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Biomedical subjects

M Sakaguchi

Publications and source records attributed to M Sakaguchi.

At least 109 records · Page 6Linked to original sources
PubMed2000

A REIC gene shows down-regulation in human immortalized cells and human tumor-derived cell lines.

Normal human cells stop proliferation after a certain number of cell divisions. This phenomenon is called cellular aging. The fact that the senescence phenotype is dominant and the immortal one is recessive indicates that immortalization of human cells may be caused by loss of functions of certain genes in normal cells. Based on this evidence, several cDNA clones whose expression was down-regulated during the immortalization process of human cells were isolated by the representative difference analysis (RDA) system in our laboratory. One of them, which was named REIC, was expressed to a lower degree in three human immortalized cell lines as compared with their parental normal counterparts. In addition, the expression of REIC was markedly lower in eight human tumor-derived cell lines (Hep3B and HuH-7 hepatocellular carcinomas, HuH-6 Clone 5 hepatoblastoma, HuCCT-1 cholangiocarcinoma, A549 lung cancer, HaCaT immortalized keratinocyte, HeLa cervical carcinoma, and Saos-2 osteosarcoma). In contrast, among the human tissues examined, the heart and brain, which contain a large number of post-mitotic cells, showed the highest expression of REIC. The full-length REIC cDNA revealed that the predicted protein is 350 amino acids in length and possesses coiled-coil tertiary structures in each of the amino- and carboxyl-termini. Furthermore, a search of the protein database revealed a match of this gene product with Dkk-3, which is a novel inhibitor of Wnt oncogene. These results indicate that the REIC cloned by us may function as a tumor suppressor.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Transplantation of highly differentiated immortalized human hepatocytes to treat acute liver failure.

BACKGROUND: Temporary support of a damaged liver by a bioartificial liver (BAL) devise is a promising approach for the treatment of acute liver failure. Although human primary hepatocytes are an ideal source of hepatic function in BAL, shortage of human livers available for hepatocyte isolation is the limiting factor for the use of this modality. A clonal human hepatocyte cell line that can grow economically in culture and exhibit liver-specific functions should be an attractive solution to this problem. METHODS: To test this alternative, primary human fetal hepatocytes were immortalized using Simian virus 40 large T antigen. To investigate the potential of the immortalized cells for BAL, we transplanted the cells into the spleen of adult rats and performed a 90% hepatectomy 12 hr later. RESULTS: One of the cloned human liver cell lines, OUMS-29, showed highly differentiated liver functions. Intrasplenic transplanting of 20x10(6) OUMS-29 cells protected the animals from hyperammonemia and the associated hepatic encephalopathy. Survival was significantly prolonged in 90% of hepatectomized rats receiving OUMS-29 cells. CONCLUSIONS: A highly differentiated immortalized human hepatocyte cell line, OUMS-29, was able to provide metabolic support during acute liver failure induced by 90% hepatectomy in rats. Essentially unlimited availability of OUMS-29 cells may be clinically useful for BAL treatment.

Ammonia↗
PubMed2000

Establishment of a human hepatoma cell line, HLE/2E1, suitable for detection of p450 2E1-related cytotoxicity.

By transfection of an expression vector of human cytochrome P450 2E1 (CYP2E1) into a human hepatoma cell line (HLE), a new cell line (HLE/2E1) that stably expresses activity of CYP2E1 has been established. The HLE/2E1 cell line expressed a higher level of CYP2E1 messenger ribonucleic acid than did the mother HLE cell line. CYP2E1 enzyme activity determined by a p-nitrophenol oxidation assay was also higher in HLE/2E1 cells than in HLE cells. In addition, the enzyme activity of the HLE/2E1 cells was increased by ethanol treatment. Exposure to acetaminophen (APAP) or buthionine sulfoximine (BSO) caused a greater decrease in viability of the HLE/2E1 cells than that of the HLE cells, as determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay. The cytotoxicity of APAP or BSO to HLE/2EI cells was inhibited by the addition of ethanol or vitamin E. However, the cytotoxicity of both APAP and BSO was enhanced by 24-h preincubation of HLE/2E1 cells with ethanol. These results show that this cell line provides a useful model for studying catalytic properties of CYP2E1 and cytotoxic mechanisms of chemicals metabolized by CYP2E1.

Acetaminophen↗
PubMed2000

Transient eosinophilia by HIV infection.

We describe a case of early human immunodeficiency virus infection characterized by transient eosinophilia without an elevated immunoglobulin E concentration, allergic symptoms, or atopic dermatitis. Possible mechanisms of the eosinophilia are discussed.

Adult↗
PubMed2000

Sensitization to gelatin in children with systemic non-immediate-type reactions to varicella vaccines.

