Biomedical subjects

M Kanzaki

Publications and source records attributed to M Kanzaki.

At least 55 records · Page 3Linked to original sources

PubMed1999

Involvement of Smad proteins in the differentiation of pancreatic AR42J cells induced by activin A.

AIMS/HYPOTHESIS: Activin A induces differentiation of amylase-secreting pancreatic AR42J cells into endocrine cells. This study assesses the role of Smad proteins in the actions of activin A in AR42J cells. METHODS: The expression of Smad proteins was determined by northern blotting. Phosphorylation and translocation of Smad2 was measured by transfecting flag-tagged Smad2. Involvement of Smad2 was examined by transfecting cDNA encoding N-terminal and C-terminal domains of Smad2. RESULTS: The mRNAs for Smad2 and Smad4 were abundantly expressed whereas the expression of mRNA for Smad1 and Smad3 was very low. Activin A induced serine-phosphorylation and the subsequent accumulation of the Smad2 in nuclei. Transfection of the N-terminal domain of Smad2, which acts as a dominantly negative mutant (Smad2-N), blocked the morphological changes induced by activin A whereas the C-terminal domain of Smad2, which acts as a constitutively active mutant (Smad2-C), reproduced the activin-induced morphological changes. Similarly, Smad2-N blocked apoptosis induced by activin A and Smad2-C induced apoptosis. Activin A converted AR42J into insulin-secreting cells in the presence of hepatocyte growth factor and introduction of Smad2-N inhibited the differentiation. Smad2-C, however, did not induce differentiation in the presence of hepatocyte growth factor. CONCLUSIONS/INTERPRETATION: Activation of the Smad2 pathway is necessary and sufficient to induce apoptosis and morphological changes. Although activation of the Smad2 pathway is required, it is not solely sufficient for the differentiation of AR42J into endocrine cells.

Activins↗

PubMed1999

Synip: a novel insulin-regulated syntaxin 4-binding protein mediating GLUT4 translocation in adipocytes.

Insulin-stimulated glucose transport and GLUT4 translocation require regulated interactions between the v-SNARE, VAMP2, and the t-SNARE, syntaxin 4. We have isolated a novel syntaxin 4-binding protein, Synip, which specifically interacts with syntaxin 4. Insulin induces a dissociation of the Synip:syntaxin 4 complex due to an apparent decrease in the binding affinity of Synip for syntaxin 4. In contrast, the carboxyterminal domain of Synip does not dissociate from syntaxin 4 in response to insulin stimulation but inhibits glucose transport and GLUT4 translocation. These data implicate Synip as an insulin-regulated syntaxin 4-binding protein directly involved in the control of glucose transport and GLUT4 vesicle translocation.

Adipocytes↗

PubMed1999

Translocation of a calcium-permeable cation channel induced by insulin-like growth factor-I.

Calcium plays a critical part in the regulation of cell growth, and growth factors stimulate calcium entry into cells through calcium-permeable channels. However, the molecular nature and regulation of calcium-permeable channels are still unclear at present. Here we report the molecular characterization of a calcium-permeable cation channel that is regulated by insulin-like growth factor-I (IGF-I). This channel, which we name growth-factor-regulated channel (GRC), belongs to the TRP-channel family and localizes mainly to intracellular pools under basal conditions. Upon stimulation of cells by IGF-I, GRC translocates to the plasma membrane. Thus, IGF-I augments calcium entry through GRC by regulating trafficking of the channel.

3T3 Cells↗

PubMed1999

Klinefelter's syndrome in the male infertility clinic.

