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Biomedical subjects

H Ishikawa

Publications and source records attributed to H Ishikawa.

At least 19 recordsLinked to original sources
PubMed1995

8-Hydroxyadenine (7,8-dihydro-8-oxoadenine) induces misincorporation in in vitro DNA synthesis and mutations in NIH 3T3 cells.

An oligodeoxyribonucleotide containing 8-hydroxyadenine (OH8Ade) was chemically synthesized and single- and double-stranded c-Ha-ras gene fragments with OH8Ade at the second position of codon 61 were prepared. The single-stranded ras gene fragment was used as a template for in vitro DNA synthesis with the Klenow fragment of Escherichia coli DNA polymerase I, Taq DNA polymerase, rat DNA polymerase beta and mouse DNA polymerase alpha. The former two enzymes exclusively incorporated dTMP opposite OH8Ade. The DNA polymerases alpha and beta misinserted dGMP, and dAMP and dGMP, respectively. The c-Ha-ras gene was constructed using the double-stranded ras gene fragment containing OH8Ade and was transfected into NIH 3T3 cells. The gene with OH8Ade induced focus formation, indicating that OH8Ade elicited point mutations in cells. When c-Ha-ras genes present in transformed cells were analyzed, an A-->G transition and an A-->C transversion were detected. These results indicate that OH8Ade induced misincorporation in in vitro DNA synthesis and mutations in mammalian cells.

3T3 Cells↗
PubMed1995

Gamma delta T cells play an important role in hsp65 expression and in acquiring protective immune responses against infection with Toxoplasma gondii.

Previously, we reported that the expression of hsp65 within and on host macrophages correlates closely with protection against infection with Toxoplasma gondii in mice. Herein, we propose that gamma delta T cells play a crucial role in the induction of hsp65 and also in the protective immune response to T. gondii. Intraperitoneal inoculation with this protozoan resulted in hsp65 being expressed on and in host peritoneal macrophages and resulted in an increase of T cells bearing the gamma delta receptor with Thy-1+ and Thy-1- phenotypes in the peritoneal cavity and spleen. When mice were depleted of gamma delta T cells by the administration of a mAb, hsp65 expression was markedly decreased. In contrast, the expression of this protein was rather enhanced and gamma delta T cells were prominently expanded in mice depleted of alpha beta T cells. The protection in mice treated with the mAb paralleled the magnitude of hsp65 expression. Mice depleted of gamma delta T cells died most frequently in the early stages of infection, whereas most of those depleted of alpha beta T cells survived the early stages of lethal infection with T. gondii. However, the latter group of mice did not definitely control the T. gondii infection in its late stages. IFN-gamma was not essential for either the expression of hsp65 or the resistance induced by gamma delta T cells, as demonstrated in mice treated with mAb to murine IFN-gamma. These findings indicated that gamma delta T cells having both the Thy-1+ and Thy-1- phenotypes contribute to hsp65 expression within and on macrophages in an IFN-gamma-independent manner. This, in turn, plays a role in the development of protective immunity during the early stage of this parasitic infection.

Animals↗
PubMed1995

Modulation by bovine angiogenin of tubular morphogenesis and expression of plasminogen activator in bovine endothelial cells.

Angiogenin is a potent angiogenic molecule in the chick chorioallantoic membrane assay and rabbit corneal assay. However, no angiogenic activity has been reported in in vitro system. In this study, we isolated and purified angiogenin from bovine milk, and the biological effect of the bovine angiogenin on bovine endothelial cells was examined in in vitro angiogenesis models. Bovine angiogenin significantly stimulated both cell migration and formation of tube-like structures in the collagen gel by bovine aortic endothelial cells. Angiogenin at 10-100 ng/ml stimulated about 2-fold higher the tube formation when basic fibroblast growth factor (bFGF) at 10 ng/ml stimulated about 3-fold over the control. Tubular morphogenesis stimulated by bFGF or angiogenin was almost completely blocked in the presence of aprotinin, an inhibitor of serine proteases. Angiogenin up-regulated both mRNA level and activity of urokinase type plasminogen activator, a key mediator of angiogenesis. Moreover, both bFGF and bovine angiogenin induced a marked increase of c-fos mRNA level at 30 min after stimulation. These novel effects of bovine angiogenin are to be discussed in relation to its structure specificity.

