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Biomedical subjects

Y Mitsui

Publications and source records attributed to Y Mitsui.

At least 109 records · Page 6Linked to original sources
PubMed1998

Ca2+-nitric oxide-cGMP signaling in rabbit parotid acinar cells.

Guanosine 3',5'-cyclic monophosphate (cGMP) is a second messenger generated in response to hormones or neurotransmitters in various tissues and cells. In parotid acinar cells, the activation of muscarinic cholinergic and beta-adrenergic receptors induces an increase in intracellular cGMP. However, the mechanism of cGMP production in parotid acinar cells has not been well elucidated. cGMP production is induced by the activation of guanylyl cyclases, which are directly activated by nitric oxide (NO). NO plays an important role as an inter- and intracellular signal molecule in various organs and cells. Biosynthesis of NO is catalyzed by NO synthase (NOS), and NO generation is controlled by the regulation of NOS activity, for example by Ca2+. We have studied the regulation of NOS activity, NO generation and cGMP production in rabbit parotid acinar cells, and have demonstrated a functional Ca2+-NO-cGMP signaling pathway.

Animals↗
PubMed1997

Hyperoxia induces the differentiated neuronal phenotype of PC12 cells by producing reactive oxygen species.

Neurite extension of PC12 cells induced by nerve growth factor (NGF) is a well-known model of neuronal differentiation. In this study, the incubation of PC12 cells in a 50% O2 atmosphere (hyperoxia) caused neurite extension. In these cells, amounts of differentiation-marker proteins, tyrosine hydroxylase and neurofilament M increased. The effects of hyperoxia were inhibited by ascorbic acid or N-acetyl-cysteine, antioxidant reagents, suggesting the involvement of reactive oxygen species (ROS). In support of this, artificial generation of free radicals induced the same effects as hyperoxia. In these cells, total phosphorylation of cellular proteins was enhanced similar to NGF-treated cells. These results suggest that hyperoxia enhances the signal for neuronal differentiation by producing ROS, resulting in the induction of the differentiated neuronal phenotype of PC12 cells.

Animals↗
PubMed1997

Elevated levels of mortalin expression in human brain tumors.

We have performed immunohistochemical studies of mortalin in normal and tumor human brain sections. In normal brain sections, the expression was seen mainly as being confined to neurons. Normal astrocytes showed undetectable expression of this unique member of the heat shock 70 protein family. Three grades of astrocyte tumors (low-grade astrocytoma, anaplastic astrocytoma, and glioblastoma), however, showed an increasing number of mortalin-positive cells. Other types of brain tumors, such as meningiomas, neurinomas, pituitary adenomas, and metastases, also showed elevated levels of mortalin expression compared to those in the normal brain. Mortalin has earlier been reported to have differential intracellular distribution in normal and transformed cells in vitro. Therefore, we substantiated the present study with immunofluorescence localization of the protein in normal and glioblastoma cells. The observations indicated that the tumors might be expressing a nonpancytosolic mortalin. An increase in number of mortalin-positive cells with malignant progression of brain tumors and its correlation with Ki-67 (a cell proliferation marker)-positive cells further suggested an involvement of nonpancytosolic mortalin(s) in malignant transformation of cells in vivo.

Animals↗
PubMed1997

Decrease in amplified telomeric sequences and induction of senescence markers by introduction of human chromosome 7 or its segments in SUSM-1.

Introduction of human chromosome 7 by microcell-mediated chromosome transfer suppresses indefinite division of SUSM-1, an in vitro established human fibroblast line. This cell line has unusually long telomeric sequences although it lacks detectable telomerase activity. Thus, we examined whether such telomeric sequences change upon introduction of chromosome 7 or its segments. In the microcell hybrids that stopped dividing by introduction of chromosome 7, the telomeric sequences were found to be lost or markedly diminished. Introduction of various fragments (2-40 Mb) of chromosome 7 contained in radiation hybrids gave similar results. On the other hand, the telomeric sequences were not altered significantly in the unsuppressed hybrids, a revertant of one suppressed clone, or subclones of SUSM-1 used as controls. In the suppressed microcell hybrids, the distribution of a mortality marker, mortalin, was changed to the cytosolic type of mortal cells from the immortal type of perinuclear fibres. Also, senescence-associated beta-galactosidase was induced to a level similar to that of normally senesced diploid fibroblasts. These results suggest that human chromosome 7 induces senescence in SUSM-1 by suppressing its telomere maintenance mechanism, which does not depend on telomerase.

