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Biomedical subjects

T Osawa

Publications and source records attributed to T Osawa.

At least 109 records · Page 6Linked to original sources
PubMed1998

Bifid ribs observed in the third and the fourth ribs.

Three cases of bifid ribs were found in two cadavers during routine dissections at the Iwate Medical University School of Dentistry. All of the cases were found in the third or the fourth rib. The distal parts of the osseous rib bifurcated with an angle of 60 degrees and both of the branches had their own costal cartilage. The costal cartilage fused again to form the trunk which was connected to the sternum. The space between the two branches was filled with presumably normal intercostal muscles. Blood supply was maintained by a small branch from the interthoracic artery to the upper branches. However, the intercostal nerves did not branch toward the upper branch but only ran along the lower margins of the lower branches of the bifid ribs.

Aged↗
PubMed1997

Protein modification by lipid peroxidation products: formation of malondialdehyde-derived N(epsilon)-(2-propenol)lysine in proteins.

Malondialdehyde (MDA), a naturally occurring dialdehyde produced in the membrane lipid peroxidation, is known to react with lysine residues of proteins, but the MDA-lysine adducts generated in the proteins have not been characterized adequately. In the present study, we provide evidence that the enaminal-type MDA-lysine adduct, N(epsilon)-(2-propenal)lysine, is formed in human low-density lipoprotein (LDL) upon reaction with MDA or Cu2+. We found that the incubation of N(alpha)-acetyllysine with MDA generated N(alpha)-acetyl-N(epsilon)-(2-propenal)lysine as the predominant product. In addition, a polyclonal antiserum raised against the MDA-modified protein was found to contain antibody populations that could be purified by affinity gel prepared by covalent attachment of N(alpha)-acetyl-N(epsilon)-(2-propenal)lysine. It was concluded that the affinity-purified anti-N(epsilon)-(2-propenal) lysine antibody was highly specific to the enaminal derivative of both lysine residues and phosphatidylethanolamine, based on the observations that (i) MDA was the only aldehyde which generated immunoreactive materials in proteins; (ii) among structurally defined MDA-lysine adducts tested, the antibody recognized the enaminal adduct only; and (iii) immunoreactivity to N-(2-propenal)serine was still significant but much weaker than its reactivity to N-(2-propenal)ethanolamine. Furthermore, analysis of antibody recognition sites with a variety of N-(2-propenal)alkylamines revealed that the mono-specific antibody recognized the N-2-propenal-N-ethyl moiety [-(CH2)2-NH-CH=CH-CHO] of enaminal adducts. Determination by a competitive enzyme-linked immunosorbent assay demonstrated that N(epsilon)-(2-propenal)lysine accounted for 33.7 and 3.1% of the lysine residues that disappeared during in vitro incubation of LDL with MDA and Cu2+, respectively. These results suggest that N(epsilon)-(2-propenal)lysine represents a major form of MDA covalently attached to proteins.

Antibodies, Monoclonal↗
PubMed1997

Selective inhibition of platelet-derived growth factor (PDGF) receptor autophosphorylation and PDGF-mediated cellular events by a quinoline derivative.

This report describes the biological effects of our original compound, Ki6783 ((3,4-dimethoxy)-4-phenoxy-6,7-dimethoxyquinoline), a potent and selective inhibitor of platelet-derived growth factor (PDGF) receptor autophosphorylation. This compound strongly inhibited autophosphorylation of the PDGF beta-receptor in cultured rat glomerular mesangial cells (MC) bearing this receptor (IC50 0.1 microM), although it did not inhibit autophosphorylation of other growth factor receptors even at 100 microM. In a cell-free kinase experiment, it showed selective inhibition of PDGF beta-receptor tyrosine kinase. A kinetic study of the compound to this tyrosine kinase revealed a competitive mode of action to ATP. [3H]Thymidine incorporation and cell proliferation of MC were inhibited by Ki6783 in a dose-dependent manner after Ki6783 and PDGF-BB were added to the culture medium. Furthermore, this compound normalized the fibrotic cell shape of v-sis-transformed NIH3T3 cells, which grow in an autocrine manner via the PDGF receptor. These effects could be explained by the inhibition of intracellular signal transduction triggered by PDGF receptor autophosphorylation, in which activation of mitogen-activated protein kinase occurs. These results suggest that Ki6783 is one of the more potent and selective inhibitors of PDGF receptor autophosphorylation and that it may be useful in ameliorating cell abnormalities due to excess action of PDGF and its receptor systems in several diseases.

