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Biomedical subjects

M Oishi

Publications and source records attributed to M Oishi.

At least 163 records · Page 9Linked to original sources
PubMed1994

A novel regulatory sequence affecting the constitutive expression of tissue plasminogen activator (tPA) gene in human melanoma (Bowes) cells.

In order to identify a possible enhancer-like regulatory sequence for the human tissue plasminogen activator (tPA) gene, various DNA segments in the 5'-flanking region were ligated to the chloramphenicol acetyltransferase (CAT) reporter gene driven by the tPA gene promoter, and their CAT enhancing activities were measured following transfection to human melanoma-derived Bowes cells that highly express the gene. Major and minor activities were detected in two adjacent upstream sequences, 160 base pairs (bp) (-2288 to -2129) and 102 bp (-2390 to -2289), respectively, and the former was subjected to further analysis. The CAT enhancing activity of the 160-bp sequence was greatly affected by its position and orientation in the constructs and the sequence also functioned weakly with the SV40 promoter. Deletion of any small portion from the sequence abolished the CAT enhancing activity, suggesting that the entire sequence is required for the activity. This sequence did not show a further CAT enhancing activity in Bowes cells treated with the inducers phorbol 12 myristate 13-acetate and dexamethasone and did not function in HeLa or HT1080 cells under any conditions. Taken together, the 160-bp sequence is likely to be responsible for the constitutive and/or cell type-specific expression of the tPA gene in human cells.

Base Sequence↗
PubMed1994

Induction of specific protein tyrosine phosphatase transcripts during differentiation of mouse embryonal carcinoma (F9) cells.

We have investigated the pattern of PTPase transcript expression during in vitro differentiation of mouse embryonal carcinoma (F9) cells. While the transcripts of most PTPases were unchanged or undetected during embryonal differentiation induced by retinoic acid, several PTPase transcripts exhibited distinct patterns of induction. Mutant cells defective in differentiation did not display the induction of some of these PTPase transcripts. Interestingly, three out of the four PTPase transcripts induced were the same PTPase transcripts induced during in vitro erythroid differentiation of mouse erythroleukemia (MEL) cells [(1994) J. Biol. Chem. 269, 4709-4712] [corrected]. The possible role played by specific PTPases in cell differentiation is discussed.

Animals↗
PubMed1994

Receptor-binding affinities of human epidermal growth factor variants having unnatural amino acid residues in position 23.

Ile-23 of human epidermal growth factor (hEGF) has been indicated, by mutagenesis and NMR studies, to be directly recognized by the receptor. In the present study, an unnatural phenylalanine analog, either 2-azaphenylalanine (2aF), 3-azaphenylalanine (3aF), 4-azaphenylalanine (4aF), or 4-fluorophenylalanine (4fF), was incorporated by an in vivo protein synthesis system into position 23 of [Phe23]hEGF, which retains appreciable receptor-binding affinity (about 20% of the wild type). We compared the receptor-binding affinities of the variants with that of [Phe23]hEGF and found that substitution of Phe-23 with 2aF or 3aF raised the affinity, while substitution with 4aF or 4fF remarkably reduced the affinity. The tertiary structure of [Phe23]hEGF was not significantly affected by the substitution of Phe-23 with 2aF, as shown by the two-dimensional nuclear magnetic resonance analysis. In addition, the substitution of residue 23 with His or Tyr produced an hEGF variant with a slightly higher receptor-binding affinity than that of [Phe23]hEGF. Our results suggest that the receptor has an asymmetric hydrophobic pocket for recognition of the side chain in position 23 of hEGF. Furthermore, on the receptor surface, this pocket seems to be adjacent to a less hydrophobic region with a hydrogen-bond acceptor and donor. Thus, the use of unnatural amino acids in addition to the 20 natural ones allows analyses of the structure-function relationship of a protein at a higher resolution than conventional site-directed substitution by only natural amino acid residues.

Amino Acids↗
PubMed1994

A differential cloning procedure of complex genomic DNA fragments.