The clinical features of patients with Klinefelter's syndrome attending a male infertility clinic have been investigated in order to consider their assisted reproduction treatment options. Over 12 years, a total of 148 patients with sterility due to azoospermia had Klinefelter's syndrome. Eight patients were shown by fluorescence in-situ hybridization (FISH) on metaphase spreads to be mosaic (46,XY/47,XXY), and 140 patients showed only 47,XXY. Small testes were observed in 95% of patients and gynaecomastia was seen in 12.4%. Half of the patients showed hypergonadotrophic hypogonadism, while others showed normogonadism (usually hypergonadotrophic). Spermatozoa were observed in semen from one patient with mosaicism and one without. Three-colour FISH revealed hyperploidy in 2.7% and 2.3% of these spermatozoa respectively. Multiple-site testicular biopsies in five recent patients were performed and yielded a specimen with round and elongated spermatids in one patient with 47,XXY karyotype. This sample was cryopreserved for future intracytoplasmic sperm injection. At follow-up, 46% of couples had chosen artificial insemination with donor sperm, and none had chosen adoption. Two patients developed testicular tumours, one a mature teratoma and the other a Leydig cell tumour. Two patients required androgen replacement therapy.

Adult↗

PubMed1999

Growth hormone regulates steroidogenic acute regulatory protein expression and steroidogenesis in Leydig cell progenitors.

Gonadal development and differentiation is dependent in part on GH, as GH deficiency has been implicated as a cause of lowered fertility and spermatogenic cessation in humans and some biological models. In this study, we demonstrate that GH receptor messenger RNA (mRNA) is preferentially expressed in progenitor Leydig cells (PLCs) isolated and purified from 21-day-old rats. GH induces significant increases in the levels of steroidogenic acute regulatory protein (StAR), 3beta-hydroxysteroid dehydrogenase (3beta-HSD) expression, and androgen production in PLCs. Additionally, the cytokine interferon-gamma (IFNgamma) markedly inhibits GH-stimulated StAR mRNA and protein levels. When cells are cultured with both GH and IFNgamma, IFNgamma decreases the stimulating effect of GH on androgen production. Treatment of PLCs with cycloheximide does not prevent the GH-induced StAR mRNA, indicating that GH induction of StAR transcripts does not require de novo protein synthesis. In contrast, the induction of 3beta-HSD mRNA by GH is altered by cycloheximide treatment. H7, a serine/threonine kinase inhibitor, completely abrogates the increases in StAR mRNA by GH, whereas the tyrosine kinase inhibitor genistein does not. Moreover, GH further enhances StAR and 3beta-HSD mRNA expression in isolated adult rat Leydig cells despite their increased basal expression subsequent to maturational acquisition of these steroidogenic components. These data provide the first demonstration of the direct effects of GH on testicular steroidogenesis during progenitor Leydig cell differentiation.

3-Hydroxysteroid Dehydrogenases↗

PubMed1999

[The use of the Wallstent prosthesis in the treatment of superior vena cava syndrome].

Five patients (4 men and one woman) aged 48-82 years who were suffering from superior vena cava (SVC) syndrome had Wallstent implanted percutaneously to relieve the obstruction. After stent placement, all cases received anticoagulant therapy with heparin. However one transient stent thrombosis occurred, stent placement resulted in complete relief of all patients. Wallstent placement could be an extremely useful treatment for SVC syndrome and should be the treatment of choice in all cases where SVC obstruction occurs or recurs, following chemotherapy or radiotherapy.

Aged↗

PubMed1999

[Clinical trial of fadrozole hydrochloride for postmenopausal patients with recurrent breast cancer].

Eleven recurrent postmenopausal breast cancer patients with osseous or lung metastases were received fadrozole hydrochloride at a dose of 1 mg twice a day for more than 8 weeks. The median disease-free interval of these 11 patients with metastasis was 74 months. Out of 11 evaluable cases, 2 PR, 6 long-NC and 3 PD were observed. The overall response rate was 18.2% and the long-NC rate was 54.5%. The average overall duration of responses and long-NC were 567 days and 573 days, respectively. There was no adverse drug reaction. A combination therapy with fadrozole hydrochloride 2 mg daily and cyclophosphamide 100 mg orally on days 1-14 was given to 14 postmenopausal patients with recurrent breast cancer. The median disease-free interval of these 14 patients with metastasis was 33 months. There were 2 CR, 3 PR, 4 long-NC, 2 NC and 3 PD. The overall response rate and long-NC rate were 35.8% and 28.6%, respectively. The average overall duration of responses and long-NC were 700 days and 443 days, respectively. The adverse drug reactions were anorexia (Grade 2) and neutropenia (Grade 1 and 2). These results suggested that a combination therapy with fadrozole hydrochloride and cyclophosphamide can be effective and contribute to survival time in the treatment of postmenopausal breast cancer.