Allantois↗
PubMed1995

Synthesis and biological evaluation of a series of new parenteral optically active 3-[[(N-alkylpyridinium-4'-yl)thio]methyl]-2-oxaisocephems.

The preparation and biological evaluation of a series of 7-[2-(2-aminothiazol-4-yl)-2-(Z)-[(cyclopentyloxy)imino]acetamido] optically active 2-oxaisocephems, substituted at the 3-position with [(N-alkylpyridinium-4'-yl)thio]methyl groups, are described. The resulting family of parenteral compounds displays a broad spectrum of in vitro antibacterial activity. These compounds exhibit increased activity against Gram-positive organisms including methicillin-resistant Staphylococcus aureus and Enterococcus faecalis which are resistant to most cephalosporins with a similar level of Gram-negative activity to that of the third-generation antibiotics. In vivo efficacy of new antibacterial agents in this investigation is excellent against both Gram-positive and Gram-negative bacteria as compared with reference compounds. The in vitro and in vivo antimicrobial activity and the structure-activity relationships are presented.

Animals↗
PubMed1995

Tissue-specific expression of the 230-kDa bullous pemphigoid antigen gene (BPAG1). Identification of a novel keratinocyte regulatory cis-element KRE3.

The 230-kDa bullous pemphigoid antigen gene (BPAG1) is expressed exclusively in basal keratinocytes of epidermis. In this study, we have identified a novel cis-element, keratinocyte responsive element 3 (KRE3), at position -216 to -197 of the human BPAG1 gene. A promoter-CAT construct containing this element had approximately 50-fold higher expression than a similar construct devoid of this sequence when tested in transient transfections of cultured human keratinocytes. However, there was no effect on the low base-line level of expression in cultured skin fibroblasts. KRE3 contains a palindromic sequence 5'-CAAATATTTG-3', and mutations in this sequence significantly reduced the promoter activity. Gel mobility shift assays with an oligomer containing KRE3 sequence demonstrated binding activity with nuclear proteins isolated from keratinocytes. One of the DNA/protein complexes was clearly specific, since competition with > 12.5-fold excess of the unlabeled oligomer resulted in disappearance of this band. No specific binding activity was noted with nuclear proteins extracted from fibroblasts. Thus, KRE3 appears to serve as the binding site for keratinocyte-specific trans-activating factor(s), and KRE3 may thus confer the tissue-specific expression to the BPAG1 gene.

Autoantigens↗
PubMed1995

EDA-containing fibronectin is synthesized from rheumatoid synovial fibroblast-like cells.

OBJECTIVE: To identify the cells that synthesize EDA-containing fibronectin (FN) and examine the role of EDA+FN in the pathogenesis of rheumatoid joint lesions. METHODS: Localization of EDA+FN and c-Fos protein in rheumatoid joints was studied immunohistochemically by utilizing antibodies for EDA+FN and c-Fos. Expression of EDA+FN was studied by immunoelectron microscopy and in situ hybridization. The amount of EDA+FN was measured by enzyme-linked immunosorbent assay. RESULTS: EDA+FN was specifically localized in the synovial lining layer of synovium with active rheumatoid arthritis (RA) (n = 17), but not in that with osteoarthritis (n = 4) or with inactive fibrous RA (n = 2). EDA+FN messenger RNA was localized in the synovial lining layer. EDA+FN was immunoelectron microscopically localized in the synovial lining fibroblast-like (type B) cells. EDA+FN was also detected at the cartilage-pannus junction and on the surface of RA cartilage. Double staining showed that EDA+FN colocalized with c-Fos protein in the rheumatoid synovial lining layer. Quantification of EDA+FN showed that it was highly concentrated in rheumatoid synovial fluids. CONCLUSION: EDA+FN is synthesized by the synovial lining fibroblast-like (type B) cells in situ in rheumatoid synovium, and appears to be expressed in association with activated or transformed states of synovium.

