Biomedical subjects

S Shichijo

Publications and source records attributed to S Shichijo.

At least 55 records · Page 3Linked to original sources

PubMed1995

Expression of MAGE-1, MAGE-2, MAGE-3/-6 and MAGE-4a/-4b genes in ovarian tumors.

MAGE genes encoding tumor-rejection antigens recognized by cytotoxic T lymphocytes are expressed at the mRNA level in various malignant tumors. We have investigated the expression of genes MAGE-1, -2, -3/-6 and -4a/-4b at the mRNA level in malignant and non-malignant ovarian tumors as well as in normal ovaries by reverse transcription-polymerase chain reaction. MAGE-1, -2, -3/-6 and -4a/-4b were expressed in 12, 5, 11 and 4 of 58 malignant tumors, respectively. The majority of these MAGE-mRNA-positive tumors were histologically surface-epithelial-stromal tumors, in particular serous adenocarcinomas. They mostly consisted of either advanced-stage or recurrent tumors. In contrast, neither benign tumors nor normal ovaries expressed any of the MAGE genes investigated. A 46-kDa MAGE-1 protein was identified in MAGE-1-mRNA-positive serous adenocarcinomas by immunoblot analysis with polyclonal anti-MAGE-1 antibody. These results provide important information for specific immunotherapy of ovarian serous adenocarcinomas with MAGE gene products.

Amino Acid Sequence↗

PubMed1995

Establishment of an enzyme-linked immunosorbent assay (ELISA) for measuring cellular MAGE-4 protein on human cancers.

The MAGE genes encoding tumor-rejection antigens are expressed on various human cancers. An enzyme-linked immunosorbent assay (ELISA) was established for measuring cellular MAGE-4 protein (MAGE-4a and/or -4b) expressed on human tumor cells using a monoclonal antibody (mAb) and polyclonal Ab to recombinant MAGE-4b protein. Both the R5 mAb (IgG1) and the polyclonal Ab recognized a 45 kDa protein in extracts of MAGE-4 mRNA positive cancers, and showed no apparent cross-reactivity to the other MAGE gene products (MAGE-1, -2, -3, -6, and -12) by the immunoblot analyses. The R5 mAb and the polyclonal Ab primarily recognized one (the position 119-133) and two oligopeptides (the positions 119-133 and 259-273), respectively, among a series of 31 different MAGE-4b oligopeptides. The amino acid sequences of these two peptides were identical to those of MAGE-4a and -4b, but differed from those of all the other MAGE proteins (MAGE-1, -2, -3, -6, and -12). Substitution of glycine for amino acid in position 123 (arginine, R), 124 (lysine, K), 126 (R) or 128 (K) in a MAGE-4b oligopeptide of the position 119-132 severely decreased the reactivity of the R5 mAb to the oligopeptide. This ELISA also showed no apparent cross-reactivity with the other MAGE gene products (MAGE-1, -2, -3, -6, and -12). The minimum detectable level of MAGE-4 protein was determined to be 10 pg/well (100 pg/ml). The results suggest that this ELISA is a reliable and quantitative method to measure cellular MAGE-4 protein that is a potential target molecule for specific immunotherapy of human cancers.

Amino Acid Sequence↗

PubMed1995

Identification of MAGE-1 and MAGE-4 proteins in spermatogonia and primary spermatocytes of testis.

The MAGE genes encoding tumor rejection antigens recognized by CTLs are expressed at the mRNA level in various cancers and in the testis but not in the other normal tissues. The expression of MAGE-1 or MAGE-4 protein in the testicular cells was studied immunohistochemically with the antibodies to the recombinant MAGE-1 or MAGE-4 protein. Both proteins were identified in the nucleus and cytoplasm of spermatogonia and in primary spermatocytes but not in spermatids or Sertoli's cells. Therefore, MAGE proteins are normal tissue antigens compartmentalized in particular testicular cells playing an important role in the early phase of the spermatogenesis.

Aged↗

PubMed1995

Sequence analysis of the MAGE gene family encoding human tumor-rejection antigens.

The MAGE multigene family, which includes the MAGE-1 and -3 genes that encode tumor-rejection antigens on HLA-A1 recognized by cytotoxic T-lymphocytes (CTL), is preferentially expressed at the mRNA level on human malignant cells, but not on normal cells. However, little is known about the MAGE-4, -41 and -6 genes. In this study, we have amplified 1040 bp (MAGE-1), 1061 bp (MAGE-3 and -6) and 1064 bp (MAGE-4 and -41) cDNA fragments, including the entire coding sequences (927-951 bp), using the reverse transcription-polymerase chain reaction (RT-PCR) method followed by nucleotide (nt) sequencing. One member had greater than 80 or 66% homology with the other members at the nt or deduced amino acid (aa) levels, respectively. Higher homology was found between MAGE-3 and -6 (98% at the nt level) and also between MAGE-4 and -41 (98%). The results of this investigation demonstrated high homology, as well as the clear differences between the members of the MAGE family at the coding sequence level.