Aged↗

PubMed1998

Studies on the betacellulin receptor in pancreatic AR42J cells.

Betacellulin is a member of the epidermal growth factor family and converts pancreatic AR42J cells into insulin-producing cells. This study was conducted to characterise the receptor for betacellulin in AR42J cells. AR42J cells expressed two classes of binding sites for radioactive iodine labelled betacellulin, with Kd values of 4.6 x 10(-11) mol/l and 3.0 x 10(-10) mol/l. The binding of [125I]betacellulin was inhibited by unlabelled betacellulin in a dose-dependent manner, but epidermal growth factor was 50 fold less effective than betacellulin. Affinity cross-linking showed a [125I]betacellulin-binding protein with a molecular weight of approximately 180 KDa. When this protein was immunoprecipitated with antibody against epidermal growth factor receptors ErbB-1, ErbB-2, ErbB-3 or ErbB-4, it was immunoprecipitated only by the anti-ErbB-1 antibody. When the [125I]betacellulin-labelled proteins were immunoprecipitated with a combination of the four ErbB antibodies, and the unprecipitated proteins were then immunoprecipitated with anti-phosphotyrosine antibody, a 190 KDa protein was observed. Betacellulin induced the tyrosine phosphorylation of ErbB-1, ErbB-2 and ErbB-4. Finally, while 100 pmol/1 betacellulin converted all of the AR42J into insulin-producing cells in the presence of activin A, 10 nmol/l epidermal growth factor induced differentiation in only about 30 % of the cells. Higher concentrations of epidermal growth factor were less effective. Neu differentiation factor in the presence or absence of epidermal growth factor was ineffective. These results indicate that betacellulin binds to ErbB-1 and possibly another protein with a molecular weight of 190 KDa. The latter betacellulin-binding protein may be involved in the differentiation-inducing activity of betacellulin.

Animals↗

PubMed1998

Stem cell factor in human seminal plasma as a marker for spermatogenesis.

OBJECTIVES: To measure the level of stem cell factor (SCF) in human seminal plasma to determine whether SCF may be useful in evaluating the ability to produce sperm and search the role of SCF in the testes. METHODS: We measured the level of SCF in seminal plasma obtained from 108 males, including idiopathic azoospermia due to germ cell aplasia (n = 10), oligospermia (n = 50), asthenospermia (n = 31), and normospermia (n = 1 7). The expression of SCF messenger ribonucleic acid in the human testis was evaluated by reverse transcriptase polymerase chain reaction (RT-PCR). The correlation between its level and clinical findings was also evaluated. RESULTS: RT-PCR showed a larger form that encoded the soluble protein and a smaller form that encoded the membrane-associated form of SCF in the human testis. The similar ratio of the larger form to the smaller one was observed both in the testis of normal and oligospermic men. The level of SCF is significantly correlated with the sperm count (r = 0.214; P < 0.05). CONCLUSIONS: The level of SCF in seminal plasma appeared to predict the ability to produce sperm. Thus, this factor may play an important role in spermatogenesis.

Humans↗

PubMed1998

Activation of calcium-permeable cation channel by insulin in Chinese hamster ovary cells expressing human insulin receptors.

