Biomedical subjects

K Miki

Publications and source records attributed to K Miki.

At least 289 records · Page 16Linked to original sources

PubMed1991

The nucleotide and deduced amino acid sequences of porcine liver proline- beta-naphthylamidase. Evidence for the identity with carboxylesterase.

A cDNA clone for porcine liver proline-beta-naphthylamidase was isolated and sequenced. The deduced amino acid sequence of 567 residues was highly homologous with those of carboxylesterases (EC 3.1.1.1) previously reported for other species. In addition, proline-beta-naphthylamidase purified from porcine liver was shown to have strong activity towards p-nitrophenylacetate, a representative substrate for carboxylesterases. These results suggest that proline-beta-naphthylamidase is identical with carboxylesterase.

Amino Acid Sequence↗

PubMed1991

Structural and immunological evidence for the identity of prolyl aminopeptidase with leucyl aminopeptidase.

Prolyl aminopeptidase (EC 3.4.11.5) has been assumed to be a unique enzyme catalyzing specifically the removal of unsubstituted NH2-terminal L-prolyl residues from various peptides and to be distinct from leucyl aminopeptidase (EC 3.4.11.1). In the present study, prolyl aminopeptidases were purified to apparent homogeneity from pig small intestine mucosa and human liver and their NH2-terminal amino acid sequences were determined together with that of pig kidney leucyl aminopeptidase. The NH2-terminal 24-residue sequence of pig intestinal prolyl aminopeptidase was shown to be identical with that of pig kidney leucyl aminopeptidase. The NH2-terminal sequence of human liver prolyl aminopeptidase was also shown to be very similar to that of pig kidney leucyl aminopeptidase. Further, pig intestinal prolyl aminopeptidase and pig kidney leucyl aminopeptidase were immunologically indistinguishable. These lines of evidence strongly suggest that prolyl aminopeptidase is identical with leucyl aminopeptidase.

Amino Acid Sequence↗

PubMed1991

Evaluation of x-ray diffraction data from protein crystals by use of an imaging plate.

A system has been developed which uses an imaging-plate diffraction apparatus to obtain structure factors for protein crystals. The latter is connected to a graphics workstation through an Ethernet. Pixel data from an imaging plate are transferred via the Ethernet to a workstation and processed to give the structure factors. The quality and precision of the diffraction data were examined from the point of view of spatial uniformity, linearity with exposure, decay with time, and reproducibility. X-rays from an 55Fe source and Debye-Scherrer rings from Si powder were used for calibration purposes. Finally, diffraction data were obtained and evaluated for lysozyme and cytochrome c-553 using this system. The results were satisfactory in terms of sensitivity and precision.

Protein Conformation↗

PubMed1991

Quantitative and sustained suppression of renal sympathetic nerve activity by left atrial distension in conscious dogs.

In order to investigate the interrelations between left atrial pressure (Pla) and renal sympathetic nerve activity (RSNA) and heart rate (HR), Pla was increased by a balloon both in conscious sham-operated and cardiac-denervated dogs. RSNA was decreased and HR was increased with increasign Pla in sham-operated dogs. The reflex changes in RSNA and HR induced by the stimulation of atrial receptors persisted for at least 15 min. There was a consistent relationship between Pla and RSNA which could be described by a single exponential equation: % delta RSNA = -80.0(1-e-0.395 delta Pla), or by a linear equation when delta Pla was less than 6 mmHg; % delta RSNA = -13.5 delta Pla-2.25. The relationship between Pla and HR can be described by a single exponential equation: delta HR = 86.5(1-e-0.125 delta Pla). A significant linear relationship between RSNA and HR was obtained during a graded left atrial distention (% delta RSNA = -1.08 delta HR-17.7). In cardiac-denervated dogs, RSNA tended to increase and HR remained constant during the left atrial distention. These results indicate that left atrial receptors regulate RSNA and HR in both a quantitative and sustained manner in conscious sham-operated dogs.