We have developed a differential cloning procedure designed for cloning of anonymous restriction DNA fragments whose molecular sizes differ between two genomic DNA preparations from higher organisms. The procedure, which was extensively revised from the original one, consists of several steps as summarized below. (i) Digestion of two DNA preparations (target and reference DNA) with the same restriction enzyme (4 base cutter). (ii) Biotinylation of target DNA and conversion of reference DNA to nonamplifiable form by terminal dephosphorylation. (iii) Electrophoresis of the two DNA preparations through a synthetic gel with a large excess of reference DNA as a competitor. (iv) In-gel alkaline dissociation of DNA, followed by reassociation (in-gel competitive reassociation). (v) Elution of DNA from the gel and PCR after adapter ligation and adsorption of DNA onto streptavidin-coated matrix. By repeating these steps, we attained substantial enrichment (approximately 10,000-fold) of DNA fragments which were originally present at one copy or less per complex mammalian genome. The details of the procedure and its unique characteristics in cloning of altered genomic DNA fragments, particularly from mammalian genome, are discussed.

Adenine Phosphoribosyltransferase↗
PubMed1994

Enrichment of human DNAs that flank poly(dA).poly(dT) tracts by triplex DNA formation.

Human DNA fragments which contain poly(dA).poly(dT) tracts and their immediate flanking regions were enriched by means of triplex DNA formation. Human DNA fragments were mixed with biotinylated (dT)34 in the presence of Mg2+ and the triplex DNA [(dT)34.poly(dA).poly(dT)] was adsorbed onto streptavidin-coated magnetic beads and the DNA fragments which formed triplexes were eluted from the beads with a buffer containing EDTA. A control experiment using a plasmid with a poly(dA).poly(dT) tract indicated that DNA fragments with the tract could be enriched over 60-fold after one cycle of the treatment. After PCR amplification, the sample was subjected to the next cycle of the affinity enrichment. After four cycles, we obtained a human genomic DNA library of clones with inserts ranging from 500 to 1000 bp among which 86% had at least one poly(dA).poly(dT) tract. No less than 60% of the clones were close or distant members of the Alu family. Sequence determination of 25 clones revealed that the length of the poly(dA).poly(dT) tracts was 14 to 37 (average 28) bp and that they were located close to either end of the fragments. While 15 clones were identified as Alu family homologues and one as a human L1 family member, nine clones were of unknown origin. None of these nine clones were a highly repeated sequence nor a part of transcriptionally active regions.

Bacterial Proteins↗
PubMed1994

Construction of a normalized cDNA library by introduction of a semi-solid mRNA-cDNA hybridization system.

We report a novel procedure to construct a normalized (equalized) cDNA library. By introduction of the highly efficient self-hybridization system between a whole mRNA population and their corresponding cDNA immobilized on latex beads, which involves relatively simple manipulations, we were able to generate an mRNA population in which the copy number of abundant species was reduced while that of rare species was enriched. In a typical experiment, after several cycles of self-hybridization on the beads, the ratio of the most to the least abundant marker mRNA species dropped by a factor of 300 (from 10,000 to 30) while the complexity and length of mRNAs in the population remained unchanged. The procedure should provide a potent tool for the expression cloning of cDNA and also facilitate the construction of whole cDNA catalogs from specific tissues (or cell types) from higher organisms.

Animals↗
PubMed1994

Cell cycle-dependent regulation of the phosphorylation and metabolism of the Alzheimer amyloid precursor protein.