The present study was conducted to examine the ability of insulin receptor to activate the calcium signaling system in Chinese hamster ovary (CHO) cells expressing human insulin receptor (CHO-IR cells). In these cells, insulin evoked the elevation of cytoplasmic free calcium concentration, [Ca2+]c, measured by using fura-2. Insulin-induced increase in [Ca2+]c was blocked by reducing the extracellular calcium concentration to 1 microM or by adding nickel chloride, an inorganic inhibitor of calcium entry. Insulin did not elevate [Ca2+]c in parental CHO cells or in CHO cells expressing mutant insulin receptor lacking an ATP-binding site. When the transmembrane calcium current was measured by perforated whole-cell patch clamp, adding insulin to the bath solution markedly augmented the inward calcium current. In a cell-attached patch, a single channel activity appeared when insulin was included in the pipette. In contrast, insulin added outside the patch was ineffective. The current/voltage relationship demonstrated that insulin activated a voltage-independent calcium-permeable cation channel with a single-channel conductance of 10 pS. Exposing CHO-IR cells to pertussis toxin abolished the subsequent insulin effect on [Ca2+]c and activation of the calcium-permeable channel. Mastoparan activated the 10-pS calcium-permeable cation channel. In an inside-out patch, insulin activated the calcium-permeable channel when the bath solution contained both GTP and ATP. Nonhydrolyzable ATP could substitute for ATP. These results indicate that in CHO-IR cells, insulin elevates [Ca2+]c by activating the 10-pS calcium-permeable cation channel. Activation by the insulin receptor involves pertussis toxin-sensitive G protein.

Adenosine Triphosphate↗

PubMed1998

Lactogenic hormone-inducible phosphorylation and gamma-activated site-binding activities of Stat5b in primary rat Leydig cells and MA-10 mouse Leydig tumor cells.

The signal transducer and activator of transcription Stat5b has been implicated in signal transduction pathways for a number of cytokines and growth factors, including GH and PRL. Although these lactogenic hormones have the potential to enhance gonadotropin-induced steroidogenesis, the role of GH and PRL in the testis has long been and remains the subject of controversy. In this report we provide, to our knowledge, the first evidence of Stat5b protein expression in the testis and characterize the activation of Stat5b by these lactogenic hormones in primary rat progenitor, immature and adult Leydig cells, and mouse MA-10 Leydig tumor cells. In MA-10 cells, both GH and PRL mediate tyrosine phosphorylation of Janus kinase (JAK) 2 and Stat5b and induce DNA-binding activity of Stat5b. GH enhances both PIE (PRL-inducible element) and Fc gammaRI gamma-activated sites (GAS), but PRL modulates only PIE GAS. In primary Leydig cells isolated from 18-day-old rats, GH, but not PRL, activates cytoplasmic Stat5b and induces the binding of translocated nuclear Stat5b to GAS elements. Although Stat5b protein is expressed in both Percoll- and elutriator-purified adult rat Leydig cells, neither GH nor PRL treatment results in Stat5b-DNA binding. Our studies indicate that the MA-10 cell has the capacity to bind both GH and PRL and provides a useful model system with which to study the distinct testicular roles of these hormones. Moreover, our findings suggest that progenitor and immature Leydig cells are functional targets for GH in the immature rat, suggestive of a role for GH-Stat5b in testicular development. Our data indicate that lactogenic hormone-inducible transcriptional activation may target distinct gene expression in a signaling cascade(s) involving Stat5b but also imply coordinate control by multiple Leydig cell factors.

Animals↗

PubMed1998

Identification and regulation of testicular interferon-gamma (IFNgamma) receptor subunits: IFNgamma enhances interferon regulatory factor-1 and interleukin-1beta converting enzyme expression.