Amino Acid Sequence↗

PubMed1995

Detection of MAGE-4 protein in lung cancers.

Expression of genes of the MAGE family, which encode tumor-rejection antigens recognized on HLA-AI and -Cw1601 by cytotoxic T lymphocytes (CTL), was investigated in lung cancers at the mRNA [MAGE-1, -2, -3/-6, and -4 (4a and/or 4b)] and protein (MAGE-4) levels. MAGE-1, -2, -3/-6 and -4 genes were expressed, respectively, at the mRNA level in 6, 7, 20 and 7 of 53 lung cancers (50 non-small-cell lung cancers and 3 small-cell lung cancers) by the reverse transcription-polymerase chain reaction (RT-PCR) method. Polyclonal antibody (Ab) and monoclonal antibody (MAb) against recombinant MAGE-4b protein were developed to detect MAGE-4 protein. Both the polyclonal Ab and the R5 MAb recognized a 45-kDa protein in extracts of MAGE-4 mRNA-positive lung cancers, and showed no apparent cross-reactivity with the other MAGE gene products except with MAGE-4a by immunoblot analyses and transfection experiments. MAGE-4 protein was detected on 13 of 44 (30%) lung cancers (18 to 55,989 pg/mg) by ELISA with the polyclonal Ab and R5 MAb. These 13 lung cancers consisted of 6 of 6 MAGE-4 mRNA-detectable and 7 of 38 MAGE-4 mRNA-undetectable lung cancers. Histologically, these comprised 7 of 10 squamous-cell carcinomas, 4 of 30 adenocarcinomas and 2 of 3 small-cell lung cancers. The proportions of MAGE gene-positive samples, at both the mRNA and protein levels, correlated with the size of the primary tumors and with regional node involvement. These results should provide important information on specific immunotherapy of lung cancers using MAGE gene products.

Animals↗

PubMed1995

Nonsteroidal anti-inflammatory drugs differentially regulate cytokine production in human lymphocytes: up-regulation of TNF, IFN-gamma and IL-2, in contrast to down-regulation of IL-6 production.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are well known as immunoregulators, but the mechanisms of their action are not fully explained by the inhibition of PG synthesis. We have investigated the effect of NSAIDs on cytokine production in human PBMC and T cell clones (TCC). NSAIDs up-regulated TNF, IFN-gamma and IL-2 production at both the mRNA and protein levels, and IL-12 expression at the mRNA level. In contrast, NSAIDs down-regulated IL-6 production both at the mRNA and protein levels, and down-regulated IL-4 mRNA expression. The modulation at the mRNA level became detectable 1 h after culture. This modulation was also observed at the level of TCC. Indomethacin (IM) enhanced TNF production in all the eight TCC that were established from a patient with human T lymphotrophic virus type 1 uveitis or pulmonary sarcoidosis, and suppressed IL-6 production in six of the eight TCC, without affecting their low levels of PGE2 production. IM also enhanced TNF and suppressed IL-6 production, respectively, in both IL-2-activated PBMC and IL-2-dependent NK cell line, with the inhibition of their high levels of PGE2 production. Culture with PGE2 alone suppressed TNF production by three of the six TCC and NK cell lines, which was neutralized by addition of IM. It had, however, no effect on TNF production by the remaining three TCC or IL-2-activated PBMC. The effects of PGE2 on IL-6 production also varied among TCC.(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed1995

Expression of the MAGE gene family in human lymphocytic leukemia.