Accumulation of the amyloid A beta peptide, which is derived from a larger precursor protein (APP), and the formation of plaques, are major events believed to be involved in the etiology of Alzheimer's disease. Abnormal regulation of the metabolism of APP may contribute to the deposition of plaques. APP is an integral membrane protein containing several putative phosphorylation sites within its cytoplasmic domain. We report here that APP is phosphorylated at Thr668 by p34cdc2 protein kinase (cdc2 kinase) in vitro, and in a cell cycle-dependent manner in vivo. At the G2/M phase of the cell cycle, when APP phosphorylation is maximal, the levels of mature APP (mAPP) and immature APP (imAPP) do not change significantly. However, imAPP is altered qualitatively. Furthermore, the level of the secreted extracellular N-terminal domain (APPS) is decreased and that of the truncated intracellular C-terminal fragment (APPCOOH) is increased. These findings suggest the possibility that phosphorylation-dependent events occurring during the cell cycle affect the metabolism of APP. Alterations in these events might play a role in the pathogenesis of Alzheimer's disease.

Alzheimer Disease↗
PubMed1994

Induction of specific protein tyrosine phosphatase transcripts during differentiation of mouse erythroleukemia cells.

We reported previously that most of the phosphotyrosine-containing cellular proteins were quickly dephosphorylated at the very early stage of erythroid differentiation of mouse erythroleukemia (MEL) cells. These and other experimental results implicated a specific protein tyrosine phosphatase(s) (PTPase(s)) involved in the commitment of the erythroid differentiation. We have investigated the pattern of transcripts of PTPases during MEL cell differentiation and found that while the transcripts of most PTPases were unchanged or undetected in the cells, transcripts for two PTPases (PTP beta 2 and RIP) exhibited distinct patterns of induction at a very early stage of differentiation. Some of the mutant cells defective in differentiation did not show the induction of these PTPase transcripts. We discuss the possible role played by the PTPases in the commitment of MEL cell differentiation.

Acetamides↗
PubMed1994

Distribution of apoptosis-mediating Fas antigen in human skin and effects of anti-Fas monoclonal antibody on human epidermal keratinocyte and squamous cell carcinoma cell lines.

Fast antigen is a cell surface protein that mediates apoptosis. Using immunohistological, flow cytometry and electron microscopic analyses, we investigated the expression of Fas antigen on various skin tissues, and on cultured SV40-transformed human epidermal keratinocyte cell line KJD and human skin squamous cell carcinoma cell line HSC. The Fas antigen was widely distributed in skin components such as the keratinocytes in the lower portion of the epidermis, epidermal dendritic cells, endothelial cells, fibroblasts, apocrine glands, eccrine sweat glands, sebaceous glands, some normal melanocytes and infiltrating lymphoid cells. It was also strongly expressed on the keratinocytes of lichenoid eruptions seen in lupus erythematosus and lichen planus, and on the spongiotic or acanthotic epidermis seen in chronic eczema, adult T-cell leukaemia/lymphoma (ATLL) and atopic dermatitis. Its expression was closely correlated with lymphoid infiltrating cells and it was strongly expressed in lymphoid neoplastic cells, particularly ATLL cells, and fibroblasts seen in dermatofibroma. However, the antigen was not detected on basal cell epithelioma cells, some malignant melanomas or any junctional naevi. The cell lines KJD and HSC strongly expressed the Fas antigen, and crosslinking of the Fas antigen by an anti-Fas monoclonal antibody induced apoptosis of these cell lines. These results indicate that the apoptosis-mediating Fas antigen may play an important role in normal skin turnover and cell differentiation, in immune regulation of skin tumours, and in the pathogenesis of various skin diseases.

Antibodies, Monoclonal↗
PubMed1994

Construction of an equalized cDNA library from human brain by semi-solid self-hybridization system.

A cDNA library enriched in clones for mRNA species (an equalized cDNA library) was constructed from human brain employing semi-solid self-hybridization system and cDNA amplification by in vitro transcription. Sequence analysis of the clones in the cDNA library indicated that (1) clones representing abundant species such as those of mitochondrial origin and expression sequence taq (EST) are drastically reduced in their proportion compared to the original library and (2) clones representing rare mRNA species or which do not have matched sequences in the database increased substantially in their proportion. cDNA libraries thus constructed, therefore, are a good source for cloning of cDNA for rare mRNA species from specific mammalian tissues and also for direct expression cloning since the inserts maintained their original molecular sizes after equalization.