Interferon-gamma (IFNgamma) transmits its signal through a specific cell surface receptor (IFNgammaR), which consists of a primary ligand binding alpha-chain (IFNgammaR alpha) and a signaling beta-chain (IFNgammaR beta). Recent studies identified the cytokines IFNgamma, interleukin-6 (IL-6), IL-1alpha, and tumor necrosis factor-alpha in testicular cells. Therefore, we: 1) examined the expression of IFNgammaR alpha and IFNgammaR beta subunits in freshly isolated and purified rat testicular cells; 2) examined the differential regulation of receptor components by cytokines using primary cultures of Sertoli cells; 3) identified the cell signaling pathway components of testicular IFNgammaR; and 4) characterized the functional role of testicular IFNgamma using primary Sertoli cells. We demonstrated the messenger RNAs for both chains of IFNgammaR in rat testicular cells using Northern hybridization analysis. Western blot analysis and immunocytochemistry showed that both specific IFNgammaR protein subunits were present in cultured primary Leydig and Sertoli cells prepared from the testes of immature rats. The expression of both IFNgammaR component messenger RNAs in cultured Sertoli cells was increased by its specific ligand (IFNgamma), as well as IL-1alpha and tumor necrosis factor-alpha, in both a time- and dose-dependent manner. IFNgamma-activation of the Janus (JAK) tyrosine kinases, JAK1 and JAK2 proteins, indicate that IFNgammaR, expressed in the Sertoli cell, is functional. Moreover, IFNgamma modulates the expression of interferon regulatory factor (IRF)-1 and IL-1beta converting enzyme genes in Sertoli cells. Thus, our data are suggestive of a role(s) for IFN-gamma in the regulation of distinct gene expression and cell-specific sensitivity to apoptosis in the testis.

Animals↗

PubMed1998

Detection of Listeria monocytogenes in humans, animals and foods.

The Listeria monocytogenes-carrying rates were 100% for listeriosis patients and 1.3% for healthy humans. The L. monocytogenes contamination rates for retail sliced beef (34.2%) and pork (36.4%) were significantly higher (p < 0.05) than those for cattle (2.0%) and pigs (0.8%) and for cattle (4.9%) and swine (7.4%) carcasses. The percentages of serotypes 1/2a, 1/2b and 4b which are most dominant in human patients were high in isolates from fresh (90.0%) and processed (100%) fish and shellfish and imported natural cheese (96.7%).

Adolescent↗

PubMed1998

Treatment of patients with retrograde ejaculation in the era of modern assisted reproduction technology.

PURPOSE: We determined a rational strategy for treatment of patients with retrograde ejaculation in the era of modern assisted reproduction technology. MATERIALS AND METHODS: In 7 consecutive patients medical treatment or retrieval of spermatozoa from the bladder was performed at a male infertility clinic. RESULTS: Antegrade ejaculation was restored in 3 patients, and spermatozoa were retrieved from the bladder and used for assisted reproduction in 3. Spermatozoa with good oolemma penetrating ability were collected by seminal vesicle massage. CONCLUSIONS: Modern assisted reproduction technology is a powerful treatment option for retrograde ejaculation when combined with a technique to retrieve spermatozoa of good quality from the bladder.

Adult↗

PubMed1998

[Recent advances in growth factor receptor research].

Growth factors regulate cell growth, differentiation, migration, and apoptosis by stimulating the intrinsic protein kinase activities of their specific cell surface receptors. Most growth factors such as PDGF, EGF, FGF and IGF-I, bind to receptors with tyrosine kinase activity but members of the TGF-beta supergene family bine to those containing serine/threonine kinase activity. Recently, there has been substantial progress in the research field of growth factor receptors and their signaling mechanisms. This review briefly summerizes the tyrosine kinase receptors and also describes recent works in receptors for the TGF-beta supergene family and their signaling mechanisms.

Animals↗

PubMed1997

Thermodynamic Studies of the Adsorbed Films and Micelles of Sodium Taurodeoxycholate.

