Isolation and characterization of light actinide metallofullerenes.
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Journal Article↗
Biomedical subjects
M Sakaguchi
Publications and source records attributed to M Sakaguchi.
Identification of a phosphoprotein that is downregulated in immortalized human fibroblasts.
Many lines of evidence indicate that the immortalization step is critical for the neoplastic transformation of normal human cells. Once normal human cells have been immortalized, they are relatively easily transformed into neoplastic cells. In order to understand these phenomena, patterns of protein phosphorylation in proliferating normal human fibroblast cell strains and their immortalized cell lines were compared by using two-dimensional polyacrylamide gel electrophoresis. It was found that the expression and phosphorylation levels of the human heat shock protein 27 (HSP27) were predominantly downregulated in the immortalized cells compared with those in their normal counterparts. In the normal cells, HSP27 expression and phosphorylation were markedly increased by physiological and nonphysiological stresses, such as serum addition, treatment with a carcinogenic agent like 4-nitroquinoline-1-oxide, and a high osmotic pressure. This may be a normal defense against acute changes of cellular environment and cytotoxic effects. However, these stresses had no effects on the expression and phosphorylation of HSP27 in the immortalized cells. These results suggest that an abnormal regulation of HSP27 expression and phosphorylation may be one of the reasons for easy neoplastic transformation of the immortalized cells by the treatment with carcinogenic agents.
Amino Acid Sequence↗
Release of IL-4 and IL-5 from human peripheral blood mononuclear cells co-cultured with Japanese cedar pollen antigen in vitro.
The Japanese cedar pollen (JCP) is a major allergen with respect to pollinosis in Japan. It is believed that interleukin-4 (IL-4) and interleukin-5 (IL-5) derived from lymphocytes and other cells play a pivotal role in allergic reactions. We investigated whether the JCP antigen stimulates the release of these cytokines by peripheral blood mononuclear cells (PBMCs). PBMCs from eight adults (five adults with JCP pollinosis and three adults without JCP pollinosis) were co-incubated with purified JCP antigens. IL-4 was released in response to JCP antigens in six of the eight subjects at 24 h and in three subjects at 48 h. IL-4 release at 24 h occurred in all five subjects with JCP pollinosis but in only one of the three subjects without pollinosis. IL-5 was released in response to the JCP antigen in five of the eight subjects at 24 h and 48 h, including four of the five subjects with JCP pollinosis and one of the three subjects without pollinosis. These results suggest that PBMCs were more likely to release IL-4 and IL-5 in the presence of JCP pollinosis.
Adult↗
Polyvalent recombinant Marek's disease virus vaccine against poultry diseases.
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Animals↗
Establishment of a human hepatocyte line (OUMS-29) having CYP 1A1 and 1A2 activities from fetal liver tissue by transfection of SV40 LT.
Immortalized human hepatocytes that can retain functions of drug-metabolizing enzymes would be useful for medical and pharmacological studies and for constructing an artificial liver. The aim of this study was to establish immortalized human hepatocyte lines having differentiated liver-specific functions. pSVneo deoxyribonucleic acid, which contains large and small T genes in the early region of simian virus 40, was introduced into hepatocytes that had been obtained from the liver of a 21-wk-old fetus. Neomycin-resistant immortalized colonies were cloned and expanded to mass cultures to examine hepatic functions. Cells were cultured in a chemically defined serum-free medium, ASF104, which contains no peptides other than recombinant human transferrin and insulin. As a result, an immortal human hepatocyte cell line (OUMS-29) having liver-specific functions was established from one of the 13 clones. Expression of CYP 1A1 and 1A2 messenger ribonucleic acid by the cells was induced by treatment with benz[a]pyrene, 3-methylcholanthrene, and benz[a]anthracene. OUMS-29 cells had both the polycyclic aromatic hydrocarbon receptor (AhR) and AhR nuclear translocator. Consequently 7-ethoxyresorufin deethylase activity of the cells was induced time- and dose-dependently by these polycyclic aromatic hydrocarbons. This cell line is expected to be instrumental as an alternative method in animal experiments for studying hepatocarcinogenesis, drug metabolisms of liver cells, and hepatic toxicology.
