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Biomedical subjects

K Horie

Publications and source records attributed to K Horie.

At least 109 records · Page 6Linked to original sources
PubMed1994

The pharmacological profile of cloned and stably expressed alpha 1b-adrenoceptor in CHO cells.

Using Chinese hamster ovary (CHO) cells stably expressing the alpha 1B-adrenoceptor (CHO alpha 1B cells) as a model, we investigated whether the transfected cells that express alpha 1B subtype of adrenoceptor can show the pharmacologic characteristics as previously defined in native tissues. Radioligand binding studies with 2-[beta-(4-hydroxy-3-[125I]iodophenyl)ethylamino-methyl]tetralone ([125I]HEAT) in CHO alpha 1B cells showed the similar Ki values of the alpha 1-adrenoceptor selective drugs as previously observed in rat liver and spleen, and that pretreatment with chlorethylclonidine markedly inactivated the binding sites (94.7-98.6%). In CHO alpha 1B cells alpha 1-adrenoceptor agonists caused a dose-dependent increase in transients of cytosolic Ca2+ concentrations ([Ca2+]i), and the potency order of antagonists in inhibiting norepinephrine-induced [Ca2+]i response was similar to that observed in radioligand binding assays. In summary, the present study shows that the ligand binding property, the pharmacological characteristics and the intracellular transduction mechanisms of alpha 1B-adrenoceptors stably expressed in CHO cells appear to be the same as those defined in native tissues. Thus they can be a useful model system for further characterization of the receptor as well as for the development of specific ligands.

Adrenergic alpha-Agonists↗
PubMed1994

Role of oxygen radicals and IL-6 in IL-1-dependent cartilage matrix degradation.

It has been suggested that IL-1 produces cartilage matrix degradation by metalloproteinases such as collagenase and that such degradation is regulated by metalloproteinase inhibitors. In the present study, the effects of IL-6 and oxygen radical scavengers on cartilage matrix degradation were studied. Superoxide dismutase, catalase, or methionine all significantly inhibited cartilage matrix degradation both in IL-1 beta-stimulated and unstimulated experimental conditions. Both 10 mM EDTA and 100 nM tissue inhibitor of metalloproteinase (TIMP) significantly inhibited cartilage matrix degradation. The addition of methionine significantly inhibited collagenase activity produced in the culture supernatants of chondrocytes stimulated with IL-1 beta. IL-6 significantly suppressed cartilage matrix degradation produced spontaneously or by IL-1 beta stimulation in chondrocytes. IL-6 inhibited superoxide production by chondrocytes both in IL-1 beta-stimulated or unstimulated conditions. These results suggest that oxygen radicals are involved in cartilage matrix degradation mediated by both paracrine and autocrine IL-1 mechanisms and that oxygen radical-mediated activation of collagenase in chondrocytes may explain the mechanisms of how oxygen radicals are involved in cartilage matrix degradation. IL-6 inhibited superoxide production in chondrocytes and thus inhibited cartilage matrix degradation.

Arthritis, Rheumatoid↗
PubMed1994

Characteristics of ceftibuten uptake into Caco-2 cells.

The characteristics of ceftibuten uptake into Caco-2 cells grown in a collagen-coated dish were examined. Ceftibuten showed stereoselective and pH-dependent uptake. The pH-dependency of ceftibuten was more marked than that of cefaclor or cephalexin, but all three antibiotics showed maximal uptake at pH 5.5. Ceftibuten uptake was linear for the initial 1 hr and then reached a plateau. The initial uptake (15 min) was markedly reduced by the addition of 2,4-dinitrophenol or FCCP (a protonophore), or by lowering the incubation temperature. The uptake of ceftibuten into the brush-border membrane vesicles prepared from cultured Caco-2 cells showed an overshoot in the presence of an H(+)-gradient. These findings indicated that the uptake of ceftibuten was energy-dependent, especially H(+)-gradient-dependent. Uptake inhibition by various compounds was compared using Caco-2 cells. Amino acids and a tetrapeptide did not inhibit uptake, whereas di- or tri-peptides were effective inhibitors. These observations suggest that ceftibuten is taken up by a carrier-mediated transport system(s) for dipeptides. Various antibiotics differed in their ability to inhibit uptake, with cyclacillin showing maximum inhibition. Differences in the inhibitory effect may be accounted for by the heterogeneity (multiplicity) of the transport systems.

