstoichiometric magnetite. Thirty-eight non-small cell lung cancer patients participated in the study; each received 5 mL of ferumoxides, injected around the tumor intraoperatively. Fifteen minutes after injection, lung resection and lymph node dissection were carried out. The magnetic force within the lymph nodes was measured using a highly sensitive handheld magnetometer ex vivo. All lymph nodes were also subjected to conventional histological analysis. RESULTS: The rate of detection of sentinel lymph nodes was 81.6% (31/38). The accuracy, sensitivity, and false-negative rates were 96.8% (30/31), 85.7% (6/7), and 14.3% (1/7), respectively. CONCLUSION: Intraoperative sentinel lymph node mapping using ferumoxides and a highly sensitive magnetometer is a safe, accurate, and sensitive way to detect sentinel lymph nodes in non-small cell lung cancer patients.

Adult↗

PubMed2003

Soy protein isolate enhances hepatic copper accumulation and cell damage in LEC rats.

In a series of experiments, the effects of soy protein isolate (SPI), defatted soy (DFS) or SPI supplemented with L-methionine (SPIM) were examined in the Long-Evans rat with a cinnamon coat color (LEC rat), a model animal of Wilson's disease with a hereditary defect in the Atp7b gene resulting in defective copper metabolism and copper accumulation in hepatocytes. Milk casein in the control AIN-93G diet (20 g/100 g) was totally or 60% replaced by the soy products, SPI, DFS or SPIM (L-Met added to be equal to that in the control diet) beginning when rats were 6 wk old. Copper and iron concentrations in SPI and DFS were measured and the concentrations of these metals in the salt mix were adjusted so that test and the control diets had the same final concentrations. Food intake did not differ among groups. Rats were euthanized when they became moribund with jaundice. Survival time in the SPI diet group was shorter (14.0 +/- 0.8 wk) than in the control group (19.1 +/- 1.7 wk) (P < 0.001), and that in the DFS diet group was intermediate (16.0 +/- 1.7 wk). Survival time in the SPIM diet group did not differ from that of the SPI diet group. Copper concentrations in the livers of rats in the SPI and SPIM diet groups were approximately 80% higher than in rats fed the control diet. Liver iron concentrations did not differ among the groups. The results, including histological analyses, indicate that SPI enhances copper uptake into the liver cells and promotes liver cell damage in LEC rats. However, this did not occur in the livers of F344 rats with wild-type Atp7b. Recommendations to individuals suffering from Wilson's disease to avoid consuming soy protein may be warranted.

Administration, Oral↗

PubMed2003

Connexin26-mediated gap junctional communication reverses the malignant phenotype of MCF-7 breast cancer cells.

A growing body of evidence indicates that the gap junction (GJ) plays a pivotal role in tumor suppression by exerting cell-cell communication. It has, however, been reported that expression of connexin26 (Cx26) protein is induced in human ductal carcinomas of the breast and that its amount increases in proportion to the grade of malignancy. We thus examined the effects of overexpressed Cx26 on growth characteristics in GJ-deficient human MCF-7 breast cancer cells that maintain the phenotype of early-stage cancers. MCF-7 cells were transfected with Cx26 cDNA, and several clones of stable transformants exhibiting a high level of cell-cell communication were established. When they were examined in terms of various growth characteristics in vitro, the proliferation rate and the saturation density were drastically reduced in Cx26-transfected clones compared with the mock-transfectant. The anchorage-independent growth capacity was also decreased by 50-75% after transfection of Cx26. Furthermore, the cell migration toward growth factors and cell invasion into Matrigel in a Boyden chamber were suppressed to 5-10% and 20-60%, respectively, of the control in Cx26-transfected clones. When implanted into the mammary fat pads of nude mice in the presence of an excess of 17beta-estradiol, Cx26-transfected clones tended to show slower tumor growth than the mock-transfectant, although the difference was not statistically significant. Our results strongly suggest that the induction of Cx26 protein observed in human breast cancers, reported previously, may not be very relevant to the development of breast cancers, and that Cx26 can function as a tumor suppressor in breast cancer cells.