Base Sequence↗
PubMed1994

Differences in IL-2 receptor levels between mycosis fungoides and cutaneous type adult T-cell leukemia/lymphoma in the early stages of the disease.

Skin lesions in mycosis fungoides (MF) and cutaneous-type adult T-cell leukemia/lymphoma (c-ATL) are sometimes difficult to differentiate. Our previous investigations showed that the alpha-chain of interleukin-2 receptor (IL-2R alpha) secretion is enhanced in both acute and chronic type ATL. This finding suggested increased levels of IL-2R alpha in c-ATL that might differentiate it from MF. In this study, we report several observations on IL-2R alpha levels in MF and c-ATL. 1) Plasma levels of IL-2R alpha determined by enzyme-linked immunosorbent assay were higher in patients with c-ATL (tumor group, 17,571 +/- 5,919 U/ml; non-tumor group, 4,546 +/- 1,788 U/ml) than those of patients with MF (tumor group, 2,352 +/- 861 U/ml; plaque group, 530 +/- 174 U/ml). 2) The percentages of IL-2R alpha-positive cells in skin lesions of c-ATL were 30.5 +/- 13.4% in the non-tumor group and 62.7 +/- 12.0% in the tumor group, which were significantly higher than those of MF, plaque group (12.7 +/- 5.9%), and tumor group (3.4 +/- 2.3%). 3) Further, there was a significant correlation between plasma IL-2R alpha levels and the number of IL-2R alpha-positive cells in skin lesions in patients with c-ATL (r = 0.654, p < 0.0001) and MF (r = 0.537, p < 0.01) on an individual basis; whereas no increased IL-2R alpha levels in peripheral blood mononuclear cells were observed in patients with either MF or c-ATL. 4) Skin lesions of c-ATL had much higher levels of IL-2R alpha genomic RNA than those of MF in their initial stages when measured by DNase/reverse transcription/polymerase chain reaction and liquid hybridization analysis. These findings suggest that increased plasma IL-2R alpha levels in patients with either MF or c-ATL may be linked to cellular IL-2R alpha levels in the skin lesions, and thus reflect the difference between MF and c-ATL.

Adult↗
PubMed1994

Langerhans cell histiocytosis presenting as a varicelliform eruption over the entire skin.

A boy with skin eruptions resembling varicella and specific for Langerhans cell histiocytosis (LCH) is reported. At his initial visit when he was four months old, vesiculopustular lesions were present over the entire body; these had first appeared on the third day post partus. Histopathological, immunohistochemical, and electron microscopical examination confirmed the Langerhans cell phenotype and Birbeck granules in the responsible cells. He also had hydronephrosis, recurrent fever, and cutaneous bacterial infections. His parents refused further medical treatment and he died of diarrhea with cachexia about two years later. LCH may present diagnostic difficulties by manifesting as a skin eruption which resembles varicella.

Chickenpox↗
PubMed1994

Crow-Fukase syndrome: a case associated with vasospastic angina.

The patient is a 60-year-old man who developed numbness of the extremities, paralysis, hyperpigmentation of the skin, hypertrichosis, anasarca and chest pain at the age of 58 years. The diagnosis of Crow-Fukase syndrome was made and myeloma was not found. Prednisolone therapy was effective but chest pain reappeared every morning when prednisolone was tapered to 30 mg alternate day. Coronary arteriogram showed no stenosis but administration of acetylcholine into the coronary artery produced ST elevation in electrocardiogram, chest pain and coronary artery stenosis which were relieved by administration of nitrates into the coronary artery.

Acetylcholine↗
PubMed1993

Characterization of the 6-aminohexanoate-dimer hydrolase from Pseudomonas sp. NK87.