Animals↗
PubMed1995

Effect of histamine and substance P on the rabbit and human iris sphincter muscle.

BACKGROUND AND METHODS: In an attempt to clarify the functional action of histamine and substance P on atropine-resistant miosis, we isolated rabbit and human iris sphincter muscles and investigated their mechanical properties using the isometric tension recording method. RESULTS: Substance P dose-dependently contracted the rabbit iris sphincter, but had no effect on the human iris sphincter. In the rabbit iris sphincter, histamine reduced the amplitude of twitch contraction evoked by field stimulation but had no effect on carbachol-induced contraction. Thioperamide, but not mepyramine or cimetidine, partially antagonized the histamine-induced reduction in the amplitude of twitch contractions. In the human iris sphincter, on the other hand, histamine dose-dependently provoked contraction and the amplitude of histamine-induced contraction was affected neither by atropine nor by indomethacin. CONCLUSIONS: These results provide evidence that histamine has strong contractile effect on the human iris sphincter muscle; the rabbit iris sphincter muscle, however, apparently lacks functional histamine receptors. In rabbits, exogenously applied histamine only activates H3 receptors located on the cholinergic nerve terminal, hence the excitatory neuro-effector transmission is suppressed. Thus, histamine may have an important roles in atropine-resistant miosis in humans, but not in rabbits.

Animals↗
PubMed1995

Occurrence of fasciculated microtubules at nodes of Ranvier in rat spinal roots.

Fascicles of microtubules, previously considered to be unique to the initial segments of myelinated axons, were found at nodes of Ranvier of sensory and motor axons in rat spinal roots, though their occurrence was limited to the proximal portion of the axons. No fasciculated microtubules were noticed in the internodes of myelinated axons or in unmyelinated axons. In transverse sections, microtubules in fascicles were characteristically cross-linked by short filamentous strands at a centre-to-centre distance of 36-40 nm. In sensory axons, the density of fasciculated microtubules at the node of Ranvier was 45-48% at the level of the dorsal root ganglion and decreased progressively as thin sections were made more distal to the ganglion in both directions. In motor axons, microtubule-fascicles involved about 24% of the microtubules in the ventral rootlets, but they were rarely seen in distal portions of the ventral roots. No fasciculation of microtubules was discerned in the sciatic and saphenous nerves. These findings suggest that cross-linking protein(s) for fasciculation, which are synthesized in the perikaryon and primarily used for the initial segment, may be further transported to fasciculate microtubules at some proximal nodes of Ranvier.

Animals↗
PubMed1995

Primary sclerosing cholangitis with marked eosinophilic infiltration in the liver.

A 16-year-old boy was diagnosed as having primary sclerosing cholangitis (PSC), based on retrograde cholangiography showing mixed features of narrowing and dilatation of the common hepatic and intrahepatic bile ducts. However, periductal fibrosis was not observed in the needle biopsy liver specimen. The liver biopsy specimen obtained 11 years previously, at the onset of the disease had disclosed a marked infiltration of eosinophils in the portal tract with eosinophilic catinonic protein immunostaining, with marked eosinophilia (54%) being noted. In Japanese reports, eosinophilia of more than 7% was reported in 13 of 32 (40.6%) PSC patients. However, the early stage of PSC, with marked eosinophilia and eosinophilic infiltration in the liver, such as in the present case, has rarely been reported. The findings in this case suggest that eosinocytes are related to the pathogenesis of PSC.

Adolescent↗
PubMed1995

Liver with hypoechoic nodular pattern as a risk factor for hepatocellular carcinoma.