Cell Line↗
Early and long-term results of coronary artery bypass grafting in dialysis patients.
OBJECTIVE: We evaluated the surgical outcome of coronary artery bypass grafting in dialysis patients. METHODS: We retrospectively analyzed 25 consecutive dialysis patients requiring isolated coronary artery bypass grafting. Their mean age was 61.6 years and average dialysis duration 80.4 months. RESULTS: Eight patients (32%) had emergency or urgent surgery. The mean number of bypassed vessels was 3.3 +/- 1.2. Internal thoracic artery grafts were used in 19 patients and gastroepiploic artery grafts in 5. Two (8%) died during hospitalization and complications occurred in 11 (44%). No cerebral complications were observed. All survivors showed ameliorated symptoms and improved overall function. Of 14 late deaths, 5 were cardiac-related, with 2 involving obvious myocardial ischemic events. Actuarial survival, including hospital deaths, at 1 year was 70.9%, at 3 years was 43.5%, and at 5 years was 34.8%. Cardiac-death-free survival was 70.2% at 3 years and 70.2% at 5 years. CONCLUSION: Coronary artery bypass grafting in dialysis patients is associated with a higher incidence of complications but has acceptable surgical mortality and effectively relieves angina symptoms.
Aged↗
Transgenic medaka overexpressing a melanin-concentrating hormone exhibit lightened body color but no remarkable abnormality.
Melanin-concentrating hormone (MCH) is a cyclic heptadecapeptide that concentrates melanin granules in the melanophores and lightens the body color of a fish. To investigate the utility of MCH as a reporter gene, a transgenic medaka strain overexpressing the MCH gene was established and its phenotypic features were examined. The salmon MCH gene driven by cytomegalovirus promoter was injected into 100 fertilized eggs of the HNI-1 medaka strain, which exhibits black body color. One F(0) female transmitted the transgene and a lightened body color phenotype to the F(1) generation. A homozygous transgenic strain was established by crossing F(2) fish homozygous for the transgene. Expression of the transgene was detected in several organs by Northern blotting. The melanin granules of transgenics were highly shrunk. Bioassay using scales confirmed the secretion of MCH into blood, and the MCH concentration was estimated between 0.5 and 5 microM. Development, growth, feeding behavior, and reproduction of transgenics did not differ significantly among transgenic and nontransgenic siblings. The result whereby enhanced MCH expression induced a change in body color, but no remarkable abnormality, suggests the usefulness of MCH as a novel reporter gene with unique features.
Journal Article↗
Different angiotensin II-forming pathways in human and rat vascular tissues.
We studied the angiotensin II-forming pathways in extracts from human and rat vascular tissues. In the extract from human artery, angiotensin I mainly converted to two products, angiotensin-(1-9) and angiotensin II, while in the extract from rat artery, the major angiotensin I products were angiotensin II and angiotensin-(5-10). The concentrations of angiotensin II and angiotensin-(1-9) generated in the human extract (1 mg protein/ml) after incubation for 30 min were 3.2 and 2.5 nmol, respectively, and that of angiotensin II and angiotensin-(5-10) generated in the rat extract (1 mg protein/ml) were 0.28 and 2.3 nmol, respectively. In the extract from human vascular tissues, the angiotensin II formation was inhibited by 8% with lisinopril and by 95% with chymostatin. The other product, angiotensin-(1-9) was inhibited completely by carboxypeptidase inhibitor. In the extract from rat vascular tissues, the angiotensin II formation was suppressed to 4% by lisinopril, but not by chymostatin. The angiotensin-(5-10) formation was completely inhibited by chymostatin. These findings suggest clearly that human vascular tissues contain two angiotensin II-forming enzymes, angiotensin-converting enzyme and chymase, but rat vascular tissues have no chymase-dependent angiotensin II-forming pathway.
