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M Hatano

Publications and source records attributed to M Hatano.

At least 19 recordsLinked to original sources
PubMed1992

Hepatocyte growth factor down-regulates the alpha-fetoprotein gene expression in PLC/PRF/5 human hepatoma cells.

Hepatocyte growth factor (HGF) is a potent mitogen for hepatocytes; however, in certain human hepatoma cell lines, the growth is inhibited by HGF. In the present study, the effect of HGF on the alpha-fetoprotein (AFP) gene expression was analyzed in PLC/PRF/5 human hepatoma cells. HGF did not inhibit cell proliferation, but dose-dependently suppressed AFP secretion at the concentrations of 10 ng/ml or less. By Northern blot analysis, the levels of AFP mRNA were suppressed by HGF, whereas the levels of beta-actin mRNA used as a control did not show any significant changes. In the transient chloramphenicol acetyltransferase plasmid transfection assays, the AFP promoter activity was repressed by HGF, in contrast, the AFP enhancer activity was not affected by HGF. These results suggest that the AFP gene expression is down-regulated by HGF through the suppression of its promoter activity in human hepatoma cells.

Animals↗
PubMed1992

Role of I-A molecules in early stages of B cell maturation.

The role of the Ia molecule in the early phase of B cell development remains controversial. In contrast to previous studies, we have detected minute amounts of Ia (I-A) molecule on early B lineage (B220+IgM-) cells from normal bone marrow, using ELISA. The presence of the I-A molecule even on pro-B cells was deduced from experiments in which a monoclonal anti-I-A antibody completely blocked the generation of pre-B cells from B progenitor (B220-) cells in stromal cell-dependent B cell culture. Inasmuch as this antibody did not inhibit the maturation of pre-B cells to IgM+ B cells in culture, the I-A molecule on early B lineage cells probably plays a role in their maturation. We also examined the role of the I-A molecule in early B cell development, using transgenic mice harboring the antisense DNA to I-A beta-chain gene. The amount of I-A molecule on splenic B cells from the young transgenic mice decreased in the presence of abundant amounts of the antisense RNA. B cell development was perturbed in spleen from the transgenic mice. Stromal cell-dependent B cell cultures from these mice clearly showed that the maturation of B lineage cells was delayed at a very early stage of development (B220- to B220+). We propose that the I-A molecule on early B lineage cells may play an essential role in their maturation.

Animals↗
PubMed1992

Role of Glu318 at the putative distal site in the catalytic function of cytochrome P450d.

Most microsomal P450s have a conserved "threonine cluster" composed of three Thrs (Thr319, Thr321, Thr322 for P450d) at a putative distal site. An ionic amino acid at 318 is also well conserved as Glu or Asp for most P450s. To understand the role of these conserved polar amino acids at the putative distal site in the catalytic function of microsomal P450, we studied how mutations at this site of P450d influence the activation of molecular oxygen in the reconstituted system. Catalytic activity (0.02 min-1) toward 7-ethoxycoumarin of the Glu318Ala mutant of P450d was just 6% of that (0.33 min-1) of the wild type, while those of Glu318Asp, Thr319Ala, and Thr322Ala were comparable to or even higher than that of the wild type. Consumption rates of O2 and formation rates of H2O2 of those mutants varied in accord with the catalytic activities. Especially, the efficiency (0.5%) of incorporated oxygen atom to the substrate versus produced H2O2 for the Glu318Ala mutant was much lower than that (3.7%) of the wild type, while that (58.8%) for the mutant Glu318Asp was 16-fold higher than that of the wild type. In addition, the autoxidation [Fe(II)---- Fe(III)] rate (0.074 s-1) of the Glu318Ala mutant was much lower than those (0.374-0.803 s-1) of the wild type and other mutants. Thus, we strongly suggest that Glu318 plays an important role in the catalytic function toward 7-ethoxycoumarin of microsomal P450d.

