Biomedical subjects

J Furuyama

Publications and source records attributed to J Furuyama.

At least 73 records · Page 4Linked to original sources

PubMed1990

Amplification and expression of a multidrug resistance gene in human glioma cell lines.

Two human glioma cell lines were examined for multidrug resistance (MDR). A vincristine (VCR)-resistant glioma cell line showed a cross resistance to Adriamycin (doxorubicin, ADR) and etoposide (VP-16) to varying extents, suggesting the presence of MDR; the resistance to VCR was considerably decreased by calcium entry blockers. On the other hand, another VCR-sensitive glioma cell line exhibited no cross resistance to ADR or VP-16. Double minute chromosomes and homogeneously staining regions as well as clonal aberrations of chromosome 7 were not observed in cytogenetic studies of multidrug-resistant and multidrug-sensitive glioma cell lines. In Northern and Southern blot analyses, MDR gene 1 (MDR1) messenger ribonucleic acid (mRNA) was shown to be overexpressed without any amplification of the MDR1 gene in multidrug-resistant glioma cell lines as compared to multidrug-sensitive glioma cell lines. It would be reasonable to suggest that amplification of the MDR1 gene may not be a sine qua non for acquisition of MDR and that the MDR1 mRNA level may be correlated with the extent of MDR.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1990

[Molecular cloning and expression of Mycobacterium tuberculosis Aoyama B peptide antigen genes in Escherichia coli--a gene encoding 60 kD antigen (AT201) and immunological activity of recombinant peptides (15 and 60 kD)].

To obtain recombinant peptides related to PPDs, we constructed a genomic library from the DNA of Mycobacterium tuberculosis Aoyama B, a standard strain in Japan to manufacture PPDs, using plasmid vectors pUC18 series. Seven clones reacting the anti-PPDs-rabbit-serum on immunoblotting were obtained, and restriction map was analysed. A nucleotide sequence and a putative open reading frame (ORF) of pAT01, encoding 15kD peptide, as well as the mode of expression was reported previously). In this study, nucleotide sequence of 60 kD peptide gene was determined, and the comparative database analysis (GENBANK) revealed a striking level of homology to so called mycobacterial heat shock protein. The expression mode of pAT201 encoding 60 kD, as well as pAT01 encoding 15 kD peptide, indicated that these peptides were not hybrid proteins with the lacZ gene product, but they were consisted of peptides only mycobacterial source. Therefore, 15 kD and 60 kD directly were subjected to immunological studies. The peptides were extracted from E. coli, carrying pAT01 or pAT201, purified through series of DEAE chromatography and followed by Detoxi-Gel to remove LPS. 15 kD peptide behaved almost similar to PPDs both in the DTH skin reaction and the lymphocyte proliferation response on guinea pigs or rats in respect to sensitivity. However, 60 kD was unique in that, and it behaved like a general mitogen. We discussed role of 60 kD peptide, comparing with the common antigen, generally found in most species of bacteria as the heat shock protein.

Antigens, Bacterial↗

PubMed1989

B cell response pathways regulated by IL-5 and IL-2. Secretory microH chain-mRNA and J chain mRNA expression are separately controlled events.

We have examined the effect of IL-5 and/or IL-2 on the expression of the secretory form of microH chain (microsecond) and J chain mRNA in a homogeneous neoplastic B cell clone, in which proliferation, IL-2R up-regulation and entry into the IgM secretory state are separately controlled events. The IL-5 signal triggers a partial induction of CL-3 cells into the IgM secretory state, characterized by a striking increase of microsecond mRNA expression and an increase in the ratio of the secretory to membrane forms of microH chain mRNA, with a modest increase of J chain mRNA. In contrast, amplification of J chain mRNA is accomplished by the late-acting B cell differentiation stimulus, IL-2, acting on IL-5-pretreated CL-3 cells or acting simultaneously with IL-5 on CL-3 cells. Such dually stimulated cells now are fully induced into IgM secreting cells. These results define the relative roles of IL-5 and IL-2 in B cell differentiation by showing important regulatory effects at the mRNA level. In addition, these results substantiate that appearance of mRNA for J chain, a molecule key to the formation of pentameric IgM, is a limiting factor for high level IgM secretion. The separate control of microsecond and J chain mRNA found in CL-3 cells stimulated with IL-5 and IL-2 elucidates a molecular mechanism by which these two lymphokines synergize in the development of CL-3 cells into IgM secreting cells.