3T3 Cells↗
PubMed1997

Cellular response to the redox active lipid peroxidation products: induction of glutathione S-transferase P by 4-hydroxy-2-nonenal.

Membrane lipid peroxidation is known to produce various aldehydic compounds which cause a wide range of biological effects including heart disease, aging, and cancer. To investigate the effect of lipid peroxidation products on the expression of glutathione S-transferases (GSTs), which catalyze the conjugation of reactive chemicals with glutathione and play an important role in protecting cells, normal rat liver epithelial cells (RL34) were exposed to a variety of aldehydic compounds. We found that the GST activity in RL34 cells was induced by alpha,beta-unsaturated aldehydes, such as acrolein (1.3-fold), crotonaldehyde (1.3-fold), 4-hydroxy-2-hexenal (HHE) (1.4-fold), and 4-hydroxy-2-nonenal (HNE) (1.7-fold). The induction of GST activity by HNE was time-dependent, reaching a plateau after 16 h. The immunoblot analysis using the polyclonal antibodies against GST isozymes demonstrated that GST-P (pi-class), a well-known tumor marker, was significantly induced 16 h after the HNE treatment. Also, immunostaining for the presence of GST-P confirmed the enhanced expression of GST-P in the cytoplasm of the cells. Northern blot analysis revealed that the HNE treatment of RL34 cells for 1 h enhanced the expression of GST-P mRNA, which returned to the control level after 16 h. These data suggest that the induction of GST-P by HNE may represent an important cellular defense mechanism against oxidative injury.

Aldehydes↗
PubMed1997

Protein modification by a Maillard reaction intermediate methylglyoxal. Immunochemical detection of fluorescent 5-methylimidazolone derivatives in vivo.

Methylglyoxal (MG), an endogenous metabolite that increases in diabetes, is a common intermediate in nonenzymatic glycation (Maillard reaction) in vivo. Here we describe the immunochemical approach to the detection of MG adducts in proteins in vitro and in atherosclerotic lesions of human aorta in vivo. The reaction of protein (bovine serum albumin) with MG led to selective loss of arginine and lysine residues, accompanied by the formation of 5-methylimidazolone (N delta-(5-methylimidazolon-2-yl)ornithine) and imidazolysine (1,3-di-lysino-4-methylimidazole) derivatives, respectively. The anti-5-methylimidazolone antibody was prepared by immunizing rabbits with a MG-keyhole limpet hemocyanin conjugate and purifying the serum on an affinity gel prepared by covalent attachment of the 5-methylimidazolone derivative. The antibody cross-reacted with the proteins treated with not only MG but trioses, such as hydroxyacetone, dihydroxyacetone, and glyceraldehyde. The immunohistochemical analysis revealed that atherosclerotic lesions of human aorta contained 5-methylimidazolone derivatives whose distributions were identical to those of advanced glycation end products (AGEs) detected by the anti-AGE antibody.

Animals↗
PubMed1997

Biomarker evidence of DNA oxidation in lung cancer patients: association of urinary 8-hydroxy-2'-deoxyguanosine excretion with radiotherapy, chemotherapy, and response to treatment.

Ratios of urinary 8-hydroxy-2'-deoxyguanosine to urinary creatinine (8-OHdG/creatinine) have been considered as a good biological indicator of DNA oxidation. Urinary 8-OHdG/creatinine levels of lung cancer patients were evaluated by enzyme-linked immunosorbent assay using a monoclonal antibody N45.1 during radiotherapy and chemotherapy. An increase in urinary 8-OHdG/creatinine was found in non-small-cell carcinoma (non-SCC) patients during the course of radiotherapy. SCC patients showed higher levels of urinary 8-OHdG/creatinine than the controls. Furthermore, SCC patients with complete or partial response to the chemotherapy showed a significant decrease in urinary 8-OHdG/creatinine while patients with no change or progressive disease showed an increase.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1997

Oxidative modification of tryptophan residues exposed to peroxynitrite.