Surface tension of aqueous solutions was measured for sodium taurodeoxycholate, as a typical example of the bile salt compounds, in the temperature range 20 to 35 degreesC at 2.5 degreesC intervals and concentration range 0 to 7 mmol kg-1. We examined thermodynamic quantities obtainable from the surface tension measurements according to the thermodynamic relations given by K. Motomura [J. Colloid Interface Sci. 64, 348 (1978)]. Sodium taurodeoxycholate was strongly adsorbed and formed the saturated adsorbed film at low concentrations. However, the gaseous/expanded phase transition does not take place in the film. The thermodynamic quantities associated with adsorption did not change as markedly at the critical micelle concentration as those observed for typical surfactants. It was suggested that molecular interactions between sodium taurodeoxycholate molecules in aqueous solutions and adsorbed films are too weak to induce critical changes in the thermodynamic quantities. Copyright 1997 Academic Press.

Journal Article↗

PubMed1997

Regulation of the expression of follistatin in rat hepatocytes.

To elucidate the regulation of follistatin production in the liver, we studied changes in steady-state follistatin mRNA levels in cultured rat hepatocytes. Activin A stimulated follistatin mRNA levels in a time- and concentration-dependent manner. The stimulatory effect of activin A on follistatin mRNA was significant at 2 h, maximal at 6 h and declined thereafter. Incubating the cells with EGF increased follistatin mRNA levels at 48 h and later. The EGF-induced increase in follistatin mRNA was markedly inhibited by exogenous follistatin in the culture medium, which blocks the action of activin A synthesized in hepatocytes, suggesting that endogenous activin A at least partly mediated the effect of EGF. We also examined the effects of transforming growth factor-beta (TGF-beta), glucagon and alpha-adrenergic agonist, phenylephrine, on follistatin mRNA levels. TGF-beta increased the follistatin mRNA to levels similar to those caused by activin A. Phenylephrine and glucagon also increased follistatin mRNA levels but the effects were transient and weaker than those caused by activin A. Finally, follistatin mRNA levels were markedly increased in remnant liver 3 h after 70% hepatectomy. The mRNA remained elevated for up to 72 h. These results indicate that the expression of mRNA for follistatin is positively controlled by activin A, TGF-beta and other hormones or neurotransmitters. The stimulatory effect of EGF on follistatin mRNA is mediated by activin A released from hepatocytes.

Activins↗

PubMed1997

Activation of the calcium-permeable cation channel CD20 by alpha subunits of the Gi protein.

When the calcium-permeable cation channel CD20 is expressed in Balb/c 3T3 cells, it is activated by insulin-like growth factor-I (IGF-I) via the IGF-I receptor (Kanzaki, M., Nie, L., Shibata, H., and Kojima, I. (1997) J. Biol. Chem. 272, 4964-4969). The present study was conducted to investigate the role of G proteins in the regulation of the CD20 channel. In the excised patch clamp mode, activation of the CD20 channel by IGF-I required GTP, Mg2+, and ATP in the bath solution, and removal of either GTP or ATP attenuated the activation. Non-hydrolyzable ATP could substitute for ATP, and guanyl-5'-yl thiophosphate blocked the activation of the channel by IGF-I. The CD20 channel was also activated by guanosine 5'-3-O-(thio)triphosphate, and ATP was not required for the activation. Addition of a preparation of Gi/Go holoprotein purified from bovine brain activated the CD20, and the beta-adrenergic receptor kinase peptide did not affect the number of channel openings induced by the G protein. The CD20 channel was stimulated by the GTP-bound form of recombinant Gi2 alpha subunit purified from Sf9 cells. The Gi3 alpha subunit was less effective, and the Gi1 alpha subunit had no effect. Purified recombinant beta1gamma2 subunits did not affect the activity of the channel. Finally, IGF-I-induced activation of CD20 was inhibited by an antibody against Gi2 alpha subunit. These findings indicate that the CD20 channel expressed in Balb/c 3T3 cells is activated by the IGF-I receptor via the alpha subunits of heterotrimeric G proteins.

3T3 Cells↗