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Biomedical subjects

E Kawashima

Publications and source records attributed to E Kawashima.

At least 55 records · Page 3Linked to original sources
PubMed1992

Functional characterization of neurokinin-1 receptors on human U373MG astrocytoma cells.

The neurokinin-1 (NK-1, substance P) receptor belongs to the class of seven transmembrane domain (7-TM) receptors that interact with cellular effector systems via guanine nucleotide binding regulatory proteins (G-proteins). In this study, coupling mechanisms of functional NK-1 receptors endogenously expressed in a human astrocytoma cell line (U373MG) were analyzed. Stimulation with substance P (SP) resulted in 1) a rapid increase in inositol 1,4,5-trisphosphate (IP3) synthesis; 2) a rise in cytosolic free calcium concentration ([Ca2+]i); 3) induction of immediate early gene transcription as monitored by c-fos and c-jun expression; and 4) a significant increase in de novo DNA synthesis. Thus, the functional responses induced by stimulation of NK-1 receptors on U373MG strongly correlate with those observed after treatment of primary astrocytes with SP and make U373MG cells a useful in vitro model system for the analysis of NK-1 receptor function on astrocytes in vivo.

Astrocytoma↗
PubMed1992

The effect of acute hypoxia on left ventricular function with special reference to diastolic function--an analysis using ultrasonic method.

In order to evaluate the effect of acute hypoxia on left ventricular (LV) contractility and diastolic function, hemodynamics and LV wall motion were investigated in anesthetized open-chest paced dogs using M-mode or pulsed Doppler echocardiography. Animals were ventilated with 10% oxygen (Hypo 1) and 6.3% oxygen (Hypo 2). LV contractility and diastolic functions were enhanced under "Hypo 1" and at an early phase of "Hypo 2". However, LV functions, both systolic and diastolic, were simultaneously reduced in the presence of hypercapnic acidosis by "Hypo 2". Peak velocities of diastolic rapid filling flow (R) and atrial contraction flow (A) were increased under "Hypo 1", but showed a biphasic change (an increase and a subsequent decrease) under "Hypo 2". The ratio of A/R, known as an index of LV diastolic function, was not altered under hypoxia alone or even under hypercapnic acidosis. Even when hypoxia seems to enhance LV contractility, LV function has already begun to be depressed with a reduction of pH. This seems, however, to be compensated for by LV dilatation and increase in preload, or preservation of left atrial performance.

Acidosis↗
PubMed1991

Recombinant bovine neurokinin-2 receptor stably expressed in Chinese hamster ovary cells couples to multiple signal transduction pathways.

Neurokinins are a family of neuropeptides with widespread distribution mediating a broad spectrum of physiological actions through three distinct receptor subtypes: NK-1, NK-2, and NK-3. We investigated some of the second messenger and cellular processes under control by the recombinant bovine NK-2 receptor stably expressed in Chinese hamster ovary cells. In this system the NK-2 receptor displays its expected pharmacological characteristics, and the physiological agonist neurokinin A stimulates several cellular responses. These include 1) transient inositol 1,4,5-trisphosphate (IP3) formation and Ca2+ mobilization, 2) increased out put of arachidonic acid and prostaglandin E2 (PGE2), 3) enhanced cyclic AMP (cAMP) generation, 4) increased de novo DNA synthesis, and 5) an induction of the "immediate early" genes c-fos and c-jun. Although NK-2 receptor-mediated IP3 formation involves activation of a pertussis toxin-insensitive G-protein, increased cAMP production is largely a secondary response and can be at least partially attributed to autocrine stimulation by endogenously generated eicosanoids, particularly PGE2. This is the first demonstration that a single recombinant neurokinin receptor subtype can regulate, either directly or indirectly, multiple signal transduction pathways and suggests several potential important mediators of neurokinin actions under physiological conditions.

Animals↗
PubMed1991

Seroepidemiological survey of lymphocytic choriomeningitis virus in wild house mouse (Mus musculus) in Yokohama Port, Japan.