The aim of this study was to clarify the mechanism of loss of Trp residues in proteins exposed to peroxynitrite. The Trp residues in bovine serum albumin and collagen IV were decreased by peroxynitrite treatment. To identify the degradation products of the Trp residue by peroxynitrite, tert-butoxycarbonyl-L-tryptophan (Boc-Trp) was used as a model of the Trp residue in proteins, and the products formed from peroxynitrite-treated Boc-Trp were then isolated. Boc-Trp decreased with an increase in peroxynitrite concentration. N-Formylkynurenine, oxindole, and hydropyrroloindole were identified as major products. The formation of these products may account for the losses of Trp residues in proteins by peroxynitrite.

Animals↗
PubMed1997

Induction of cell death by chimeric L-selectin-Fas receptors.

In the present study, we have established a system where engagement of an adhesion molecule triggers a death signal into cells. L-selectin, which is a well characterized adhesion receptor involved in the initial adhesion between lymphocyte and endothelium, was fused to the intracellular domain of an apoptosis-inducing molecule, Fas. Ligation of the chimeric receptors with a carbohydrate ligand for L-selectin, fucoidin or a mAb that recognizes the lectin domain of L-selectin, induced apoptosis in receptor-expressing cells. However, ligation with an anti-L-selectin mAb reactive with a non-ligand binding site did not induce apoptosis, indicating that stimulation through the lectin domain of L-selectin in the chimeric receptor leads to signal delivery. Upon activation L-selectin shows a unique proteolytic cleavage at the membrane proximal site on the extracellular (EC) domain, of which the significance is also unclear. We found that truncations in the EC domain which abrogate the proteolytic cleavage of L-selectin did not influence induction of apoptosis, suggesting that the cleavage on the EC domain itself is not important for the signaling function of the chimeric receptor. This is the first demonstration that an adhesion signal can be converted to a signal that leads to apoptotic cell death.

Animals↗
PubMed1997

Covalent attachment of 4-hydroxy-2-nonenal to erythrocyte proteins.

It has been established that the covalent modification of proteins occurs in vivo as a consequence of reaction with reactive lipids such as 4-hydroxy-2-nonenal (HNE). The fact that HNE occurs in blood under physiological as well as pathophysiological conditions suggests that erythrocytes undergo modification by HNE. Here we describe the immunochemical characterization of HNE-treated erythrocytes by demonstrating the Michael-type HNE addition to both membrane and cytosolic proteins. Exposure of erythrocytes to HNE (0-0.5 mM) for 2 h resulted in HNE labeling of multiple membrane proteins. Pretreatment of erythrocytes with a sulfhydryl reagent, N-ethylmaleimide (NEM), resulted in a significant decrease of HNE attached to the proteins, suggesting that HNE primarily reacts with the sulfhydryl groups of erythrocyte membrane proteins, whereas enhanced HNE labeling of the membrane proteins was observed when the erythrocytes were pretreated with H2O2 (0.1-5 mM) for 15 min. On the other hand, highly selective modification of a 30 kDa protein was observed in the hemolysates of erythrocytes treated with HNE. The protein, which represents a major intracellular target of HNE in erythrocytes, was identified as carbonic anhydrase, based on the observations that (i) a reverse-phase HPLC analysis of the chloroform/ethanol extract of HNE-treated erythrocytes detected two major proteins, which cross-reacted with anti-carbonic anhydrase antibody as well as with the anti-HNE adducts antibody, (ii) the chloroform-ethanol extraction of authentic carbonic anhydrase gave a similar HPLC pattern, and (iii) the HNE treatment of erythrocytes resulted in the partial inhibition of the carbonic anhydrase activity.

Aldehydes↗
PubMed1997

Postpartum adrenal, pituitary and ovarian functions in dairy cows.

Postpartum stress and the resumption of pituitary and ovarian functions were investigated in 29 dairy cows during the first month after calving. Adrenocorticotropic hormone (ACTH, 25 i.u., IM) challenge tests for adrenal function at 8 and 22 days postpartum and gonadotropin releasing hormone (GnRH, 50 micrograms IM) challenge tests for pituitary function at 7 and 21 days postpartum were conducted. Ovarian function, particularly the interval to first ovulation, was evaluated through weekly rectal palpation and ultrasonographic examination, as well as thrice weekly milk progesterone levels. Elevated basal cortisol levels were found in 24.1% and 20.7% of cows at 8 and 22 days postpartum, respectively. Adrenal response to ACTH was similar at 8 and 22 days postpartum with low, medium and high peak responses in 13.8%, 62.1% and 24.1% of cows, respectively. A moderate correlation was observed between basal and peak cortisol levels. Peak pituitary luteinizing hormone (LH) response was low in 65.5%, medium in 27.6% and high in 6.9% of cows at 7 days postpartum. Pituitary responsiveness improved by day 21 with low in 34.5%, medium in 37.9% and high in 27.6% of cows. About a week after calving, a negative correlation between peak cortisol and peak LH levels was observed. First ovulation occurred in 28%, 48% and 17% of postpartum cows during the second, third and fourth week, respectively. These results indicate a negative effect of peak cortisol levels on pituitary responsiveness to GnRH stimulation during the first week, altered adrenal and pituitary function in a third of the cows until the third week and restoration of ovarian function in most cows (93%) a month after parturition.