BACKGROUND: We recently found that four children who experienced systemic immediate-type reactions to varicella vaccine with gelatin had anti-gelatin IgE. We also found systemic non-immediate-type allergic reactions, which mainly consist of systemic cutaneous signs, appearing several hours or more after the vaccination. OBJECTIVE: To investigate the relationship between immune responses to gelatin and non-immediate-type reactions to gelatin-containing varicella vaccines, we measured anti-gelatin IgE and IgG in the sera of the children with these allergic reactions. METHODS: Serum samples were taken from 21 children who showed non-immediate-type reactions to varicella vaccines. As a positive control, serum samples were taken from 33 children who showed immediate-type reactions to varicella vaccine and had anti-gelatin IgE. As a negative control, serum samples were taken from 50 children who showed no reaction to the vaccine. We then examined anti-gelatin IgE and IgG in sera of the children. RESULTS: Of 21 children with non-immediate-type reactions, two (10%) had anti-gelatin IgE and six (29%) had anti-gelatin IgG. In the positive control group, all 33 children with immediate-type reactions had anti-gelatin IgG as well as IgE. In the negative control group, all 50 children who showed no allergic reaction to varicella vaccines had neither anti-gelatin IgE nor IgG. CONCLUSIONS: These results suggest that the possibility exists that some non-immediate-type reactions to varicella vaccine are caused by immune reactions to gelatin.

Angioedema↗
PubMed2000

Identification and characterization of a group 2 conifer pollen allergen from Chamaecyparis obtusa, a homologue of Cry j 2 from Cryptomeria japonica.

BACKGROUND: Not only Cryptomeria japonica (Japanese cedar) pollen but also that of Chamaecyparis obtusa (Japanese cypress) induces the allergic symptoms of Japanese cedar pollinosis. However, allergens from C. obtusa pollen have not been as well characterized as those from C. japonica pollen. OBJECTIVE: We sought to identify and characterize a homologue of the second major allergen of C. japonica pollen, Cry j 2, from the pollen of C. obtusa. METHODS: An allergen homologous to Cry j 2 was identified in C. obtusa pollen extract by immunoblot analysis, probed with anti-Cry j 2 monoclonal antibodies and purified by a series of column chromatographic steps. RESULTS: The allergen isolated from the extract showed a slightly diffuse band of 45 kDa and closely spaced double-bands of 42 and 45 kDa on SDS-PAGE, under reducing and non-reducing conditions, respectively; the bands were approximately 5-7 kDa larger than those of Cry j 2. In 24 of 30 residues, the N-terminal amino acid sequence of the allergen was identical with corresponding sequence in Cry j 2. Most patients with pollinosis who were IgE antibody-positive to Cry j 2 were shown to be IgE antibody-positive to this allergen, and the IgE antibody levels to both allergens were highly correlated. CONCLUSION: The results indicate that the allergen isolated from C. obtusa pollen in this study is a homologue of Cry j 2. The allergen was designated as Cha o 2 according to the WHO/IUIS Allergen Nomenclature Subcommittee recommendation.

Allergens↗
PubMed2000

Common antigenicity between Japanese cedar (Cryptomeria japonica) pollen and Japanese cypress (Chamaecyparis obtusa) pollen, I. H-2 complex affects cross responsiveness to Cry j 1 and Cha o 1 at the T- and B-cell level in mice.

Common antigenicity among two purified Japanese cedar pollen allergens (Cry j 1 and Cry j 2) and one Japanese cypress pollen allergen (Cha o 1) was explored at the T-cell and B-cell level in mice of different H-2 haplotypes. Cry j 2 did not show any common antigenicity with Cry j 1 or Cha o 1. B10.S (H-2S) mice immunized with Cry j 1 or Cha o 1 generated T cells and antibodies reactive to both antigens, indicating the common antigenicity of these antigens. C57BL/6 (H-2b) mice were non-responders to Cry j 1. BALB/c (H-2d) mice immunized with Cry j 1 or Cha o 1 and C57BL/6 mice immunized with Cha o 1 generated T cells that were only reactive with the respective immunogen, but produced antibody reactive to both Cry j 1 and Cha o 1, indicating that Cry j 1 and Cha o 1 share their B-cell epitope but not their T-cell epitope. This finding may provide a clue for the clarification of the T-cell and B-cell epitopes of Cry j 1 and Cha o 1, even though the data are influenced by H-2 complex restriction in mice. Considering that H-2 complex restriction affects cross responsiveness to Cry j 1 and Cha o 1 at the T- and B-cell level in mice, we assessed the possible situation in humans exposed sequentially to Japanese cedar pollen and Japanese cypress pollen.

Allergens↗
PubMed2000

Common antigenicity between Japanese cedar (Cryptomeria japonica) pollen and Japanese cypress (Chamaecyparis obtusa) pollen, II. Determination of the cross-reacting T-cell epitope of cry j 1 and cha o 1 in mice.

