Development of hyperthyroidism in donor and recipient after allogeneic bone marrow transplantation.
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Adolescent↗
Biomedical subjects
T Naoe
Publications and source records attributed to T Naoe.
[Simultaneous analysis of c-myc protein expression and cell cycle with monoclonal antibody and flow cytometry].
C-myc protein plays an important role in the regulation of cell proliferation and differentiation. In this paper, c-myc protein and DNA were doubly stained and analysed simultaneously with flow cytometry (FCM). At first, three fixatives, ethanol, methanol and paraformaldehyde, were examined using HL-60 cells, among which 50% ethanol was found to be optimal for each staining. After fixation, the cells were stained with a monoclonal antibody against c-myc protein, followed by DNA staining with PI. Simultaneous analysis demonstrated that c-myc protein was constantly expressed through the cell cycle and that the protein amount at the G2 + M phases was 1.5 times higher than that at the G1 phase. Further, the differentiation study with TPA and RA revealed that the growth, S phase and also c-myc protein expression were suppressed during the differentiation.
Cell Cycle↗
Cytogenetic and cellular characteristics of a human embryonal rhabdomyosarcoma cell line, RMS-YM.
A human tumour cell line, designated RMS-YM, was established from a childhood rhabdomyosarcoma. The monolayer cells were polygonal, round or spindle-shaped. The cells became multilayered and formed many focal piles when confluent. RMS-YM became stable with a doubling time of about 30 h and has been maintained for 104 passages to date. Tumourigenicity of the cells was confirmed by heterotransplantation into nude mice. Morphological features were similar to those of the primary tumour, and myofibrils were found by electron microscopy. The expression of desmin and human myoglobin, and high levels of striated muscle system specific enzymes were recognised. Chromosomal analysis revealed possible gene amplification in the form of homogeneously staining regions. Oncogene analysis was performed on the primary tumour and the cell line, but neither N-myc nor N-ras genes were amplified, nor were Ki-ras, Ha-ras or N-ras genes mutated at the 12th, 13th and 61st codons. The RMS-YM cell line may provide a system to identify novel genes which are amplified in rhabdomyosarcoma.
Animals↗
A case of lupus nephritis showing rectal erosion with cytomegalovirus infection.
A case of lupus nephritis showing rectal erosion with cytomegalovirus infection is described. The patient revealed nephrotic syndrome. She complained of skin ulcer with livedo reticularis on the right breast, purpura with thrombocytopenia, hepatic dysfunction, and bloody stool. Renal biopsy demonstrated diffuse proliferative lupus nephritis with prominent wire loops. Intestinal fiberscopy showed rectal erosion which was a cause of bloody stool, and rectal biopsy revealed cytomegalic inclusions characteristic of cytomegalovirus infection. Immunosuppression due to combined therapy with prednisolone and mizoribine could have led to reactivation of latent cytomegalovirus infection, resulting in the rectal erosion with bloody stool.
Adult↗
Disappearance of minimal residual lymphoblastic leukemia cells 6 months after allogeneic bone marrow transplantation without GVHD.
The recent development of the polymerase chain reaction (PCR) has enabled us to determine the hypervariable sequence of immunoglobulin heavy chain known as complementarity determining region (CDR)-III. We amplified the leukemia-specific CDR-III from common acute lymphoblastic leukemia (cALL) cells using the PCR and determined its sequence. To detect minimal residual leukemia (MRL) cells, a second round PCR was performed with clone-specific primers corresponding to 5' and 3' ends of CDR-III to detect MRL cells. A million-fold diluted leukemia cells were clearly detected. Using stepwise diluted materials, the number of residual cells was semiquantitatively estimated. In one patient with cALL, induction chemotherapy resulted in a hematologically complete remission with only a 2-log reduction of the leukemia cells. MRL cells at a level of 10(-6) were also detected within 1 month after bone marrow transplantation (BMT), but leukemia cells were not detectable 6 months after BMT.
Adult↗
Treatment of myelodysplastic syndromes with orally administered N-(2S, 3R)-3-amino-2-hydroxy-4-phenylbutyryl-L-leucine (ubenimex).