The DNA base sequence of the Pseudomonas sp. NK87 gene (P-nylB) for 6-aminohexanoate-dimer hydrolase (P-EII), a xenobiotic-compound-degrading enzyme, was determined. It has an open reading frame of 1188 bp, initiated by ATG and terminated by TAG, and coding for 396 amino acids. The base sequence of the open reading frame has 53% sequence similarity to that of the gene for the same enzyme of Flavobacterium sp. KI72 (F-nylB) and 35% sequence similarity with respect to the deduced amino acid sequence. The P-EII enzyme was purified from an Escherichia coli clone in which the P-EII gene was highly expressed. The P-EII enzyme was inhibited by a serine protease inhibitor, diisopropyl fluorophosphate, as was the F-EII enzyme. Double reciprocal plots obtained from various concentrations of 6-aminohexanoate-dimer indicated that the kcat value of the P-EII enzyme (9.2 s-1) was approximately half that of the F-EII enzyme (19 s-1), and the P-EII enzyme had higher affinity toward this substrate (Km for P-EII, 0.6 mM; Km for F-EII, 15 mM). The P-EII enzyme had a temperature optimum of 48 degrees C, and a pH optimum of 7.5. It is speculated that since the P-nylB and F-nylB genes are more diverged from each other than the corresponding nylA genes, the latter may have evolved more recently.

Amidohydrolases↗
PubMed1993

Serum levels of interleukin 6 in patients with pustulosis palmaris et plantaris.

Serum interleukin 6 (IL-6) was measured in patients with pustulosis palmaris et plantaris by a fluorescent sandwich enzyme-linked immunosorbent assay (ELISA) method. Increased production of IL-6 was observed in 8 cases out of 10 showing numerous pustules in their active stage. Six cases were measured before and after tonsillectomy, and statistically significant reduction in the levels of IL-6 paralleled an improvement in the disease activity. It is suggested that IL-6 may be one of the mediators which link tonsillectomy to the activity of pustulosis palmaris et plantaris.

Adult↗
PubMed1993

Chromosome 7 suppresses indefinite division of nontumorigenic immortalized human fibroblast cell lines KMST-6 and SUSM-1.

Using nontumorigenic immortalized human cell lines KMST-6 (KMST) and SUSM-1 (SUSM), we attempted to identify the chromosome that carries a putative senescence-related gene(s). These cell lines are the only ones that have been established independently from normal human diploid fibroblasts following in vitro mutagenesis. We first examined restriction fragment length polymorphisms on each chromosome of these immortalized cell lines and their parental cell lines and found specific chromosomal alterations common to these cell lines (a loss of heterozygosity in KMST and a deletion in SUSM) on the long arm of chromosome 7. In addition to these, we also found that introduction of chromosome 7 into these cell lines by means of microcell fusion resulted in the cessation of cell division, giving rise to cells resembling cells in senescence. Introduction of other chromosomes, such as chromosomes 1 and 11, on which losses of heterozygosity were also detected in one of the cell lines (KMST), to either KMST or SUSM cells or of chromosome 7 to several tumor-derived cell lines had no effect on their division potential. These results strongly suggest that a gene(s) affecting limited-division potential or senescence of normal human fibroblasts is located on chromosome 7, probably at the long arm of the chromosome, representing the first case in which a specific chromosome reverses the immortal phenotype of otherwise normal human cell lines.

Animals↗
PubMed1993

Large and small coils in magnetic stimulation of the proximal portion of the facial nerve.

Magnetic stimulation (Magstim model 200) was performed with a large coil (14 cm in diameter) and a small coil (7 cm in diameter) in 12 healthy persons. Muscle potentials were obtained from ipsilateral orbicularis oris and nasalis muscles when magnetic stimulation was performed by placing a large or small magnetic coil at T6 or by placing a large magnetic coil at C4. The latency obtained by placing a magnetic coil at T6 was 1.2-1.3 ms (with a large coil) and 1.3-1.4 ms (with a small coil) longer, and the latency obtained by placing a large magnetic coil at C4 was 1.3-1.4 ms longer than the latencies obtained by stimulating at the stylomastoid foramen. The proximal portion of the facial nerve is considered to be stimulated transcranially when magnetic stimulation is performed by placing either a large or small coil at T6 or by placing a large coil at C4.

Adult↗