B-Lymphocytes↗

PubMed1989

A testis-specific antigen of the C57BL/6 mouse.

Spleen cells from female mice of the C57BL/6 strain iso-immunized with an homogenate from 3-week-old mice testis were fused with P3U1 cells. After cloning, two hybridomas, producing IgM, were obtained. Tissue specificity of the monoclonal antibody (Moab) in ascitic fluid was investigated by indirect immunofluorescence. Moab 1A1 reacted specifically with the cytoplasm of spermatogonia, spermatocytes and spermatids, but not with spermatozoa. Testicular antigen recognized by Moab 1A1 (Moab1A1-TA) was prepared by tissue sonication and then subjected to gel filtration. Moab1A1-TA detected in the void volume by ELISA was analyzed by SDS-PAGE and Western blotting. Immuno-staining of membrane filters revealed a broad area within the 45-205 kD range. Moab1A1-TA was treated with proteolytic enzymes, but no changes were observed after Western blotting. Thus, Moab1A1-TA was further digested by peptidases and glycolytic enzymes, electrophoresed using cellulose acetate membranes and immuno-stained with Moab 1A1. Evidence obtained from these experiments strongly suggests that Moab1A1-TA consists of an acidic peptide and a carbohydrate molecule. The antigenicity would be included in the carbohydrate epitope. Moreover, partial digestion of Moab1A1-TA by keratanase indicates that the lacto-series structure is included in the antigenic carbohydrate moiety.

Animals↗

PubMed1989

Molecular cloning and nucleotide sequence of the gramicidin S synthetase 1 gene.

The entire gene for gramicidin S synthetase 1 (GS 1) was cloned into the plasmid vector pUC18, and the nucleotide sequences of the GS 1 gene and its flanking region were determined. The full-length clone was 4,539 base pairs long and had an open reading frame of 3,294 nucleotides coding for 1,098 amino acids. The calculated molecular weight of 123,474 agreed with the apparent molecular weight of 120,000 found in SDS-PAGE of GS 1 from B. brevis. The nucleotide sequence of GS 1 gene was highly homologous to that of tyrocidine synthetase 1. The overall similarity between the deduced amino acid sequences of the two genes was 57.5%. The gene product of clone GS309 was easily purified to an essentially homogeneous state by ammonium sulfate fractionation followed by DEAE-Sepharose CL-6B, Ultrogel AcA-34, and second DEAE-Sepharose CL-6B column chromatography. The purified protein catalyzed the D-phenylalanine-dependent ATP-32PPi exchange reaction which is specific for GS 1 activity, and the specific activity of the purified product was nearly the same as the purified GS 1 from B. brevis. The product also showed a weak phenylalanine racemase activity.

Amino Acid Isomerases↗

PubMed1989

[Familial spontaneous pneumothorax in three siblings including identical twins].

Two identical twin brothers and their sister who suffered from spontaneous pneumothorax (ages at onset, 18, 22 and 20 years) were admitted to our hospital. Their symptoms and clinical courses were similar and all three were finally treated by thoracotomy. No other members of their family had any history of spontaneous pneumothorax. None of these cases presented any abnormal level of serum ACE or alpha 1-antitrypsin. In order to establish the monozygosity of the brothers more accurately, we examined restriction fragment length polymorphisms (RFLPs) which were shown as the band patterns of hypervariable minisatellite regions in human genomes. However there have been few reports of spontaneous pneumothorax in identical twins, but the onset of the disease tends to be the same as in our cases, suggesting a genetical background of spontaneous pneumothorax.