The MAGE gene family, encoding tumor-rejection antigens recognized by cytotoxic T lymphocytes, is frequently expressed in human solid cancers. However, its expression in leukemia has not been well studied. We have investigated MAGE gene expression at the mRNA level in human leukemia. The MAGE gene family was expressed in 17 of 34 (50%) examples of T cell leukemia (12/21 patients' peripheral blood mononuclear cells and 5/13 cell lines), in 7 of 16 (44%) cases of B cell leukemia (1/8 and 6/8 respectively), but in none of 23 myelomonocytic leukemia cases (0/16 and 0/7), as evaluated by the primers common to the MAGE-1, -3, -4 (-4a and/or -4b), and -6 genes and the semi-quantificative reverse transcription/polymerase chain reaction method. None of a panel of normal lymphoid cells expressed the MAGE gene family. As revealed by the primers specific for each of the MAGE genes, the MAGE-1, -2, -3, -4 or -6 gene was expressed in 8, 8, 6, 2, or 6 respectively out of 23 types of leukemia cell lines. Expression of the MAGE-1 protein in both the cell lines and patients' cells was confirmed by immunoblot analysis with the polyclonal antibody to recombinant MAGE-1 protein. Cellular MAGE-4 protein in the cell lines was measured by an enzyme-linked immunosorbent assay with the polyclonal and monoclonal antibodies to recombinant MAGE-4b protein. In summary, the MAGE gene family was found to be expressed in the substantial proportion of T cell leukemias, but in no case of myelomonocytic leukemia. Antigens coded by the MAGE gene family could be important molecules for understanding specific immunity against lymphocytic leukemia.

Acute Disease↗

PubMed1995

Expression of MAGE-1 gene by esophageal carcinomas.

Expression of the MAGE genes encoding tumor-rejection antigens on HLA-A1 and -Cw1601 recognized by cytotoxic T lymphocytes was investigated in esophageal carcinomas at the mRNA level by the semiquantitative reverse transcription-polymerase chain reaction method. MAGE-1 and -2 genes, but not MAGE-3, -3/-6 and -4a/-4b genes, were expressed in substantial proportions of the primary esophageal carcinomas and their metastatic lymph nodes. The proportion of MAGE-positive samples in the primary esophageal carcinomas correlated with the T factor of the TNM classification (pT1: 2 of 12 tumors, pT2: 1 of 6, pT3: 12 of 29, and pT4: 7 of 18). These results have important implications for specific immunotherapy of esophageal carcinomas using MAGE-1 gene product.

Antigens, Neoplasm↗

PubMed1995

Adenosine deaminase isoenzyme levels in patients with human T-cell lymphotropic virus type 1 and human immunodeficiency virus type 1 infections.

In serum, the enzyme adenosine deaminase (ADA) is known to be divided into two isoenzymes, ADA1 and ADA2, which have different molecular weights and kinetic properties. The present study investigated ADA isoenzyme levels in the sera of patients infected with retroviruses associated with adult T-cell leukemia (ATL), human T-cell lymphotropic virus type 1 (HTLV-1)-associated myelopathy (HAM), and AIDS, ADA isoenzyme activities were found to be significantly (P < 0.001) higher in the sera of patients with ATL, HAM, and AIDS than in the sera of healthy controls. In the case of the ADA subtypes in the sera of patients with ATL, ADA1 activity was significantly (P < 0.001) elevated in patients with the acute and lymphoma types of ATL compared with that in patients with the chronic and smoldering types of ATL. ADA2 activity was significantly elevated in the sera of patients with the acute, lymphoma, and chronic types of ATL (P < 0.001) compared with that in patients with smoldering ATL and HTLV-1 carriers. In the case of patients with human immunodeficiency virus type 1 (HIV-1) infection, ADA1 and ADA2 activities in the sera of patients with AIDS and HIV-1 antibody-positive individuals were significantly (P < 0.001) higher than those in the sera of HIV-1 antibody-negative individuals. A significant elevation in ADA2 activity was also seen in the sera of AIDS patients (P < 0.01) compared with that in the sera of HIV-1 antibody-positive individuals. These results suggest that the magnitude of elevation of ADA isoenzyme levels in serum correlates well with the clinical conditions of the patients with these diseases. Measurement of the activities of ADA isoenzymes may therefore provide an additional parameter for distinguishing the subtypes of ATL and may prove to be useful as prognostic and therapeutic monitors in diseases associated with HTLV-1 and HIV-1 infections.

Acquired Immunodeficiency Syndrome↗

PubMed1993

A monoclonal antibody (H227) recognizing a new epitope of 4F2 molecular complex associated with T cell activation.

