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Biomedical subjects

J Fujimoto

Publications and source records attributed to J Fujimoto.

At least 91 records · Page 5Linked to original sources
PubMed1999

Hepatocyte growth factor promotes migration of human hepatocellular carcinoma via phosphatidylinositol 3-kinase.

Hepatocyte growth factor (HGF) is known to be a potent mitogen and motogen for epithelial cells. Hepatocellular carcinoma (HCC) often metastasizes, and the c-Met/HGF receptor is highly expressed by HCC cells. The aim of this study was to investigate the signaling pathways associated with the motogenic effect of HGF on HCC cells via c-Met. HCC cell lines (Hep3B, HepG2, PLC, and Huh-7) and HCC cells harvested from patients were used for the Boyden chamber assay of chemotactic activity as well as for immunoprecipitation and immunoblotting studies. HGF stimulated the motility of Hep3B, HepG2, and Huh-7 cells in a dose-dependent manner in association with tyrosine phosphorylation of c-Met and activation of phosphatidylinositol 3-kinase (PI3-K). A tyrosine kinase inhibitor (genistein) and a PI3-K inhibitor (wortmannin) prevented the migration of HCC cells. However, migration was not prevented by calphostin C, an inhibitor of protein kinase C (PKC), which is a downstream target of phospholipase Cgamma (PLCgamma). HGF also stimulated the migration of HCC cells obtained from three patients, while wortmannin prevented the migration of these cells. These results indicate that HGF stimulates the migration of HCC cells through the tyrosine phosphorylation of c-Met via activation of PI3-K.

Androstadienes↗
PubMed1999

Adaptation to a starch environment and regulation of alpha-amylase in Drosophila.

The adaptation to glucose and starch foods in six species, D. melanogaster, D. virilis, D. saltans, D. funebris, D. levanonensis and D. americana, was studied by measuring productivity. D. melanogaster and D. virilis adapted more to the starch environment than to the glucose environment, while D. saltans adapted more to the glucose environment than to the starch environment. D. funebris, D. levanonensis, and D. americana did not distinctly adapt to either environment. In addition, the regulation of amylase in the six species was investigated by measuring the levels of amylase activity with glucose and starch food environments. The levels of amylase activity in D. levanonensis and D. saltans were substantially low, indicating that these species cannot utilize starch as a carbon source. The starch-adapted species, D. melanogaster and D. virilis, showed higher levels of amylase activity with the starch environment and higher inducibility. These results suggest that changing the regulation of amylase is important for the adaptation to a starch environment in Drosophila.

Adaptation, Physiological↗
PubMed1999

Hepatocyte growth factor gene therapy of liver cirrhosis in rats.

Liver cirrhosis is the irreversible end result of fibrous scarring and hepatocellular regeneration, characterized by diffuse disorganization of the normal hepatic structure of regenerative nodules and fibrotic tissue. It is associated with prominent morbidity and mortality, and is induced by many factors, including chronic hepatitis virus infections, alcohol drinking and drug abuse. Hepatocyte growth factor (HGF), originally identified and cloned as a potent mitogen for hepatocytes, shows mitogenic, motogenic and morphogenic activities for a wide variety of cells. Moreover, HGF plays an essential part in the development and regeneration of the liver, and shows anti-apoptotic activity in hepatocytes. In a rat model of lethal liver cirrhosis produced by dimethylnitrosamine administrations, repeated transfections of the human HGF gene into skeletal muscles induced a high plasma level of human as well as enodogenous rat HGF, and tyrosine phosphorylation of the c-Met/HGF receptor. Transduction with the HGF gene also suppressed the increase of transforming growth factor-beta1 (TGF-beta1), which plays an essential part in the progression of liver cirrhosis, inhibited fibrogenesis and hepatocyte apoptosis, and produced the complete resolution of fibrosis in the cirrhotic liver, thereby improving the survival rate of rats with this severe illness. Thus, HGF gene therapy may be potentially useful for the treatment of patients with liver cirrhosis, which is otherwise fatal and untreatable by conventional therapy.

Animals↗
PubMed1999

Expression of platelet-derived endothelial cell growth factor (PD-ECGF) and its mRNA in uterine cervical cancers.

