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Biomedical subjects

J Fujimoto

Publications and source records attributed to J Fujimoto.

At least 19 recordsLinked to original sources
PubMed1996

Estrogen activates migration potential of endometrial cancer cells through basement membrane.

The migration potential through a basement membrane in an endometrial cancer cell line, such as Ishikawa, HEC-1-A or HHUA cell, in terms of strength, was enhanced by estradiol, but not modified by progesterone, medroxyprogesterone acetate (MPA), danazol or tamoxifen alone, by which estradiol-enhanced migration potential was inhibited. The order of the level of estrogen receptor was Ishikawa > HEC-1-A > HHUA cells. Therefore, it is suggested that the invasiveness of endometrial cancer cells might be activated by estradiol via estrogen receptors, but inactivated by progesterone, MPA, danazol or tamoxifen as an antiestrogen action, and that endometrial cancer cells could become invasive in the estrogen-predominant milieu, and the antiestrogenic agents could protect it.

Antineoplastic Agents, Hormonal↗
PubMed1995

Overexpressions of c-fos/jun mRNA and their oncoproteins (Fos/Jun) in the mouse uterus treated with three natural estrogens.

To further understand hormonal carcinogenesis of natural estrogens (estrone, 17 beta-estradiol (E2) and estriol), we determined the expressions of c-fos/jun mRNA, and their oncoproteins (Fos/Jun) with intracellular localization in the uterus of ovarectomized mice treated with these estrogens. Mid-term chronic, as well as short-term assays were examined. Of three estrogens examined, mid-term chronic E2-treatment significantly increased the expression of c-fos/jun mRNA, and their oncoproteins (Fos/Jun). These were most prominently expressed in glandular cells of E2-treated mouse endometrium. Therefore, mid-term chronic E2-treatment might partially induce glandular cell transformation of uterine endometrium via overexpression of Fos/Jun.

Animals↗
PubMed1995

Increase of solubility of foreign proteins in Escherichia coli by coproduction of the bacterial thioredoxin.

Eukaryotic proteins are frequently produced in Escherichia coli as insoluble aggregates. This is one of the barriers to studies of macromolecular structure. We have examined the effect of coproduction of the E. coli thioredoxin (Trx) or E. coli chaperones GroESL on the solubility of various foreign proteins. The solubilities of all eight vertebrate proteins examined including transcription factors and kinases were increased dramatically by coproduction of Trx. Overproduction of E. coli chaperones GroESL increased the solubilities of four out of eight proteins examined. Although the tyrosine kinase Lck that was produced as an insoluble form and solubilized by urea treatment had a very low autophosphorylating activity, Lck produced in soluble form by coproduction of Trx had an efficient activity. These results suggest that the proteins produced in soluble form by coproduction of Trx have the native protein conformation. The mechanism by which coproduction of Trx increases the solubility of the foreign proteins is discussed.

Animals↗
PubMed1995

Reduced cell motility and enhanced focal adhesion contact formation in cells from FAK-deficient mice.

The intracellular protein tyrosine kinase FAK (focal adhesion kinase) was originally identified gy its high level of tyrosine phosphorylation in v-src-transformed cells. FAK is also highly phosphorylated during early development. In cultured cells it is localized to focal adhesion contacts and becomes phosphorylated and activated in response to integrin-mediated binding of cells to the extracellular matrix, suggesting an important role in cell adhesion and/or migration. We have generated FAK-deficient mice by gene targeting to examine the role of FAK during development. Mutant embryos displayed a general defect of mesoderm development, and cells from these embryos had reduced mobility in vitro. Surprisingly, the number of focal adhesions was increased in FAK-deficient cells, suggesting that FAK may be involved in the turnover of focal adhesion contacts during cell migration.

Animals↗
PubMed1995

Possible role of protein kinase C in the regulation of intracellular stability of focal adhesion kinase in mouse 3T3 cells.

Effects of various types of protein kinase inhibitor on the adhesion and spreading of BALB/c mouse 3T3 cells and on the phosphorylation and stability of focal adhesion kinase (FAK) in the cells were studied. Inhibitors of protein tyrosine kinases, methyl 2,5-dihydroxycinnamate and herbimycin A, inhibited tyrosine-phosphorylation of FAK and the adhesion of 3T3 cells to fibronectin. Among inhibitors of serine/threonine kinases tested, calphostin C, a specific inhibitor of protein kinase C, inhibited cell spreading rather than cell adhesion, and it induced the decrease of intracellular FAK within 30 min. Inhibitors of tyrosine kinase, A kinase, G kinase, and myosin light chain kinase did not induce such a rapid and specific decrease of FAK. When calphostin C (20 microM) was added to sub-confluent monolayer cultures, serine-phosphorylation of FAK was inhibited by 67% within 2 h, and decrease in the amount of FAK and rounding up of the cells began after 4 h. Label-chase experiments indicated that about 60% of 35S-labeled FAK degraded within 1-2 h after addition of calphostin C to monolayer cultures. These results indicated that serine-phosphorylation of FAK induced by protein kinase C was important in the regulation of metabolic stability of FAK.

