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Biomedical subjects

Naoko Watanabe

Publications and source records attributed to Naoko Watanabe.

At least 19 recordsLinked to original sources
PubMed2007

Effect of rapamycin on hepatocyte function and proliferation induced by growth factors.

BACKGROUND: Rapamycin is a specific inhibitor of the mammalian target of rapamycin (mTOR). The effect of rapamycin on proliferation and cellular function was studied in hepatocytes stimulated by hepatocyte growth factor (HGF) or transforming growth factor-alpha (TGFalpha). METHODS AND RESULTS: When isolated rat hepatocytes were cultured at low density, the addition of HGF or TGFalpha increased DNA synthesis but did not affect albumin or fibrinogen concentrations in the medium. In contrast, in hepatocytes cultured at high density, the albumin and fibrinogen concentrations, but not DNA synthesis, were increased by HGF or TGFalpha. The HGF- or TGFalpha-induced increase in DNA synthesis and in albumin or fibrinogen concentrations was suppressed by the addition of rapamycin, as well as wortmannin, a phosphatidylinositol-3 kinase inhibitor. CONCLUSION: HGF and TGFalpha stimulate proliferation and function of hepatocytes depending upon the conditions, and rapamycin inhibited these stimulatory effects, possibly by inhibiting the mTOR-dependent signaling pathway.

Albumins↗
PubMed2006

Reproductive effects and bioconcentration of 4-nonylphenol in medaka fish (Oryzias latipes).

The objective of this study was to investigate the reproductive success (fertility and fecundity) of medaka (Oryzias latipes) exposed to 4-nonylphenol (4-NP) in water and the viability of their F(1) offspring. In addition, we measured the bioconcentration of 4-NP in eggs. After a 21-d exposure to 100microg/l 4-NP, medaka showed reduced egg production and fertility. Hepatic vitellogenin levels were increased significantly in males treated with 10, 50 and 100microg/l of 4-NP. In the F(1) generations, the hatchability and time to hatching of embryos in the 100microg/l treatment group were adversely affected, and 2-7microg 4-NP/g egg was found in spawned eggs (the bioconcentration factor: 30-100). These results indicate that (1) 4-NP adversely affected the reproduction (fecundity and fertility) of adult medaka, (2) 4-NP accumulated in eggs through maternal transfer and (3) these levels of 4-NP were associated with adverse effects in the F(1) offspring.

Animals↗
PubMed2006

Use of the SST-REX method for the identification of genes expressed at the condensation stage of chondrogenic cell line ATDC5.

In endochondral ossification, chondrogenesis precedes bone development. Cartilage differentiation is initiated by the formation of chondrogenic cell condensates. Thus, it is essential to investigate genes expressed at the condensation stage for a better understanding of chondrogenesis and ossification. To this end, we constructed a cDNA library from the mRNA fraction derived from a chondrogenic cell line (ATDC5) at the cell condensation stage by using the signal sequence trap by retrovirus-mediated expression (SST-REX) method. We obtained 486 factor (IL-3)-independent clones by screening 5.7 x 10(3) clones. DNA sequencing analysis of the clones identified genes encoding 157 known proteins and 4 novel proteins. These 4 genes encoding novel proteins were expressed not only in chondrogenic ATDC5 cells but also in osteogenic MC3T3-E1 and myogenic C2C12 cells. The mRNA expression level of 1 of the 4 clones increased at the calcification stage of the differentiation of MC3T3-E1 cells. The results demonstrate that the SST-REX method is a useful experimental system to identify genes involved in the complicated mechanisms of bone formation.

Animals↗
PubMed2006

Chronic eosinophilic leukemia with the FIP1L1-PDGFRalpha fusion gene in a patient with a history of combination chemotherapy.

Hypereosinophilic syndrome (HES) was diagnosed in December 2000 in a 43-year-old man on the basis of persistent eosinophilia (11.7 x 10(9)/L) and a normal karyotype of the bone marrow cells. He had developed intra-abdominal non-Hodgkin's lymphoma and in 1992 had received 3 courses of combination chemotherapy with doxorubicin (Adriamycin), cyclophosphamide, vincristine, methotrexate, bleomycin, and prednisolone. The patient was orally given prednisolone (10 mg/day) and cyclophosphamide (50 mg/day) as HES treatment without a subsequent improvement of the eosinophilia. In May 2003, anemia (hemoglobin, 7.9 g/dL) and thrombocytopenia (65 x 10(9)/L) manifested with progressive eosinophilia (21.0 x 10(9)/L) and a small number of blasts. The patient became febrile and was admitted in July 2003. Cytogenetic reexamination of the bone marrow cells disclosed the deletion of 4q12, indicating the presence of a fusion of the Fip1-like 1 (FIP1L1) gene to the plateletderived growth factor receptor alpha (PDGFRalpha) gene and consequently the clonal nature of his hematopoietic cells. DNA sequence analysis demonstrated that the breakpoints of the FIP1L1 and PDGFRalpha genes were present in exon 9 and exon 12, respectively. Treatment with imatinib mesylate (300 mg/day) promptly brought about complete remission. Although a number of similar eosinophilic cases have been reported, our patient may be the first such patient with a history of chemotherapy.