N-(2S, 3R)-3-amino-2-hydroxy-4-phenylbutyryl-L-leucine (ubenimex) was administered orally, to patients with myelodysplastic syndromes (MDS) and acute leukemia derived from MDS, in a multi-institute study. Out of 77 patients evaluated, one achieved a complete remission, three a good response and two a partial response while 71 failed to respond to a daily oral administration of 30 mg ubenimex. The overall response rate was 7.8% (95% confidence limits; 3.6-16.0%); 7.0% (3.0-15.4%) in 71 MDS and 16.6% (3.0-56.3%) in acute six leukemias derived from MDS. Responses continued for six to 24 (median 10.5) weeks. No serious hematologic, biochemical or clinical toxicity was encountered, except for gastro-intestinal (GI) toxicity in one patient. The present study demonstrated ubenimex not to be generally beneficial for patients with MDS, and not to be recommended as a standard treatment for the disease.
Acute Disease↗
New enzyme-linked immunosorbent assay for glycocalicin in plasma.
A new sandwich-type enzyme-linked immunosorbent assay for quantifying glycocalicin, a proteolytic fragment of platelet membrane glycoprotein Ib, is described. The assay is based on the use of two monoclonal antibodies raised against glycoprotein Ib and involves the avidin-biotin technique. The detection limit is 7 micrograms/L and the range of glycocalicin determined in plasma is 0.01 to 1 mg/L. Assay time is 2 h. The intra-assay CV ranged from 3.6% to 5.2%, the interassay CV from 5.4% to 8.0%. Analytical recovery of purified glycocalicin added to a plasma pool averaged 96%. In 36 healthy subjects, the mean glycocalicin concentration in plasma was 0.36 (SD 0.07) mg/L (2.7 nmol/L). We conclude that this assay is suitable for measuring glycocalicin in plasma and is also more sensitive and precise than the previously published immunoassays based on competitive binding assay.
Adolescent↗
[Is allogeneic bone marrow transplantation the first choice for acute leukemia in first remission? From chemotherapeutical point of view].
Since 1984 several prospective randomized clinical trials have been reported that the disease-free survival for BMT in first remission was superior to chemotherapy for AML. However, there raise two main controversies about these conclusions. First, in last several years intensive consolidation therapy was applied and nearly 50% of patients are reported to be alive 5 years. Second 10% to 50% BMT scheduled patients were excepted mainly because of their condition. On the contrary, about ALL, even these several years, age is the still main risk factor independently of therapy. Reports with good 5 year disease free survival all due to their patients' youth. The prognosis of ALL patients age 30 or more are poor and BMT seems to be a first choice in first-remission in patients under age 40. Only large-scale prospective randomized study among patients with or without HLA-identical donors will give the answer to this important issue.
Acute Disease↗
Glycine to aspartic acid mutations at codon 13 of the c-Ki-ras gene in human gastrointestinal cancers.
Point mutations of c-ras genes were analyzed in human gastrointestinal cancers. DNA obtained from the tissues was amplified by polymerase chain reaction and then analyzed by dot blot hybridization assay with oligonucleotide probes to detect mutations at codons 12, 13, and 61 of c-Ki-ras, c-Ha-ras, and c-N-ras. In two of 25 cases of stomach cancer point mutations at codon 13 of c-Ki-ras were found. In colorectal cancer, eight of 30 cases showed mutations: four cases of codon 12 and one case at codon 13 of c-Ki-ras and two cases at codon 61 and one case at codon 13 of c-N-ras. These results may indicate involvement of a wide variety of c-ras gene point mutations, in addition to those at codon 12 of c-Ki-ras, in oncogenesis of human gastrointestinal cancers. In all three mutations of c-Ki-ras at codon 13 which had been seldom found in human cancers, glycine to aspartic acid mutations due to identical G to A transition at the second nucleotide were observed.
Aspartic Acid↗
Autoantibodies to c-myc nuclear protein products in autoimmune disease.
