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Biomedical subjects

T Gomi

Publications and source records attributed to T Gomi.

At least 91 records · Page 5Linked to original sources
PubMed1993

[Acetazolamide effect on vascular response in crossed cerebellar diaschisis as measured by 99mTc-HMPAO SPECT].

We studied the effect of acetazolamine (DIAMOXR) on vascular response in areas with crossed cerebellar diaschisis (CCD) using consecutive 99mTc-hexamethyl-propyleneamine oxime (HMPAO) SPECT studies before and after DIAMOX administration with a subtraction method. Regions of interest were drawn over the bilateral cerebellar cortices and interhemispheric asymmetry indices (IAI) were obtained. Eleven of 14 patients with CCD at baseline showed decrease of IAI after DIAMOX administration. Although there is a possibility of underestimation of increase in perfusion in unaffected cerebellar hemisphere, this result suggests a greater increase in perfusion in the affected cerebellar hemisphere as compared with in the contralateral unaffected one and suggests more dilatation of the arterioles in areas with CCD post-DIAMOX than in areas without CCD. However nonsignificant correlation between IAI at baseline and difference of IAI from baseline to post-DIAMOX did not support the hypothesis of more dilatation post-DIAMOX of more constricted arterioles.

Acetazolamide↗
PubMed1992

Rat guanidinoacetate methyltransferase: mutation of amino acids within a common sequence motif of mammalian methyltransferase does not affect catalytic activity but alters proteolytic susceptibility.

1. Manual alignment of amino acid sequences of mammalian S-adenosylmethionine-dependent methyltransferases of known sequence revealed the presence of 2 homologous regions. 2. The sequence of the region at the C-terminal side is unique to mammalian methyltransferases, and in guanidinoacetate methyltransferase this sequence occurs at residues 159-165. 3. Mutagenesis of 5 conserved residues in this sequence did not affect the catalytic activity but altered tryptic susceptibility at Arg20.

Amino Acid Sequence↗
PubMed1992

Hemoglobin Andrew-Minneapolis (beta 144 (HCl) lysine----asparagine) in a Japanese family.

Hemoglobin Andrew-Minneapolis, an abnormal hemoglobin with an asparagine residue substituted for a lysine at position 144 of the beta-globulin chains, was identified in three members of two generations in a Japanese family. The carriers of hemoglobin Andrew-Minneapolis showed false high levels of hemoglobin A1c, as measured by the standard cation exchange high performance liquid chromatography, and a moderate tendency for erythrocytosis. This family is the first report of this abnormal hemoglobin in Japan.

Adult↗
PubMed1991

Recombinant rat guanidinoacetate methyltransferase: structure and function of the NH2-terminal region as deduced by limited proteolysis.

Recombinant rat liver guanidinoacetate methyltransferase, a monomeric protein with Mr 26,000, is inactivated upon incubation with low concentrations of trypsin. Examination of the reaction products by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high-performance liquid chromatography followed by amino acid analysis and sequencing of isolated peptides reveals that the inactivation is due to the cleavage of the NH2-terminal segment after Arg20. The cleaved peptide is not tightly associated with the rest of the protein. The rate of inactivation is not affected by the presence of either S-adenosylmethionine (AdoMet) or guanidinoacetate, but a substantial retardation of inactivation is observed when both substrates are present. The cleavage at Arg20 is also slowed by cross-linking Cys15 and Cys90 by a disulfide bond. An equilibrium binding study shows that guanidinoacetate methyltransferase in the free form binds AdoMet but not guanidinoacetate. The trypsin-modified enzyme, despite having no catalytic activity, can weakly bind AdoMet and guanidinoacetate in the presence of AdoMet. Chymotrypsin rapidly hydrolyzes the peptide bond after Trp19, and elastase cleaves the bond after Ala24, leading in both cases to loss of activity. The results obtained in this study suggest that the portion of the methyltransferase around residues 19-24 is highly exposed to the solvent and flexible. The results also indicate that the NH2-terminal region is not directly involved in substrate binding but plays a role in catalysis.

Amino Acid Sequence↗
PubMed1991

Intermediate and mechanism of hydroxylation of o-iodophenol by salicylate hydroxylase.