We developed and characterized a monoclonal antibody (mAb), H227, recognizing a new epitope of human 4F2 molecular complex whose roles remain to be identified. Both staining patterns and molecular sizes recognized by H227 mAb were the same to those of 4F2 mAb. Thus, both H227 and 4F2 mAbs were reactive to monocytes, thymocytes, and activated lymphocytes and immunoprecipitated the 125-kDa molecular weight membrane protein under nonreducing conditions and 85-kDa heavy-chain and 40-kDa light-chain proteins under reducing conditions. These two mAbs immunoprecipitated only the 85-kDa heavy-chain protein when the cell lysate was initially treated with dithiothreitol. Sequential immunoprecipitation proved their cross-reactivity. Both the mAbs inhibited phytohemagglutinin- or concanavalin A-induced proliferation of peripheral blood mononuclear cells (PBMC). In contrast to these similarities, the pretreatment of cells with one mAb failed to block the reactivity to the other, suggesting that their epitopes were different from each other. Furthermore, only H227 mAb augmented phorbol myristate acetate (PMA)-induced PBMC proliferation in association with increase in interleukin 2 receptor expression. In summary, H227 mAb recognizes a new epitope of 4F2 heavy chain that might be involved in the PMA-induced T cell activation pathway and therefore shall be a useful tool for understanding the roles of the 4F2 molecular complex.

Antibodies, Monoclonal↗

PubMed1993

A novel antigen (H47 Ag) on human lymphocytes involved in T cell activation.

Surface molecules involved in human T cell activation were investigated using a newly developed monoclonal antibody (H47 mAb). H47 antigen (Ag) recognized by H47 mAb was expressed on approximately 10% of resting T cells (mostly CD4-CD8+), 30% of phorbol 12-myristate 13-acetate (PMA)-activated T cells (both CD4+CD8- and CD4-CD8+), and most NK, B cells, and monocytes in the peripheral blood mononuclear cells (PBMC). H47 mAb respectively immunoprecipitated a 100 or 120-kD molecular weight (MW) membrane protein of T cells and monocytes under nonreducing or reducing conditions, suggesting that H47 Ag consists of a single polypeptide that has intramolecular disulfide bonds. H47 mAb significantly enhanced PMA-induced proliferation of PBMC in a monocyte-independent fashion. H47 mAb, however, failed to enhance T cell proliferation induced by anti-CD3 mAb, anti-CD2 mAb, or phytohemagglutinin (PHA). H47 mAb also enhanced PMA-induced interleukin-2 receptor (IL-2R) expression and IL-2 synthesis, but did not induce a change in intracellular free calcium ([Ca2+]i) of T cells. These results suggest that H47 Ag is a new membrane molecule involved in PMA-induced T cell activation.

Animals↗

PubMed1992

Multiple intracellular signaling pathways of the neuropeptide substance P receptor.

The rat substance P (SP) receptor cDNA has been transfected into cultured rat KNRK cells, and a stable cell line expressing functional SP receptors established. Upon stimulation with SP, these cells responded by simultaneously activating two signaling pathways: the mobilization of intracellular Ca2+ and the raising of cyclic adenosine triphosphate (cAMP) levels. Both Ca2+ and cAMP responses were elicited in a similar dose-dependent manner with half maximal concentrations of approximately 5 x 10(-10) M. Following ionomycin treatment SP-dependent Ca2+ responses were abolished, whereas cAMP responses were preserved. Forskolin eliminated the SP-dependent cAMP elevation, however, the SP-induced Ca2+ mobilization remained unchanged. Furthermore, treatment with phorbol esters had no significant effect on either of the two SP-induced responses. Thus it appears that the SP receptor is capable of independently activating Ca2+ mobilization and cAMP pathways. These results may provide new insights for further understanding the diverse activities of SP in various systems in vitro and in vivo.

Animals↗

PubMed1992

Comparison between natural and immunized anti-GM1 antibodies analyzed by marker releasing rate assay with liposome.

Naturally occurring rabbit anti-GM1 was compared with immunized induced anti-GM1 antibody in the same individual rabbits by the characterization of immune potency determined by complement-dependent damage to liposomes containing GM1 as antigen. Antigen-antibody reaction was the rate determining step in this assay. The maximum rate was measured by the first derivative conversion monitoring in spectrophotometry and the maximum level of marker released was measured by the conventional method described previously. When the concentration of sera required to get fifty percent of maximum values in each assay were compared, the ratios (rate assay/conventional assay) were 5.7 to 10.0 in natural antibodies and 3.2 to 2.3 in immunized induced antibodies, respectively. Furthermore, the maximum rate of marker release from liposome was not correlated with the maximum level of marker released depended on the sources of antibodies. These results suggest that natural antibody, even the titer was high in conventional liposome assay, has different characteristics compared with the immunized induced antibody.

Animals↗

PubMed1991

Histamine effects on the 5-HT1c receptor expressed in Xenopus oocytes.

