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Biomedical subjects

K Morikawa

Publications and source records attributed to K Morikawa.

At least 163 records · Page 9Linked to original sources
PubMed1993

Bromocriptine effects on plasma luteinizing hormone and its responses to gonadotropin-releasing hormone in normal men.

Secretory changes in plasma pituitary luteinizing hormone (LH) after administration of a dopaminergic drug were studied in five normal men. Each subject received orally 5 mg of bromocriptine (Brc) daily for 8 weeks. Each received gonadotropin-releasing hormone (GnRH) stimulation tests at the beginning (control), and at 2, 4, 6, and 8 weeks after initiation of Brc treatment. We referred the basal plasma LH and a ratio of maximally GnRH-responded plasma LH to its basal level (R-Max) as indicators of secretory alterations of the LH. Mean basal levels of plasma LH in the five subjects at the beginning and those of R-Max were 3.4 +/- 2.3 (SD) mIU/mL and 8.5 +/- 2.9 units, respectively. Statistically, both the mean values of plasma LH and R-Max during the control period did not differ significantly from those obtained after Brc treatment, although mean basal levels of plasma prolactin during the control period (15.6 +/- 4.6 ng/mL) decreased significantly (p < 0.05) after initiation of treatment. A low dose of Brc administered to normal men for 8 weeks does not significantly influence pituitary secretion of LH.

Adult↗
PubMed1993

Induction of CD5 antigen on human CD5- B cells by stimulation with Staphylococcus aureus Cowan strain I.

We have examined whether the CD5 phenotype could be induced on human B cell surfaces by the polyclonal B cell stimulator, Staphylococcus aureus Cowan strain I (SAC). Fresh tonsillar B cells were prepared by Percoll density gradient from E- cells. The proportion of CD5+ B cells in the 50/60% and 60/70% interface high-density fractions varied between 1.2 and 10.2% depending on the tonsil preparations when they were placed on the in vitro culture 12-60 h prior to flow cytometric analysis. The expression of CD5 antigen obviously increased in the presence of SAC (1:10(5) v/v). The percentage of CD5+ B cells varied from tonsil to tonsil, from 25.1 to 65.9% in a series of experiments. The CD5+ B cells were found both among CD23+CD25+CD71+ and CD23-CD25-CD71- B cells. The level of CD5 expression was related to the cell size enlargement. The addition of anti-CD5 antibody in the culture blocked the CD5 induction by SAC without interfering with the expression of other activation markers. A time-course study showed that CD5 antigen appeared to be induced on the cell surface during the G0 to G1 phase transition in the cell cycle. When CD5+ and CD5- B cells were separated by magnetic isolation, the CD5- B cells showed DNA synthesis to the stimulation by SAC and expressed CD5 antigen on their cell surface. These results suggest that human CD5- B cells can express the CD5 phenotype by stimulation with the polyclonal B cell stimulator, SAC.

Antibodies, Monoclonal↗
PubMed1993

Structural study of mutants of Escherichia coli ribonuclease HI with enhanced thermostability.

Systematic replacement of the amino acid residues in Escherichia coli ribonuclease HI with those in the thermophilic counterpart has revealed that two mutations, His62-->Pro (H62P) and Lys95-->Gly (K95G), increased the thermostability of the protein. These single-site mutant proteins, together with the mutant proteins His62-->Ala (H62A), Lys95-->Asn (K95N) and Lys95-->Ala (K95A), were crystallized and their structures were determined at 1.8 A resolution. The crystal structures of these mutant proteins reveal that only the local structure around each mutation site is essential for the increase in thermostability. For each mutant protein, the stabilization mechanism is considered to be as follows: (i) H62P is stabilized because of a decrease in the entropy of the unfolded state, without a change in the native backbone structure; (ii) K95G is stabilized since the strain caused by the left-handed backbone structure in the typical 3:5 type loop is eliminated; and (iii) K95N is slightly stabilized by a hydrogen bond formed between the side-chain N delta-atom of the mutated aspargine residue and the main-chain carbonyl oxygen within the same residue.

Enzyme Stability↗
PubMed1993

Impaired G-CSF production at post-transcriptional level in a patient with chronic idiopathic neutropenia.