Amino Acid Sequence↗
PubMed1992

[Relationship between types of hypertension and patterns of urinary catecholamine excretion in Sipple's syndrome].

We report here a case of Sipple's syndrome, and we also analyze the relationship between the type of hypertension and urinary excretion of catecholamines in Sipple's syndrome based on the literature in Japan. One hundred and fourteen cases of Sipple's syndrome have been reported in Japan. The hypertension of patients with Sipple's syndrome shows a ratio of fitfull hypertension to continual hypertension of 6 to 1, whereas the ratio is 1 to 1.5 in patients whose pheochromocytoma is not accompanied by Sipple's syndrome. The patients with Sipple's syndrome, being pheochromocytoma can be classified into the adrenaline (U-AD) dominant type and noradrenaline (U-NA) dominant type based on the catecholamine excretion in the urine. The U-AD predominant (U-AD/U-NA greater than 0.4) patients mostly reveal fitfull hypertension, while patients with continual hypertension hardly show U-AD predominant.

Adrenal Gland Neoplasms↗
PubMed1992

Role of Glu318 and Thr319 in the catalytic function of cytochrome P450d (P4501A2): effects of mutations on the methanol hydroxylation.

Polar amino acids in the (putative) distal site are well conserved in P450s. For example, Glu318 for P450d is well conserved as either Glu or Asp for P450s, and Thr319 for P450d is also conserved for P450s. We have studied how mutations at Glu318 and Thr319 of P450d influence the catalytic activity toward methanol associated with the activation of O2. Catalytic activities of Glu318Asp, Glu318Ala, and Thr319Ala mutants toward methanol were 60, 25, and 38%, respectively, compared with that of the wild type. O2 consumption and NADPH oxidation rates of each mutants varied corresponding to the catalytic activities. However, surprisingly, efficiency (16-40%) of incorporated O to the substrate vs. consumed O2 for the Glu318Ala and Thr319Ala mutants were higher than that (9%) of the wild type. In addition, H2O2, which is produced from uncoupling for the wild-type P450d, was not observed for reaction of the Glu318Ala and Thr319Ala mutants. It seemed that consumed O2 was partially reduced to 2 mol of H2O by 4-electron transfer from NADPH for the wild-type and Thr319Ala mutant. However, for the two Glu318 mutants, it appeared that the consumed O2 was not reduced in the same way. It was thus suggested that the conserved Glu318 and Thr319 of P450d are not essential for the activation of O2 in the methanol oxidation. Role of the water molecule or the methanol molecule in the catalytic function was implied.

Amino Acid Sequence↗
PubMed1992

Effect of mutations at Lys250, Arg251, and Lys253 of cytochrome P450 1A2 on the catalytic activities and the bindings of bifunctional axial ligands.

Some eukaryotic cytochromes P450 (P450s) have a series of ionic amino acids, corresponding to Lys250, Arg251, and Lys253 residues in the P450 1A2 sequence. To understand the roles of those ionic amino acids in the catalytic function of P450, three single mutants, Lys250Leu, Arg251Leu, and Lys253Leu of P450 1A2 were obtained from yeast (Saccharomyces cerevisiae) expression system. Turnover numbers of the Arg251Leu mutant in dealkylation reactions of methoxy- and ethoxyresorufin catalyzed by the P450 reconstituted system were remarkably increased by sixfold compared to those of the wild type. The Lys250Leu and Lys253Leu mutants also showed turnover numbers higher than those of the wild type by three- to fourfold. Those catalytic activities were inhibited competitively by pyridine derivatives, nitrogenous axial ligands to the P450 heme. From those findings together with other spectral data, it was suggested that the ionic site of Lys250, Arg251, and Lys253 may be somehow located near the substrate recognition site and/or near the axial-ligand access channel of this enzyme.

Amino Acid Sequence↗
PubMed1992

Accelerated decay of the cell bound C4b2a complex by serum of patients with membranoproliferative glomerulonephritis and acute poststreptococcal glomerulonephritis.