Adolescent↗

PubMed1988

Intragenic suppression in the uvrD gene of Escherichia coli. I. Temperature-sensitive uvrD mutations.

A temperature-sensitive uvrD mutant, HD323 uvrD4, was isolated from the uvrD mutant HD4 uvrD3. The temperature sensitivity of the uvrD4 gene product was reversible. The suppressor mutation uvrD44 which rendered the uvrD3 mutant temperature-sensitive could be separated from the uvrD3 mutation by replacing the PstI fragment, which encodes the C-terminal half of the UvrD protein. The uvrD44 mutation was found to make host bacteria lethal at non-permissive temperatures only when cloned on a low copy vector pMF3. The nucleotide sequence of the uvrD3 and uvrD4 mutant genes was determined. The nucleotide change found in the uvrD3 at +1235, GAA to AAA, only alters the amino acid sequence from Glu at 387 to Lys. The uvrD44 has another nucleotide change at +1859, GAA to AAA (Glu at 595 to Lys), which is considered to be the suppressor mutation uvrD44.

Adenosine Triphosphatases↗

PubMed1988

Femininity control at the XXth Universiade in Kobe, Japan.

In 1985, we were in charge of the femininity control tests for the participants of the XXth Universiade in Kobe. At that time, we chose as screening tests checking both X chromatin from buccal mucous membrane and Y chromatin from hair roots. The number of female competitors who attended the Universiade, except tennis, gymnastics, and fencing, was 631. Among them, 108 had already had the certificate of femininity so we examined 523 female competitors. All but three of them were normal for femininity, i.e., X chromatin positive and Y chromatin negative. Two of the three exceptions were Y chromatin positive and X chromatin negative. Chromosomal analyses from their peripheral lymphocytes showed a pattern concordant with a 46,XY karyotype. The other was both X and Y chromatin positive, but we could find Y chromatin on only one slide out of three. Since this person refused to have a blood sample taken, we can only suppose her karyotype as being 46,XX/46,XY or 46,XX/47,XXY. This last case would not have been found, if we had checked only the X chromatin. From our experience, we conclude that for femininity control, it is safer to screen the Y chromatin. We would strongly advise the use of both X and Y chromatin tests for femininity control.

Female↗

PubMed1988

[Recent trends in studies on carcinogenesis in familial adenomatous polyposis].

Familial adenomatous polyposis is rare disease, but it is considered as the most important clinical model for studying cancer control and carcinogenesis in general, by its extremely high risk for colorectal cancer as well as for malignancies of diverse organs which is transmitted by autosomal dominant mode. According to Knudson, the mutation for a dominantly inherited cancer susceptibility may be the first step in a recessive change in the tumor cell and same gene may be involved in both familial and non-familial cases of the tumor. The recent studies by Bodmer et al and by Solomon et al which reported on FAP locus in chromosome 5 and on loss of heterozygosity of this locus in the colorectal cancer have made an important break through in proving the Knudson's hypothesis. The epidemiological, clinical and pathological observations on FAP including our studies has been discussed in the view of these recent development of molecular biology.

Adenoma↗

PubMed1988

Biochemical studies on lymphoblastoid cells with inherited N-acetyl-glucosamine 1-phosphotransferase deficiency (I-cell disease).

Lymphoblastoid cells transformed by Epstein-Barr virus from peripheral lymphocytes of normal individuals and I-cell disease (ICD) patients were used for the enzymic study of lysosomal hydrolases and N-acetylglucosamine 1-phosphotransferase. ICD lymphoblastoid cells secreted a larger amount of hydrolases into medium than normal cells, although the intracellular hydrolases were not deficient in ICD cells. The stimulating effect of 10 mM ammonium chloride on secretion of hydrolases was found only with normal cells, and not with ICD cells, indicating that the hydrolase molecule bearing mannose 6-phosphate was secreted. The ICD lymphoblastoid cells retained the enzymologic characteristics of both lysosomal hydrolases and N-acetylglucosamine 1-phosphotransferase seen in ICD fibroblasts, which allows us to study the pathophysiology of ICD in cells other than fibroblasts.