We have previously detected common antigenicity between Cry j 1 and Cha o 1 in B10.S mice. B10.S mice immunized with Cry j 1- or Cha o 1-generated T cells and antibodies reactive to both allergens. In the present study, we investigated the cross-reacting and Cry j 1-specific T-cell epitopes in B10.S mice. Lymph node cells from B10. S mice immunized with Cry j 1 recognized Cry j 1 p111-130, p211-230, and p310-330 as well as Cha o 1 p209-228. The existence of the cross-reacting T-cell epitope in Cry j 1 and Cha o 1 was confirmed by the response of newly established p211-230-specific and Cha o 1 p209-228-specific T-cell lines. The minimum peptide sequence (p213-224) of the cross-reacting T-cell epitope was identical in Cry j 1 and Cha o 1. These findings clearly demonstrate that common antigenicity at the T-cell level between Japanese cedar and cypress pollen allergens was caused by the existence of an identitical-cell epitope in Cry j 1 and Cha o 1.

Allergens↗
PubMed2000

Inhibition of immunoglobulin E response to Japanese cedar pollen allergen (Cry j 1) in mice by DNA immunization: different outcomes dependent on the plasmid DNA inoculation method.

To develop a new immunotherapy for Japanese cedar (Cryptomeria japonica; CJ) pollinosis, we evaluated the use of DNA immunization by inoculating mice with plasmid DNA encoding Cry j 1 as a CJ pollen major allergen (pCACJ1). Repeated intramuscular (i.m.) inoculation of BALB/c mice with pCACJ1 produced anti-Cry j 1 antibody responses, which were predominately of the immunoglobulin G2a (IgG2a) type. Furthermore, this inoculation suppressed immunoglobulin E (IgE) and IgG1 antibody responses to subsequent alum-precipitated Cry j 1 injections. Splenic T cells isolated from mice inoculated with pCACJ1 i.m. secreted interferon-gamma (IFN-gamma), but not interleukin (IL)-4, in vitro upon stimulation with Cry j 1 as well as with p277-288, a peptide corresponding to the T-cell epitope of Cry j 1. In contrast, inoculation of BALB/c mice with pCACJ1 by gene gun injection caused response predominantly of the IgG1 type, and enhanced production of anti-Cry j 1 IgE antibodies to subsequent alum-precipitated Cry j 1 injections. Splenic T cells isolated from pCACJ1-innoculated mice by gene gun injection secreted both IFN-gamma and IL-4 in vitro, upon stimulation with Cry j 1 as well as with p277-288. These findings suggest that i.m. inoculation with pCACJ1 effectively elicits Cry j 1-specific T helper 1 (Th1)-type immune responses, resulting in inhibition of the IgE response to Cry j 1.

Allergens↗
PubMed2000

Identification of a nuclear export signal in MKK6, an activator of the carp p38 mitogen-activated protein kinases.

Carp homologues of p38 mitogen-activated protein kinase (MAPK) and its activator MAPK kinase 6 (MAPKK6, referred to as MKK6) were identified. There exist at least two distinct carp p38s, cp38a and cp38b, both of which consist of 361 amino acids. The transcript of c38a was exclusively expressed in the ovary, whereas that of cp38b was ubiquitously expressed. Western blot analysis with anti-(phosphorylated MAPK) Ig specific to the active p38 or JNK has shown that p38 was activated in response to hypertonic stress (1 M sorbitol) in epithelioma papilosum cyprini carp epithelial cells (EPC) and that the activation of p38 proceeded faster to the maximal level than that of JNK. Carp homologue (cMKK6) of p38 activator MKK6 consists of 404 amino acids. It was expressed ubiquitously but was most abundant in the ovary. An in vitro kinase assay demonstrated that cMKK6 is an upstream activator of cp38 and cp38b in carp because it specifically phosphorylated and activated cp38a and cp38b. Interestingly, we found that cMKK6 has a nuclear export signal (NES) sequence in its N-terminal region although upstream activators of stress-activated MAPKs, p38 and JNK, do not in other animals. The NES sequence facilitated nuclear export of cMKK6 and ovalbumin. Leucine residues in the sequence were crucial for the NES activity, as the activity was lost on replacement of the leucines to alanines. The existence of an NES in cMKK6 implies the requisite of strict regulation of the p38 MAPK pathway in carp. The abundance of these components for the stress-activated pathway in the ovary might be related to ectogenetic early development.

Amino Acid Sequence↗
PubMed2000

A new method for simultaneous measurements of mast cell proteases in human vascular tissue.