3T3 Cells↗
PubMed1995

Anaplastic large cell lymphomas expressing the novel chimeric protein p80NPM/ALK: a distinct clinicopathologic entity.

Anaplastic large cell lymphoma (ALCL) is a subtype of non-Hodgkin's lymphoma characterized by the CD30+ large neoplastic cells and sometimes carries a t(2;5)(p23;q35). Recently, we found a novel hyperphosphorylated 80-kD protein tyrosine kinase, p80, in ALCLs with t(2;5). Subsequent cDNA cloning showed p80 to be a fusion protein of two genes, the novel tyrosine kinase gene and the nucleophosmin gene, in accordance with the sequence of the NPM/ALK gene (Morris et al, Science 263:1281, 1994). Meanwhile, the clinicopathologic features of p80-carrying ALCLs have remained unclear. Paraffin sections of 105 cases of ALCL were immunostained using anti-p80 antibody, and 30 of them were shown to express p80. Clinicopathologic comparison between p80-positive and -negative ALCLs showed that p80-positive cases occurred in a far younger patient age group (16.2 +/- 12.9 years; p80-negative cases, 51.0 +/- 22.3 years; P < .0001) and the patients showed a far better 5-year survival rate (79.8%; p80-negative group, 32.9%; P < .01). These data showed that p80-positive ALCL is a distinct entity both clinically and pathogenetically and should be differentiated from p80-negative ALCL.

Adolescent↗
PubMed1995

Focal adhesion kinase is not essential for in vitro and in vivo differentiation of ES cells.

Focal adhesion kinase, FAK, is a unique protein tyrosine kinase found in cellular focal adhesions. It is widely expressed and highly phosphorylated during embryogenesis. To examine the function of FAK in cell differentiation, we made FAK-deficient embryonic stem (ES) cells by homologous recombination. However, FAK-deficiency did not interfere with differentiation of the ES cells into cells of three germ layers when implanted subcutaneously into nude mice or when treated with retinoic acid in vitro, nor was there any evidence of defects in hematopoiesis in vitro.

Animals↗
PubMed1995

Acute hepatitis in rats expressing human hepatitis B virus transgenes.

The molecular mechanisms responsible for hepatocyte death and the events leading to viral clearance in hepatitis B virus (HBV) infections are not well understood. Elucidation of the mechanisms involved have been complicated by the difficulty of infecting human hepatocytes with HBV in vitro and the lack of an appropriate animal model. We report an animal model of human HBV infection by in vivo transfection. We have directly introduced a replication-competent, cloned HBV construct into rat liver by using a membrane fusion-promoting cationic lipid. HBV mRNA and 3.2-kb HBV DNA were expressed in the liver by this in vivo transfection method. In the majority of rats, HBV virions and hepatitis B e antigen were found in the blood 3-7 days after transfection, after which antibody to the e antigen appeared. Two to three weeks after the transfection, glutamic-pyruvic transaminase levels were elevated in serum, hepatocyte death and lymphocyte infiltration were observed in the vicinity of the portal vein of liver, and HBV virions were no longer detected in the serum. Thus, transfection of HBV into rats resulted in histological and serological changes comparable to HBV-induced acute hepatitis in humans. In contrast, no hepatocellular injury was observed in T-lymphocyte-deficient nude rats transfected with the same HBV construct, and viremia was substantially prolonged, providing direct evidence that T lymphocytes play an essential role in liver cell injury and in the clearance of HBV. This rat hepatitis model will be useful for studying pathogenesis of HBV infection.

Acute Disease↗
PubMed1995

Acute onset of diabetic pathological changes in transgenic mice with human aldose reductase cDNA.