From 1985 to 1989, a total of 129 mice was captured from 7 piers of Yokohama port. Of these, 9 (7.0%) were positive to lymphocytic choriomeningitis virus (LCMV) antigen in indirect fluorescence antibody test. Six out of 31 mice (19.4%) in 1985 and 3 out of 23 mice (13.0%) captured in 1986 were positive. All the mice (74) captured in 1988 and 1989 were negative. Although 7 out of 17 mice (41.2%) in Osanbashi-Shinko pier and 2 out of 23 (8.7%) in Honmoku pier were positive in 1985 and 1986, all mice captured in other piers were negative. This is the first report detecting LCMV antibody in wild house mice in Japan.

Animals↗
PubMed1991

[Clinical studies on attic retraction].

Attic retraction was studied in 327 ears of 264 patients in whom photographs of the tympanic membrane were taken from 1983 to 1989. Attic retraction was investigated clinically on the basis of findings of the tympanic membrane in reference to the photographs and ear X-ray findings, including CT. Attic retraction was also observed in 4.5% of cases without inflammation, but its severity was mild. The incidence of attic retraction in cases of adhesive otitis media was 42.7% in mild adhesive otitis media (adhesive grade I) and 60.8% in severe adhesive otitis media (adhesive grade II), both incidences were significantly higher than the incidence of 23.8% in cases of otitis media with effusion. Thus, the incidence of attic retraction increased as the grade of adhesion became severer. In cases of otitis media with effusion, the incidence was significantly higher in the adult group (32.3%) than in the child group (17.4%). In many of the ears with otitis media with effusion, adhesive grade I and chronic otitis media, the severity of attic retraction was mild. There were more cases of milder retraction in the child group than in the adult group. Attic retraction underwent change with time in 2.8% of cases, and cholesteatoma occurred in 1.8% of cases. No improvement in attic retraction was achieved by insertion of a ventilation tube. Bilateral attic retraction was observed in 56.9% of the child group and 30.2% of the adult group, and the difference was statistically significant. Ear X-ray findings by Schüller's method revealed many ears with poorly developed mastoid cells and poor pneumatization.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1991

[A case of right atrial myxoma--the availability of transesophageal echocardiography in the detection of right atrial myxoma].

A 72-year-old woman had experienced palpitation and fatigue during exertion for two months and was referred to our hospital from her nearby hospital. On physical examination, a systolic murmur was heard in the left fourth intercostal space. A chest X-ray film showed cardiac enlargement (CTR 64%). An ECG showed elevated P waves in leads II, III. Transthoracic echocardiography revealed a large oval heterogeneous mass in the right atrium. Transesophageal echocardiography (TEE) revealed the right atrial mass clearly, which was attached to the atrial septum with a short wide stalk. This mass prolapsed from the right atrium into the right ventricle in diastole. And there were some cysts in the homogeneous high-echoic lesion. The chest CT and MRI also showed the mass in the right atrium. However, these images were not clear. Surgical excision of the mass was undertaken. A solid mass measuring 75 x 50 x 45 mm was attached to the fossa ovalis with a wide short stalk. There were several cysts in the mass. Pathological examination showed myxomatous tissue. In this case, TEE was the most valuable means for evaluation of the right atrial mass.

Aged↗
PubMed1990

Human immunodeficiency virus DNA amplification and serology in blood donors.