Adult↗
IgE reactivity and cross-reactivity to Japanese cedar (Cryptomeria japonica) and cypress (Chamaecyparis obtusa) pollen allergens in dogs with atopic dermatitis.
The natural occurrence of Japanese cedar (Cryptomeria japonica) pollinosis has been reported in dogs with atopic dermatitis. However, the reactivity to Japanese cypress (Chamaecyparis obtusa) pollen allergens in these dogs has not been reported. The present study was designed to investigate the reactivity to Japanese cypress pollen allergens in dogs sensitized to Japanese cedar pollen allergens. In 19 dogs with specific IgE to C. japonica pollen allergen, we measured the specific IgE to C. obtusa pollen allergen and examined the reactivity to the allergen by intradermal test. Of the 19 dogs, 18 had specific IgE to crude and purified major allergens (Cha o 1) of C. obtusa pollen. Most of the dogs showed a positive reaction to C. obtusa pollen allergens in the intradermal test. Allergenic cross-reactivity between Cha o 1 and Cry j 1 (a major allergen in C. japonica pollen) was observed by the ELISA inhibition method. Dogs sensitized to Japanese cedar pollen allergens demonstrate reactivity to Japanese cypress pollen allergens.
Allergens↗
Two-dimensional gel electrophoretic studies on the cellular aging: accumulation of alpha-2-macroglobulin in human fibroblasts with aging.
To understand the mechanisms that control life span and age-related phenotypes, we used two-dimensional (2D) gel electrophoresis to study the intracellular proteins whose amounts change during the process of cellular aging. We found that the amount of an alpha-2-macroglobulin (A2M) fragment derived from culture medium increased in the cells with aging. A2M is linked to Alzheimer's disease both genetically and functionally. This is the first report of accumulation of an A2M fragment in senescent fibroblasts. We also studied 2D gel profiles of human fibroblasts immortalized by treatment with either 60Co gamma rays or 4-nitroquinoline 1-oxide. As immortalized cells overcome cellular senescence to gain an unlimited life span, the proteins whose amounts change after immortalization may be relevant to the age-related phenotypes. 2D gel analysis revealed that the A2M fragment was down-regulated in the immortalized cells, compared with their normal counterparts, regardless of their passage. We also found that the other four proteins increased in amount with aging and decreased in amount after immortalization. Our results suggest: (1) the A2M incorporation into the cells is increased in the process of cellular aging; and (2) A2M may be linked to the age-related phenotypes that were lost during the process of immortalization of human cells.
Cell Line↗
Up-regulation of S100C in normal human fibroblasts in the process of aging in vitro.
S100 proteins belonging to the EF-hand Ca(2+)-binding protein family regulate a variety of cellular processes via interaction with different target proteins. Several diseases, including cancer and Alzheimer's disease, are related to a disorder of multifunctional S100 proteins, which are expressed in cell- and tissue-specific manners. We previously demonstrated that S100C could move to and accumulate in the nuclei of normal human fibroblasts but not in the nuclei of immortalized and neoplastic cells. In addition, we found that its nuclear accumulation resulted in suppression of DNA synthesis in normal cells at a confluent stage. In the present study, we investigated whether S100C was associated with cellular senescence in vitro. We found that S100C expression increased in normal human fibroblasts in the process of aging in culture and was accompanied by accumulation of its protein in the nuclei of senescent fibroblasts. In addition, the nuclear accumulation of S100C increased expression of a cyclin-dependent kinase inhibitor p21(Sdi1), a strong inhibitor of cell growth. These findings suggest that an increase in the cells having nuclear accumulation of S100C is closely related to the process of cellular senescence of normal human fibroblasts.
Cell Division↗
Specific IgE and IgG to gelatin in children with systemic cutaneous reactions to Japanese encephalitis vaccines.