Serum from patients with membranoproliferative glomerulonephritis (MPGN) and acute poststreptococcal glomerulonephritis (APSGN) accelerated the decay of the cell bound C4b2a (C42) and C4b hemolytic activity relative to pooled normal human serum (pNHS) after 5 min incubation at 30 degrees C in EDTA-GVB. The accelerated decay of the C42 hemolytic activity was heat stable (56 degrees C 30 min) and was inhibited by monoclonal antibody against human C4 binding protein (MoAb:C4BP) or C4 binding protein (C4BP) depleted serum. C4 nephritic factor (C4NeF) was employed to stabilize the labile classical pathway C3 convertase C42 complex. Serum from patients with MPGN and APSGN reduced the C4NeF stabilizing activity. Sera from 32 of 46 patients with MPGN and all of 7 patients with APSGN reduced the C42 hemolytic activity relative to 50 normal human serum (NHS) after 5 min incubation at 30 degrees C in EDTA-GVB, and there was no relationship with the serum concentration of C4BP. In vivo, accelerated decay of C42 convertase might interfere with the clearing and processing mechanism of circulating immune complexes (IC) by reducing deposition of C3b on the IC lattice.

Acute Disease↗
PubMed1992

Binding study of metal ions to S100 protein: 43Ca, 25Mg, 67Zn and 39K n.m.r.

The interactions of the S100 protein (S100) with metal cations such as Ca2+, Mg2+, Zn2+ and K+ were studied by the metal n.m.r. spectroscopy. The line widths of 43Ca, 25Mg, 67Zn and 39K n.m.r. markedly increased by adding all S100s. A broad 43Ca n.m.r. band of Ca(2+)-S100a solution was not affected by Zn2+ and K+, while it was greatly decreased by adding Mg2+. The 43Ca n.m.r. spectra of Ca(2+)-S100a0 and -S100b solutions consisted of two slow-exchangeable signals which corresponded to Ca2+ bound to two environmentally different sites of the S100a0. These two 43Ca n.m.r. signals were not affected by Zn2+ and K+. The line width of broad 25Mg n.m.r. band of the Mg(2+)-S100 solution greatly decreased by adding Ca2+, while it did not change by adding Zn2+ and K+. Further, the addition of Ca2+, Mg2+ and K+ did not affect the line width of the 67Zn n.m.r. of the Zn(2+)-S100 solutions. These findings suggest that: (1) Mg2+ binds to all S100s, and at least one of the Mg2+ binding sites of S100 molecule is the same as the Ca2+ binding site; (2) Zn2+ binds to S100s, although the binding site(s) is/are different from Ca(2+)- or Mg(2+)-binding site(s), and the environment of Zn2+ nuclei will not change even though Ca2+ binds to S100s.

Animals↗
PubMed1992

Effects of L-NG-monomethyl arginine on the cyclic GMP formations in rat mesenteric arteries.

To evaluate the contribution of L-arginine as a precursor of the endothelium-derived relaxing factor (EDRF) on vascular cyclic GMP formation, we examined the effects of L-NG-monomethyl arginine (L-NMMA), and analog of L-arginine, on basal and acetylcholine (ACh)-, sodium nitroprusside (SNP)- and atrial natriuretic peptide (ANP)-induced cyclic GMP formations in rat mesenteric arteries. The mesenteric arteries were perfused with Krebs-Henseleit solution containing 0.2 mM isobutyl methyl xanthine. The effluents from the arteries were collected before and after infusions of graded doses of ACh, SNP or ANP in the absence or presence of 100 microM L-NMMA, and the levels of cyclic GMP were measured. Basal and ACh-induced cyclic GMP formations in the mesenteric arteries were significantly inhibited in the presence of L-NMMA, whereas a concomitant infusion of 300 microM L-arginine restored the inhibition of basal as well as ACh-induced cyclic GMP formations. L-NMMA did not affect SNP- and ANP-stimulated cyclic GMP formations, respectively. These results suggest that L-arginine is necessary for not only the stimulated cyclic GMP formation but also the basal cyclic GMP formation in the mesenteric arteries, whereas the SNP- and ANP-stimulated cyclic GMP formations in the arteries are independent of L-arginine.