Adsorption↗

PubMed1988

[XYY syndrome: report of three cases].

Three cases of XYY syndrome (YY syndrome) were experienced. Two cases had such rare associated anomalies as hypospadias and cryptorchism. Pathogenesis, incidence, clinical symptoms, gonadal function and fertility of the patients with XYY syndrome are briefly discussed.

Child↗

PubMed1987

Demonstration of up-regulated IL 2 receptor expression on an in vitro cloned BCL1 subline.

We have established BCL1 CL-3 cells capable of responding to B15-TRF and interleukin 2 (IL 2). This clone has both high affinity and low affinity receptors for IL 2 (IL 2R), but IL 2 by itself did not stimulate either proliferation or immunoglobulin (Ig) secretion. B15-TRF, which possesses both growth and differentiation activity, causes an increase in size of CL-3 cells and renders CL-3 cells responsive to IL 2, including an increased expression of IL 2R (eight-fold to 10-fold) and the differentiation of CL-3 cells into Ig secretion (60 to 80% of cultured cells). CL-3 cells pretreated with B15-TRF for 12 hr become competent to respond to IL 2 by up-regulation of IL 2R within 12 hr. In contrast CL-3 cells pretreated with IL 2 for 12 hr required 24 hr B15-TRF stimulation to result in IL 2R up-regulation. Thus the ordered action of B15-TRF and IL 2 is the most effective operational pathway for the up-regulation of IL 2R. This IL 2-mediated IL 2R up-regulation and induction of Ig synthesis depends upon the concentration of IL 2 in the culture. Both responses seem to be caused by IL 2 molecules bound to high affinity IL 2R. However, the possibility of involvement of low affinity IL 2R can not be vigorously excluded. In fact the level of IL 2 required for a response is far higher than that needed for activated T cell proliferation. This cloned BCL1 subline promises to be a useful tool for studying the regulation and mechanisms of B cell responses.

Animals↗

PubMed1987

Differential hypersensitivity of xeroderma pigmentosum lymphoblastoid cell lines to ultraviolet light mutagenesis.

Survival and mutation after u.v. light irradiation were examined in four human lymphoblastoid cell lines; one cell line with normal excision-repair capacity (HH4), two excision-repair-deficient xeroderma pigmentosum (XP) cell lines from patient XP3BE (complementation group C) and XP7NI (group A), and one cell line from an XP heterozygote (XPF7NI, father of XP7NI). Relative to HH4 and XPF7NI, both XP3BE and XP7NI were more sensitive to the cytotoxic effect of u.v. by virtue of a diminished shoulder and a steeper slope in the survival curve determined by growth curve extrapolations. The mutagenesis for 6-thioguanine resistance (TGr) was measured by the limiting dilution technique using 96 well microtiter plates. Even at non-cytotoxic fluences, u.v. light induced TGr mutations in HH4 and XPF7NI with no apparent threshold, and the mutagenic response of XPF7NI was no different than that of HH4. Both XP cell lines were more sensitive than normal cells to u.v.-induced mutation. However, XP7NI cells were extremely hypermutable by u.v.; i.e. 3.3 J/m2 induced an approximately 500-fold increase of mutant fraction with the background of 2 X 10(-6). XP7NI cells remained much more mutable by u.v. even when plotted against survival, whereas the mutant fractions for XP3BE and HH4 followed the same line. These results imply that the mechanism of u.v.-induced mutation in XP7NI cells may intrinsically be different from that in XP3BE, XP heterozygote or normal cells, or that potentially mutagenic lesions are repaired much less efficiently in XP7NI cells than are potentially lethal lesions, as compared with XP3BE, XPF7NI and HH4 cells.

Cell Line↗

PubMed1986

Establishment of cell lines derived from ataxia telangiectasia and xeroderma pigmentosum patients with high radiation sensitivity.