Angiogenesis contributes to the growth and secondary spreading of solid tumours. Platelet-derived endothelial cell growth factor (PD-ECGF) is identified as such an angiogenic factor. In the present study, the prognosis of the patients with high PD-ECGF uterine cervical cancers was worse than those with low PD-ECGF cancers, and PD-ECGF expression correlated with cellular proliferation and with vascular density and venous invasion in uterine cervical cancers. Therefore, PD-ECGF might contribute to the growth of uterine cervical cancers via angiogenesis related to vascular spreading. Furthermore, PD-ECGF and its mRNA had a wide range and were highly expressed in uterine cervical cancers, especially squamous cell carcinoma, regardless of clinical stage. Therefore, PD-ECGF in uterine cervical cancers might play a role of basic angiogenesis in all processes of advancing of uterine cervical cancers. This indicates that 5'-deoxy-5-fluorouridine might be highly effective in squamous cell carcinoma of the cervix, which possesses a high activity of thymidine phosphorylase to convert 5'-deoxy-5-fluorouridine to 5-fluorouracil, and that some angiogenic inhibitors of new capillary formation might be effective in the inhibition of tumour growth and spreading associated with angiogenesis.

Adult↗
PubMed1999

Expression of vascular endothelial growth factor (VEGF) and its mRNA in uterine cervical cancers.

To know the potential of growth, invasion and metastasis of uterine cervical cancer associated with neovascularization, localization of vascular endothelial growth factor (VEGF) and microvessel density in tumours were determined by immunohistochemical staining, the levels of VEGF subtypes were determined by Western blot analysis and by a sandwich enzyme immunoassay, and the levels of VEGF subtype mRNAs were determined by reverse transcription polymerase chain reaction (RT-PCR)-Southern blot analysis in uterine cervical cancers. The relation between VEGF subtype expressions and microvessel density, histological types and clinical stages of uterine cervical cancers was analysed. The expression of VEGF was seen dominantly in the cancer cells, and correlated with microvessel density in uterine cervical cancers. Among the four subtypes of VEGF, the populations of VEGF165 and VEGF121 were dominant in normal uterine cervices and uterine cervical cancers. The levels of VEGF and VEGF165 and VEGF121 mRNAs were remarkably higher in some stage II and III/IV adenocarcinomas of the cervix than in other cases, including normal cervices. Therefore, the elevation of VEGF165 and VEGF121 might contribute to the relatively late advancing via angiogenic activity in some adenocarcinomas of the cervix.

Adult↗
PubMed1999

Synthesis and utilization of 13C(8)-enriched purines.

A new and more efficient method is presented for the synthesis of 13C(8)-enriched adenine. In addition, we present for the first time the synthesis of 13C(8)-enriched 2-aminopurine and purine. All three analogues have been converted to the corresponding ribonucleosides. The adenine analogue has been further converted to the 2'-deoxy-nucleoside and incorporated into a synthetic oligonucleotide. Data is presented demonstrating the utility of 13C-enrichment in heteronuclear isotope-edited NMR spectra.

Base Sequence↗
PubMed1999

Shiga toxins induce apoptosis in pulmonary epithelium-derived cells.

The effect of Shiga toxins (Stxs) produced by Escherichia coli on human lung epithelial cells was investigated. Specific antibodies for Stxs positively stained lung tissue from a patient who died of hemolytic uremic syndrome associated with Stx-producing E. coli (STEC) infection, indicating the deposition of Stxs in the lung. Binding experiments with normal lung tissue revealed apparent Stx binding to both vascular endothelium and to portions of the pulmonary epithelium. CD77-positive lung carcinoma cell lines, which are derived from lung epithelium, showed binding to Stx and a high susceptibility to Stxs, as determined by MTT assay. Consistent with our previous reports on renal tubular epithelium, apoptosis is involved in the Stx-mediated cytotoxicity of these lines. These data indicate that lung epithelium is another target for Stxs, and Stx-mediated injury to lung epithelial cells is thought to play an important role in the pathogenesis of pulmonary involvement associated with STEC infection.

Antigens, CD↗
PubMed1999

Expression of oestrogen receptor alpha and beta mRNA in corpus luteum of human subjects.

To investigate the role of oestrogen receptor beta (ERbeta) in the function of human ovarian corpus luteum, the levels of luteal ERalpha and ERbeta mRNA were determined using competitive reverse transcription-polymerase chain reaction-Southern blot analysis. The expression of ERalpha and ERbeta mRNA was detected in all luteal samples analysed. Luteal ERalpha and ERbeta mRNA levels were significantly lower (P<0.01 and P<0.05 respectively) at the late secretory phase than those at the early and mid-secretory phases of the endometrium. The ratio of ERalpha to ERbeta mRNA levels showed no change during the secretory phase of the endometrium. This study demonstrates that ERbeta is co-expressed with ERalpha in human corpus luteum and is likely to play a biological role in the regulation of steroidal action of the corpus luteum with ERalpha.

Adult↗
PubMed1999

Expression of oestrogen receptor-alpha and -beta in ovarian endometriomata.

