Biomedical subjects

M Amano

Publications and source records attributed to M Amano.

At least 73 records · Page 4Linked to original sources

PubMed1998

CD1 expression defines subsets of follicular and marginal zone B cells in the spleen: beta 2-microglobulin-dependent and independent forms.

We have used multicolor FACS analysis, immunohistology, and functional assays to study the expression of CD1 on B cell subsets from normal and beta 2m-/- mice. Two B cell subpopulations were identified that express high levels of CD1 in normal mice: splenic marginal zone B cells (IgMhigh IgDlow CD21high CD24intermediate CD23- CD43-) and a newly identified subpopulation of follicular B cells. The latter cells are unusual, because they are IgDhigh CD23+, like follicular B cells, but express high levels of CD21 and IgM, an expression pattern that is associated with marginal zone B cells. Therefore, the high-level expression of CD1 and CD21 was found to be closely associated on splenic B cells. Immunohistology confirmed the expression of CD1 on marginal zone B cells and on clusters of B cells in splenic follicles. Both the high-level CD1 expression by these cells and the low-level CD1 expression by subpopulations of B cells in the spleen, lymph node, peritoneal cavity, and bone marrow were markedly reduced in beta 2m-/- mice. Despite this, a CD1-restricted T cell clone proliferated vigorously in response to LPS-activated spleen cells that had been obtained from both beta 2m-/- and wild-type mice. This response was inhibited by the 3C11 anti-CD1 mAb. These results show the heterogeneity of B cell subsets in their expression of the beta 2m-dependent form of CD1. They further suggest that a beta 2m-independent form of CD1 is expressed on B cells that can stimulate T cells; however, this form is not easily visualized with the anti-CD1 mAb used here.

Animals↗

PubMed1998

Comparison of the structural characteristics of chromosome VI in Saccharomyces sensu stricto: the divergence, species-dependent features and uniqueness of saké yeasts.

Previous studies have revealed that chromosome VI of saké yeasts is much larger than that of the other strains of Saccharomyces cerevisiae. Southern analysis using segments of chromosome VI of a laboratory strain as probes suggested that the nucleotide sequence of a major portion of this chromosome is conserved, but considerable diversity was found in the distal parts in the other strains. Physical maps also indicated that differences in length of chromosome VI were mainly due to differences in its ends. NotI was found to generate 9 kb and/or 16 kb fragments from the left telomere of chromosome VI in most saké yeasts, but no fragment in the case of AB972. SfiI produced one or two 30-50 kb fragments from the right end of this chromosome in all saké yeasts tested, but produced a 20 kb fragment in the case of AB972. All S. cerevisiae strains not employed in saké brewing were the same as AB972 in these respects. S. paradoxus had one NotI site in chromosome VI, while S. bayanus had two, one of which is possibly common to both species. The SfiI site mentioned above was present in chromosome VI of all species, while that of S. bayanus and S. paradoxus each had a second site distinct from the other. Chromosome VI of S. pastorianus was not distinguishable from that of S. bayanus.

Blotting, Southern↗

PubMed1998

Phosphorylation of vimentin by Rho-associated kinase at a unique amino-terminal site that is specifically phosphorylated during cytokinesis.

We found that vimentin, the most widely expressed intermediate filament protein, served as an excellent substrate for Rho-associated kinase (Rho-kinase) and that vimentin phosphorylated by Rho-kinase lost its ability to form filaments in vitro. Two amino-terminal sites on vimentin, Ser38 and Ser71, were identified as the major phosphorylation sites for Rho-kinase, and Ser71 was the most favored and unique phosphorylation site for Rho-kinase in vitro. To analyze the vimentin phosphorylation by Rho-kinase in vivo, we prepared an antibody GK71 that specifically recognizes the phosphorylation of vimentin-Ser71. Ectopic expression of constitutively active Rho-kinase in COS-7 cells induced phosphorylation of vimentin at Ser71, followed by the reorganization of vimentin filament networks. During the cell cycle, the phosphorylation of vimentin-Ser71 occurred only at the cleavage furrow in late mitotic cells but not in interphase or early mitotic cells. This cleavage furrow-specific phosphorylation of vimentin-Ser71 was observed in the various types of cells we examined. All these accumulating observations increase the possibility that Rho-kinase may have a definite role in governing regulatory processes in assembly-disassembly and turnover of vimentin filaments at the cleavage furrow during cytokinesis.