Animals↗

PubMed1991

A second group of hepatitis C viruses.

cDNA clone 11-7 was isolated by immunoscreening a cDNA library that was prepared from a pooled plasma of non-A non-B hepatitis (NANBH) patients using expression vector lambda gt11. This cDNA corresponds to known nucleotide positions 3983-4745 of the genome of hepatitis C virus (HCV). This clone was used as a probe for screening the HCV-related cDNAs in a cDNA library similarly prepared by using lambda gt10. As a result, six more cDNA clones were isolated and analyzed for their nucleotide sequences. The results strongly suggested that there are at least two groups of HCV, group I and group II. According to our classification, the prototype HCV and clone 11-7 belong to group I HCV, and their nucleotide and deduced amino acid sequences were diverged from those of group II HCV. Genetic variation observed in the nucleotide and the amino acid sequences between the two groups resembles that in the NS3 region of the genome between Japanese encephalitis virus and West Nile fever virus. Polypeptides produced in Escherichia coli carrying a clone 11-7 or a group II cDNA clone E reacted with antibodies in the blood of 12 or 4 out of 14 individual chronic NANBH patients, respectively. Our data clearly indicate the existence of a second group of HCV.

Amino Acid Sequence↗

PubMed1991

Immunological and virological characterization of improved construction of recombinant vaccinia virus expressing rinderpest virus hemagglutinin.

We constructed a recombinant vaccinia virus (RVV) expressing rinderpest virus (RPV) hemagglutinin (H) by modifying the promoter region of the original RVV. The promotor region was modified at three points, i.e., an outframe ATG was eliminated, the sequence between the promoter and initiation codon was shortened and the base sequence just upstream of the initiation codon was changed. As compared with the original RVV, the modified RVV was found to produce a remarkably large amount of H protein in infected rabbit kidney cells cultured in vitro and to induce high titers of anti-RPV-H antibodies in rabbits. The median protective doses in rabbits of the modified and of the original RVVs were 10(2) pfu and 10(3.5) pfu, respectively, indicating that the modified RVV was at least 10-times more effective in protection than the original. The neurovirulence of the modified RVV and the parental LC16mO strain was roughly at the same level, and was much lower than that of WR strain. The modified RVV was as heat-stable as the original one. These results indicate that the modified RVV could be a candidate rinderpest vaccine for further examinations including cattle.

Animals↗

PubMed1991

Regulation of plaque size and host range by a vaccinia virus gene related to complement system proteins.

A vaccinia virus variant, LC16m8, and its parental Lister (Elstree) strain (LO) were employed to identify the viral gene(s) responsible for plaque size and host range: the large-plaque-forming LO strain but not the small-plaque-forming LC16m8 strain can actively proliferate in Vero (YTV) cells. Previously, we suggested that some particular gene(s) present in the HindIII D fragment of LO DNA was responsible for these biological activities. In the present experiment, the mapping of the putative gene was done by introducing various subfragments of the HindIII D fragment of LO DNA into the gene of LC16m8 strain and screening of the resultant virus variants for the capability of forming large plaques or of proliferating well in Vero cells. The results indicated that an open reading frame (called ps/hr gene) in LO HindIII D fragment was responsible for either plaque size or host range. This gene encoded a polypeptide of 317 amino acids related to the regulators of complement activation (RCA) gene family of mammals. Thus, the present genetic analysis provided direct evidence for a previously unrecognized function of RCA-related proteins encoded by the virus.

Amino Acid Sequence↗

PubMed1991

Crystallization and preliminary X-ray diffraction studies of a flavoprotein, FP390, from a luminescent bacterium, Photobacterium phosphoreum.

A flavoprotein, FP390, obtained from a luminescent bacterium, Photobacterium phosphoreum, in the purification of luciferase has been crystallized by the vapor-diffusion procedure. Crystals obtained from polyethylene glycol 4000 solutions, whose X-ray photographs show powder diffraction patterns, were unsuitable for further crystallographic work. However, tetragonal crystals grown from potassium phosphate solution well diffracted X-rays beyond 3 A resolution. The space group of this crystal is P4(1)22 or P4(3)22 with unit-cell dimensions of a = b = 76.8 and c = 241 A. Assuming two or three molecules in an asymmetric unit, the value for the crystal volume per unit molecular mass, Vm, is calculated as 3.3 or 2.2 A3/Da, respectively. A total of 13,555 independent reflections for the native crystal was collected up to 3 A resolution using a Weissenberg camera attached to the synchrotron radiation source, the merging R factor being 0.077 for 79,335 measurements.