The contribution of oestrogen receptor (ER) isoforms, ER-alpha and ER-beta, in oestrogen-dependent development and growth of ovarian endometriomata, is unknown. Therefore, we examined the expression of ER-alpha and ER-beta in ovarian endometriomata and normal uterine endometrium. ER-alpha and ER-beta were shown to be dominantly expressed in the nuclei of the epithelial lining cells of ovarian endometrioma and of the glandular cells of normal uterine endometrium. ER-beta was expressed at a much lower level than ER-alpha in the glandular cells of normal uterine endometrium, while ER-beta was expressed at a slightly lower level than ER-alpha in the epithelial lining cells of ovarian endometrioma. In normal uterine endometrium, ER-beta mRNA was expressed at a much lower level than ER-alpha mRNA, and the expression pattern of ER-beta mRNA during the menstrual cycle was similar to that of ER-alpha mRNA. On the other hand, ER-beta mRNA expression was significantly higher and over a much greater range in ovarian endometriomata (P < 0.05) than in normal uterine endometrium during the menstrual cycle, while ER-alpha mRNA expression was relatively lower and more random. Therefore, in ovarian endometriomata, oestrogen action via ER-alpha cascades seems to be partially damaged, as the expression of ER-alpha mRNA does not respond to endocrinological alterations during the menstrual cycle, while the relative over-expression of ER-beta might be related to a unique oestrogen-dependent growth and spreading of ovarian endometriomata.

Adult↗
PubMed1999

PTEN/MMAC1 mutations in hepatocellular carcinomas: somatic inactivation of both alleles in tumors.

Allelic loss of loci on chromosome 10q occurs frequently in hepatocellular carcinomas. Somatic mutations of the PTEN/MMAC1 gene on this chromosome at 10q23 were recently identified in sporadic cancers of the uterus, brain, prostate and breast. To investigate the potential role of PTEN/MMAC1 gene in the genesis of hepatocellular carcinomas, we examined 96 tumors for allelic loss on 10q and also for subtle mutations anywhere within the coding region of PTEN/MMAC1 gene. Allelic loss was identified in 25 of the 89 (27%) tumors that were informative for polymorphic markers in the region. Somatic mutations were identified in five of those tumors: three frameshift mutations, a 1-bp insertion at codon 83-84 in exon 4 and two 4-bp deletions, both at codon 318-319 in exon 8; two C-to-G transversion mutation, both at -9 bp from the initiation codon in the 5' non-coding region of exon 1. No missense mutation was observed in this panel of tumors. In most of the informative tumors carrying intragenic mutations of one allele, we were able to detect loss of heterozygosity as well. These findings suggest that two alleles of the PTEN/MMAC1 gene may be inactivated by a combination of intragenic point mutation on one allele and loss of chromosomal material on the other allele in some of these tumors.

Alleles↗
PubMed1999

Immunohistochemical detection of human telomerase reverse transcriptase in normal mucosa and precancerous lesions of the stomach.

Telomerase activity confers cell immortality through stabilization of the chromosome, participating in the development of a majority of human cancers. Human telomerase reverse transcriptase (TERT) has been identified as a catalytic subunit of telomerase, and is overexpressed in most gastric carcinomas. We immunohistochemically examined the expression of TERT in normal gastric mucosa and candidate precancerous lesions such as intestinal metaplasia and adenoma. In non-neoplastic gastric mucosa including intestinal metaplasia and normal fundic mucosa, weak but significant expression of TERT was detected in nuclei of epithelial cells located in the lower two-thirds of the glands (wider than the proliferative zone). The telomerase activity was found in a half of gastric adenomas, whose levels of the activity were about 10% of those in gastric carcinomas. TERT protein was expressed in the nuclei of the adenoma cells at moderate levels, that were not necessarily comparable with the telomerase activities. These findings overall suggest that TERT expression may be one of the prerequisites for telomerase activation in an early stage of stomach carcinogenesis.

Adenoma↗
PubMed1999

Monoclonal antibody to Shiga toxin 2 which blocks receptor binding and neutralizes cytotoxicity.