To investigate the role of human aldose reductase (hAR) in the pathogenesis of diabetic complications, we generated transgenic mice carrying hAR cDNA driven by the murine MHC class I molecule promoter (hAR-Tg). Northern and Western blot analyses and immunoassay of hAR revealed that both hAR mRNA and the protein were expressed in all tissues tested. Thrombosis in renal vessels and fibrinous deposits in Bowman's capsule were observed in 6-week-old hAR-Tg mice fed a normal diet. Ingestion of a 30% glucose diet for 5 days caused sorbitol concentrations in the liver, kidney, and muscle of hAR-Tg mice to be elevated significantly. Seven-week-old hAR-Tg mice fed a 20% galactose diet for 7 days developed cataracts and occlusion of the retinochoroidal vessels, in addition to pathological changes in the kidney. Despite an elevated aldose reductase level in hAR-Tg mice and their intake of an aldose diet, no histopathological changes were found in other tissues, including the brain, lungs, heart, thymus, spleen, intestine, liver, muscle, spinal cord, or sciatic nerve. Results suggest that target organs of diabetic complications, such as the kidney, lens, and retina are sensitive to damage associated with a high level of AR expression, but other organs are not; the susceptibility of each organ to diabetic complications is determined by not only hAR but also other factors.

Aldehyde Reductase↗
PubMed1995

Are human herpes viruses or measles virus associated with esophageal achalasia?

In order to test the hypothesis that esophageal achalasia may be due to neurotropic viral damage to the esophageal myenteric plexus, esophageal tissue with or without achalasia was analyzed by polymerase chain reaction for the presence of human herpes virus DNA or measles virus RNA. The DNA and RNA were extracted from the esophageal muscle of 12 patients with achalasia and six patients with upper esophageal carcinoma. Peripheral blood mononuclear cells from eight adult volunteers and two samples of umbilical blood mononuclear cells were also used as controls. PCR amplification with a pair of primers specific for herpes simplex type 1 and 2 viruses identified 92-bp fragments in nearly all specimens, including those without achalasia. Each 92-bp fragment was confirmed to be identical to a single herpes simplex virus sequence by automated DNA sequence analysis. No amplification for five other herpes viruses or measles virus was detected. Therefore, a specific viral etiology for achalasia was not identified in this study.

Adult↗
PubMed1995

Danazol decreases transcription of estrogen receptor gene in human monocytes.

1. Administration of danazol for over one month reduced the levels of estrogen receptor (ER) and its mRNA to approximately 50 and 20%, respectively in monocytes. 2. Danazol did not alter the degradation rate of ER mRNA in monocytes. 3. Danazol decreased the transcription rate of ER gene to approximately 50% in monocytes in a run-on assay. 4. Danazol may release estrogen predominance via the reduction of transcription for ER gene, which leads to the reduction of ER mRNA and ER expressions in monocytes.

Adult↗
PubMed1995

Expression of sex hormone-binding globulin mRNA in human endometrial cancers.

To more fully understand the role of sex hormone-binding globulin (SHBG) on the intracellular steroidal action in endometrial cancers, we investigated the expression of SHBG mRNA as the substitute of SHBG expression in human endometrial cancers. In the present study, the levels of SHBG mRNA were analyzed using competitive reverse transcription-polymerase chain reaction (RT-PCR)-Southern-blot analysis. The higher level of SHBG mRNA tended to be expressed in the normal secretory and late proliferative phase endometrium > early proliferative phase endometrium > well differentiated adenocarcinoma of the endometrium (G1) > moderately differentiated adenocarcinoma (G2) > poorly differentiated adenocarcinoma (G3), in the order shown. These studies indicate that endometrial cancer cells might synthesize intracellular SHBG to conserve their estrogen-dependent properties. Further, it indicates that endometrial cancer cell synthesis of SHBG mRNA is lost as these cells undergo de-differentiation.

Adenocarcinoma↗
PubMed1995

Estrogen induces c-Ha-ras expression via activation of tyrosine kinase in uterine endometrial fibroblasts and cancer cells.

Endometrial fibroblasts derived from uterine endometrium as controls and endometrial cancer cells (Ishikawa and HHUA cells) were used to analyze the manner of induction of c-Ha-ras transcripts in endometrial cancers, some of which are estrogen-dependent in growth. Estrogen increased c-Ha-ras expression and tyrosine kinase (TK) activity in fibroblast and Ishikawa cells, but not in HHUA cells. Progesterone diminished c-Ha-ras expression and tyrosine kinase (TK) activity induced by estradiol in the fibroblasts, but not in Ishikawa cells, which persistently overexpressed c-Ha-ras. In these cells, epidermal growth factor (EGF) increased c-Ha-ras expression as did estradiol. Pretreatment with tyrphostin, an inhibitor of TK, abolished estrogen-inducible overexpression of c-Ha-ras. The combination of both estradiol and EGF at maximum effective concentration exerted no additive or synergistic effect on induction of c-Ha-ras expression. In conclusion, persistent activation of TK might lead to overexpression of c-Ha-ras in some endometrial cancer cells under estrogen predominant milieu, which might be associated with the transformation or growth potential.