Salicylate hydroxylase [EC 1.14.13.1] from Pseudomonas putida catalyzes the hydroxylation of salicylate, and also o-aminophenol, o-nitrophenol, and o-halogenophenols, to catechol. The reactions with these o-substituted phenols comprise oxygenative deamination, denitration, and dehalogenation, respectively. The reaction stoichiometry, as to NADH oxidized, oxygen consumed, and catechol formed, is 2 : 1 : 1, respectively. The mechanisms for the deiodination and oxygenation of o-iodophenol were investigated in detail by the use of I(+)-trapping reagents such as DL-methionine, 2-chlorodimedone, and L-tyrosine. The addition of the traps did not change the molar ratio of catechol formed to NADH oxidized, nor iodinated traps produced were in the incubation mixture. The results suggest that I+ was not produced on the deiodination in the hydroxylation of o-iodophenol. On the other hand, L-ascorbate, L-epinephrine, and phenylhydrazine increased the molar ratio. o-Phenylenediamine decreased it, being converted to phenazine. This suggests that o-benzoquinone is formed in the oxidation of o-iodophenol as a nascent product. The quinone was detected spectrophotometrically by means of the stopped-flow method. Kinetic analysis of the reactions revealed that o-benzoquinone is reduced nonenzymatically to catechol by a second molecule of NADH. A mechanism of elimination for the ortho-substituted groups of substrate phenols by the enzyme is proposed and discussed.

Hydroxylation↗
PubMed1991

Electron-microscopic observations of the alveolar brush cell of the rat.

The alveolar brush cells in the rat lung were observed by transmission electron microscopy. The general structural features of the brush cells in this report were basically in agreement with earlier descriptions. However, this report is the first to detect long, thick microvilli with a unique form protruding into the alveolar lumen in parallel with the basement membrane, not vertically to the basement membrane. Moreover, intranuclear inclusions in the nucleus were observed for the first time in the alveolar brush cells in this study.

Animals↗
PubMed1991

Hydroxylation of o-halogenophenol and o-nitrophenol by salicylate hydroxylase.

Salicylate hydroxylase [EC 1.14.13.1] from Pseudomonas putida catalyzed the formation of catechol from substrate analogues such as o-nitro-, o-amino-, o-iodo-, o-bromo-, and o-chloro-phenol by removing the ortho-substituted groups. They are converted into nitrite, ammonia, and halide ions, respectively. Kinetic parameters of these reactions were determined by spectrophotometric and polarographic methods. Hydroxylation of o-nitro- or o-iodophenol proceeds with the unusual stoichiometry of 2:1:1 for consumed NADH, O2-uptake, and catechol formed. Other ortho-substituted phenols examined also gave the same results. Like salicylate, these substrates perturb the absorption spectrum of salicylate hydroxylase in the visible region, indicating the formation of enzyme.substrate complexes. Titration experiments with ortho-substituted phenols gave the dissociation constants of the complexes. The complexes were quantitatively reduced with NADH or dithionite without detectable formation of the intermediates. The fact that one atom of 18O2 was incorporated into the produced catechol in hydroxylation of o-nitrophenol indicates that the reaction is of monooxygenase nature. It is concluded that salicylate hydroxylase cleaves the C-N and C-X bonds of ortho-substituted phenols.

Catechols↗
PubMed1991

Interleukin 1 alpha, tumor necrosis factor alpha, and interferon gamma in psoriasis.

Although recent studies have suggested that a variety of cytokines released by keratinocytes and inflammatory leukocytes could contribute to induction or persistence of the inflammatory processes in psoriasis, it remains unclear how production of these cytokines is regulated in psoriatic patients. To elucidate the biologic relevance of these cytokines to the pathogenesis of psoriasis, we investigated serum levels of interleukin 1 alpha, tumor necrosis factor alpha, and interferon gamma in 21 patients with psoriasis vulgaris, together with 21 healthy controls. The mean serum levels of interleukin 1 alpha and tumor necrosis factor alpha were not significantly different from those in controls, while those of interferon gamma were significantly elevated in the patients with psoriasis. Serum levels of interleukin 1 alpha correlated negatively with clinical disease severity expressed as psoriasis area and severity index score and with duration of psoriasis. In contrast, interferon gamma levels were related, although not significantly, to disease severity. In addition, an inverse correlation was noted between the interleukin 1 alpha levels and interferon gamma levels. These results indicate that interleukin 1 alpha and interferon gamma may be relevant to the induction and perpetuation, respectively, of the inflammatory responses in psoriasis, and that these cytokines, which have similar biologic properties, may strictly regulate one another's production in vivo.

Adult↗
PubMed1990

Site-directed mutagenesis of rat liver S-adenosylhomocysteinase. Effect of conversion of aspartic acid 244 to glutamic acid on coenzyme binding.