BACKGROUND/AIMS: Ultrasonography should be used for screening of hepatocellular carcinoma, but there are few reports on the relationship between liver ultrasonographic findings and the development of hepatocellular carcinoma (HCC). Using prospective follow-up studies, we examined the role of liver with a hypoechoic nodular pattern as a high-risk factor in HCC. METHODS: The study was performed by follow-up on 593 patients with chronic liver disease recorded at our hospital. The ultrasonographic pattern of the liver parenchyma was classified either as a small or large hypoechoic nodular pattern or as a nonnodular pattern. Patients were followed up from the time of initial ultrasonographic examination (1985-1987) until January 1, 1991. RESULTS: During the follow-up period (average, 4.2 years, range, 0.3-6.0 years), 62 patients were found to have HCC (12%). Patients whose livers showed small or large hypoechoic nodular pattern had a significantly higher risk of HCC than did patients whose livers showed a nonnodular pattern (rate ratios were 14.0 and 20.0, respectively, adjusted for age, sex, hepatitis virus markers, ICG R15, alpha-fetoprotein concentration, and ultrasonographic pattern of the liver). CONCLUSIONS: Liver showing a hypoechoic nodular pattern is a major risk factor in HCC.

Adult↗
PubMed1995

Combined effects of maternal age and delayed fertilization on the frequency of chromosome anomalies in mice.

Animal experiments indicate that both maternal age and delayed fertilization increase the incidence of chromosome errors in the offspring. In the present study, therefore, we examined how both factors act in combination. We set up normal (0 h delay) and delayed fertilization (6 h delay) subgroups in three different age groups (young, 3-4 months; middle-aged, 9-11 months; and old, 13-15 months), and compared the incidence of chromosome anomalies and other reproductive effects among them. Although the present study confirmed two previous findings that aneuploidies increase with advancing maternal age, and that polyploidies increase with delayed fertilization in mice, it was not clear whether maternal age and delayed fertilization affected the occurrence of aneuploidies either additively or multiplicatively.

Animals↗
PubMed1995

Interventional trial for colorectal cancer prevention in Osaka: an introduction to the protocol.

We established a protocol for an interventional randomized controlled trial for prevention of colorectal cancer by attaching special importance to feasibility. The subjects were patients with multiple colorectal tumors. Two regimens were formulated for prevention of colorectal cancer. One was dietary guidance alone (Regimen I), and the other was dietary guidance plus eating wheat bran biscuits (Regimen II). The main end points of the trial were examinations for recurrence of colorectal tumors after 2 and 4 years. The target number of patients was 200 in total, i.e. 100 for each group. During the 18 months from the beginning of recruiting of subjects (up to November 1994), 28 (97%) of the 29 patients recruited for Regimen I and 32 (97%) of the 33 patients recruited for Regimen II agreed to participate in the trial. The trial progressing well.

Aged↗
PubMed1995

Structural analysis of TRAS1, a novel family of telomeric repeat-associated retrotransposons in the silkworm, Bombyx mori.

We characterized TRAS1, a retrotransposable element which was inserted into the telomeric repetitive sequence (CCTAA)n of the silkworm, Bombyx mori. The complete sequence of TRAS1, a stretch of 7.8 kb with a poly(A) tract at the 3' end, was determined. No long terminal repeat (LTR) was found at the termini of the element. TRAS1 contains gag- and pol-like open reading frames (ORFs) which are similar to those of non-LTR retrotransposons. The two ORFs overlap but are one nucleotide out of frame (+1 frameshift). Most of the approximately 250 copies of TRAS1 elements in the genome were highly conserved in the structure. Chromosomal in situ hybridization showed that TRAS1 elements are clustered at the telomeres of Bombyx chromosomes. A phylogenetic analysis using the amino acid sequence of the reverse transcriptase domain within the pol-like ORF revealed that TRAS1 falls into one lineage with R1, which is a family of non-LTR retrotransposons inserted into the same site within the 28S ribosomal DNA unit in most insects. TRAS1 may have been derived from R1 and changed the target specificity so that TRAS1 inserts into the telomeric repetitive sequence (CCTAA)n. Southern hybridization and Bal 31 exonuclease analyses showed that TRAS1 elements are clustered proximal to the terminal long tract of (CCTAA)n. TRAS1 is a novel family of non-LTR retrotransposons which are inserted into the telomeric repetitive sequences as target sites.

Amino Acid Sequence↗
PubMed1995

Viability, development and incidence of chromosome anomalies of preimplantation embryos from XO mice.