Adult↗
PubMed2005

The herbal medicine inchin-ko-to (TJ-135) induces apoptosis in cultured rat hepatic stellate cells.

Use of herbal remedies in the treatment of various diseases has a long tradition in Eastern medicine and the liver diseases are not an exception. In their use, lack of elucidation of mechanism(s) as well as randomized, placebo-controlled clinical trials has been a problem. Recently, we and others reported that inchin-ko-to (TJ-135), one of herbal remedies, suppressed hepatic fibrosis in animal models. In the course of clarifying the mechanism, we directed our focus on hepatic stellate cells (HSCs), playing a pivotal role in hepatic fibrosis, and found that rat HSCs cultured with TJ-135 changed their morphology to star-like configuration with thin, slender and dendritic processes with fewer stress fibers, which might be the features in apoptosis. In fact, TJ-135 induced HSC apoptosis in a time- and concentration-dependent manner as judged by the nuclear morphology, quantitation of cytoplasmic histone-associated DNA oligonucleosome fragments and caspase 3 activity. In HSCs treated with TJ-135, increased expression of p53 and decreased expression of Bcl-2 and phosphorylated Akt and Bad were determined. HSC apoptosis is shown to be involved in the mechanisms of spontaneous resolution of rat hepatic fibrosis and the agent which induces HSC apoptosis has been shown to reduce experimental hepatic fibrosis in rats. Thus, the induction of HSC apoptosis could be the mechanism how TJ-135 works on the resolution of hepatic fibrosis. Our current data may shed light on the novel effect of the herbal remedy.

Animals↗
PubMed2005

Toxicity to early life stages and an estrogenic effect of a bisphenol A metabolite, 4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene on the medaka (Oryzias latipes).

In a recent study, it was reported that 4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene (MBP), a metabolite of bisphenol A (BPA; 2,2-bis(4-hydroxyphenyl)propane), showed estrogenic activity in several in vitro assays, and the estrogenic activity of MBP was higher than that of BPA. In this study, we have investigated the early life stage toxicity and estrogenic effect of MBP on medaka (Oryzias latipes). The 96-h median lethal concentration value of MBP and BPA with 24-h-old larvae was estimated to be 1,640 and 13,900 microg/l, respectively. The hatchability of fertilized eggs exposed to MBP and BPA over 14 days was significantly decreased at doses of 2,500 microg/l and 12,500 microg/l, respectively. Moreover, to compare the potency of estrogenic activity in vivo, male medaka were exposed to various concentrations of MBP and BPA for 21 days. The lowest-observed-effect concentrations of MBP and BPA for hepatic vitellogenin induction in male medaka were estimated to be 4.1 and 1,000 microg/l, respectively. These results suggest that MBP has high toxicity for early life stages of the medaka, and that the estrogenic activity of MBP was about 250-fold higher than that of BPA to male medaka.

Animals↗
PubMed2005

Relating desorption and biodegradation of phenanthrene to SOM structure characterized by quantitative pyrolysis GC-MS.

A set of four soils was extensively studied for composition, sorption/desorption, and biodegradation to investigate linkages among these three inter-related factors. Composition of soil organic matter (SOM) was quantitatively characterized using pyrolysis GC-MS; although CO2 dominated the total ion chromatogram for all soils, each soil produced a distinctively different pyrogram and 1.4-4.8% of the soil carbon was quantified as one of 205 pyrolysis marker compounds using external standards. Amorphous and free iron and aluminum contents were determined as potential indicators of the reactivity of the mineral phase, which may influence the configuration and accessibility of SOM domains within soil. We observed a statistically significant positive correlation between two-site model fast fraction f values derived from mineralization and desorption rate studies, which suggests that desorption limits biodegradation. However, no statistically significant correlation was observed between two-site model fast and slow rate constants (k(f), k(s)) for the two processes. No evidently strong correlations were found between functional parameters (organic carbon normalized distribution coefficient Koc, hyeteresis index HI, and two-site model parameters f, k(f), and k(S) for both desorption and biodegradation and maximum rate and extent of biodegradation) and SOM structural descriptors (pyrolysis results). Lack of strong correlations may suggest (i) that multiple SOM structures are collectively responsible for desorption resistance or (ii) that the pyrolysis GC-MS method used in this study was unable to identify relevant structures. In contrast, amorphous and free iron and aluminum contents showed statistically significant correlations with K(OC) and HI values, indicating the potential importance of underlying mineral phases in determining desorption and biodegradation rates.