Bacterial Proteins↗

PubMed1991

Protective immunity against bovine leukaemia virus (BLV) induced in carrier sheep by inoculation with a vaccinia virus-BLV env recombinant: association with cell-mediated immunity.

The effects of vaccination of sheep with a recombinant vaccinia virus (rVV) expressing the bovine leukaemia virus (BLV) envelope glycoprotein (gp60) were studied by determining BLV titres in peripheral blood leukocytes after vaccination and challenge. The proliferation of BLV was suppressed markedly, not only when rVV was inoculated prior to challenge with BLV, but also when it was inoculated after challenge. These results indicate that vaccination with rVV induces protective immunity that can suppress the growth of BLV in carrier animals. Since rVV induced a strong anti-BLV delayed-type hypersensitivity response without producing detectable levels of binding or neutralizing antibodies, and there was no apparent correlation between the humoral immune response and BLV proliferation, a cell-mediated immune response was assumed to play a major role in protective immunity.

Animals↗

PubMed1991

Characterization of bullous pemphigoid antibodies by use of recombinant bullous pemphigoid antigen proteins.

The seroreactivity of patients with bullous pemphigoid (BP) to recombinant proteins representing sequences in the carboxyl domain of the murine 230-kD BP antigen (BPA) was determined. Sera from 133 patients with BP, 20 patients with pemphigus, and 21 normal subjects were examined by Western blotting by using two recombinant proteins: RP120 (MW = 120 kD), representing the C-terminal half of the 230-kD BPA, and RP60 (MW = 60 kD), representing the C-terminal quarter. These RP120 and RP60 were recognized by 84% and 61%, respectively, of the BP sera that reacted with the 230-kD BPA in epidermal extract, and not by any of pemphigus or normal sera. Furthermore, these RP120 and RP60 were not recognized by any BP sera that reacted only with the 170-kD BPA, which is known to be another major BPA. These findings indicate that one or more of the major antigenic regions localizes in the carboxyl-half domain of the 230-kD BPA, and also suggest that the 230-kD BPA may be distinct from the 170-kD BPA.

Animals↗

PubMed1991

Inhibitory effect of 8-chloro-cyclic adenosine 3',5'-monophosphate on cell growth of gastric carcinoma cell lines.

A cAMP analogue, 8-chloro-cAMP (8-Cl-cAMP), selectively binds to site 1 receptor of type II regulatory subunit (RII) of cAMP-dependent protein kinase. The effects of 8-Cl-cAMP on human gastric carcinoma cell lines were studied. Twenty microM 8-Cl-cAMP clearly inhibited cell growth in six cell lines (TMK-1, KATO-III, MKN-7, -28, -45, and -74) but not in MKN-1. Cell population in the G1 phase was increased in KATO III cells, which were more responsive to 8-Cl-cAMP, while cell cycle progression in TMK-1 and MKN-1 cells was apparently not influenced by 8-Cl-cAMP. The various changes induced by 8-Cl-cAMP were further analyzed in TMK-1 cells. Decrease of type I regulatory subunit (RI) of cAMP-dependent protein kinase and translocation of RII from cytosol to nucleus were induced by 8-Cl-cAMP treatment. 8-Cl-cAMP increased the level of cAMP-response element (CRE) binding protein in addition to inducing FOS mRNA, whose promoter contains CRE. 8-Cl-cAMP decreased the expression of mRNA for transforming growth factor-alpha (TGF-alpha), while the expression of epidermal growth factor receptor was not changed. Expression of HRAS and MYC mRNAs was slightly increased, whereas the amounts of HRAS and MYC proteins remained unchanged. Our results overall suggest that 8-Cl-cAMP might be a useful tool for antitumor therapy of gastric cancers and that cell growth inhibition by 8-Cl-cAMP might account for the decrease of TGF-alpha expression by tumor cells.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1991

Methylation and expression of human pepsinogen genes in normal tissues and their alteration in stomach cancer.