2,4-Dinitrophenol↗
PubMed1994

Structures of replacement vectors for efficient gene targeting.

The relationship between DNA structure of replacement vectors and gene targeting efficiency was studied using positive-negative selection. The vectors contained pBR322 DNA, a bacterial neomycin-resistance gene (neo) for positive selection, a herpes simplex virus (HSV) thymidine kinase gene (tk) for negative selection, and a mouse genomic fragment, including exons 1 to 3 of the transthyretin (ttr) gene. The neo gene that confers G418 resistance was inserted into the second ttr exon, and the HSV-tk gene that confers gancyclovir (GANC) sensitivity was added to the 3' end of the ttr fragment. The vectors were linearized by digesting with restriction enzyme(s) and transfected into mouse embryonal carcinoma F9 cells. In this system, the enrichment by GANC selection as well as the frequency of gene targeting was increased by placing the pBR322 DNA at the 3' end of the HSV-tk gene. Adding one more HSV-tk gene at the 5' end of the ttr fragment did not increase the enrichment by GANC selection. This enrichment factor was also increased by reducing the size of the ttr fragment present between the two selection markers. However, it decreased the frequency of gene targeting and, overall, it did not increase the efficiency of isolating targeted clones. When structures of the vector DNA fragments present in 20 G418-resistant and GANC-resistant non-targeted clones were examined by Southern blot analysis, the inefficiency of GANC selection proved to be mostly caused by exonucleolytic degradation of HSV-tk genes progressing from ends of the vectors.

Animals↗
PubMed1994

A survey of genes expressed in undifferentiated mouse embryonal carcinoma F9 cells: characterization of low-abundance mRNAs.

As a first step to catalogue mRNAs present in mouse embryonal carcinoma F9 cells, 879 clones corresponding to low-abundance mRNAs were selected from among 2,896 randomly picked up clones of undifferentiated F9 cDNA libraries, using DNA probes complementary to poly(A)+RNAs prepared from undifferentiated F9 cells and to ones prepared from mouse fibroblast L cells. Five-hundred and eighty-two of the 879 clones were partially sequenced, and the subsequent homology search revealed that 201 corresponded to 180 known genes or known DNA sequences, which include not only housekeeping genes but also various tissue-specific genes. Interestingly, at least 24 of the 180 genes are development-related genes in mammals. Among these 24, those for midkine (growth and/or differentiation factor) and interferon-beta are reportedly up-regulated, and those for ECA39 (target for c-Myc regulation), REX-1 (zinc finger protein), and OCT-3 (POU-domain transcription factor) are down-regulated during the development of mouse embryonal carcinoma cells. Thirty-seven of the 582 clones matched the 36 previously reported unidentified ESTs (expressed sequence tags) and the remaining 344 corresponded to 329 novel ESTs. Therefore, partial sequencing of F9 cDNA clones corresponding to low-abundance mRNAs in F9 cells not only provides valuable information concerning development-related genes in mammals, but also many novel ESTs useful for studying mammalian genomes.

Animals↗
PubMed1994

Local environmental change from the G- to F-form of the actin molecule detected on anisotropy decay measurement.

The fluorescence intensity has been reported to increase 10 to 25 times when N-(1-pyrene)-iodoacetamide (PIAA)-conjugated actin polymerizes from the G- to the F-form. To elucidate the molecular mechanism underlying this process, we measured the time-averaged anisotropy of PIAA-actin in both the G- and F-forms. The anisotropy ratio of PIAA-G-actin (0.137 +/- 0.008) was smaller than that of PIAA-F-actin (0.221 +/- 0.008). Similar results were obtained when N-(1-pyrene)-3-iodopropionamide (PIPA), a PIAA analogue with an extra carbon-chain in its reactive group, was conjugated with actin. The anisotropy ratio increased from 0.096 +/- 0.042 to 0.199 +/- 0.035 when PIPA-actin was transformed from the G- to the F-form. Further more, we measured the anisotropy decay of PIAA-actin in both the G- and F-forms. Least-square fitting revealed that the decay pattern was well fitted the wobbling-in-cone model. In the G-form, the pyrene of PIAA actin diffused in a cone region with a vertical half angle of 33.4 degrees. This value decreased to 25.9 degrees when the actin was transformed to the F-form. Because PIAA and PIPA were conjugated at Cys-374 of actin, our results suggest that a small cleft exists in the actin molecule in the vicinity of Cys-374, and this cleft becomes narrow upon polymerization, resulting in an increase in fluorescence intensity.