BACKGROUND: Systemic allergic reactions to Japanese encephalitis (JE) vaccine that include urticaria, angioedema, and rash have been reported. In Japan, children who suffered from allergic immediate-type reactions to JE vaccine had antigelatin IgE in their sera. However, the immunologic mechanism of allergic nonimmediate-type reactions that consist of cutaneous signs appearing several hours or more after JE vaccination has not been defined. METHODS: Serum samples were taken from 28 children who showed allergic nonimmediate-type cutaneous reactions to JE vaccine. Furthermore, serum samples were taken from 10 children who showed allergic immediate-type reactions with cutaneous signs and/or respiratory symptoms to JE vaccine. We have defined an immediate-type reaction as one occurring within 1 h after vaccination. RESULTS: Of 10 children who showed immediate-type reactions, all had antigelatin IgE and IgG. Of 28 children who showed systemic nonimmediate-type reactions, one had antigelatin IgE and nine (32%) had antigelatin IgG. The child who had antigelatin IgE showed urticaria 2 h after JE vaccination. CONCLUSION: These results suggest that some children who showed allergic nonimmediate-type reactions to JE vaccine were sensitized to gelatin.
Antibody Specificity↗
Anaphylaxis to Japanese encephalitis vaccine.
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Adolescent↗
Seasonal changes of humoral and cellular immune responses to Japanese cedar (Cryptomeria japonica) pollen allergens in Japanese monkeys (Macaca fuscata) with pollinosis.
The natural occurrence of Japanese cedar [Cryptomeria japonica (CJ)] pollinosis has been reported in Japanese monkeys (Macaca fuscata). The present study was designed to investigate seasonal changes in immunological reactions to CJ pollen allergens in monkeys with CJ pollinosis. Blood samples were collected from six monkeys with CJ pollinosis before and after CJ pollen season. Seasonal changes in specific IgE and IgG to major allergens (Cry j 1 and Cry j 2) were observed before and after CJ pollen season. The humoral responses decreased significantly before CJ pollen and increased after CJ pollen season. Similar seasonal changes in peripheral blood mononuclear cells proliferative responses to CJ allergens were observed before and after CJ pollen season. These humoral and cellular immune responses might serve as a biomarker for assessing new immunotherapies for monkeys with pollinosis.
Allergens↗
Preclinical evaluation of an immunotherapeutic peptide comprising 7 T-cell determinants of Cry j 1 and Cry j 2, the major Japanese cedar pollen allergens.
BACKGROUND: Peptide immunotherapy is a new approach to treating allergic diseases, but a therapeutic peptide for Japanese cedar pollinosis has not yet been developed. OBJECTIVE: The aim of this study is to prepare and preclinically evaluate a hybrid peptide comprising 7 T-cell determinants of Cry j 1 and Cry j 2, the major Japanese cedar pollen allergens. METHODS: The recombinant hybrid peptide was prepared after immunodominance of 7 T-cell determinants was confirmed by means of PBMC proliferation assay in 113 volunteers with pollinosis. The hybrid peptide was compared with a mixture of the 7 T-cell determinants in a dose-dependent PBMC proliferation assay in 6 volunteers with pollinosis. PBMC proliferation and binding activity of serum IgE antibody against the hybrid peptide, Cry j 1, and Cry j 2 were investigated in 48 volunteers with pollinosis. RESULTS: The hybrid peptide induced T-cell proliferation with an average 100-fold lower concentration than a mixture of the 7 peptides. PBMCs from 44 (92%) of 48 volunteers proliferated against the hybrid peptide, with significant correlation (r = 0.87) in T-cell proliferation against Cry j 1 and Cry j 2. No serum IgE antibodies specific to Cry j 1 or Cry j 2 bound to the hybrid peptide. CONCLUSION: A hybrid peptide comprising 7 T-cell determinants has the potential for inducing T-cell proliferative responses that is superior to the potential of a mixture of the T-cell determinants and comparable with that of Cry j 1 and Cry j 2. The hybrid peptide will be of use in specific immunotherapy against Japanese cedar pollinosis.
Adult↗
Anaphylaxis to gelatin-containing rectal suppositories.