A monoclonal antibody (MAb) was raised against Shiga toxin 2 (Stx2) of Escherichia coli O157:H7. MAb VTm1.1 belonged to the immunoglobulin G1 subclass and had a kappa light chain, and it could neutralize the cytotoxic activity of Stx2 and variants derived from patient strains but not that of variants derived from animals. MAb VTm1.1 was shown to bind to the B subunit of these neutralized Stx2s by Western blotting. Comparison of B-subunit amino acid sequences and reactivities to these Stxs suggested six amino acids (Ser30, Ser53, Glu56, Gln65, Asn68, and Asp69) that were candidates for the MAb VTm1.1 epitope. Consequently, five Stx2 mutants (S30N, S53N, E56H, Q65K, and N68Ter) were prepared by site-directed mutagenesis to determine which residue is essential for the epitope. All of these mutants showed cytotoxicity almost equal to that of the wild-type Stx2. Of the five Stx2 mutants, only E56H could not be neutralized by MAb VTm1.1. Western blot analysis also showed that MAb VTm1.1 could not bind to the E56H B subunit. These results indicated that Glu56 is an important residue recognized by MAb VTm1. 1. Immunofluorescence analysis further indicated that MAb VTm1.1 inhibits the binding of Stx2 to its receptors. MAb VTm1.1 could be a useful therapeutic agent for Shiga toxin-producing E. coli infection.

Amino Acid Sequence↗
PubMed1999

Evidence for the synthesis of corticosteroid-binding globulin in human placenta.

We demonstrated the expression of corticosteroid-binding globulin (CBG) in human placenta using reverse transcription-polymerase chain reaction-Southern blot analysis and immunohistochemical and immunoblotting studies. In the RT-PCR-Southern blot analysis, only one predicted PCR product was detected without nonspecific products in all samples of human placenta and 3A (tPA-30-1) human placental cells. In Western blot analysis, polyclonal anti-CBG antibodies recognized a protein of approximately 55 kD in the protein extracts prepared from 3A (tPA-30-1) cells. Additionally, CBG mRNA expression was demonstrated by in situ hybridization in the syncytiotrophoblasts. Immunohistochemical studies performed on the placenta demonstrated the presence of specific immunoreactivity in the syncytiotrophoblast layer surrounding the chorionic villi. These findings suggest that CBG is synthesized in human placenta during pregnancy in addition to its synthesis in the liver.

Adult↗
PubMed1999

Dominant expression of progesterone receptor form B mRNA in ovarian endometriosis.

This study was designed to examine the biological implications of progesterone receptor form A (PR-A) and B (PR-B) mRNA expressions in human ovarian endometriosis (ectopic endometrium). A high ratio of PR-B to PR-AB (PR-A+PR-B) mRNA expression was found in 8 of 14 cases of endometriosis, compared with the ratio in eutopic endometrium. The mean ratio in ectopic endometria was significantly (p < 0.01) higher than in eutopic endometria. The ratio in eutopic and ectopic endometria showed no significant change during the menstrual cycle. The mean ratio in ectopic endometria in the proliferative and secretory phases of the endometrium was significantly (p < 0.01) higher than in eutopic endometria. In conclusion, PR-B mRNA was relatively highly expressed in some endometriomas, which might lead to aberrations in the control of progestational effects involving responsiveness to sex steroidal growth regulation.

Adult↗
PubMed1999

Angiogenesis in endometriosis and angiogenic factors.

Among angiogenic factors, VEGF secreted from activated macrophages under the influence of ovarian steroids, IL-8 expressed in endometrial stromal cells, and basic FGF expressed in endometriotic tissue and PD-ECGF expressed in lining epithelial cells independently of the sex steroidal milieu might contribute to the characteristic advancement of angiogenic lesions in endometriosis in individual manners. Copyrightz1999S.KargerAG,Basel

Adult↗
PubMed1999

Progestin regulation in tumor growth of female genital tract cancers.

The irregular response to progestins directly in tumor growth might be caused by dominant negative progesterone receptor (PR) mutants and the damage to PR-A expression. Progestin treatment as an anti-angiogenic therapy would be less effective in the PR-mutated tumors. Therefore, various anti-angiogenic inhibitors must be used in progestin-refractory and progestin-dependent tumors.

Endothelial Growth Factors↗
PubMed1999

The detection of Shiga toxins in the kidney of a patient with hemolytic uremic syndrome.

Infection of Shiga toxin (Stx)-producing Escherichia coli induces hemolytic uremic syndrome (HUS) in 10 to 15% of cases in infants and young children. Although the endothelial cell damage induced by Stx is widely believed to be a primary event of renal dysfunction in HUS, the precise mechanism remains to be elucidated. We were able to examine the kidney obtained at autopsy of a child who died after HUS associated with Stx-producing Escherichia coli O157:H7 infection, and immunohistochemistry indicated the deposition of Stxl and Stx2 in a portion of the distal tubular epithelia. To our knowledge, this is the first report to show the presence of Stx in human tissue of a patient with HUS, and the results obtained in this study provide evidence that Stx indeed migrates into the kidney and binds to renal tubules during Stx-producing Escherichia coli infection.

Bacterial Toxins↗