Animals↗
PubMed1997

The rescuing effect of nerve growth factor is the result of up-regulation of bcl-2 in hyperoxia-induced apoptosis of a subclone of pheochromocytoma cells, PC12h.

The rat pheochromocytoma cell line PC12 is useful for studying neuronal cell differentiation since this cell line differentiates into neuron-like cells in response to nerve growth factor (NGF). We demonstrated that PC12h cells, a subclone of PC12 cells, died under hyperoxia (50% O2). This cell death did not occur in the presence of antioxidant reagents. In the dead cells, DNA fragmentation and chromatin condensation were observed, suggesting that hyperoxia-induced apoptosis via reactive oxygen species (ROS). NGF effectively suppressed this hyperoxia-induced apoptosis. Accordingly, the amounts of bcl-2, a proto-oncogene product, increased in the cells rescued from apoptosis by NGF. Furthermore, bcl-2 antisense oligonucleotide canceled this rescuing effect of NGF. The present findings indicate that NGF rescues PC12h cells from hyperoxia-induced apoptosis via up-regulation of bcl-2.

Animals↗
PubMed1997

Transcriptional squelching by ectopic expression of E2F-1 and p53 is alleviated by proteasome inhibitors MG-132 and lactacystin.

The transcription factors p53 and E2F-1 play important roles in the control of cell cycle progression. In transient transfection experiments, expression of E2F-1, other E2F family members, or p53 squelched transcription from cotransfected plasmids in a dose-dependent manner. Although the proteasome inhibitors MG-132 and lactacystin markedly increased the level of expression of E2F-1 and p53, these inhibitors completely alleviated squelching by both proteins. Several observations indicate MG-132 alleviates squelching by influencing the conformation of newly synthesized p53 and E2F-1:MG-132 increased the fraction of wild type p53 bound by a monoclonal antibody which preferentially recognizes mutant conformers of p53, increased binding of hsp70 to p53 and inhibited nuclear accumulation of both p53 and E2F-1, but not the pocket protein p107. The protease inhibitors ALLN and ALLM did not influence expression of E2F-1 or p53, nor did they alleviate squelching by either transcription factor. Because MG-132 and lactacycstin are highly specific inhibitors of the proteasome protease, our results suggest that the proteasome influences post-translational processes involved in proper folding and cytoplasmic clearing of E2F-1 and p53.

Acetylcysteine↗
PubMed1997

Analysis of expressed sequence tags (ESTs) of the parasitic protozoa Entamoeba histolytica.

A directional cDNA library constructed from mRNA of the trophozoite of Entamoeba histolytica HM-1:IMSS strain was used for the generation of expressed sequence tags (ESTs). From 5' ends of the distinct cDNA clones, 105 ESTs were obtained. Of these, 30 clones (29%) were previously known E. histolytica genes. Forty-five clones (42%) had matches with entries for other organisms in the databases. These new E. histolytica genes constituted a broad range of transcripts distributed among cytoplasmic structural and regulatory proteins, enzymes, nuclear and other proteins, and proteins of unknown function. Thirty clones (29%) had no significant database matches and thus potentially represent E. histolytica-specific genes. These data of E. histolytica genes identified by nucleotide sequencing indicate the value of the adoption of genome sequencing strategies for the rapid increase in knowledge of organisms causing dysentery and liver abscess.

Animals↗
PubMed1997

Expression analysis of mortalin, a unique member of the Hsp70 family of proteins, in rat tissues.