Biodegradation, Environmental↗
PubMed2005

Decreased fidelity in replicating CpG methylation patterns in cancer cells.

The unmethylated or methylated status of individual CpG sites is faithfully copied into daughter cells. Here, we analyzed the fidelity in replicating their methylation statuses in cancer cells. A single cell was clonally expanded, and methylation statuses of individual CpG sites were determined for an average of 12.5 DNA molecules obtained from the expanded population. By counting the deviation from the original methylation patterns inferred, the number of errors was measured. The analysis was done in four gastric cancer cell lines for five CpG islands (CGI), and repeated six times (total 1,495 clones sequenced). HSC39 and HSC57 showed error rates <1.0 x 10(-3) errors per site per generation (99.90-100% fidelity) for all the five CGIs. In contrast, AGS showed significantly elevated error rates, mainly due to increased de novo methylation, in three CGIs (1.6- to 3.2-fold), and KATOIII showed a significantly elevated error rate in one CGI (2.2-fold). By selective amplification of fully methylated DNA molecules by methylation-specific PCR, those were stochastically detected in KATOIII and AGS but never in HSC39 and HSC57. When methylation of entire CGIs was examined for eight additional CGIs, KATOIII and AGS had frequent methylation, whereas HSC39 and HSC57 had few. KATOIII and AGS had four and eight times, respectively, as high expression levels of DNMT3B as HSC39. These data showed that some cancer cells have decreased fidelity in replicating methylation patterns in some CGIs, and that the decrease could lead to methylation of the entire CGIs.

Breast↗
PubMed2005

Changes of glucose metabolism and skin-collagen neogenesis in vitamin B6 deficiency.

The mechanism of pellagrous changes in skin caused by a deficiency of vitamin B6 was studied in respect to neogenesis of proline in skin collagen and glucose metabolism. In vitamin B6 deficiency the insulin/glucagon coefficient in serum decreased significantly from 3.02 to 2.32, indicating a metabolic change towards gluconeogenesis. A deficiency of vitamin B6 caused a decrease in the levels of vitamin B6-dependent enzymes, such as ornithine aminotransferase, alanine aminotransferase, and aspartate aminotransferase, which also contribute to gluconeogenesis. Because the conversion of ornithine to proline via pyrroline-5-carboxylate was suppressed due to the decrease in ornithine aminotransferase activity, the amount of proline in the skin collagen fraction also decreased significantly in vitamin B6-deficient rats compared with the pair-fed control. These results suggest that the pellagrous lesions in vitamin B6-deficiency are caused by an impaired synthesis of proline from ornithine, which results in the suppression of collagen neogenesis in the skin.

Animals↗
PubMed2005

Excellent response of chemotherapy-resistant B-cell-type chronic lymphocytic leukemia with meningeal involvement to rituximab.

A 70-year-old woman was diagnosed with B-cell-type chronic lymphocytic leukemia (B-CLL) in May 2001. Initial white blood cell (WBC) count was 37 x 10(9)/l and most of the cells were mature small lymphocytes. Surface antigen analysis of these lymphocytes revealed positive reactions for CD19, 20, 25, 5, and lambda-light chain. Despite her Rai stage-0 status, various treatments were ineffective, including cyclophosphamide; fludarabine; 6-mercaptopurine; a combination of vincristine, cyclophosphamide, prednisolone, and adriamycin; and etoposide. Her WBC count increased, ranging from 150 to 450 x 10(9)/l, with marked splenomegaly, and symptoms of meningitis, such as headache, ophthalmalgia, hearing disturbance, and abnormal behavior, being manifested. The WBC count in the cerebrospinal fluid was elevated to 134/microl. The surface phenotype of these cells was identical to that of circulating lymphocytes, indicating meningeal involvement of leukemia, a rare complication in B-CLL. At the time of this WBC elevation, 24% of circulating lymphocytes had prominent nucleoli, indicating progression of the disease to CLL/prolymphocytic leukemia. Her symptoms disappeared after repeated intrathecal injections of methotrexate and dexamethazone. After four courses of treatment of the refractory B-CLL with rituximab, an anti-CD20 monoclonal antibody, the WBC count returned to normal levels and the splenomegaly disappeared. She is currently well, with sustained remission, as of April 2004.