Animals↗
PubMed1992

[Elevated production of GM-CSF by peripheral blood mononuclear cells stimulated with mite antigens in children with bronchial asthma].

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine which can stimulate the proliferation and functions of eosinophils and neutrophils. We have studied whether peripheral blood mononuclear cells (PBMC), obtained from children with stable asthma, can produce GM-CSF through stimulation with crude Dermatophagoides farinae (Df) antigen in vitro. The patients had strongly positive RAST results for house dust mite Df. Levels of GM-CSF in PBMC culture supernatant were assayed by an enzyme immunoassay. When stimulated with Df antigen, patient's PBMC released significantly increased levels of GM-CSF compared with unstimulated ones. PBMC of non-allergic controls, however, did not. The addition of polymyxin B, an inactivator of endotoxin, did not suppress the Df-antigen-induced production of GM-CSF. Levels of GM-CSF produced by PBMC stimulated with Df antigens were significantly higher in the patients than in the controls. These results indicate that PBMC of children with Df-allergic asthma, unlike those of non-allergic children, can produce elevated levels of GM-CSF by stimulation with Df antigens.

Animals↗
PubMed1992

Early morphological changes of tubules in rats with chronic renal failure.

We investigated the early changes of tubules and effect of the oral adsorbent, AST-120, on the early changes of tubules in rats with chronic renal failure. Sprague-Dawley rats were divided into two groups with and without AST-120, after 3/4 nephrectomy. Although there were no significant differences in levels of blood urea nitrogen, serum creatinine, creatinine clearance, inulin clearance, para-aminohippuric acid clearance and urinary N-acetyl-beta-D-glucosaminidase at week 8 between the two groups, the amount of 24-hour urinary protein excretion and the direct systolic blood pressure at week 8 were significantly decreased in the group with AST-120. Examinations by light microscopy at week 8 revealed that proteinaceous casts in the tubules, tubular dilatation and infiltration of monocytes into the interstitium in the group with AST-120 were less prominent than those in the group without AST-120. A significant difference in numbers of proteinaceous casts was noted at week 8 between the two groups. In rats with chronic renal failure at the early stage, it is concluded that the formation of proteinaceous casts, resulting in tubular damage, is increased and that AST-120 delays the occurrence of proteinaceous casts by delaying the increase in urinary protein excretion.

Adsorption↗
PubMed1991

Absorption spectral study of cytochrome P450d-phenyl isocyanide complexes: effects of mutations at the putative distal site on the conformational stability.

Interactions of phenyl isocyanide (PheNC) with purified engineered cytochrome P450d wild type and putative distal mutants, Glu318Asp and Glu318Ala, were studied with optical absorption spectra. The wild type and the mutant Glu318Asp were purified as the high-spin state, while the mutant Glu318Ala was purified as the oxygen-bound low-spin form. Thus, it is suggested that Glu318 is important to make the appropriate heme environment of P450d. Spectral dissociation constants (0.19-0.39 mM) of the ligand for the ferric mutants were lower than that (0.74 mM) of the wild type. These dissociation constants were changed by adding a substrate, 7-ethoxycoumarin. The reduced wild type-PheNC complex showed a Soret peak at 451 nm, while the reduced mutant-PheNC complexes showed two peaks at 451 and 423 nm. The 451-nm peak of the complexes decreased with the concomitant increase of a new peak at 433 nm at room temperature. Thus, it was suggested that P450d can take two conformationally different forms from the characteristic spectral features. The Soret spectral conversions which followed the first-order kinetics were analyzed by changing the temperature. The activation energy (69 kcal/mol) for the conversion for the wild type was higher than those (37-50 kcal/mol) for the mutants. The activation energy for the wild type further increased (by 55%) by adding the substrate, while those for the mutants were essentially unchanged by adding the substrate. We discuss the important role of Glu318 at the putative distal site of P450d in the packing or the conformational stability of the putative distal site of the P450d molecule.