Four human fibroblast cell lines, three of which were derived from a patient with ataxia telangiectasia and the other from a patient with xeroderma pigmentosum, were established after transfection with cloned SV40 DNA. These 4 cell lines showed some phenotypes characteristic of neoplastically transformed cells, and had a human karyotype with heteromorphisms identical to those of the parental fibroblasts. Their sensitivity to the cytotoxic effects of gamma-rays or ultraviolet irradiation was as high as those of their parental fibroblasts.

Ataxia Telangiectasia↗

PubMed1984

Different properties in lymphoblastoid cell lines from patients with Bloom syndrome.

In 3 lymphoblastoid cell lines derived from 3 patients with Bloom syndrome (BS), the baseline frequency of sister chromatid exchanges (SCEs), chromosomal breakage, sensitivity to ethylmethane-sulfonate (EMS), and bromodeoxyuridine (BrdUrd) were examined. The SCE frequency of 2 BS lines (EB-BS-NoKi-2 and EB-BS-AkSak) was about 2 to 2.5 times those of the normal cell lines, while that of another BS line (EB-BS-2KA) was about 10 to 11 times. The net increase in the number of SCEs in EB-BS-NoKi-2 and EB-BS-AkSak lines induced by EMS was similar to that of normal cell lines, but it was high in the BS-2KA line. Sensitivity to BrdUrd was examined using SCE induction at different concentrations of BrdUrd and by cell cycle analysis. EB-BS-NoKi-2 and EB-BS-AkSak lines were no more sensitive than normal cell lines, while the EB-BS-2KA line was more sensitive than controls. High frequency of chromosomal breakage was found only in the EB-BS-2KA line. These results suggest that 2 types of cells exist in the B-lymphoblastoid cells of BS.

B-Lymphocytes↗

PubMed1984

Cytidine deaminase levels in cultured mammalian cell lines measured by the growth tests and enzyme assays.

We developed a test medium for cytidine deaminase in order to examine the distribution of this enzyme in cultured cell lines. The growth of various mammalian cell lines was tested in culture medium containing 2 microM pyrazofurin and 100 microM cytidine. Enzymological assays for the enzyme also were made spectrophotometrically with cell extracts. A good correlation was found between results of cell growth tests and the levels of enzyme activity. Twelve of twenty cell lines were killed in the test medium, but the remaining lines showed good growth. The levels of enzyme activities were lower in the former lines than in the latter. The critical level of enzyme activity required to support cell growth was approximately 30 units per mg protein. These findings indicate that culture medium containing 2 microM pyrazofurin and 100 microM cytidine serves as a test medium for cytidine deaminase. The possibility that the cytidine deaminase may be useful in determining the embryonic origin of cultured cell lines is discussed, based on the growth properties of various cultured cell lines in the test medium.

Amides↗

PubMed1983

Loss of high frequency of sister chromatid exchanges in Epstein-Barr virus-established lymphoblastoid cell lines from two patients with Bloom's syndrome.

Seven lymphoblastoid cell lines were established through transformation by Epstein-Barr virus of peripheral blood lymphocytes from two patients with Bloom's syndrome (BS), the parents of a patient, and normal controls. High baseline levels of sister chromatid exchanges (SCEs) in peripheral blood lymphocytes of BS were reduced to about 10% of their initial value in BS lymphoblastoid cell lines, and the elevation of SCE frequencies induced by ethylmethanesulfonate was the same as in controls.

Adolescent↗

PubMed1983

The oral manifestations of 4p- syndrome.

The oral manifestations of 80 reported cases of 4p- syndrome are analyzed and a new case is reported. Results indicate that the common pattern of cheilognathopharyngeal disorders in 4p- syndrome is an isolated cleft palate or a high-arched palate with micrognathia. Cases of cleft lip are only about 10% of the total. The significance of other reported aberrant phenomena cannot yet be estimated.

Abnormalities, Multiple↗