Amino Acid Sequence↗

PubMed1998

Identification of the major allergens in wheat flour responsible for baker's asthma.

Baker's asthma, a typical occupational allergic disease, is a serious problem in the food industries. In this study, purification and identification of major allergens recognized by IgEs in sera of allergic patients were performed. Major immunoreactive proteins were purified from the albumin fraction by gel filtration on a Toyopearl HW-50 column followed by reverse-phase HPLC. The N-terminal amino acid sequences and molecular masses measured by MS indicated that the major immunoreactive proteins are members of the alpha-amylase inhibitor family, 0.19 and 0.28. Significant leukotriene release by each purified protein was observed in cell-associated stimulation tests, suggesting in vivo activity of these antigens. Carbohydrate analyses of major allergens indicated that they are monoglycosylated but not N-glycosylated in spite of the presence of a potential N-glycosylation site. Recombinant 0.19 expressed in Escherichia coli showed the same reactivity with IgE as native wheat 0.19 in Western blotting and ELISA using methyl vinyl ether maleic anhydride co-polymer as an immobilizing reagent, suggesting that the allergenic epitopes are located in the peptide portions.

Allergens↗

PubMed1998

Subsets of transgenic T cells that recognize CD1 induce or prevent murine lupus: role of cytokines.

T cells with T cell receptor (TCR) transgenes that recognized CD1 on syngeneic B cells stimulated B cells to secrete immunoglobulins in vitro. The CD4+, CD8+, or CD4-CD8- T cells from the spleen of the TCR transgenic BALB/c donors induced lupus with anti-double stranded DNA antibodies, proteinuria, and immune complex glomerulonephritis in irradiated BALB/c nude mice reconstituted with nude bone marrow. Injection of purified CD4-CD8- T cells from the marrow of transgenic donors prevented the induction of lupus by the transgenic T cells. Transgenic T cells that induced lupus secreted large amounts of interferon (IFN)-gamma and little interleukin (IL)-4, and those that prevented lupus secreted large amounts of IL-4 and little IFN-gamma or IL-10.

Animals↗

PubMed1998

Rho-kinase phosphorylates COOH-terminal threonines of ezrin/radixin/moesin (ERM) proteins and regulates their head-to-tail association.

The ezrin/radixin/moesin (ERM) proteins are involved in actin filament/plasma membrane interaction that is regulated by Rho. We examined whether ERM proteins are directly phosphorylated by Rho-associated kinase (Rho-kinase), a direct target of Rho. Recombinant full-length and COOH-terminal half radixin were incubated with constitutively active catalytic domain of Rho-kinase, and approximately 30 and approximately 100% of these molecules, respectively, were phosphorylated mainly at the COOH-terminal threonine (T564). Next, to detect Rho-kinase-dependent phosphorylation of ERM proteins in vivo, we raised a mAb that recognized the T564-phosphorylated radixin as well as ezrin and moesin phosphorylated at the corresponding threonine residue (T567 and T558, respectively). Immunoblotting of serum-starved Swiss 3T3 cells with this mAb revealed that after LPA stimulation ERM proteins were rapidly phosphorylated at T567 (ezrin), T564 (radixin), and T558 (moesin) in a Rho-dependent manner and then dephosphorylated within 2 min. Furthermore, the T564 phosphorylation of recombinant COOH-terminal half radixin did not affect its ability to bind to actin filaments in vitro but significantly suppressed its direct interaction with the NH2-terminal half of radixin. These observations indicate that the Rho-kinase-dependent phosphorylation interferes with the intramolecular and/ or intermolecular head-to-tail association of ERM proteins, which is an important mechanism of regulation of their activity as actin filament/plasma membrane cross-linkers.

3T3 Cells↗

PubMed1998

PyNPase expression in human colon cancer.