The significance of indeterminate screening antibody test for human immunodeficiency virus (HIV) serology is still difficult to evaluate, especially in low-risk populations. One hundred twenty-seven blood donors with an initially reactive screening test for HIV antibodies were enrolled in this study. The sera of 95 of these blood donors were reactive on repetition of the test, and none had detectable circulating p24 antigen. Western blot (WB) analysis of the repeatedly reactive sera was as follows: 9 positive, 31 indeterminate, and 55 negative. One of the blood donors with indeterminate WB later presented a seroconversion. On subsequent control 3 to 12 months later, the sera from donors with indeterminate or negative WB did not present any parameters that may indicate a seroconversion. DNA was purified from citrated blood collected from the 127 blood donors at the time of the initial antibody screening. Five micrograms of each DNA sample corresponding to 7 x 10(5) nucleated white blood cells was amplified by polymerase chain reaction (PCR) in the presence of oligonucleotides (primers) corresponding to a highly conserved segment of the pol gene. The detection of amplified DNA was achieved by dot blot and Southern blot using appropriate 32P-labeled oligonucleotides. Ten DNA samples were positive, 9 corresponded to blood donors with a positive HIV serology, and 1 to the blood donor who later presented a seroconversion. These results confirm the sensitivity of the PCR for the diagnosis of HIV infection; they also suggest that repetition of the serology at 3- to 12-month intervals is a valuable procedure for the control of HIV infection status in blood donors.

Base Sequence↗
PubMed1990

Cloning and nucleotide sequence of the Salmonella typhimurium pepM gene.

The pepM gene coding for a methionine-specific aminopeptidase was cloned from Salmonella typhimurium and its nucleotide sequence determined. The gene encoded a 264 amino acid protein that was homologous to a similar protein from Escherichia coli. The sequence of an overproducer mutant allele, pepM100, contained a single base change in the likely--35 region of the pepM promoter that increased its homology to the consensus promoter sequence. A region downstream from the pepM coding sequence contained extensive inverted repeats and was homologous to sequences found elsewhere in both Salmonella and other bacterial species.

Alleles↗
PubMed1990

[A study of ultrasonic scaling in combination with povidone-iodine solution. (1)].

The effects of ultrasonic scaling in combination with povidone-iodine solution were examined clinically. The results obtained were as follows: 1. Three days after instrumentation, significant improvement was found in all experimental groups. Thereafter the improvement was at the same level and backing in each group. 2. Clinical findings, with exception of plaque index, showed improvement with ultrasonic scaling in combination with povidone-iodine solution in comparison with ultrasonic scaling alone and root planing.

Combined Modality Therapy↗
PubMed1989

Effects of synthetic and naturally occurring flavonoids on mitogen-induced proliferation of human peripheral-blood lymphocytes.

Examination was made of the effects of 17 synthetic and naturally occurring flavonoids on human lymphocyte proliferation in the presence of concanavalin A as a mitogen. Twelve of the flavonoids examined were mono-hydroxy or methoxy derivatives. The mitogen-induced response of lymphocytes was evaluated from the extent of the incorporation of [3H]thymidine into cells in vitro. All the compounds showed inhibitory effects; 4.5-77.7% of [3H]thymidine incorporation was blocked by an 1.0 micrograms/ml concentration. The viability of lymphocytes before and after treatment, as assessed by a dye exclusion test, indicated no change, and thus the flavonoids may inhibit DNA synthesis. The flavonoids possessing 5-hydroxyl, 5-methoxyl and 6-methoxyl groups, and those with cyclohexyl instead of phenyl substituent (i.e. 2-cyclohexyl-benzopyran-4-one), showed the greatest inhibition. The inhibitory effect of any one of them was less than one half that of prednisolone, but essentially the same or somewhat exceeding that of bredinine or azathioprine. It would thus appear that the well-known anti-inflammatory effects of flavonoids may possibly arise in part from the inhibition of the proliferative response of lymphocytes.

Chemical Phenomena↗
PubMed1989

Expression of human and murine interleukin-5 in eukaryotic systems.