We examined the viability and developmental status of XO embryos at preimplantation stage (day 3 of gestation) by assessing blastocyst formation and counting cell number using our XO mouse colony. We also examined the incidence of chromosome anomalies. Embryos from XO mice (XY, XX and XO) developed more slowly (in cell numbers and blastocoele formation) than those from XX mice (XY and XX). XO embryos also tended to develop more slowly than XX embryos in the XO group. Although litter size at the preimplantation stage of gestation was almost twice as large (12.7) as those at mid-gestation (7.6) and near-term (7.2) in this colony, the adjusted XY:XX:XO ratio (2.8:2.0:1.0) did not differ greatly. This indicates that almost half of the embryos must have been eliminated during the first half of gestation in the XO group, probably regardless of sex chromosome complements. Thus, we consider that maternal XO sex chromosome constitution is disadvantageous for the intrauterine development of the embryo during the early period of gestation. This may be related to precocious aging of XO mice. Further, we confirmed that a high incidence of abnormal karyotypes occurs in embryos from our XO mouse colony.

Animals↗
PubMed1995

A compartmentalized bias for T-cell receptor V beta usage in summer-type hypersensitivity pneumonitis.

Several monoclonal antibodies specific to the human T-cell receptor (TCR) variable (V) region were used to examine whether T-cell oligoclonality plays a key role in the development of the T-cell-mediated lung disease, summer-type hypersensitivity pneumonitis (SHP). The usage of TCR V-regions was examined using bronchoalveolar lavage (BAL) and matched peripheral blood lymphocytes (PBL) in patients with SHP. In 18 SHP patients, the majority of lymphocytes in both BAL and PBL was CD3+ cells (94.2 vs. 63.9%, respectively), which was similar to that in normals (n = 7). Of CD3+ lymphocytes in a SHP patient, the percentage of CD4+ cells in BAL was reduced compared to those in paired PBL, whereas the percentage of CD8+ cells was significantly elevated. In contrast, the percentage of CD4+ cells was much higher than that of CD8+ cells in both BAL and PBL of normal subjects. While a majority of CD3+ T cells expressed TCR alpha/beta+ in BAL and PBL from both SHP patients and normal controls, a marked increase of the expression of TCR gamma/delta+ was observed in PBL in 4 out of 18 SHP patients. A markedly increased prevalence (> 3 SD) of specific TCR alpha/beta+ V-region families in BAL was detected in 5 of 18 patients with SHP as compared to normal controls, whereas no specific increase was found in PBL. Increased V beta 8 was detected in 3 cases; V beta 6 was observed in 1 patient, and V beta 5 in another patient.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Pulmonary hypertension in rats. 1. Role of bromodeoxyuridine-positive mononuclear cells and alveolar macrophages.

The role of activated cells determined by staining with bromodeoxyuridine (BrdU) and the role of alveolar macrophages as a source of cytokines were investigated in monocrotaline (MCT)-induced pulmonary hypertension (PH) in rats. BrdU was mainly incorporated by mononuclear cells; their number increased strikingly prior to the development of PH. Cytokines such as interleukin (IL)-1, IL-6 and tumor necrosis factor were produced during the development of PH. The amount of IL-6 showed significant correlation (p < 0.05) with the number of BrdU-positive cells, which, in turn, correlated with the extent of PH. These results indicate that BrdU-positive cells and cytokines produced by alveolar macrophages may play an important role in the pathogenesis of MCT-induced PH in rats.

Animals↗
PubMed1995

Pulmonary hypertension in rats. 2. Role of interleukin-6.

We administered human interleukin (IL)-6 to rats to examine whether it is implicated in the development of pulmonary hypertension (PH). The rats injected with IL-6 developed PH as determined by the weight ratio of the right ventricle to the left ventricle+septum. This ratio decreased with the appearance of IgG anti-human IL-6 antibody. Histologically we observed in the lungs, luminal occlusion of small muscular arteries, capillaries filled with fibrin thrombi, and localized hemorrhage. IL-6 increased the number of platelets, and the number of platelets correlated with the extent of PH. Thus, it seems likely that (IL-6) is involved in the formation of PH in this model.

Animals↗