Although colorectal cancer tissue is rich in pyrimidine nucleoside phosphorylase (PyNPase), there is no consensus as to whether cancer cells or stromal cells predominately express PyNPase. We micro-dissected OCT compound embedded frozen tissue sections into epithelial and stromal components and then analyzed the extracted samples separately. The PyNPase expression level was higher in stromal cells than in cancer cells and the difference increased with inflammation induced by the immunostimulator OK432. These results suggest that stromal cells are the major PyNPase source in colorectal cancer.

Adult↗

PubMed1998

Age-dependent accumulation of heavy metals in baikal seal (Phoca sibirica) from the Lake Baikal.

Concentrations of Fe, Mn, Zn, Cu, Cd, and Hg were determined in the liver, kidney, and muscle of 60 Baikal seals collected from Lake Baikal in 1992 to investigate age-dependent accumulation. Among essential elements, Fe concentrations in the muscle, liver, and kidney increased with age, suggesting development of diving ability. The concentrations of Mn, Zn, and Cu decreased with age, especially at immature stages. Toxic elements such as Hg and Cd decreased in adult males and thus the male-female difference was clearly observed in their concentrations, which differed from patterns usually found in marine mammals. Such accumulation patterns were due to difference in the feeding rates between males and females under low exposure to Hg and Cd. In addition, a greater excretion of Hg than that of Cd through molting and parturition was estimated.

Aging↗

PubMed1998

Two differing salmon GnRH precursor mRNAs are co-expressed in the brain of sockeye salmon.

The localization of two salmon-type gonadotropin-releasing hormone (sGnRH) precursors, pro-sGnRH-I (short type) and pro-sGnRH-II (long type), was investigated by using in situ hybridization techniques in the brain of the landlocked sockeye salmon, Oncorhynchus nerka. We used 30-mer oligonucleotide probes complementary to pro-sGnRH-I and pro-sGnRH-II cDNA. No significant differences were observed in the localization of sGnRH neurons expressing pro-sGnRH-I and pro-sGnRH-II mRNAs; both were expressed in the olfactory nerve, the olfactory bulbs, the regions between the olfactory bulb and telencephalon, the ventral telencephalon, the preoptic area, and the hypothalamus. Almost all sGnRH neurons examined co-expressed both precursors. The expression of two sGnRH precursors in the same neuron and the wide distribution of such neurons in the brain suggest that there are no functional differences between the two precursors.

Animals↗

PubMed1998

Ontogenic development of salmon GnRH and chicken GnRH-II systems in the brain of masu salmon (Oncorhynchus masou).

Ontogenic development of salmon gonadotropin-releasing hormone (GnRH) and chicken GnRH-II systems in masu salmon (Oncorhynchus masou) was examined. Salmon GnRH was first detected by radioimmunoassay in the embryo on day 36 after fertilization. Salmon GnRH-immunoreactive fibers were detected initially by immunocytochemistry in the vicinity of the olfactory placode of the embryo (day 36) and were distributed widely in the brain as well as in the pituitary gland of fish just after hatching (day 80). Salmon GnRH-immunoreactive neuronal somata were first detected about 6 months after fertilization in the rostroventral brain area, ranging from the olfactory nerve to the preoptic area. Salmon GnRH neuronal somata were detected earlier by in situ hybridization than by immunocytochemistry. Neuronal somata expressing salmon GnRH mRNA were first detected in the vicinity of the olfactory epithelium on day 40 and then were seen to be migrating from the olfactory epithelium, along the olfactory nerve, on day 60 and in the transitional area between olfactory nerve and olfactory bulb on day 80. In the brain, these neurons were first detected in the ventral olfactory bulb on day 80, and thereafter they were also detected in the caudal brain regions. The chicken GnRH-II system was detected later than the salmon GnRH system; chicken GnRH-II was first detected by radioimmunoassay on day 57, and chicken GnRH-II-immunoreactive fibers were first detected on day 67. Chicken GnRH-II-immunoreactive neuronal somata were not detected during the observation period. These results suggest that salmon GnRH neurons derive from the olfactory placode and then migrate into the brain and that salmon GnRH is synthesized before chicken GnRH-II.

Animals↗

PubMed1998

Primary cancer of the small intestine and mutational analysis of the K-ras and p53 genes.