A cDNA coding for murine interleukin-5 (IL-5) was isolated from the EL4.ExC5 cell line. With the exception of a single amino acid substitution at position 79 (Arg----His), it is identical to a published sequence. The coding sequence for human IL-5 was synthesized chemically, allowing the introduction of strategically located restriction enzyme cleavage sites. Both cDNAs were expressed in various eukaryotic systems. Deletion of the 3' untranslated region of the murine IL-5 gene led to a 5- to 10-fold increase in expression in Xenopus laevis oocytes and in NIH-3T3 cells. The highest production, however, was obtained in Sf9 cells using a baculovirus vector. Human IL-5 was obtained from transformed Saccharomyces cerevisiae as a secreted, mature form using an in-frame fusion to the leader sequence of alpha-mating type factor, and was purified to homogeneity. In all cases mentioned, IL-5 was found to be glycosylated, and its biological activity was dependent on a 40- to 50-kD homodimer configuration, linked together by disulfide bridges. Deglycosylation did not affect the biological activity. Recombinant human IL-5 is biologically active on some human B-CLL cells (proliferation in the presence of IL-2) and on murine BCl1 cells (proliferation) at a low specific activity (about 1-2 x 10(3) U/mg) and on human eosinophils (eosinophil peroxidase assay) at a high specific activity (at least 5 x 10(6) U/mg). Recombinant murine IL-5 from Sf9 cells has a specific activity of 1-2 x 10(7) U/mg in the BCl1 proliferation assay. An additive effect is seen in the presence of murine granulocyte-macrophage colony-stimulating factor (GM-CSF) and a synergistic effect in the presence of murine IL-4.

Animals↗
PubMed1988

DNA containing the base analogue 2-aminoadenine: preparation, use as hybridization probes and cleavage by restriction endonucleases.

The base analogue 2-aminoadenine (2,6-diaminopurine, D) has been introduced at selected positions into synthetic oligodeoxyribonucleotides and DNA by the combined use of chemical and enzymatic methods. 2-aminoadenine substitution for adenine introduces changes in the minor groove of DNA and creates an additional hydrogen bond in the Watson-Crick base pair with thymine. Oligonucleotide hybridization probes containing 2-aminoadenine showed increased selectivity and hybridization strength during DNA-DNA hybridization to phage or genomic target DNA. Properties of the base analogue with respect to DNA modifying enzymes were examined. 2-aminoadenine was used to probe minor groove determinants during the treatment of DNA by 12 restriction endonucleases. Inhibition of cleavage was found for several restriction enzymes.

2-Aminopurine↗
PubMed1987

Uromodulin (Tamm-Horsfall glycoprotein): a renal ligand for lymphokines.

The protein portion of the immunosuppressive glycoprotein uromodulin is identical to the Tamm-Horsfall urinary glycoprotein and is synthesized in the kidney. Evidence that the glycoproteins are the same is based on amino acid sequence identity, immunologic cross-reactivity, and tissue localization to the thick ascending limb of Henle's loop. Nucleic acid sequencing of clones for uromodulin isolated from a complementary DNA bank from human kidney predicts a protein 639 amino acids in length, including a 24--amino acid leader sequence and a cysteine-rich mature protein with eight potential glycosylation sites. Uromodulin and preparations of Tamm-Horsfall glycoprotein bind to recombinant murine interleukin-1 (rIL-1) and human rIL-1 alpha, rIL-1 beta, and recombinant tumor necrosis factor (rTNF). Uromodulin isolated from urine of pregnant women by lectin adherence is more immunosuppressive than material isolated by the original salt-precipitation protocol of Tamm and Horsfall. Immunohistologic studies demonstrate that rIL-1 and rTNF bind to the same area of the human kidney that binds to antiserum specific for uromodulin. Thus, uromodulin (Tamm-Horsfall glycoprotein) may function as a unique renal regulatory glycoprotein that specifically binds to and regulates the circulating activity of a number of potent cytokines, including IL-1 and TNF.

Amino Acid Sequence↗
PubMed1987

Secondary structure at the bacteriophage G4 origin of complementary-strand DNA synthesis: in vivo requirements.