Actins↗
PubMed1994

Expression of leukemia inhibitory factor in human endometrium and placenta.

Leukemia inhibitory factor (LIF), a cytokine that induces macrophage differentiation in the murine M1 myeloid leukemia cell line, is essential for blastocyst implantation in mice. However, its expression and the role it plays in the human uterus are unknown. To clarify these issues, we examined LIF gene expression in the human uterus by Northern blot hybridization and by a quantitative reverse transcription-polymerase chain reaction (RT-PCR) method. Analysis of LIF mRNA showed two hybridization bands, with estimated mRNA sizes of about 4.0-kb pairs and 1.8-kb pairs. LIF mRNA was detected at high levels in endometrial tissue and decidua, but at low levels in the chorionic villus in first trimester and term placenta. In the secretory phase, the endometrial tissue showed higher LIF expression than in the proliferative phase (9.5-fold; p < 0.01). The endometrial tissues were separated into a stroma-enriched fraction (SF) and an epithelium-enriched fraction (EF), and the LIF mRNA levels in each fraction were examined by quantitative RT-PCR. These levels were higher in the EF than in the SF (3.3-fold; p < 0.05). These findings suggest that, in humans, LIF plays a role in uterine function during the menstrual cycle, as well as during pregnancy.

Base Sequence↗
PubMed1994

High incidence of uterine inversion in mast cell-deficient osteopetrotic mutant mice of mi/mi genotype.

Mutations at either W or mi (microphthalmia) loci in the mouse can lead to a deficiency in melanocytes and mast cells. In addition, W mutants can be anemic and sterile, whereas mi mice are osteopetrotic because of a monocyte/macrophage/osteoclast defect. Since c-kit receptor tyrosine kinase is the gene product of the W locus and mi mutation has been suggested to affect the transduction of signals from the c-kit and c-fms receptors, we here examined the effect of mi mutation on fertility. Testes and ovaries from mi/mi mice were histologically normal, and the pattern of c-kit protein expression was not different from that of +/+ mice. Homozygous mutant crosses (mi/mi x mi/mi) were fertile, but inversion of the uterus occurred in 86% of the deliveries. In some cases, the placenta was found still attached to the inverted uterus after delivery. Decidual cells were present and expressed c-kit protein normally in the placenta of mi/mi mice. The inversion was also observed in mi/mi females mated to +/+ males. No uterine inversion was noted when +/mi females were crossed with mi/mi or +/mi males, suggesting that the genotype of the mother but not of the father or fetus is important for the pathogenesis. The numbers and body weights of mi/mi newborns were less than those of +/mi littermates. Mast cells were absent, but c-kit-positive cells were present, in the uterine muscle layers of pregnant mi/mi mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Percutaneous transluminal coronary angioplasty of "superdominant" left anterior descending artery. A case report.

A seventy-one-year-old woman suffering from angina pectoris had a superdominant left anterior descending artery with a 95% stenosis just after it extended the apex. This superdominant artery was demonstrated angiographically by the findings that it ran in the posterior interventricular sulcus and reached the crux of the heart. Percutaneous transluminal coronary angioplasty for the stenosis beyond the apex was successfully performed. After the procedure, she was relieved from chest pain.

Aged↗
PubMed1994

Morphological changes in the mouse popliteal lymph node after local injection of dextran sulfate.