Aspartic acid 244 that occurs at the putative NAD(+)-binding site of rat liver S-adenosylhomocysteinase was replaced by glutamic acid by oligonucleotide-directed mutagenesis. The mutant enzyme was purified to homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Gel permeation chromatography showed that the purified mutant enzyme was a tetramer as is the wild-type enzyme. In contrast to the wild-type enzyme, which possesses 1 mol of tightly bound NAD+ per mol of enzyme subunit, the mutant enzyme had only 0.05 mol of NAD+ but contained about 0.6 mol each of NADH and adenine per mol of subunit. The mutant enzyme, after removal of the bound compounds by acid-ammonium sulfate treatment, exhibited S-adenosylhomocysteinase activity when assayed in the presence of NAD+. From the appearance of activity as a function of NAD+ concentration, the enzyme was shown to bind NAD+ with a Kd of 23.0 microM at 25 degrees C, a value greater than 280-fold greater than that of the wild-type enzyme. In the presence of a saturating concentration of NAD+, the mutant enzyme showed apparent Km values for substrates similar to those of the wild-type enzyme. Moderate decreases of 8- and 15-fold were observed in Vmax values for the synthetic and hydrolytic directions, respectively. These results indicate the importance of Asp-244 in binding NAD+, and are consistent with the idea that the region of S-adenosylhomocysteinase from residues 213 to 244 is part of the NAD+ binding site. This region has structural features characteristic of the dinucleotide-binding domains of NAD(+)- and FAD-binding proteins (Ogawa, H., Gomi, T., Mueckler, M. M., Fujioka, M., Backlund, P.S., Jr., Aksamit, R.R., Unson, C.G., and Cantoni, G.L. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 719-723).

Adenosylhomocysteinase↗
PubMed1989

Human liver serine dehydratase. cDNA cloning and sequence homology with hydroxyamino acid dehydratases from other sources.

Rat liver serine dehydratase cDNA was used to screen a human liver cDNA library in lambda gt11. One positive clone occurred in every 5,000 clones. Fifteen positive clones were plaque purified. The largest cDNA obtained contained an open reading frame of 987 base pairs, and 5' and 3' noncoding regions of 89 and 317 base pairs, respectively. The deduced amino acid sequence, with a calculated Mr of 34,615, was similar to that of rat liver serine dehydratase except for the absence of a segment consisting of 36 amino acid residues. In vitro transcription/translation with the cDNA resulted in the formation of a polypeptide with an Mr of approximately 35,000, which cross-reacted with the anti-rat serine dehydratase antibody. These results suggest that the human serine dehydratase is structurally cognate with the rat enzyme. Moreover, portions of the sequence postulated to be essential for activity in microbial threonine dehydratases are found in the mammalian serine dehydratases, suggesting that hydroxyamino and dehydratases may have originated from a common ancestor.

Amino Acid Sequence↗
PubMed1989

Expression of rat liver S-adenosylhomocysteinase cDNA in Escherichia coli and mutagenesis at the putative NAD binding site.

The cDNA for rat liver S-adenosylhomocysteinase has been cloned, and the nucleic acid sequence has been determined. By comparison of the deduced amino acid sequence for S-adenosylhomocysteinase with that of the dinucleotide binding region for other proteins, the sequence from amino acids 213 to 244 in rat liver S-adenosylhomocysteinase was proposed to be part of the NAD binding site (Ogawa, H., Gomi, T., Mueckler, M. M., Fujioka, M., Backlund, P. S., Jr., Aksamit, R. R., Unson, C. G., and Cantoni, G. L. (1987) Proc. Natl. Acad. Sci. U. S. A. 84, 719-723). A vector has been constructed that expresses S-adenosylhomocysteinase in Escherichia coli in the presence of isopropyl beta-D-thiogalactopyranoside by inserting the coding sequence of rat liver S-adenosylhomocysteinase cDNA downstream from the lac promoter of plasmid pUC118. The enzyme that is produced comprises as much as 10% of the soluble cellular proteins. The purified enzyme is a tetramer, contains 4 mol of tightly bound NAD, and has kinetic properties indistinguishable from those of the liver enzyme. Tryptic peptide mapping and NH2-terminal sequence analysis indicate that the recombinant enzyme is structurally identical to the liver enzyme except for the absence of the NH2-terminal blocking group. The rat liver enzyme has a blocked NH2-terminal alanine residue (Ogawa, H., Gomi, T., Mueckler, M. M., Fujioka, M., Backlund, P. S., Jr., Aksamit, R. R., Unson, C. G., and Cantoni, G. L. (1987) Proc. Natl. Acad. Sci. U. S. A. 84, 719-723). By oligonucleotide-directed mutagenesis mutant vectors have been generated that express proteins in which each of the glycines in the Gly-Xaa-Gly-Xaa-Xaa-Gly sequence of the putative NAD binding site (residues 219-224) was changed to valine. Immunoblot analysis of extracts of the cells transformed with these vectors reveals the presence of immunoreactive proteins with the subunit molecular weight of S-adenosylhomocysteinase. The mutant proteins have no catalytic activity, contain no bound NAD, and do not form the same quaternary structure as the recombinant S-adenosylhomocysteinase.