The bacteriophage G4 origin of complementary strand DNA synthesis, G4 ori, contains several regions of potential secondary structure. In this study, we ask whether DNA secondary structure is important for G4 ori function in vivo. Point mutations were generated within a region of potential secondary structure so as to disrupt intrastrand base pairing. These mutations led to a strong temperature-dependent reduction in ori function in vivo. A double point mutation which introduces the same base substitutions without destabilizing intrastrand base pairing did not cause a temperature-dependent disruption in ori function. The double mutant did display a slight temperature-independent reduction in ori function compared to the wild-type G4 ori. Based on these findings, we conclude that DNA secondary structure, as well as recognition of specific sequences, is required for G4 ori activity in vivo.

Coliphages↗
PubMed1986

The genes for tumor necrosis factor (TNF-alpha) and lymphotoxin (TNF-beta) are tandemly arranged on chromosome 17 of the mouse.

We have isolated clones containing the gene for tumor necrosis factor (TNF-alpha) from a mouse genomic library. Four out of five clones containing the TNF-alpha gene also hybridized to a human lymphotoxin (TNF-beta) probe. We constructed a restriction enzyme cleavage map of a 6.4 kb region from one of the genomic clones. From partial sequencing data and hybridizations with exon-specific oligonucleotide probes, we conclude that this region contains the mouse TNF-alpha and TNF-beta genes in a tandem arrangement, that they are separated by only about 1100 bases, and that their intron-exon structure is very similar to that seen in man. We probed genomic blots of DNA from human/mouse hybrids containing single mouse chromosomes for the presence of the mouse TNF genes. The results show that the genes are located on mouse chromosome 17, which also contains the major histocompatibility complex. Therefore, both the mouse and the human TNF genes are tandemly arranged and located on the same chromosome as the MHC.

Animals↗
PubMed1985

Molecular cloning and expression of human tumor necrosis factor and comparison with mouse tumor necrosis factor.

U-937 cells, a monocytic line derived from a human histiocytic lymphoma, were induced for human tumor necrosis factor (TNF) secretion into the medium and were used for the preparation of TNF mRNA. Biological activity of the latter was quantified in a Xenopus laevis oocyte injection system. TNF mRNA was enriched by gradient centrifugation and this size-fractionated mRNA was used for synthesis of cDNA and inserted into the unique PstI site of pAT153. A recombinant plasmid containing human TNF cDNA was selected by colony hybridization using an internal fragment of a mouse TNF cDNA clone [Fransen, L., Mueller, R., Marmenout, A., Tavernier, J., Van der Heyden, J., Kawashima, E., Chollet, A., Tizard, R., Van Heuverswyn, H., Van Vliet, A., Ruysschaert, M. R. & Fiers, W. (1985) Nucleic Acids Res. 13, 4417-4429] as a probe. The sequence of this human TNF cDNA is in agreement with the one published by Pennica et al. [Pennica, D., Nedwin, G. E., Hayflick, J. S., Seeburg, P. H., Derynck, R., Palladino, M. A., Kohr, W. J., Aggarwal, B. B. & Goeddel, D. V. (1984) Nature (Lond.) 312, 724-729]. The 157-amino-acid-long mature sequence is about 80% homologous to mouse TNF and its hydrophilicity plot is also very similar, in spite of the apparent species specificity of TNF. In contrast to mouse TNF, it contains no potential N-glycosylation site. When compared to other cytokines, like IFN-beta, IFN-gamma, or IL-2, there is a remarkably high preference for G X C pairs in the third-letter positions. Expression of the TNF cDNA in monkey COS cells or in Escherichia coli gives rise to a protein having similar biological and serological properties as natural human TNF. A human genomic clone was also identified and sequenced; it was found to be in good agreement with the one recently published by Shirai et al. [Shirai, T., Yamaguchi, H., Ito, H., Todd, C. W. & Wallace, R. B. (1985) Nature (Lond.) 313, 803-806], except for some differences in the introns and 5'-untranslated region.

Amino Acid Sequence↗