This report describes the identification of autoantibodies that react with c-myc proteins in the sera of some patients with autoimmune diseases. Sera from nine to 30 patients, including six with systemic lupus erythematosus, one with mixed connective tissue disease, one with dermatomyositis and one with autoimmune haemolytic anaemia, reacted in immunoblotting assays against a truncated human c-myc protein p42 produced by Escherichia coli. Furthermore, these sera exhibited double bands of 58,000 and 60,000 MW in the nuclear fraction of HL-60 cells with the amplified c-myc gene. These bands were the same as those detected by the anti-human c-myc protein monoclonal antibody (MYC-1) by immunoblotting assay. The binding activities to 58,000 and 60,000 MW were not reduced by DNase I digestion with the sera and could be absorbed by a p42-bound affinity column and recovered after elution.
Antibodies, Antinuclear↗
Correlation of c-myc expression with nuclear pleomorphism in human renal cell carcinoma.
The expression of the c-myc gene product in renal cell carcinomas was examined by immunostaining with monoclonal antibody (mAb) MYC-1. The effects of preservation and fixation of tissues on staining were first examined. In cryostat sections fixed with 4% buffered formalin for 15 min, staining was observed in the nucleus. On the other hand, in paraffin sections after fixation with 10% formalin, staining was observed in the cytoplasm, but not in the nucleus. Because c-myc protein has been shown to be a nuclear protein, the finding that c-myc protein was not detectable in the nucleus appeared to be due to the preservation or fixation procedures used. Therefore, cryostat sections fixed with 4% formalin were used to investigate the correlation between the reaction of MYC-1 mAb and nuclear pleomorphism in primary and metastatic renal cell carcinomas. Among 41 primary tumors, positive staining was observed in 2 of 17 tumors (12%) of grade 1, 17 of 21 (81%) of grade 2, and all 3 (100%) of grade 3. Among 17 metastatic tumors, positive staining was not observed in any of the 5 (0%) of grade 1 but was observed in 2 of 4 (50%) of grade 2 and all 8 (100%) of grade 3. Thus, the frequency of the positive reaction with MYC-1 mAb was correlated with nuclear pleomorphism in primary and metastatic renal cell carcinomas. The reaction of Ki-67 mAb, which recognized a nuclear antigen present in proliferating cells, was also correlated with nuclear pleomorphism. These findings suggest that the c-myc gene product plays a role in cell proliferation in renal cell carcinomas.
Antibodies, Monoclonal↗
Diversity of cellular molecules in human cells detected by monoclonal antibodies reactive with c-myc proteins produced in Escherichia coli.
Six clones of monoclonal antibodies, MYC-1 to -6, were prepared by using two kinds of truncated c-myc proteins, p23 and p42, produced in Escherichia coli as immunogens. Analysis with enzyme-linked immunosorbent assays and immunoblotting assays with peptides produced in Escherichia coli showed that 5 clones of monoclonal antibodies, MYC-1 to -4 and -6, were reactive with c-myc protein encoded by exon 2. The remaining one clone, MYC-5, was reactive with the portion of c-myc protein encoded by exon 3. All monoclonal antibodies were also reactive with phosphorylated c-myc protein produced by insect cells infected by the baculovirus expression vector with the human c-myc gene. With immunoblotting assays using cellular lysates, MYC-1 and -3 detected bands at the levels of 58 kDa and 60 kDa, MYC-5 detected a band at 56 kDa and MYC-6 detected bands at 68 kDa and 75 kDa. All of these bands were detectable in nuclear extracts of HL-60 and Colo320, both of which have amplified c-myc genes, and also the extract of RmycYl which is the c-myc gene transfectant into 3Yl rat cells. None of them was detectable in peripheral blood mononuclear cells and 3Yl, both of which lacked activated c-myc genes. This indicates that these nuclear proteins are either c-myc gene products or molecules closely related to the c-myc gene. The remaining two clones, MYC-2 and -4, detected a band at the level of 85 kDa in cytoplasmic extracts of all the above-mentioned cells independent of the presence of the c-myc gene. This suggests that 85 kDa protein might be irrelevant to the c-myc gene. The 56 kDa protein was detectable by MYC-5 in phytohemagglutinin-stimulated peripheral blood mononuclear cells as well as leukemic cells of some patients.