Chronic idiopathic neutropenia (CIN) is a disorder characterized by severe neutropenia and a maturational arrest of the neutrophil precursors in the bone marrow. The pathogenesis of this disorder has been obscure. We examined the production of endogenous G-CSF and the expression of G-CSFmRNA in a patient with adult type CIN. The G-CSF production by the patient's mononuclear cells was deficient in spite of the adequate accumulation of G-CSFmRNA. Our data suggests that the defect in the endogenous G-CSF production at the post-transcriptional level is likely to be an aetiological factor in CIN.

Aged↗
PubMed1993

G-CSF enhances the immunoglobulin generation rather than the proliferation of human B lymphocytes.

The effect of granulocyte-colony stimulating factor (G-CSF) on human B-cell function was studied in in vitro cultures. G-CSF alone had no effect on the proliferative response of resting B cells, but it slightly enhanced the proliferative response of these cells in the presence of polyclonal B-cell mitogen, Staphylococcus aureus Cowan strain I (SAC) at concentrations of 0.2 to 25 micrograms/ml (1.5-fold increase in the DNA synthesis). In contrast, immunoglobulin (Ig) secretion of activated B cells was increased approximately three-fold to four-fold by adding G-CSF to the cultures. The neutralization of G-CSF bioactivity with anti-G-CSF antibody abrogated this effect. Though cytoplasmic Ig-positive cells or plasma cell marker-positive cells did not change, the expression of IgM mRNA in antibody-producing B cells increased in the presence of G-CSF in the cultures. Interestingly, human B lymphocytes are shown to express the binding to biotin-conjugated G-CSF preparation, but not to biotin-conjugated GM-CSF preparation when examined by flow cytometry. These data suggest that G-CSF may influence B-cell function in special circumstances.

Antibodies↗
PubMed1993

Immunosuppressive activity of fosfomycin on human T-lymphocyte function in vitro.

Recent investigations have shown that some antibiotics also work as immunomodulators. We have recently reported that fosfomycin (FOM) has an immunomodulatory effect on human B-cell activation. FOM is a unique antibiotic which is chemically unrelated to any other known antibacterial agent. In the present study, we examined the effect of FOM on human T-cell function. FOM inhibited the proliferation of human lymphocytes induced by polyclonal T-cell mitogens in a dose-dependent manner. FOM also strongly suppressed mixed lymphocyte reaction and interleukin-2 (IL-2) production by T cells. Moreover, FOM inhibited the expressions of IL-2 receptor (CD25) and transferrin receptor (CD71) on the activated T-cell surfaces. These data suggest that FOM may block the T-cell division during the transition from G1 to S phase of the cell cycle. Combined treatment with FOM and low-dose cyclosporin A or FK506 caused additive or synergistic suppression of T-cell proliferation, but not on IL-2 receptor expression. It seems that the mode of action of FOM on T-cell function involves a specific suppression of IL-2 production.

Animals↗
PubMed1993

Immunomodulatory effect of fosfomycin on human B-lymphocyte function.

Fosfomycin (FOM) is an unique antibiotic which is chemically unrelated to any other known antimicrobial agent. Recent investigations have demonstrated that FOM inhibits histamine release from basophils. In this study, we examined the effect of FOM on human B-cell functions. FOM inhibited the proliferative response of resting B cells induced by Staphylococcus aureus Cowan 1 in a dose-dependent manner. FOM interfered with the transition from the G0 to the G1 phase of the cell cycle, leading to cell arrest. The proliferative response of in vivo-activated B cells and lymphokine-induced B-cell proliferation were also affected by FOM. In addition, FOM suppressed immunoglobulin secretion by antibody-producing B cells. Interestingly, FOM did not affect the expression of activation antigens such as the CD25 (interleukin-2 receptor) and CD71 (transferrin receptor) antigens. Moreover, FOM sustained the increased Ia expression on B-cell membranes induced by S. aureus Cowan 1 stimulation, which suggests that FOM may not block the role of B cells in antigen presentation in T-cell-B-cell interaction.

Adjuvants, Immunologic↗
PubMed1993

Stimulatory effects of HSR-803 on ileal motor activity.