Amino Acid Sequence↗
PubMed1991

Deregulation of a homeobox gene, HOX11, by the t(10;14) in T cell leukemia.

Molecular cloning of the t(10;14)(q24;q11) recurrent breakpoint of T cell acute lymphoblastic leukemia has demonstrated a transcript for the candidate gene TCL3. Characterization of this gene from chromosome segment 10q24 revealed it to be a new homeobox, HOX11. The HOX11 homeodomain is most similar to that of the murine gene Hlx and possesses a markedly glycine-rich variable region and an acidic carboxyl terminus. HOX11, while expressed in liver, was not detected in normal thymus or T cells. This lineage-restricted homeobox gene is deregulated upon translocation into the T cell receptor locus where it may act as an oncogene.

Amino Acid Sequence↗
PubMed1991

CO binding studies of engineered cytochrome P-450ds: effects of mutations at putative distal sites in the presence of polycyclic hydrocarbons.

The kinetic parameters of CO binding to genetically engineered cytochrome P-450d (P-450d) and two putative distal mutants, Glu318Asp and Thr322Ala, have been evaluated in the presence and absence of polycyclic hydrocarbons. The dissociation constant (Kd) of CO from wild-type P-450d was decreased by half (from 1.8 microM to approximately 0.9 microM) in the presence of phenanthrene or anthracene but was increased to 11 microM in the presence of 1,2:3,4-dibenzanthracene or 7,8-benzoflavone. These changed Kd values were not altered markedly by mutations at the putative distal site. In contrast, the recombination rate constants (kon) of CO to the Glu318Asp mutant in the presence of phenanthrene (15.5 X 10(5) M-1 s-1) and 7,8-benzoflavone (0.75 X 10(5) M-1 s-1) were much larger than those for the wild type. Similar but smaller increases of the kon values were observed for the Thr322Ala mutant. It was suggested that phenanthrene and anthracene distort the Fe-C-O bond and/or affect the access of CO to wild-type P-450d in an opposite way from 1,2:3,4-dibenzanthracene and 7,8-benzoflavone. Glu318 and Thr322 may be located so close to a CO binding channel in ferrous P-450d that mutations of these residues can open the sterically hindered CO channel caused by the hydrocarbons.

Animals↗
PubMed1991

Probing the role of lysines and arginines in the catalytic function of cytochrome P450d by site-directed mutagenesis. Interaction with NADPH-cytochrome P450 reductase.

To identify amino acids of cytochrome P450d (P450d) which participate in the interaction with NADPH-cytochrome P450 reductase, we changed conserved ionic amino acids of P450d to others by site-directed mutagenesis. Turnover numbers (0.032-0.008 min-1) of purified mutants Lys94-Glu, Lys99-Glu, Lys105-Glu, Lys440-Glu, Lys453-Glu, Arg455-Glu, and Lys463-Glu toward 7-ethoxycoumarin were much lower than that (0.380 min-1) of the wild type at 25 degrees C. Reduction rates (less than 0.054 s-1) of the heme of all mutants (0.1 microM) in the presence of NADPH and the reductase (0.3 microM) were much lower than that (5.9 s-1) of the wild type. Furthermore, a turnover number (0.042 min-1) of a microsomal triple mutant (Arg135-Leu + Arg136-Leu + Arg137-Leu) of a conserved Arg cluster was much lower than that (0.674 min-1) of the wild type at 37 degrees C. Thus, we suggest that Lys94, Lys99, Lys105, Lys440, Lys453, Arg455, Lys463, and perhaps the Arg cluster Arg135-Arg136-Arg137 of P450d will participate in the intermolecular electron transfer process by forming ionic bridges between the two proteins and/or by orienting appropriate geometry for electron transfer on the interfacial surface between the two proteins.

Amino Acid Sequence↗