To analyze the cross-reactivity between serotonin (5-HT) and histamine, the in vitro transcribed RNA for the 5-HT1c receptor was functionally expressed in Xenopus oocytes. 5-HT significantly increased 45Ca2+ efflux in RNA-injected oocytes, but not in uninjected and water-injected control oocytes. Furthermore, histamine and the H1 receptor agonists, but not the H2 and H3 agonists, significantly induced 45Ca2+ efflux in 5-HT1c receptor RNA-injected oocytes, but not in uninjected and water-injected oocytes. However, the H1, H2, and H3 antagonists failed to inhibit histamine-induced 45Ca2+ efflux at 10(-6) M. This finding suggests that the 5-HT1c receptor can be activated by both 5-HT and histamine, although the action of histamine is different from classic histamine pharmacology.

Animals↗

PubMed1991

[Rapid assay of hyaluronic acid in serum].

A method for measurement of hyaluronic acid (HA) level in serum was developed based on using "hyaluronic acid binding protein" (HABP)-coated polystyrene beads. After the beads and test serum being mixed, the mixture was incubated together with reaction buffer for 2 hours, and then the beads were washed. Subsequently, biotinylated HABP was added to the washed beads and incubated for 1 hour. Then peroxidase-conjugated avidin was added to the mixture and incubated again for 1 hour. After the beads being washed, a substrate solution was added to the washed beads and left for 1 hour. Then the reaction was stopped by adding 2N-H2SO4. The absorbance at 492 nm was recorded. The analytical range of HA in serum by this method was found to be between 10-800 micrograms/l, and the precision of the HA assay (CV%) was between 3.0-8.4 in the "with-in" assay (n = 10), and 4.8-8.9 in the "between" assay (n = 5). The analytical recovery of HA assay was between 92-115%. In this study, the results in screening of the serum HA level in RA patients (n = 107), OA patients (n = 16) and healthy subjects (n = 30) showed that the HA level of RA patients was demonstrated significantly higher than that of healthy subjects and OA patients.

Adult↗

PubMed1989

Assay of serum hyaluronic acid in clinical application.

A sandwich-binding protein assay to determine the concentration of hyaluronic acid (HA) in body fluids has been developed. In this method, a hyaluronic acid binding protein (HABP) was adsorbed to the surface of a solid phase, and HA bound to HABP on the solid phase was detected by biotin-conjugated HABP. The method could assay HA levels within 6 hours using precoated microwells with HABP. HA could be determined in the range of 2-500 micrograms/l by this method using 50 microliters of serum. Within-run precision (CV) was 5.2-10.2%. The specificity of HABP to HA was confirmed by the elimination of the reaction with treatment by hyaluronidase digestion. Serum HA levels (median; range) of patients with rheumatoid arthritis (34; 2-187 micrograms/l) were shown to be higher than those with osteoarthritis (1; 1-21 micrograms/ml) and healthy controls (2; 1-8 micrograms/ml). No correlation between levels of HA and rheumatoid factor was found. HA was demonstrated to be a potential diagnostic marker for rheumatoid arthritis, and this HABP assay could be useful for determination of HA in clinical laboratory tests.

Adult↗

PubMed1989

Diagnostic value of ganglioside patterns in plasma of human diseases.

A method that could be facilitated for the quantitation and qualitation of gangliosides in human plasma was developed and applied in the present study for characterization of ganglioside patterns in plasmas of patients with neoplastic diseases including gastric cancer, adult T-cell leukemia (ATL), and acute lymphocytic leukemia (ALL). As a retrovirus-infected disease with different clinical entity, human T-cell lymphotropic virus type I-associated myelopathy (HAM) was also subjected to this study. The results were compared with the patterns obtained from normal control plasmas. The analytical data revealed that GM3 increased in gastric cancer, GT1b decreased in HAM and ATL, and GD3, GM1 and GM3 decreased in ALL. There were close correlations between various human diseases and the presence of gangliosides with their specific patterns. Furthermore, gangliosides purified from plasma of patients with HAM significantly inhibited the expression of CD4 antigen on human T lymphocyte membrane. Therefore, analytical studies of plasma gangliosides could provide diagnostic and therapeutic values in retroviral infections.

Adult↗

PubMed1989

Further studies on a chemoattractant derived from Aleurites Fordii Hemsl. seed.

1. Activities stimulatory to PMN superoxide generation and chemotactic function were present in two separate gel chromatographic fractions of AFH-S and were designated AFH-S1 (500 kD) and AFH-S2 (38 kD). 2. Characterization studies of these activities revealed physiologic and pharmacologic characteristics dissimilar to those of other previously studied chemoattractants such as bacterial peptide FMLP.

Calcium↗