Adenocarcinoma↗
PubMed1995

Levels of sex hormone-binding globulin (SHBG) and corticosteroid-binding globulin (CBG) messenger ribonucleic acid (mRNAs) in ovarian endometriosis.

Recently, much evidence has indicated that sex hormone-binding globulin (SHBG) and corticosteroid-binding globulin (CBG) play a role in the intracellular action of sex steroids in target cells. In the present work, expression of SHBG mRNA and CBG mRNA was demonstrated in tissues of human normal endometrium and pelvic endometriosis, using the reverse transcription-polymerase chain reaction (RT-PCR). SHBG mRNA levels were higher in pelvic endometriosis than in normal endometrium (P < 0.02), while CBG mRNA levels were lower than in normal endometrium (P < 0.05). The SHBG mRNA/CBG mRNA ratio was significantly higher in pelvic endometriosis than in normal endometrium (P < 0.01). These findings suggest that overexpression of intercellular SHBG in endometriotic tissues results in the formation of the estrogen-predominant milieu, since SHBG-bound estrogen is considered to be protected from the metabolism in liver and available in endometrial cells, thereby assisting the development of the pelvic endometriosis.

Adult↗
PubMed1995

Expression of progesterone receptor form A and B mRNAs in gynecologic malignant tumors.

This study was designed to examine the biological implication of progesterone receptor (PR) forms A and B mRNA expressions in gynecologic cancers. The ratio of PR form A to form B in mRNA expression was approximately 1:1 in all endometria studied. The predominant expressions of form B transcript occurred in 6 out of 7 cases of advanced stages (stages III and IV) in ovarian cancers, in 5 out of 9 cases of cervical cancers, and in 5 out of 11 cases of endometrial cancers. In conclusions, the dominancy of PR form B mRNA expression might be associated with the expression of a malignant phenotype in gynecologic cancers, and advanced clinical stage in ovarian cancers, suggesting a biological marker of malignant phenotype in these three types of cancer cell.

Adult↗
PubMed1995

Induction of M-CSF receptor and its mRNA, and activation of tyrosine kinase in peripheral monocytes by oestradiol-17 beta and progesterone.

In monocytes at the secretory (oestrogen-progesterone dominant) phase of the menstrual cycle, expression of c-fms and macrophage colony-stimulating factor (M-CSF) receptor and activity of tyrosine kinase (TK) were increased by oestradiol with or without progesterone. In vivo, oestrogen may induce expression of c-fms and M-CSF receptor as well as the activation of TK in monocytes under the milieu of the secretory phase. Alternatively, cells of monocyte lineage during the secretory phase might, via various factors, obtain the potency to induce the expression and the function of M-CSF receptors, this potency being effected by oestrogen. Macrophages in peritoneal fluid in pelvic endometriosis (oestrogen predominant) might be activated during the secretory phase of the menstrual cycle, causing infertility.

Adult↗
PubMed1995

Expression of sex hormone-binding globulin mRNA in human ovarian cancers.

To know the role of sex hormone-binding globulin (SHBG) in the intracellular steroidal actions in human ovarian cancers, the expression of SHBG mRNA as a substitute for intracellular SHBG expression was investigated in normal ovarian tissues and ovarian tumors. In the present study, we used competitive reverse transcription-polymerase chain reaction-Southern blot analysis to evaluate SHBG mRNA levels. The expression of SHBG mRNA was detected in all normal ovaries and benign and malignant ovarian tumors analyzed. There were no significant differences in the mean SHBG mRNA levels among the three types of tissue. The expression in normal ovaries was significantly higher (p < 0.01) in premenopause, suggesting the predominance of a sex steroid hormone effect on ovarian SHBG synthesis. Relative overexpression of SHBG mRNA was observed in six out of 22 cases (27%) of ovarian cancer (three cases of endometrioid adenocarcinoma, two cases of serous cystadenocarcinoma and one case of mucinous cystadenocarcinoma) in comparison with normal ovaries and benign ovarian tumors. There was no difference in expression among the clinical stages of ovarian cancers. These data suggest that normal human ovaries and ovarian tumors might synthesize SHBG intracellularly, ovarian cancers might conserve an estrogen-associated property via SHBG and the regulation of intracellular SHBG expression might be changed in some cancers.

Adult↗