Stimulatory effects of HSR-803 on intestinal motor activity in vitro were studied in guinea pig ileum. HSR-803 (1 x 10(-6)-1 x 10(-4) M) increased the amplitude of longitudinal muscle contractions and increased the frequency of peristalsis in isolated segments of guinea pig ileum. The stimulatory effect in amplitude and not frequency was abolished by 1 x 10(-6) M atropine. In the Magnus method with ileal segments, HSR-803 (1 x 10(-7) - 1 x 10(-4) M) produced contractions concentration-dependently, which were inhibited by atropine (1 x 10(-8) and 3 x 10(-8) M) and 3 x 10(-7) M tetrodotoxin (TTX). In the [3H]-quinuclidinyl benzilate (QNB) binding experiment with ileal smooth muscle, HSR-803 had low affinity for acetylcholine (ACh) receptors (pKi = 4.47 +/- 0.04). In addition, HSR-803 failed to increase the spontaneous release and the electrical stimulation-induced [3H]ACh release in ileal smooth muscle. On the other hand, HSR-803 (1 x 10(-5) M) enhanced contractions induced by ACh, but had no effect on contractions induced by carbachol, which is not hydrolyzed by acetylcholinesterase (AChE). In conclusion, HSR-803 stimulated ileal motor activity. However, HSR-803 had low affinity for ACh receptors and had no influence on ACh release. It is likely that HSR-803 stimulated motor activity mainly due to prevention of ACh hydrolysis.

Acetylcholine↗
PubMed1993

[Biosynthesis and secretory regulation of pituitary prolactin].

We summarized recent advancements in biosynthesis and secretory regulation of pituitary prolactin at the cellular and molecular levels from some literatures. We singled out dopamine, estrogen and thyrotropin releasing hormone (TRH) from many factors controlling the biosynthesis and the regulation, because they are important, physiologically and pathologically. Dopamine exerts some effects on the membranes of the lactotrope inhibiting activities of cAMP, phosphatidylinositols and calcium ion channels. It inhibits prolactin gene transcription by suppressing action of the promoter region. Estrogen and TRH increase prolactin biosynthesis and the secretory regulation. Estrogen coupled with the receptors in the cytoplasma goes directly to the DNA region -1713-->-1495 being upstream to the starting site of prolactin transcription and differing from the region on which dopamine acts. TRH accelerates prolactin production and release in such steps as transcription, stabilization of mRNA, accumulation of mRNA in the cytoplasma, protein synthesis and exocytosis.

Animals↗
PubMed1993

[Surgical decompression for the C-1 and C-2 sensory roots and upper cervical cord in a case with cervical myelopathy and occipital neuralgia due to bilateral fenestration of vertebral artery: a case report].

A case of bilateral fenestration of the vertebral artery at the level of the atlas in a patient who had occipital neuralgia and cervical myelopathy is presented. MRI and vertebral angiogram demonstrated the fenestrated vertebral artery compressing the upper cervical cord. Surgical decompression for the C-1 and C-2 sensory roots and the upper cervical cord was performed. Fenestration of the vertebral artery is mostly of no clinical significance. However, considering the pathway of the fenestrated vertebral artery, it is quite possible that the fenestrated vertebral artery might compress the neural structures, resulting in some clinical problems. Although occipital neuralgia may result from a variety of causes, this case was caused by the fenestrated vertebral artery compressing the C-1 and C-2 sensory roots. The authors wish to emphasize that microsurgical vascular decompression may be the only effective treatment in such cases as well as in facial spasm and trigeminal neuralgia.

Aged↗
PubMed1993

[Crystal structure and function of pyrimidine dimer specific excision repair enzyme: T4 endonuclease V].

Bacteriophage T4 endonuclease V is an enzyme which plays an important role in pyrimidine dimer specific excision repair of DNA. This enzyme possesses two distinct catalytic activities, pyrimidine dimer glycosylase and apyrimidinic endonuclease. The three dimensional structure (3D) of the wild type enzyme was determined at 1.45A resolution by X-ray crystallography. In combination with the results of site-directed mutagenesis, the refined structure revealed that Glu23 and the surrounding basic residues constitute the catalytic center of this enzyme. Furthermore, the 3D structure of active site mutants were determined and compared with that of the wild type. The results suggest that a precise configuration of Glu23 residue is required for glycosylation and that Arg3 plays an important role in the substrate binding.