In normal human tissues, pepsinogen A mRNA was expressed only in the fundic mucosa of the stomach, whereas pepsinogen C mRNA was expressed in all regions of the stomach mucosa and also in the proximal duodenal mucosa. The distributions of these mRNAs were consistent with those of pepsinogens A and C in the gastroduodenal mucosa. Methylation analysis of DNAs from normal tissues with methylation-sensitive restriction enzymes, HpaII and HhaI, revealed that pepsinogen A and C genes are hypomethylated in tissues producing pepsinogens A and C, suggesting a role of DNA methylation in the regulation of the differential expression of the genes for the two human pepsinogens during normal differentiation. In stomach cancer tissues and cancer cell lines, the expressions of the pepsinogen genes were decreased or lost, in good accordance with their pepsinogen productions. No gross structural changes of the pepsinogen genes were observed in these cancers, but the methylation patterns of the pepsinogen genes were found to be altered in different ways in different cancers. The functional significance of the altered methylation is unknown; however, these results suggest that considerable heterogeneity of the methylation patterns occurs in human stomach cancers.

Blotting, Northern↗

PubMed1991

Mass protection program of perinatal hepatitis B virus infection in Japan and impact of an optional booster vaccination on its efficacy.

We assessed the efficacy of a government-sponsored mass protection program in Osaka, Japan, for perinatal HBV infection in infants born to HBeAg positive HBV carrier mothers. We also evaluated the impact of optional follow-up procedures in such infants, including an evaluation of anti-HBs response and a booster dose of HBV vaccine for poor responders. The results demonstrated that this mass protection program protected 94.4% of the infants from perinatal HBV infection in the Osaka area. However, the proportion of infants with an unprotective level of anti-HBs was higher in the standard group than in the follow-up group both at 1.0 and 1.5 years of age, which was also the case for HBV events. Furthermore, the present study showed that a booster dose of vaccine in poor responders was very effective in promoting an anti-HBs response. In conclusion, we recommend that a follow-up blood test to confirm a response of anti-HBs to HBV vaccine should be performed at 4-8 weeks after the third injection of HBV vaccine in infants born to HBeAg positive HBV carrier mothers. We also recommend that a booster injection of HBV vaccine should be immediately given to poor responding infants who otherwise are at a considerable risk of developing HBV infection in late infancy.

Adult↗

PubMed1991

Effect of sho-saiko-to(xiao-chai-hu-tang) on HBeAg clearance in children with chronic hepatitis B virus infection and with sustained liver disease.

We studied the effect of Sho-saiko-to (Xiao-Chai-Hu-Tang) on HBeAg clearance rate (SN rate) in fourteen children with chronic hepatitis B virus (HBV) infection and with sustained liver disease. Seven of fourteen patients (50.0%) became HBeAg negative in the average observation period of 0.47 years(0.2-0.9 years). Four of those seven patients developed anti-HBe. The annual SN rate in the She-saiko-to treated group was apparently higher than the natural annual SN rate (22.7%) of 22 untreated patients retrospectively reviewed from the onset of hepatitis. Sho-saiko-to seemed to promote clearance of HBeAg in children with chronic HBV infection and with sustained liver disease. Sho-saiko-to may be a useful drug for such patients.

Adolescent↗

PubMed1991

Sympathetic nerve activity and renal responses during continuous negative-pressure breathing in humans.