We have investigated the expression of mortalin in rat tissues by Northern analysis, RNA in situ hybridization, and immunohistochemical studies. By Northern assay, the highest level of expression was detected in brain, heart, and skeletal muscle followed by lung, liver, and kidney, and the least level of expression was detected in testis and spleen. RNA in situ and immunohistochemical studies showed that mortalin expression is significantly higher in upper nondividing layers than in the lower basal layers of skin, in neurons and nerve fibers than in surrounding glial cells in brain, in cardiomyocytes than in nonmyocytes in heart, and in interstitial secretory tissue than in germinating follicles in ovary. Such tissue- and cell-specific expression patterns of mortalin coordinates with its earlier reported antiproliferative function in normal cells. However, a deregulation of the expression is observed in rat brain tumor along with the detection of nonpancytosolic mortalin in rat glioma cell line C6. The study suggests that mortalin is involved in pathways that regulate division capacities of cells in vivo.

Animals↗
PubMed1997

Switch of coenzyme specificity of mouse lung carbonyl reductase by substitution of threonine 38 with aspartic acid.

Mouse lung carbonyl reductase, a member of the short-chain dehydrogenase/reductase (SDR) family, exhibits coenzyme specificity for NADP(H) over NAD(H). Crystal structure of the enzyme-NADPH complex shows that Thr-38 interacts with the 2'-phosphate of NADPH and occupies the position spatially similar to an Asp residue of the NAD(H)-dependent SDRs that hydrogen-bonds to the hydroxyl groups of the adenine ribose of the coenzymes. Using site-directed mutagenesis, we constructed a mutant mouse lung carbonyl reductase in which Thr-38 was replaced by Asp (T38D), and we compared kinetic properties of the mutant and wild-type enzymes in both forward and reverse reactions. The mutation resulted in increases of more than 200-fold in the Km values for NADP(H) and decreases of more than 7-fold in those for NAD(H), but few changes in the Km values for substrates or in the kcat values of the reactions. NAD(H) provided maximal protection against thermal and urea denaturation of T38D, in contrast to the effective protection by NADP(H) for the wild-type enzyme. Thus, the single mutation converted the coenzyme specificity from NADP(H) to NAD(H). Calculation of free energy changes showed that the 2'-phosphate of NADP(H) contributes to its interaction with the wild-type enzyme. Changing Thr-38 to Asp destabilized the binding energies of NADP(H) by 3.9-4.5 kcal/mol and stabilized those of NAD(H) by 1.2-1.4 kcal/mol. These results indicate a significant role of Thr-38 in NADP(H) binding for the mouse lung enzyme and provide further evidence for the key role of Asp at this position in NAD(H) specificity of the SDR family proteins.

Alcohol Oxidoreductases↗
PubMed1997

Purification of endothelin from a conditioned medium of cardiac fibroblastic cells using beating rate assay of myocytes cultured in a serum-free medium.

A conditioned medium from cardiac fibroblastic cells stimulated the beating of quiescent cardiac myocytes cultured in a serum-free medium. The aim of this study was to isolate and characterize the myocyte beat-stimulating activity of the conditioned medium of fibroblastic cells. Cardiac myocytes and fibroblastic cells were isolated individually from neonatal rats. The fibroblastic cells were grown in a growth medium until they became confluent, then serum-free conditioned medium was obtained from them. For the beating-rate assay, the cardiac myocytes were cultured in a completely serum-free medium. The beat-stimulating factor of myocytes in the conditioned medium was purified by reverse-phase liquid chromatographies and gel filtration, and was characterized by measuring the molecular weight of the activity and a pharmacological antagonistic study. The beat-stimulating activity in the conditioned medium was purified into two active fractions. Both of the activities have a molecular weight of 2.5 kDa, and the activities were abolished similarly by FR139317, an endothelin type-A receptor antagonist. These results indicate that cardiac fibroblastic cells secrete endothelin and that this may contribute in part to the functional abnormalities of the heart in patients with myocardial fibrosis.

Animals↗
PubMed1997

Nitric oxide synthase activities in mammalian parotid and submandibular salivary glands.