Amino Acid Sequence↗
PubMed1992

Role of the basic amino acid cluster and Glu-23 in pyrimidine dimer glycosylase activity of T4 endonuclease V.

T4 endonuclease V [endodeoxyribonuclease (pyrimidine dimer); deoxyribonuclease (pyrimidine dimer), EC 3.1.25.1] initiates repair of damaged DNA by hydrolysis of the N-glycosyl bond at the 5' side of a pyrimidine photodimer in double-stranded DNA. To study one of the active sites of T4 endonuclease V, systematic site-directed mutagenesis was performed on the synthetic T4 endonuclease V gene, in parallel with three-dimensional structure analysis by x-ray crystallography. The mutant proteins were evaluated for DNA glycosylase activity using an oligonucleotide duplex (14-mer) containing a single thymidine dimer as a substrate. Replacement of either Glu-23 with glutamine or asparatic acid or Arg-3 with glutamine completely abolished DNA glycosylase activity. Mutation of Arg-3 to lysine or of Arg-26 to glutamine or lysine in a basic amino acid cluster caused serious defects in DNA glycosylase activity, which are reflected in the increases in Km and decreases in kcat of DNA glycosylase activity. On the other hand, substitutions of lysine for Arg-22 or of glutamine for Arg-117 or Lys-121 resulted in increases in the Km value. The completely inactive mutant proteins, E23Q and R3Q, in which glutamine was substituted for Glu-23 and Arg-3, respectively, were further investigated by CD spectroscopy for their ability to bind the oligonucleotide substrate. It was found that the E23Q protein retained specific substrate-binding ability, whereas the R3Q protein did not. These results indicate that Glu-23 plays an important role in catalysis of the DNA glycosylase reaction, and that Arg-3 is a crucial residue for substrate binding. In addition, Arg-22, Arg-26, Arg-117, and Lys-121 in the basic amino acid cluster also participate in substrate binding. We conclude that the basic amino acid cluster in T4 endonuclease V is an essential structure for DNA glycosylase activity.

Amino Acid Sequence↗
PubMed1992

X-ray structure of T4 endonuclease V: an excision repair enzyme specific for a pyrimidine dimer.

The x-ray structure of T4 endonuclease V, an enzyme responsible for the first step of a pyrimidine-dimer-specific excision-repair pathway, was determined at a 1.6-angstrom resolution. The enzyme consists of a single compact domain classified into an all-alpha structure. This single domain has two distinct catalytic activities; it functions as a pyrimidine dimer glycosylase and as an apurinic-apyrimidinic endonuclease. The amino-terminal segment penetrates between two major helices and prevents their direct contact. The refined structure suggests the residues involved in the substrate binding and the catalysis of the glycosylation reaction.

Amino Acid Sequence↗
PubMed1992

Inhibitory effect of inaperisone hydrochloride (inaperisone), a new centrally acting muscle relaxant, on the micturition reflex.

We examined the effects of inaperisone hydrochloride (inaperisone), a new centrally acting muscle relaxant, on bladder function in anesthetized rats and isolated rat tissues. We also investigated its mechanism of action. When a balloon inserted into the bladder was expanded, rhythmic bladder contractions were observed; inaperisone (4 mg/kg i.v.) abolished these contractions, in both normal and decerebrated rats. The bladder tonus or bladder contraction induced by peripheral stimulation of the pelvic nerve was barely inhibited by inaperisone (4 mg/kg i.v.), but this dose of inaperisone abolished the efferent discharge from the pelvic nerve that accompanied the rhythmic bladder contractions. The doses of intracerebroventricularly (i.c.v.) and intrathecally injected inaperisone which abolished the rhythmic bladder contractions were 10 and 100 micrograms, respectively. The inhibitory effects of inaperisone (4 mg/kg i.v.) were not diminished by naloxone (1 mg/kg i.v.) or by bicuculline (0.5 mg/kg i.v.), but were diminished by phaclofen (30 mg/kg i.v. or 300 micrograms i.c.v.). The specific binding of [3H]baclofen to rat brain synaptosomal membranes was barely inhibited by inaperisone (up to 1 mM). From these results, it is speculated that, among other possible mechanisms, inaperisone inhibits the micturition reflex by acting indirectly on GABAB receptors in the brainstem.