Adrenal Glands↗
PubMed1997

Spice constituents scavenging free radicals and inhibiting pentosidine formation in a model system.

Many antioxidants have been found in spices and herbs, and some of them are well known as strong scavengers of active oxygen radicals. We have isolated active products, which markedly inhibited the formation of malondialdehyde (MDA from 2-deoxyribose and the hydroxylation of benzoate with the hydroxyl radical, from methanol extracts of allspice and clove. Pimentol from allspice, and biflorin and its isomer, abbreviated as clove3, from clove were identified as the active principles. These revealed strong activity as hydroxyl radical scavengers at a concentration of 2.0 microM. The antioxidative activities in an in vitro model system involving the rabbit erythrocyte membrane ghost were as strong as those of alpha-tocopherol at 200 microM. Such advanced glycation end products (AGE) as pentosidine are biomarkers of diabetes mellitus, and active oxygens have been suggested to be involved in the formation of AGE. The above-mentioned free radical scavengers effectively inhibited the formation of pentosidine in a model system of N alpha-t-butoxycarbonyl-fructoselysine and N alpha-t-butoxycarbonyl-arginine.

Animals↗
PubMed1997

Postpartum plasma PGF metabolite profile in cows with dystocia and/or retained placenta, and effect of fenprostalene on uterine involution and reproductive performance.

Objectives of this study were to show postpartum plasma PGF2 alpha metabolite (PGFM) profile, to clarify whether endogenous PGF2 alpha plays a certain role in the uterine involution in cows with dystocia and/or retained placenta, and to examine the effects of fenprostalene, a long-acting PGF2 alpha analog, on the uterine involution and reproductive performance of the cows with abnormal puerperium. A group of 27 cows with dystocia and/or retained placenta showed a massive release of PGF2 alpha after parturition as indicated by a rise of plasma concentrations of PGFM, significantly higher than 33 cows with normal puerperium. The duration of the elevated plasma PGFM concentrations in the cows with abnormal puerperium was shorter than that of the normal cows. In cows with normal puerperium, those showing relatively longer duration of elevated plasma PGFM levels needed a shorter period for postpartum uterine involution than the cows showing a shorter duration of the PGFM elevation (P < 0.01), while no such relationship was observed in cows with abnormal puerperium. In field trials, an administration of an exogenous PGF2 alpha, fenprostalene, at 7 to 10 days (78 cows) or 14 to 28 days postpartum (74 cows) was found to be effective in facilitating uterine involution and resumption of ovarian cyclicity, and improved reproductive performance. It may be concluded that a large amount of PGF2 alpha is released for a relatively shorter period in cows after dystocia and/or retained placenta and the elevation of PGFM is not responsible for the uterine involution. The administration of the exogenous PGF2 alpha was shown to be effective at improving the postpartum reproductive performance of cows with abnormal puerperium.

Abortifacient Agents, Nonsteroidal↗
PubMed1997

Antioxidative mechanism and apoptosis induction by 3-hydroxyanthranilic acid, an antioxidant in Indonesian food Tempeh, in the human hepatoma-derived cell line, HuH-7.