Adenosylhomocysteinase↗
PubMed1989

Rat liver S-adenosylhomocysteinase. Spectrophotometric study of coenzyme binding.

Rat liver S-adenosylhomocysteinase, a homotetramer, was resolved by treatment with acid ammonium sulfate into apoenzyme and NAD. The apoenzyme thus prepared retained a tetrameric structure but differed in the mobility on nondenaturing polyacrylamide gel electrophoresis. The inactive apoenzyme was reactivated upon incubation with NAD. The restoration of activity paralleled with the tight binding of NAD to apoenzyme, and full activity was obtained when 4 mol of NAD were bound per mol of apoenzyme. The kinetics of reconstitution were apparently biphasic and suggest the existence of two conformers in a slow equilibrium, one of which binds the coenzyme rapidly while the other does so very slowly, if at all. In addition to NAD, apoadenosylhomocysteinase tightly bound nicotinamide hypoxanthine dinucleotide, 3-acetylpyridine adenine dinucleotide and nicotinic acid-adenine dinucleotide. NADP was not bound. Catalytic activity was found only with the enzyme reconstituted with NAD or nicotinamide hypoxanthine dinucleotide. The spectral change observed on interaction of apoadenosylhomocysteinase with NAD was similar to those seen with adenine nucleotides, and was largely approximated by the addition of dioxane to aqueous solutions of adenine nucleotides. By comparison of the difference spectra, it is suggested that the adenine portion of the coenzyme is bound in the hydrophobic pocket of the protein, and that the binding is accompanied by perturbation of tryptophan residue of the protein.

Adenosylhomocysteinase↗
PubMed1989

Fixed drug eruption. Expression of epidermal keratinocyte intercellular adhesion molecule-1 (ICAM-1).

The pathogenic mechanism of preferential localization to certain skin sites of fixed drug eruption lesions has remained unknown. Skin biopsy specimens were obtained from four patients with fixed drug eruptions at various time points after final exposure to the causative drug and were studied immunohistologically using monoclonal antibodies to intercellular adhesion molecule-1 (ICAM-1), lymphocyte function-associated antigen-1, and HLA-DR. The expression of ICAM-1 by keratinocytes was confined exactly to the involved epidermis. In contrast, the expression of HLA-DR by keratinocytes was observed not only in the involved epidermis but also in the uninvolved epidermis, although to a lesser extent. In general, the intensity of expression of ICAM-1 by keratinocytes correlated well with the degree of epidermal invasion of lymphocytes but not with the degree of dermal lymphocytic infiltration. Interestingly, in fixed drug eruption lesions, basal keratinocytes still showed intense reactivity for ICAM-1 6 to 10 days after final exposure to the causative drug, at which time the expression of HLA-DR was already down-modulated. Such a strong dissociation between the expression of ICAM-1 and HLA-DR on lesional keratinocytes was never observed at any time in the normal skin of control patients challenged with dinitrochlorobenzene. These results suggest that localized misregulated expression of ICAM-1 by the keratinocytes may be one factor that explains the preferential site-specificity characteristic of fixed drug eruptions.

Adult↗
PubMed1989

Effects of OKY-046, a selective thromboxane synthetase inhibitor, on blood pressure and thromboxane synthesis in spontaneously hypertensive rats.

The effects of OKY-046, a specific thromboxane (TX) synthetase inhibitor, on blood pressure, urinary TX excretion, TX synthesis in blood platelets, kidney slices and aortic strips, were evaluated in adult spontaneously hypertensive rats (SHR). OKY-046 was dissolved in drinking water at concentrations of 1, 10, 100 mg/dl. The average intakes of OKY-046 were 1.4 +/- 0.1, 13.0 +/- 1.1, and 147 +/- 12 mg/kg/day, in rats who took 1, 10, 100 mg/dl of OKY-046 solutions for drinking water, respectively. The systolic blood pressure was significantly decreased by 34 mmHg only with the high dose of OKY-046 (147 mg/kg/day). OKY-046 suppressed the platelet aggregability to ADP and the release of TX B2, a stable metabolite of TX A2, from blood platelets in a dose-dependent fashion. Urinary excretion of TX B2 decreased significantly in both groups treated with moderate (13.0 mg/kg/day) and high doses of OKY-046 (147 mg/kg/day). The release of TX B2 from kidney slices was decreased only by the high dose of OKY-046, while the release of TX B2 from aortic strips was not changed even by the high dose of OKY-046. OKY-046 had no effect on urinary excretion of 6-keto-prostaglandin F1 alpha, a stable metabolite of prostacyclin, or, on its release from the kidney slices and aortic strips. These results suggest that the effect of OKY-046 on TX synthesis has organ specificity and that the antihypertensive effect of this drug in SHR is related to reduced renal TX synthesis.