Aged↗
PubMed2005

Effects of nonylphenol and triclosan on production of plasma vitellogenin and testosterone in male South African clawed frogs (Xenopus laevis).

We investigated the effects of nonylphenol (NP) and triclosan (TCS) on production of vitellogenin (Vg), testosterone (T), and hepatic cytochrome P450 1A and 2B activities in male South African clawed frogs (Xenopus laevis). In a 14-d waterborne exposure test, no significant differences in the level of plasma Vg synthesis in male frogs were observed among the control, 10, 50, and 100 microg/l NP and 20, 100, and 200 microg/l TCS treatment groups. Intraperitoneal injection of male frogs with 2, 20, and 200 microg/g body weight NP resulted in no significant differences in plasma Vg levels among the control and all treatment groups. However, the levels of plasma Vg in all TCS treatment groups (intraperitoneal injection of 4, 40, and 400 microg/g body weight) were lower than that in the solvent control group, and male frogs injected with high doses of NP or TCS had lower T levels than the control group. No significant differences in hepatic cytochrome P450 1A and 2B activities were observed among the all treatment groups. Male frogs injected with 20 microg/g body weight of estradiol-17beta had significantly higher plasma Vg levels than the control group. These results suggest that profiles of plasma Vg and T production in male Xenopus laevis could be useful biomarkers for detecting hormonally active agents.

Animals↗
PubMed2005

Presence of a cytoplasmic retention sequence within the human interleukin-1alpha precursor.

Interleukin (IL)-1alpha is primarily translated as a 33 kDa molecule (IL-1alpha1-271), and then processed into a 17 kDa molecule (IL-1alpha119-271) by calpain. The precursor region of IL-1alpha (IL-1 alpha1-118) contains a nuclear localization signal (KVLKKRRL, residues 79-86). We investigated the intracellular localization of IL-1alpha fused with green fluorescent protein or beta-galactosidase. IL-1alpha1-118 was localized exclusively in the nucleus, but IL-1 alpha1-271 in both the nucleus and the cytoplasm, suggesting the presence of a cytoplasmic retention signal within the mature region of IL-1alpha. Furthermore, the intracellular localization of IL-1alpha with deletions from the C terminus, internal deletions and point mutations suggested that the cytoplasmic retention signal is located within residues 168-201.

Amino Acid Motifs↗
PubMed2005

[Renal insufficiency caused by leukemic cell infiltration in Sézary syndrome].

A 77-year-old woman was admitted to our hospital because of pneumonia and heart failure in July 2002. She had been diagnosed as having with Sézary syndrome in 1993, and had been treated with a combination of prednisolone, methotrexate, and cyclosporin A with subsequent stable disease but persistent generalized erythroderma. On admission, the white blood count was 23.9 X 10(9)/L with 28% Sézary cells, and serum creatinine levels were within normal limits. One month after admission, the pneumonia and heart failure improved remarkably with antibiotics and diuretics. However, at the same time, her renal function deteriorated with increasingly high serum creatinine levels. She died of anuria in September, 2002. An autopsy showed marked perivascular and peritubular infiltration of abnormal lymphocytes with degenerative nephrotubuli in the kidneys. This patient may be the first reported case of Sézary syndrome with renal failure caused by leukemic infiltration.

Aged↗
PubMed2004

Treatment with leucine stimulates the production of hepatocyte growth factor in vivo.

Hepatocyte growth factor (HGF) has pleiotropic effects. Up-regulation of HGF activity in vivo may be beneficial. Branched-chain amino acids (BCAAs) are known to modulate various cellular functions. When starved rats received intraperitoneal injections of valine, leucine or isoleucine, only leucine treatment increased both hepatic and circulating levels of HGF in a dose-dependent manner, up to 1.5 and 2.3 times higher, respectively, than in controls. When young growing rats with free access to food were injected with leucine once a day for a week, HGF levels and liver weights were significantly higher than those of control rats. Furthermore, 1 week of leucine treatment of adult rats resulted in elevated serum albumin levels with an increase in HGF levels. Taken together with our previous report showing that leucine stimulates HGF production by hepatic stellate cells in culture, leucine, among BCAAs, may induce an increase in HGF production by the liver in vivo.