Animals↗

PubMed2003

Characterization of atypical enteropathogenic Escherichia coli strains harboring the astA gene that were associated with a waterborne outbreak of diarrhea in Japan.

The virulence traits of the Escherichia coli strain associated with a waterborne diarrhea outbreak were examined. Forty-one of 75 students (ages 12 to 15) in Akita Prefecture, Japan, showed clinical symptoms. Seven E. coli Ouk:K-:H45 isolates were isolated from the patients as the causative agent of this outbreak. One isolate (EC-3605) showed the presence of E. coli attaching-and-effacing (eaeA) and enteroaggregative E. coli heat-stable enterotoxin-1 (astA) genes and the absence of Shiga toxin (stx1 and stx2) genes. A polymorphic enteropathogenic E. coli (EPEC) adherence factor plasmid was detected in EC-3605 with a major structural gene deletion and a regulatory gene frameshift mutation, revealing that EC-3605 represents an atypical EPEC strain harboring the astA gene. The role that atypical EPEC strains harboring the astA gene play in human disease is unclear. Our results, along with those of others, present a possibility that these strains comprise a distinct category of diarrheagenic E. coli and that astA affects the age distribution of atypical-EPEC infection.

Adolescent↗

PubMed2002

A highly specific isolation of rat sinusoidal endothelial cells by the immunomagnetic bead method using SE-1 monoclonal antibody.

BACKGROUND/AIMS: To develop a specific isolation method of hepatic sinusoidal endothelial cells (SEC), we applied the immunomagnetic method using a monoclonal antibody (SE-1) that recognizes a membranous antigen expressed only in rat SEC. METHODS: Cells were isolated by incubating mixed non-parenchymal cells, which were obtained by collagenase digestion of the liver, with SE-1-conjugated superparamagnetic polystyrene beads. The conventional Percoll method was also performed in parallel to compare with the immunomagnetic method. The isolated cells were cultured on glass coverslips coated with type I collagen in the presence of various growth factors for 6 days. RESULTS: Approximately 98% of the isolated cells were positive for SE-1 and the contamination of Kupffer cells or stellate cells was less than 1%. The purity was significantly better than that obtained by the Percoll method. The cultured cells showed typical SEC features, such as sieve plates and uptake of acetylated low-density lipoprotein. Although the cells continuously underwent apoptotic cell death after 2 days, they started robust cell growth after 3 days and were well maintained during the culture period. CONCLUSIONS: Our simple and specific isolation method enables us to culture SEC with high purity and should be useful for the biological analysis of SEC.

Animals↗

PubMed2002

Primary osteosarcoma of the uterine corpus: case report and review of the literature.

A rare case of rapidly growing osteosarcoma that developed in the uterine corpus of a 62-year-old woman is presented. The tumor occupied almost the entire pelvic cavity and extended into the abdominal cavity, with marked involvement of the intestines. Histopathologically, the tumor was composed of an osteoblastic component, accompanied by conspicuous bone formation, and a fibroblastic component. The tumor cells were positive for vimentin and osteocalcin, as well as desmin, alpha-smooth muscle actin and muscle-specific actin, but negative for h-caldesmon. The results indicated myofibroblastic differentiation in a part of the tumor. A review of 14 reported cases and our case of uterine osteosarcoma revealed that this tumor has a biologically aggressive nature, although its histopathological and immunohistochemical features are similar to those of osteosarcomas in soft tissue and bone. As the prognosis of patients with this tumor is poor, it is of importance to differentiate this tumor from other types of tumors arising from the uterine corpus.

Actins↗

PubMed2002

Characterization of enteropathogenic and enteroaggregative Escherichia coli isolated from diarrheal outbreaks.

Virulence characteristics of diarrheal outbreak-associated Escherichia coli O55:NM, O126:NM, and O111:NM were examined. The E. coli O55:NM strains were atypical enteropathogenic E. coli (EPEC), while the E. coli O126:NM and O111:NM strains should be classified as enteroaggregative E. coli (EAggEC). The contributions of EPEC and EAggEC to the human disease burden in Japan might be significantly greater than is currently appreciated.

Adolescent↗