Anesthesia↗
PubMed1992

Glutamyl-tRNA synthetase from Thermus thermophilus HB8. Molecular cloning of the gltX gene and crystallization of the overproduced protein.

The gene for the Glu-tRNA synthetase from an extreme thermophile, Thermus thermophilus HB8, was isolated using a synthetic oligonucleotide probe coding for the N-terminal amino acid sequence of Glu-tRNA synthetase. Nucleotide-sequence analysis revealed an open reading frame coding for a protein composed of 468 amino acid residues (Mr 53,901). Codon usage in the T. thermophilus Glu-tRNA synthetase gene was in fact similar to the characteristic usages in the genes for proteins from bacteria of genus Thermus: the G + C content in the third position of the codons was as high as 94%. In contrast, the amino acid sequence of T. thermophilus Glu-tRNA synthetase showed high similarity with bacterial Glu-tRNA synthetases (35-45% identity); the sequences of the binding sites for ATP and for the 3' terminus of tRNA(Glu) are highly conserved. The Glu-tRNA synthetase gene was efficiently expressed in Escherichia coli under the control of the tac promoter. The recombinant T. thermophilus Glu-tRNA synthetase was extremely thermostable and was purified to homogeneity by heat treatment and three-step column chromatography. Single crystals of T. thermophilus Glu-tRNA synthetase were obtained from poly(ethylene glycol) 6000 solution by a vapor-diffusion technique. The crystals diffract X-rays beyond 0.35 nm. The crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters of a = 8.64 nm, b = 8.86 nm and c = 8.49 nm.

Adenosine Triphosphate↗
PubMed1992

Structural details of ribonuclease H from Escherichia coli as refined to an atomic resolution.

The crystal structure of RNase H from Escherichia coli has been determined by the multiple isomorphous replacement method, and refined by the stereochemically restrained least-squares procedure to a crystallographic R-factor of 0.196 at 1.48 A resolution. In the final structure, the root-mean-square (r.m.s.) deviation for bond lengths is 0.017 A, and for angle distances 0.036 A. The structure is composed of a five-stranded beta-sheet and five alpha-helices, and reveals the details of hydrogen bonding, electrostatic and hydrophobic interactions between intra- and intermolecular residues. The refined structure allows an explanation of the particular interactions between the basic protrusion, consisting of helix alpha III and the following loop, and the remaining major domain. The beta-sheet, alpha II, alpha III and alpha IV form a central hydrophobic cleft that contains all six tryptophan residues, and presumably serves to fix the orientation of the basic protrusion. Two parallel adjacent helices, alpha I and alpha IV, are associated with a few triads of hydrophobic interactions, including many leucine residues, that are similar to the repeated leucine motif. The well-defined electron density map allows detailed discussion of amino acid residues likely to be involved in binding a DNA/RNA hybrid, and construction of a putative model of the enzyme complexed with a DNA/RNA hybrid oligomer. In this model, a protein region, from the Mg(2+)-binding site to the basic protrusion, covers roughly two turns of a DNA/RNA hybrid double helix. A segment (11-23) containing six glycine residues forms a long loop between the beta A and beta B strands. This loop, which protrudes into the solvent region, lies on the interface between the enzyme and a DNA/RNA hybrid in the model of the complex. The mean temperature factors of main-chain atoms show remarkably high values in helix alpha III that constitutes the basic protrusion, suggesting some correlation between its flexibility and the nucleic acid binding function. The Mg(2+)-binding site, surrounded by four invariant acidic residues, can now be described more precisely in conjunction with the catalytic activity. The arrangement of molecules within the crystal appears to be dominated by the cancelling out of a remarkably biased charge distribution on the molecular surface, which is derived in particular from the separation between the acidic Mg(2+)-binding site and the basic protrusion.

Amino Acid Sequence↗