A 69-year-old woman was admitted to Hokuso Shiroi Hospital because of recurrent pain in the lower right side of the abdomen. Small-intestinal cancer was strongly suspected after fluoroscopy of the small intestine. Laparotomy showed advanced cancer of the ileum, of complete annular constrictive type, 9.5 x 5cm in size. Histologically it was moderately differentiated tubular adenocarcinoma. Neither visceral nor nodal metastases were found, and the patient has been well for the 20 months since surgery. The strong resemblance between the epidemiological characteristics of small-intestinal cancers and colorectal cancers prompted us to investigate the carcinogenetic mechanisms at the molecular level. A point mutation at codon 12 of the K-ras gene was found, while no alterations were noted in the p53 gene, whose mutations are frequent in colon cancers. The carcinogenetic mechanisms of the small-intestinal cancer we experienced may thus differ from those of colon cancers.

Adenocarcinoma↗

PubMed1998

Dissociation of impairment between spatial memory, and motor function and emotional behavior in aged rats.

We investigated changes in learning and memory in aged rats, in relation to motor function and emotional behavior. Male Kbl Wistar aged rats (108-weeks-old) were divided into two groups, memory impaired and non-impaired, based on performance during six training trials in the Morris water maze task. Aged rats with a goal latency longer than the mean plus the 99% confidence limit of young rats, were regarded as memory impaired, whereas those with a goal latency within the range of the 99% confidence limit of the mean of young rats, were considered as memory non-impaired. Although the performance of the memory impaired aged rats in the standard test of the Morris water maze improved after six re-training trials to the level of the non-impaired aged rats and young rats, working memory impairment was evident. There were no differences in motor function and emotional behavior between the impaired and non-impaired aged rats. These results suggest that deficits of learning and memory in memory impaired aged rats can be dissociated from changes in motor function and emotional behavior.

Aging↗

PubMed1998

Myosin II activation promotes neurite retraction during the action of Rho and Rho-kinase.

BACKGROUND: The Rho small GTPase regulates myosin II activity through the phosphorylation of the myosin light chain (MLC) by activating Rho-kinase, which is a target of Rho. Several lines of evidence point to an important role of Rho in the action of lysophosphatidic acid (LPA) and thrombin in provoking neurite retraction in N1E-115 neuroblastoma cells. RESULTS: Here we examined whether Rho-kinase and myosin II are involved in neurite retraction in N1E-115 cells. We showed that the expression of constitutively active forms of Rho-kinase induced neurite retraction in NIE-115 cells and MLC phosphorylation in NIH 3T3 cells, whereas the expression of dominant negative forms of Rho-kinase inhibited the LPA-induced neurite retraction in N1E-115 cells and the serum-induced MLC phosphorylation in NIH 3T3 cells. The expression of mutant MLCT18D,S19D (substitution of Thr and Ser by Asp), which is known to lead to the activation of myosin ATPase and a conformational change of myosin II when reconstituted with myosin heavy chains in vitro, also promoted neurite retraction. CONCLUSION: These results indicate that Rho-kinase is involved in the LPA-induced neurite retraction downstream of Rho, and that myosin II activation promotes neurite retraction downstream of Rho and Rho-kinase.

3T3 Cells↗

PubMed1998

The epidemiology of pancreatic diseases in Japan.

We reviewed the trends and death rates of pancreatic cancer and pancreatitis in Japan over the past four decades. It is a disturbing fact that the death rate due to pancreatic cancer is rising, parallel to that of lung cancer and colon cancer and that it is affecting primarily the elderly 65 years and older in both sexes. The cause of this increase is partially attributable to improvements in the diagnosis, changes in life-style, and smoking. The death rate due to chronic and acute pancreatitis has remained constant during the past four decades.

Adult↗

PubMed1998

Functional relationship between muscles of mastication and the skull with enlarged orbit in the Baikal seal (Phoca sibirica).