6-Ketoprostaglandin F1 alpha↗
PubMed1989

Alteration of vascular thromboxane in rats with subtotal renal ablation.

To assess the roles of vascular prostaglandins in the hypertension of chronic renal failure, the release of prostacyclin and thromboxane (TX) from aorta was evaluated in male Sprague-Dawley rats, the renal mass of which was reduced by removing one kidney and two-thirds of the contralateral kidney ("5/6 nephrectomy"). Five-sixths nephrectomy was followed by significant rises in serum creatinine to 0.55 +/- 0.03 mg/dl and urea nitrogen to 42.9 +/- 3.8 mg/dl, with a concomitant rise in mean blood pressure from 121.6 +/- 1.6 mmHg to 155.3 +/- 8.4 mmHg. In 5/6 nephrectomized rats, the release of TX A2 from aorta, as measured by its stable metabolite TX B2, increased by 60% (p less than 0.01) and prostacyclin, as measured by its stable metabolite 6-keto-prostaglandin, F1 alpha (6-keto-PG F1 alpha) increased by 51% (p less than 0.05). The amounts of both TX B2 and 6-keto-PG F1 alpha released from aorta were closely related to the height of mean blood pressure. These results suggest that the enhanced vasoconstrictor TX production in the vascular walls may be relevant to hypertension in rats with subtotal renal ablation. The adaptive increase in prostacyclin production in the vascular walls may compensate for the elevation of blood pressure due to chronic renal failure in this animal model.

Animals↗
PubMed1989

Participation of the sympathetic nervous system in hypertension in rats with subtotal renal ablation.

To clarify the role of the sympathetic nervous system in the development of hypertension in chronic renal failure, plasma levels and urinary excretions of catecholamines were evaluated in male Sprague-Dawley rats. The renal mass of the rats was reduced by removing one kidney and two-thirds of the contralateral kidney (5/6 nephrectomy). Five-sixths nephrectomy was followed by significant increases in serum creatinine (to 0.55 +/- 0.03 mg/dl) and urea nitrogen (to 42.9 +/- 3.8 mg/dl). There was a concomitant increase in mean blood pressure, measured directly by an implanted aortic catheter, in comparison with control rats (155.3 +/- 8.3 versus 123.6 +/- 3.3 mmHg, P less than 0.01). Both plasma levels and urinary excretion of norepinephrine and epinephrine were elevated in the 5/6-nephrectomized rats compared with controls. Mean blood pressure correlated negatively with 24-h creatinine clearance (r = -0.66, P less than 0.05), and positively with plasma norepinephrine (r = 0.83, P less than 0.01) and urinary excretion of norepinephrine (r = 0.63, P less than 0.05). These results suggest that not only the decrease in renal function, but also hyperactivity of the sympathetic nervous system, may be involved in the pathogenesis of hypertension in rats with subtotal renal ablation.

Animals↗
PubMed1989

In vitro migration of L3T4+ cloned T cells to epidermis: possible role for keratinocyte-derived factors.

Three types of L3T4+ cloned T cells with different antigen specificities, auto-, allo-, and antigen-reactive, were characterized with respect to their migratory potential using an in vitro migration assay under agar gel. Autoreactive T cells, BB5, and alloreactive T cells, SK 1, both of which have been proved to be epidermotropic in vivo, showed specific directional migration to the epidermis, whereas no directional migration was seen with non-epidermotropic cloned T cells and freshly isolated lymph node T cells. Both BB5 and SK 1 cells were equally attracted to all the epidermal fragments tested regardless of their I-A antigens. The directional migration of BB5 cells to the epidermis was significantly inhibited by the co-cultivation with the epidermis, but not the dermis. Studies with cell lines, the conditioned media (CM), and recombinant interleukin (IL) 1, 2, and 3 revealed that BB5 cells were chemotactically attracted to a transformed keratinocyte cell line PAM212 and, to a lesser extent, to the CM from PAM212 and IL-2, but not to IL-1 and IL 3. These results suggest that epidermotropic T cells may be preferentially trapped in an area with a high concentration of keratinocyte-derived growth factors as well as IL-2.

Animals↗