Dextran sulfate 500 (DS) is known to be a potent B-cell mitogen. Injection of this compound (500 micrograms) into the footpads of mice produced characteristic changes in the draining popliteal lymph nodes, including elimination of macrophages from the lymphatic sinuses and lining the subcapsular sinus, a marked blast reaction throughout the nodal parenchyma, damage to the stroma of the outer peripheral cortex, and disorganization of all the existing primary and secondary lymph follicles. On day 6 after DS treatment, small lymphocytes bearing surface Ig were detected and distributed loosely and broadly in the deeper part of the peripheral cortex without any tendency to reorganize lymph follicles. On day 21, these small lymphocytes populated the outer and deeper layer of the peripheral cortex, sometimes forming nodular accumulations representing regenerating lymph follicles, and on day 35, various numbers of regenerated lymph follicles were present in the peripheral cortex. Tracer studies using FITC-conjugated human serum albumin showed that intraparenchymal diffusion of labeled HSA was highly blocked at the outer peripheral cortex on days 4 through 14, and that this pathway began to reopen at around 21 days. Disorganization of lymph follicles could be considered due to extensive damage to follicle stromata which became unable to bind B lymphocytes, and follicles seem to be re-organized in response to lymph-carried antigen in addition to restored follicle stromata and repopulation B lymphocytes. Macrophages had little effect on the lymph follicle formation in this study.

Animals↗
PubMed1994

Effect of pretreatment reagent for enamel on the adhesion of resin composite.

Aqueous solutions of methacryloyl tyrosine amide (MTYA), glyceryl methacrylate (GM), glutaraldehyde and succinic acid were prepared and their effectiveness as pretreatment reagents was evaluated by measuring the tensile bond strengths between resin composite and polished enamel. Pretreatment of polished enamel with MTYA.GM.S3 (A: 6% MTYA + 70% GM aqueous solution, B: 3% succinic acid aqueous solution) gave bond strengths of about 7 MPa, whereas pretreatment with MTYA.G.GM.S3 (A: 6% MTYA + 70% GM aqueous solution, B: 3% succinic acid + 2% glutaraldehyde aqueous solution) gave values of about 5 MPa. The mean bond strengths of each after 60 s of scrubbing with MTYA.GM.S3 or by MTYA.G.GM.S3 were raised to about 9.5 MPa, and the type of bond failure was fracture of the bonding agent. Although there were no resin tags, it was found that resin composite was able to adhere to polished enamel without acid etching.

Acid Etching, Dental↗
PubMed1994

Superoxide dismutase in developing mouse retina.

Superoxide dismutase (SOD) activity and the production site of H2O2 were studied in the developing mouse retina. The H2O2 production site moved from the inner to the outer region within the outer nuclear layer between the ages of 4 days through 12 days. This shift occurred in parallel with the development of the inner segments of the photoreceptor layer. SOD activity increased significantly at the age of 2-3 weeks. Both manganese-SOD (Mn-SOD) and copper-zinc SOD (Cu,Zn-SOD) activities (combined mean) in the retina of mice older than 2 weeks of age were 1.9 times greater in comparison with those of mice at the age of 2 weeks and younger. The mRNA of Cu,Zn-SOD did not change during these periods, suggesting that the translational efficiency of the mRNA and/or the stability of the SOD enzyme might increase, or the active form of the enzyme might increase during the development of the retina. The results of this study indicate that the localization and production of active oxygen change during the development of the mouse retina, and that SOD activity does exist in the retina from birth, increasing as its role in the protective system against active oxygen increases in the maturing mouse retina.

Animals↗
PubMed1994

Gene targeting by a vector with hairpin-shaped oligonucleotide caps.

To improve the efficiency of gene targeting, a linearized vector containing part of the mouse transthyretin (ttr) gene, a bacterial neomycin-resistance gene and a herpes simplex virus thymidine kinase gene was capped with hairpin-shaped oligonucleotides. The capped vector not only improved the efficiency of positive-negative selection about 3.5-fold, but also increased about 2.5-fold the ratio of ttr gene targeted clones to the selectants.

Animals↗
PubMed1993

Cloning, functional expression and tissue distribution of human cDNA for the alpha 1C-adrenergic receptor.

We have cloned human alpha 1C-adrenergic receptor from human prostate cDNA library. The deduced amino acid sequence of the clone (P2C4) encodes a protein of 466 amino acids that showed strong sequence homology to the previously cloned bovine alpha 1C-adrenergic receptor. The radioligand binding properties of P2C4 expressed in COS-7 cells were very similar to those of bovine alpha 1C-adrenergic receptor. With reverse-transcription polymerase chain reaction assay, we observed alpha 1C-adrenergic receptor transcripts in heart, brain, liver and prostate, but not in kidney, lung, adrenal, aorta and pituitary. The data show that the clone P2C4 encodes a human alpha 1C-adrenergic receptor cDNA, and the receptor subtype is expressed not widely but localized in several human tissues.