BACKGROUND: Some children--though the number is few-have been sensitized with gelatin. OBJECTIVE: To investigate the relationship between the presence of antigelatin IgE and anaphylaxis to gelatin-containing rectal suppository, we measured antigelatin IgE in the sera of the children with anaphylaxis. METHODS: Ten children showed systemic allergic reactions, including anaphylaxis, to a chloral hydrate rectal suppository containing gelatin (231 mg/dose) that had been used as a sedative. These children's clinical histories and serum samples were submitted from physicians to the National Institute of Infectious Diseases during a 2-year period from 1996 to 1997. RESULTS: Of the 10 children, 5 showed apparent anaphylaxis, including hypotension and/or cyanosis, along with urticaria or wheezing; 2 showed both urticaria and wheezing without hypotension or cyanosis; the other 3 showed only urticaria. All of the children had antigelatin IgE (mean value +/- SD, 7.9 +/- 8.4 Ua/mL). As a control, samples from 250 randomly selected children had no antigelatin IgE. These findings suggest that the 10 children's systemic allergic reactions to this suppository were caused by the gelatin component. CONCLUSION: Gelatin-containing suppositories must be used with the same caution as gelatin-containing vaccines and other medications.
Anaphylaxis↗
Chymase-dependent angiotensin II formation in the saphenous vein versus the internal thoracic artery.
OBJECTIVES: The great saphenous vein graft is known to be less patent than the internal thoracic artery graft. Recently, we reported that chymase-dependent angiotensin II formation plays an important role in the development of intimal hyperplasia in dog grafted veins. In this study we investigated the levels of angiotensin II-forming enzymes, angiotensin-converting enzyme, and chymase in human saphenous veins and internal thoracic arteries. METHODS: The saphenous vein and internal thoracic artery specimens were obtained from coronary artery bypass grafts of patients during surgical procedures (saphenous vein, n = 16; internal thoracic artery, n = 16). Activities of angiotensin-converting enzyme and chymase were determined by using the extract from the saphenous vein or internal thoracic artery. Sections of the saphenous vein or internal thoracic artery were stained with van Gieson's elastin stain and were immunostained with anti-human chymase antibody. RESULTS: The activities of angiotensin-converting enzyme in the saphenous vein and internal thoracic artery were 0.34 +/- 0.12 and 0.32 +/- 0.17 mU/mg protein, respectively, and the difference was not significant. The chymase activity in the saphenous vein was significantly higher than that in the internal thoracic artery (saphenous vein, 10.1 +/- 0.81 mU/mg protein; internal thoracic artery, 6.21 +/- 1.86 mU/mg protein). Chymase-positive cells in the saphenous vein were located in both the media and adventitia, and those in the internal thoracic artery were located only in the adventitia. The number of chymase-positive cells in the saphenous vein was about 2.6 times that in the internal thoracic artery. CONCLUSION: The chymase activity, but not the angiotensin-converting enzyme activity, was significantly higher in the saphenous vein, suggesting that the high levels of chymase activity may be related to the poorer performance of the saphenous vein for use as a bypass conduit.
Aged↗
Involvement of a 40-kDa glycoprotein in food recognition, prey capture, and induction of phagocytosis in the protozoon Actinophrys sol.
A 40-kDa glycoprotein (gp40) was identified as a Con A-binding adhesive substance of the heliozoon Actinophrys sol for immobilizing and ingesting prey flagellates. Isolation and partial characterization of gp40 showed that: 1) gp40 is a major Con A-binding protein of Actinophrys with a molecular weight of 40 kDa, and is stored in secretory granules called extrusomes; 2) gp40 was purified by Con A-affinity chromatography, and the N-terminal amino acid sequence was determined as H2N-KVLK-FEDDFDTFDLQ; 3) prey flagellates became adhered to gp40-immobilized agarose beads; 4) phagocytosis of Actinophrys was induced against gp40-immobilized agarose beads; and 5) solubilized gp40 induced exocytosis of extrusomes and cell fusion of heliozoons. These results indicate that gp40 is a multi-functional secretory protein of Actinophrys, which is required in correct targeting of the heliozoon to food organisms as well as in self-recognition.
Amino Acid Sequence↗