Nitric oxide is important as a physiological messenger molecule in various organs and cells. It is synthesized from the amino acid L-arginine by nitric oxide synthase. Here, the specific activities of nitric oxide synthase in the cytosolic fractions of rabbit, bovine, mice, rat, and guinea-pig parotid and submandibular glands were compared. Marked specific activities were detected in the rabbit and bovine parotid and submandibular glands and in the parotid of mice. The activity in rabbit parotid was highest and was similar to that in rabbit brain. The significant activities in the salivary glands were completely blocked in the absence of Ca2+ or the presence of a calmodulin inhibitor. These findings suggest that the rabbit parotid glands are useful for studying the regulation of nitric oxide generation by Ca2+/calmodulin-dependent nitric oxide synthase in salivary glands.

Animals↗
PubMed1997

Basic studies for the practical use of bitterness inhibitors: selective inhibition of bitterness by phospholipids.

PURPOSE: We examined the effects of phospholipids such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol (PI), and phosphatidic acid (PA) on human taste sensation to various substances. METHODS: The effects were evaluated psychophysically using paid volunteers. RESULTS: PA inhibited the bitterness of various substances dissolved in water without affecting sweetness, saltiness, and sourness, although its inhibitory activity was less than that of PA-LG. PI also showed inhibitory activity on bitterness, although its activity was less than PA. A soybean lecithin fraction containing high contents of PA and PI also demonstrated inhibitory activity on the bitterness of various substances. Both the incorporation of either PA or the lecithin fraction into granules containing quinine and the coating of the granules with PA or the fraction effectively inhibited the bitterness of quinine. CONCLUSIONS: The lecithin fraction is permitted for use as an additive to drugs and food and can be produced on an industrial scale. It is expected that the lecithin fraction will be used safely as a bitterness inhibitor for practical applications.

Humans↗
PubMed1997

Elucidation of the basic three-dimensional structure of type I interferons and its functional and evolutionary implications.

The scientific and personal backgrounds of the crystallographic elucidation of the three-dimensional structure of murine interferon-beta (Mu-IFN-beta) are described. This structure, elucidated in 1990, is still the only experimentally determined structure for type I IFNs. Model-building studies for various type I IFNs based on the Mu-IFN-beta structure and the arguments on the receptor-binding epitopes appearing since then are reviewed. An updated set of a table and a figure demonstrating a strong correlation between the degree of amino acid sequence variation in various cytokine proteins and that in their cognate receptor proteins is given. The origin of a remarkably larger rate of evolutionary change in amino acid sequences of cytokine proteins despite their physiologic significance is discussed in view of the cytokine network and the neutral theory of evolution.

Animals↗
PubMed1997

Mosquito vectors of bancroftian filariasis in Kwale District, Kenya.

A total of 2,906 female mosquitoes were collected over a period of one year using pyrethrum spray-sheet and human bait methods, and dissected for filaria larvae in three hinterland villages of coastal Kenya. The dominant species, Anopheles gambiae and Anopheles funestus were also found to be the main vectors. From the spray catch collections 0,9 and 1 Cx. quinquefasciatus, An. gambiae and An. funestus out of 491, 708 and 403 respectively were infective. In the same order, 4, 2 and 2 out of 512, 196 and 180 from human bait collections were infective. The results indicate that Cx.quinquefasciatus is also an important vector in this area contrary to some previous findings that it played no important role in rural hinterland areas. Differences in the results from the human bait and spray catch methods have been pointed out and the advantage of using both methods in filarial surveys indicated.

Animals↗
PubMed1997

[A case of polyneuropathy with B-CLL and HTLV-I associated myelopathy (HAM)].

We report a case of polyneuropathy associated with chronic B lymphoid leukemia (B-CLL) and HAM. A 65-year-old man had an initial onset with polyneuropathy, then myelopathy appeared. High HTLV-I antibody and anti-ganglioside antibodies were observed. The biopsy examination of the sural nerve showed mainly demyelinated changes. Muscle weakness gradually improved after cyclophosphamide therapy. Anti-ganglioside antibodies were also decreased in titer. This case suggested that the production of antiganglioside antibodies and the occurrence of polyneuropathy might be caused by both factors of HTLV-I infected T cells and B cell tumors.

Aged↗