Animals↗
PubMed2004

Lysyl oxidase is a tumor suppressor gene inactivated by methylation and loss of heterozygosity in human gastric cancers.

Lysyl oxidase (LOX) and HRAS-like suppressor (HRASLS) are silenced in human gastric cancers and are reported to have growth-suppressive activities in ras-transformed mouse/rat fibroblasts. Here, we analyzed whether or not LOX and HRASLS are tumor suppressor genes in human gastric cancers. Loss of heterozygosity and promoter methylation of LOX were detected in 33% (9 of 27) and 27% (26 of 96) of gastric cancers, respectively. Biallelic methylation and loss of heterozygosity with promoter methylation were also demonstrated in gastric cancers. Silencing of LOX was also observed in colon, lung, and ovarian cancer cell lines. As for mutations, only one possible somatic mutation was found by analysis of 96 gastric cancer samples and 58 gastric and other cancer cell lines. When LOX was introduced into a gastric cancer cell line, MKN28, in which LOX and HRASLS were silenced, it reduced the number of anchorage-dependent colonies to 57 to 61%, and the number of anchorage-independent colonies to 11 to 23%. Sizes of tumors formed in nude mice were reduced to 19 to 26%. Growth suppression in soft agar assay was also observed in another gastric cancer cell line, KATOIII. On the other hand, neither loss of heterozygosity nor a somatic mutation was detected in HRASLS, and its introduction into MKN28 did not suppress the growth in vitro or in vivo. These data showed that LOX is a tumor suppressor gene inactivated by methylation and loss of heterozygosity in gastric cancers, and possibly also in other cancers.

Animals↗
PubMed2004

Sphingosine 1-phosphate enhances portal pressure in isolated perfused liver via S1P2 with Rho activation.

Although structural changes are most important to determine vascular resistance in portal hypertension, vasoactive mediators also contribute to its regulation. Hepatic stellate cells (HSCs) are assumed to play a role in modulating intrahepatic vascular resistance based on their residence in the space of Disse and capacity to contract. Because sphingosine 1-phosphate (S1P) has been shown to stimulate HSC contractility, we wondered if S1P could regulate portal pressure. S1P at 0.5-5 microM increased portal pressure in isolated rat perfused liver. This effect was abrogated in the presence of a binding antagonist for S1P2, JTE-013. Perfusion of isolated rat liver with 5 microM S1P increased Rho activity in the liver, and co-perfusion with JTE-013 cancelled S1P-induced Rho activation. Because S1P is present in human plasma at approximately 0.2 microM, S1P might readily regulate portal vascular tone in physiological and pathological status. The antagonist for S1P2 merits consideration for treatment of portal hypertension.

Animals↗
PubMed2004

M-phase kinases induce phospho-dependent ubiquitination of somatic Wee1 by SCFbeta-TrCP.

Wee1, the Cdc2 inhibitory kinase, needs to be down-regulated at the onset of mitosis to ensure rapid activation of Cdc2. Previously, we have shown that human somatic Wee1 (Wee1A) is down-regulated both by protein phosphorylation and degradation, but the underlying mechanisms had not been elucidated. In the present study, we have identified the beta-transducin repeat-containing protein 1/2 (beta-TrCP1/2) F-box protein-containing SKP1/Cul1/F-box protein (SCF) complex (SCF(beta-TrCP1/2)) as an E3 ubiquitin ligase for Wee1A ubiquitination. Although Wee1A lacks a consensus DS(p)GXXS(p) phospho-dependent binding motif for beta-TrCP, recognition of Wee1A by beta-TrCP depended on phosphorylation, and two serine residues in Wee1A, S53 and S123, were found to be the most important phosphorylation sites for beta-TrCP recognition. We have found also that the major M-phase kinases polo-like kinase 1 (Plk1) and Cdc2 are responsible for the phosphorylation of S53 and S123, respectively, and that in each case phosphorylation generates an unconventional phospho-degron (signal for degradation) that can be recognized by beta-TrCP. Phosphorylation of Wee1A by these kinases cooperatively stimulated the recognition and ubiquitination of Wee1A by SCF(beta-TrCP1/2) in vitro. Mutation of these residues or depletion of beta-TrCP by small-interfering RNA treatment increased the stability of Wee1A in HeLa cells. Moreover, our analysis indicates that beta-TrCP-dependent degradation of Wee1A is important for the normal onset of M-phase in vivo. These results also establish the existence of a feedback loop between Cdc2 and Wee1A in somatic cells that depends on ubiquitination and protein degradation and ensures the rapid activation of Cdc2 when cells are ready to divide.

Amino Acid Sequence↗