Animals↗
A case of cALL relapse after allogeneic BMT: recurrence of recipient cell origin, initially determined as being that of donor cell origin by sex chromosome analysis.
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Adolescent↗
Southern blot analysis of Bcr, N-ras, H-ras, c-myc and myb proto-oncogenes in blast crisis of chronic myelogenous leukemia.
Proto-oncogenes were studied by Southern blotting in 12 patients with chronic myelogenous leukemia in blast crisis (CML). Cytogenetically, all cases had Philadelphia (Ph1) chromosome and 4 had additional chromosomal abnormalities including +8, +19, i(17q), double Ph1. The bcr gene was rearranged in all cases and the region of breakage points were located in the 5.8 kb Bgl II-Bam HI fragment of bcr. In a case with double Ph1 the breakage points of bcr were thought to be different from each other. We analyzed other proto-oncogenes (N-ras, H-ras, c-myc and myb) by Southern blotting, but did not find any abnormalities.
Adult↗
Production of a truncated human c-myc protein which binds to DNA.
Two kinds of truncated human c-myc proteins were produced in Escherichia coli. The human c-myc gene is composed of three exons, exons 2 and 3 having coding capacity for a protein of 439 amino acids. 252 N-terminal amino acids are encoded by exon 2, the C-terminal 187 amino acids being encoded by exon 3. One of the proteins (p42) produced in E. coli corresponds to 342 amino acids from the 98th Gln to the C-terminus, plus 21 amino acids derived from the H-ras gene at the N-terminus. The other (p23) corresponds to 155 amino acids from the 98th Gln to the 252nd Ser, plus five amino acids (Gly-Gly-Thr-Arg-Arg) at the C-terminus, plus 21 amino acids from the H-ras gene at the N-terminus. The p23 protein was produced by using cDNA in which a frame shift occurred at the boundary between exons 2 and 3. We investigated the DNA-binding activity in p42 and p23 proteins. DNA-cellulose column chromatography showed that p42 binds to DNA, whereas p23 does not. This DNA-binding activity of p42 was inhibited by antiserum prepared against p42 but not by antiserum against p23. This indicates that the DNA-binding activity of c-myc protein is localized in the portion encoded by exon 3.
Cloning, Molecular↗
Cytogenetic characterization of a T-cell line, ATN-1, derived from adult T-cell leukemia cells.
A T-cell line, ATN-1, was established by culturing peripheral blood mononuclear cells derived from a patient with adult T-cell leukemia/lymphoma (ATL/L). Identities of the patterns of chromosomal abnormalities, cell surface phenotypes, morphologic findings, rearrangement patterns of T-cell receptor beta chain gene, and an integration site of human T-cell leukemia virus I proviral genome indicated that ATN-1 was derived from original leukemic cells. Both ATN-1 and the original leukemic cells showed a variety of patterns of chromosomal abnormalities that include 3q-, 6q-, rearrangements involving 2q31, 7q11.2, 8q11, 8q24, 19p13.3, and also 14q11 and 14q32, where genes for the T-cell receptor alpha chain and the immunoglobulin heavy chain are located. Availability of a genuine ATL/L cell line with these chromosomal abnormalities may greatly facilitate the biologic analysis of ATL/L.
Antibodies, Monoclonal↗
Serological and biochemical characterization of surface antigen expressed on human myeloid cells and identification of six distinct antigen systems.
Six distinct surface antigens selectively expressed on human myeloid cells were defined with ten murine monoclonal antibodies. Four antigens (GAp95, GAp170, GAp160-220, GAp145-200) are expressed on protein molecules and one antigen (GANGL) is detected in neutral glycolipids. The biochemical nature of the remaining one (GA8.9) is unknown. GAp95 and GANGL are expressed on relatively mature myeloid cells, whereas GAp160-220 is expressed on immature myeloid cells. GAp170 and GAp145-200 are expressed on a wide range of myeloid cells. GA8.9 was found on NKM-1, a cell line of acute myelomonocytic leukemia, and also on some acute nonlymphocytic leukemia cells.
Antibodies, Monoclonal↗