1. Human mast cells contain carboxypeptidase A, chymase and tryptase. In the present study, in order to analyse the mast cell proteases simultaneously, we investigated a method for the measurement of carboxypeptidase A, chymase and tryptase in human vascular tissues. 2. Human vascular tissues were homogenized in 10 mmol/L phosphate buffer containing 0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4, 1.6 or 1.8 mol/L KCl and 0.1% non-idet P-40 and samples were then extracted. Because carboxypeptidase A and chymase convert angiotensin (Ang)I to Ang-(1-9) and AngII, respectively, the extracts were incubated with AngI in the presence of an angiotensin-converting enzyme (ACE) inhibitor. The extract prepared in buffer with over 0.8 mol/L KCl converted AngI to Ang-(1-9) and AngII. Formation of Ang-(1-9) and AngII plateaued in extracts with 1.0 and 1.2 mol/L KCl, respectively. 3. The formation of Ang-(1-9) and AngII in the extract with 1.2 mol/L KCl was inhibited by inhibitors of carboxypeptidase A and chymase, respectively, suggesting that Ang-(1-9) and AngII were generated from AngI by carboxypeptidase A and chymase, respectively. 4. Using a specific tryptase substrate, tryptase activity was detected in extract in buffer with over 0.8 mol/L KCl and reached a plateau at concentrations of KCl over 1.0 mol/L. 5. These findings show that the maximum activity of carboxypeptidase A, chymase and tryptase was detected in extracts of human homogenized vascular tissues in buffer at 1.2 mol/L KCl. The present study demonstrates a new method for the simultaneous measurement of proteases derived from mast cells in humans.

Angiotensin I↗
PubMed2000

IgE antibody to fish gelatin (type I collagen) in patients with fish allergy.

BACKGROUND: Most children with anaphylaxis to measles, mumps, and rubella vaccines had shown sensitivity to bovine gelatin that was included in the vaccines. Recently, it was found that bovine type I collagen, which is the main content in the gelatin, is a major allergen in bovine gelatin allergy. Fish meat and skin also contain type I collagen. OBJECTIVE: The present study was designed to investigate IgE antibody to fish gelatin in children with fish allergy. METHODS: Serum samples were taken from patients in 3 groups: (1) 10 patients with fish allergy and specific IgE to fish meat; (2) two patients with allergies to both fish meat and bovine gelatin and specific IgE to fish meat and bovine gelatin; and (3) 15 patients with atopic dermatitis and specific IgE to fish meat. Various fish gelatins (type I collagen) were prepared from fish skin. IgE antibody to fish gelatin was analyzed by using ELISA and immunoblotting. RESULTS: Of 10 patients with fish allergy, 3 had specific IgE to fish gelatin. Of two patients with fish allergy and bovine gelatin allergy, all had specific IgE to fish gelatin. Of 15 patients with atopic dermatitis and specific IgE to fish meat, 5 had specific IgE to fish gelatin. Furthermore, IgE from pooled serum of the patients reacted with both the alpha1 and alpha2 chains of fish type I collagen in immunoblots. There is cross-reactivity among gelatins from various fishes, but there is little cross-reactivity between fish and bovine gelatins. CONCLUSION: Some fish-sensitive patients possessed IgE antibody to fish gelatin. Fish gelatin (type I collagen) might be an allergen in subjects with fish allergy.

Adolescent↗
PubMed2000

Longer-term diabetic patients have a more frequent incidence of nosocomial infections after elective gastrectomy.

UNLABELLED: Diabetes mellitus (DM) is one of the risk factors for the development of postoperative nosocomial infections in surgical patients. We conducted this retrospective study to elucidate the perioperative risk factors for postoperative nosocomial infections in diabetic patients undergoing elective gastrectomy. Chart review was performed on diabetic and nondiabetic patients undergoing elective gastrectomy for gastric malignancy from January 1992 through April 1999. Fourteen of the 83 diabetic patients, and 23 of the 284 nondiabetic patients developed postoperative nosocomial infections. Statistical comparisons of multiple variables were made between patients with and without postoperative nosocomial infections. In diabetic patients, univariate analysis showed that longer-term DM (especially longer than 10 yr) was associated with a significantly increased risk for postoperative nosocomial infections. Multiple logistic regression analysis showed that DM lasting longer than 10 yr was an independent risk factor for postoperative nosocomial infections (odds ratio, 6.8; 95% confidence interval, 1.7 to 27.1). In nondiabetic patients, similar analysis showed that age was an independent risk factor for postoperative nosocomial infections. We conclude that patients with longer-term DM had a significantly greater incidence of postoperative nosocomial infections after elective gastrectomy. IMPLICATIONS: Postoperative nosocomial infection is one of the major problems in diabetic patients. This study demonstrated that postoperative nosocomial infections were more common in patients undergoing elective gastrectomy if they had diabetes mellitus longer than 10 yr.

Aged↗