A stretch stimulus of the cardiopulmonary receptors results in a diuresis and natriuresis in dogs due to a suppression of renal sympathetic nerve activity. In the present experiment, the stretch stimulus in humans was given by means of continuous negative-pressure breathing (CNPB), and muscle sympathetic nerve activity (MSNA), renal responses, and hormones were measured to examine whether MSNA response during CNPB correlated with the usual renal and hormonal responses for stretch stimulus of the cardiopulmonary receptors. Nine healthy males were subjected to CNPB at -11 mmHg for 60 min. MSNA in the peroneal muscle nerve fascicles was measured continuously before (pre-CNPB), during, and after CNPB (post-CNPB). A step and sustained decrease (P less than 0.05) in MSNA (30 +/- 6% for burst frequency and 37 +/- 4% for total activity from pre-CNPB level) was observed during CNPB and it returned to pre-CNPB level at post-CNPB. Urinary excretion increased by 58 +/- 18% (P less than 0.05) during CNPB, and the diuresis was entirely osmolal in nature because of a significant increase (P less than 0.05) in Na excretion and a constant free-water clearance. The plasma level of norepinephrine decreased significantly (P less than 0.05) during CNPB and remained decreased (P less than 0.05) at post-CNPB. A twofold increase (P less than 0.05) in plasma atrial natriuretic peptide and a 35 +/- 9% reduction (P less than 0.05) in plasma renin activity were observed during CNPB, whereas no change was observed in plasma aldosterone and arginine vasopressin.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1991

Phosphoramidon-sensitive endothelin converting enzyme in rat lung.

A neutral proteinase with endothelin (ET)-1 converting activity was identified in cytosol and membrane fractions prepared from rat lung, in a ratio of 1:4, respectively. The membrane-bound proteinase was solubilized by 0.5% 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS) with an increase in specific activity, and then was characterized. The solubilized proteinase was capable of converting big ET-1 to ET-1 with an optimum pH of 6.5, and the conversion was dose-dependently suppressed by phosphoramidon (IC50 = 0.5 microM). The molecular mass of the proteinase was estimated to be about 500 kDa by gel filtration in the presence of 0.5% CHAPS. These results indicate that rat lung contains a phosphoramidon-sensitive neutral proteinase catalyzing conversion of big ET-1 to ET-1. The proteinase may be involved in the biosynthetic pathway of ET-1 in the lung and/or the conversion of circulating big ET-1.

Animals↗

PubMed1991

[Morphological changes in the retina following one-shot injection of netilmicin sulfate into the vitreous].

Morphological changes of the retina following one-shot injections into the vitreous of netilmicin sulfate were investigated. The concentrations of netilmicin in the vitreous after intravitreal injection showed the curve of y = a/xb, where y is the concentrations in the vitreous body, x in the number of days since injection and a, b are constant. Concentration decreased rapidly after the injection, and almost totally disappeared after one week. Morphologically, the retina showed normal structure following injections of 100 micrograms/0.1ml and 200 micrograms/0.1ml. Following injections of over 500 micrograms/0.1ml, the outer segment of the photoreceptor disappeared and many phagosomes appeared in the retinal pigment epithelium cell. Following one shot injections of over 500 micrograms/0.1ml damage to the photoreceptor and retinal pigment epithelium was demonstrated. The above data suggested that one-shot injection of netilmicin in concentrations of up to 200 micrograms/0.1ml could be mode without retinal damage.

Animals↗

PubMed1990

Hydrocortisone-induced enhancement of expression and changes in methylation of pepsinogen genes in stomach mucosa of the developing rat.

Administration of hydrocortisone to infant rats caused a precocious increase in levels of mucosal pepsinogen and its mRNA together with morphological maturation of pepsinogen-producing cells. The increase in levels of pepsinogen mRNA was induced rapidly and was associated with increase in levels of its precursors, suggesting transcriptional regulation of pepsinogen genes by hydrocortisone. Methylation analysis with the methylation-sensitive restriction enzymes, HpaII and HhaI, revealed that hydrocortisone also induced sequential demethylation changes of CCGG and GCGC sites in and around pepsinogen genes. Most of these changes occurred after increases in transcription of the genes and did not appear to play a causal role in gene activation. Superficially, the observed demethylations corresponded to the sequential processes of morphological maturation of pepsinogen-producing cells. Thus, these changes in methylation are probably linked to hydrocortisone-induced differentiation of pepsinogen-producing cells and may reflect the mechanism in vivo for the maturation of pepsinogen genes.

Animals↗