Amino Acid Sequence↗
PubMed1993

The expression of c-kit protein in human adult and fetal tissues.

The c-kit proto-oncogene encodes a tyrosine kinase receptor and is allelic with the dominant white-spotting (W) locus of the mouse. In this study we investigated the expression of human c-kit protein in various adult and fetal human tissues immunohistochemically using anti-human c-kit monoclonal antibody. To discriminate c-kit+ cells from mast cells expressing c-kit, mast cells were identified by staining with Toluidine blue. In oogonia, spermatogonia and skin melanocytes of the fetus and in oocytes of adult ovary, c-kit expression was detected. In adult uterus, c-kit+ cells were widely distributed in the basal layer of the endometrium, myometrium and cervix, the number and distribution being almost identical to those of mast cells. In fetal uterus, c-kit+ non-mast cells clustered beneath the epithelium and a few mast cells were observed in the myometrium and subserosal layer. In both adult and fetus, c-kit+ non-mast cells were detected within smooth muscle layers of the intestine, colon and oesophagus, while mast cells were observed in the mucosal and submucosal layers of these organs. In contrast to mice, no expression of c-kit protein was detected in the human placenta and decidua. Thus, the distribution of c-kit+ cells in various tissues is similar but not identical between adult and fetus and between human and mouse.

Adult↗
PubMed1993

Effects of the molecular weight of hyaluronic acid and its action mechanisms on experimental joint pain in rats.

OBJECTIVES: It has been shown previously that hyaluronic acid (HA) has an analgesic action on bradykinin induced pain in the knee joints of rats. This study further clarifies the effects of the molecular weight of HA and its mechanism of action in the same model using HA of molecular weight 800 to 2.3 x 10(6) daltons and a bradykinin antagonist. METHODS: Bradykinin and the test HA preparations were given to rats by intra-articular injection, and the severity of pain was evaluated by a change in the walking behaviour. RESULTS: HA with a molecular weight greater than 40 kilodaltons produces analgesic effects with a simultaneous or earlier injection. The ID50 values of HA with molecular weight 40, 310, 860, and 2300 kilodaltons were greater than 2.5, 0.6, 0.07, and 0.06 mg/joint respectively. The duration of the analgesic effect of 860 and 2300 kilodalton HA was 72 hours at 10 mg/ml, whereas that of 310 kilodalton HA was short, being undetectable after 24 hours. The analgesic action of HA of 860 kilodaltons was not changed by pretreatment with four saccharide HA and inhibited by pretreatment with HA larger than six to eight saccharides, capable of binding to HA receptors. Further, HA did not interfere with the analgesic action of the bradykinin antagonist, indicating that HA does not directly bind with bradykinin receptors. CONCLUSIONS: HA with a molecular weight of greater than 40 kilodaltons produced an analgesic effect, and HA of 860 and 2300 kilodaltons produced high and long-lasting analgesia. These effects of HA appear to be caused by the interaction between HA and HA receptors.

Animals↗
PubMed1993

[Diabetic retinopathy score and fuzzy logic in therapy].

A system for determining optimal treatment of diabetic retinopathy was devised on the basis of fuzzy logic. In a previous study we devised a new retinopathy scoring system from the multivariate analysis of fluorescein angiography findings. Using this scoring system we expressed severities as language terms and assigned them membership functions which showed grade (0-1). Next, the optimal treatment was determined by defuzzification of the membership functions given to treatment programs corresponding to the severity of retinopathy. This decision system is shown as a nomogram. Users of this system can get the grade for choosing optimal treatment after they input the retinopathy score into the nomogram. Assessment of this fuzzy decision system was performed by changes in the extent of dye leakage (Le) and neovascularization (NV) in 86 eyes of 48 diabetic patients followed up for over 12 months. When the grade of choice was over 0.5, the evaluation of choosing optimal treatment was 74% by Le and 68% by NV. Le improved but NV did not improve when the grade of choice was elevated to a higher level.

Aged↗