We expected that the enlarged area of zygomatic arch, one of some skull characters adapted for enlarged eye, has an influence on form and function of muscles of mastication in the Baikal seal (Phoca sibirica). So, in this species, the Mm, masseter, temporalis, pterygoidei, digastricus were observed in the macroscopic level. The skull characters related to these muscles were also compared between the Baikal seal and a close-related species, the ringed seal (Phoca hispida). The Mm, masseter and temporalis were well-developed using the enlarged attachment area of zygomatic arch. In contrast, the M. digastricus is suggested to be not so important in the Baikal seal, because the temporal bone is not so developed as in the ringed seal. It is suggested that the Baikal seal has especially developed the Mm. temporalis and masseter using an enlarged area of zygomatic arch among Pusa species. We also suggest that the robust temporal bone is equipped to have the M. digastricus developed as a main retractor of mandibular body in the ringed seal.

Adaptation, Biological↗

PubMed1998

Using MR imaging to predict and evaluate the response of invasive cervical carcinoma to systemic chemotherapy.

OBJECTIVE: The goal of our study was to evaluate the accuracy of MR imaging in predicting the response of invasive cervical carcinoma to systemic neoadjuvant chemotherapy, to observe with MR imaging the difference in tumor response to chemotherapy between treatment cycles, and to correlate tumor size between MR imaging after chemotherapy and surgical specimen. SUBJECTS AND METHODS: Pelvic MR imaging was performed before and after chemotherapy in 41 patients with invasive cervical carcinoma. The difference in tumor reduction between the second course and the third or fourth course of treatment was statistically evaluated in 12 patients. MR images obtained after chemotherapy were compared with surgical specimens to determine residual tumor size in 10 patients. RESULTS: Tumor volume, complete replacement of cervical stroma with carcinoma, and uterine body involvement by tumor observed in the MR images before treatment were found to be significant factors in predicting tumor response. No difference in tumor reduction was found between the second course and the third or fourth course of chemotherapy. Estimates made on MR imaging of tumor size after chemotherapy were within 5 mm of the resected samples. CONCLUSION: Pelvic MR imaging proved useful in predicting and assessing the response of invasive cervical carcinoma to chemotherapy.

Adult↗

PubMed1998

[PyNPase expression and cancer progression in the colorectum].

We analyzed PyNPase expression discriminating between cancer and tumor stroma of the colorectum by Western blotting using a newly developed extraction method from microdissected tissue sections fixed with buffered formalin. Analysis of 98 colorectal cancers revealed that PyNPase was as high as 70.2 +/- 18.5 unit/mg protein in the stroma fraction (SF), whereas it was 45.1 +/- 10.5 in the cancer fraction (CF) (p < 0.0001). Vessel density was correlated with PyNPase in the SF but not in the CF. In stage IIIb, 11 cases expressing a high level of PyNPase in the CF showed poorer prognosis than 10 cases with low-level PyNPase expression (p < 0.05), although the level of PyNPase expression in the SF did not affected the patients prognosis. Immunohistochemical examination indicated that PyNPase in the SF was mainly produced by macrophages (M phi), and therefore we investigated the profile of PyNPase production by M phi. In in vitro experiments PyNPase production by M phi was greatly enhanced by stimulation with OK-432, and the culture supernatant had the ability to convert 5'DFUR to 5-FU.

Antineoplastic Agents↗

PubMed1997

Cytoskeletal rearrangements and transcriptional activation of c-fos serum response element by Rho-kinase.

The small GTPase Rho is implicated in cytoskeletal rearrangements including stress fiber and focal adhesion formation and in the transcriptional activation of c-fos serum response element. In vitro, Rho-kinase, which is activated by Rho, phosphorylates not only myosin light chain (MLC) (thereby activating myosin ATPase) but also myosin phosphatase, thus inactivating myosin phosphatase. Rho-kinase is involved in the formation of stress fibers and focal adhesions in fibroblasts. Here we show that the expression of constitutively active Rho-kinase increased the level of MLC phosphorylation. The activity of Rho-kinase was necessary for maintaining the vinculin-containing focal adhesions, whereas organized actin stress fibers were not necessary for this. The microinjection of constitutively active Rho-kinase into fibroblasts induced the formation of focal adhesions to some extent under the conditions where organized actin stress fibers were disrupted. The expression of constitutively active Rho-kinase also stimulated the transcriptional activity of c-fos serum response element. These results suggest that Rho-kinase has distinct roles in divergent pathways downstream of Rho, which include MLC phosphorylation leading to stress fiber formation, focal adhesion formation, and gene expression.

3T3 Cells↗