Recently, a new potent antioxidant was isolated from Tempeh (a traditional fermented soybean food in Indonesia) and was identified as 3-hydroxyanthranilic acid (HAA). This study deals with the antioxidant mechanism of HAA under biological systems and the cytokilling function of HAA to human malignant cells. HAA eliminated free radicals and inhibited the formation of fatty acid hydroperoxide in vitro, suggesting that HAA would serve as an antioxidant in the initial reaction in lipid oxidation systems. Actually, HAA inhibited the formation of the dominant product of membrane lipids, 12-hydroxyeicosatetraenoic acid (12-HETE) at a high concentration, while HAA accelerated 12-HETE formation at a low concentration in mammalian tissue. HAA oxidized glutathione and inhibited superoxide dismutase in vitro. Furthermore, HAA inhibited cell growth and induced apoptosis to HuH-7, a human hepatoma-derived cell line. As long as HAA is taken as a component of Tempeh, and not in large doses as a chemical, it may possibly act as a prooxidant rather than an antioxidant in vivo.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1997

Some perspectives on dietary inhibition of carcinogenesis: studies with curcumin and tea.

Topical application of curcumin inhibits chemically induced carcinogenesis on mouse skin, and oral administration of curcumin inhibits chemically induced oral, forestomach, duodenal, and colon carcinogenesis. Curcumin and other inhibitors of cyclooxygenase and lipoxygenase are thought to inhibit carcinogenesis by preventing the formation of arachidonic acid metabolites. In contrast to our expectation of a tumorigenic effect of arachidonic acid, we found that treatment of 7,12-dimethylbenz[a]anthracene-initiated mouse skin with very high doses of arachidonic acid twice daily, 5 days a week for 26 weeks, failed to result in tumors. We considered the possibility that some of the cancer chemopreventive effects of curcumin may be related to an effect of this compound on cellular differentiation, and we investigated the effect of curcumin on differentiation in the human promyelocytic HL-60 leukemia cell model system. Although curcumin alone had little or no effect on cellular differentiation, when it was combined with all-trans retinoic acid or 1alpha,25-dihydroxyvitamin D3 a synergistic effect was observed. It is possible that many dietary chemicals in fruits, vegetables, and other edible plants can prevent cancer by synergizing with endogenously produced stimulators of differentiation such as all-trans retinoic acid, 1alpha,25-dihydroxyvitamin D3, and butyrate. More research is needed to test this hypothesis. Administration of green or black tea inhibits carcinogenesis in several animal models, and tumor growth is also inhibited. Several examples were presented of chemopreventive agents that inhibit carcinogenesis in one animal model but enhance carcinogenesis in a different animal model. Greater efforts should be made to understand mechanisms of cancer chemoprevention and to determine whether a potential chemopreventive agent is useful in many experimental settings or whether it is useful in only a limited number of experimental settings.

Animals↗
PubMed1997

Quantitative immunohistochemical determination of 8-hydroxy-2'-deoxyguanosine by a monoclonal antibody N45.1: its application to ferric nitrilotriacetate-induced renal carcinogenesis model.

The DNA base-modified product 8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the most commonly used markers for the evaluation of oxidative DNA damage. A monoclonal antibody specific for 8-OHdG (N45.1) was characterized and applied in quantitative immunohistochemistry. N45.1 recognized both the modified base and deoxyribose structure of 8-OHdG and required a concentration two orders higher of 8-hydroxyguanosine as a competitor in the ELISA. In addition, N45.1 did not cross-react with the original four deoxyribonucleosides, other DNA base-modified products such as 8-hydroxy-2'-deoxyadenosine and O6-methyl-2'-deoxyguanosine, or urine components such as uric acid, creatine, and creatinine. A ferric nitrilotriacetate-induced rat renal carcinogenesis model was used for the evaluation of quantitative immunohistochemistry. The 8-OHdG index of quantitative immunohistochemistry, as analyzed by NIH image freeware, correlated reasonably well with the 8-OHdG amount determined by high-performance liquid chromatography with an electrochemical detector-except for a difference in peak time, which could be attributed to the selection of target location. The present method has advantages over the high-performance liquid chromatography/electrochemical detector, gas chromatography/mass spectrometry, and 32P-postlabeling methods in that it allows localization of 8-OHdG to be specified without the risk of artifactual production of 8-OHdG during the DNA extraction and hydrolytic processes.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1997

Characterization of a specific polyclonal antibody against 13-hydroperoxyoctadecadienoic acid-modified protein: formation of lipid hydroperoxide-modified apoB-100 in oxidized LDL.

Lipid hydroperoxide may react with protein or amino phospholipid without secondary decomposition. We prepared a polyclonal antibody to lipid hydroperoxide-modified proteins using 13S-hydroperoxy-9Z, 11E-octadecadienoic acid-modified keyhole limpet hemocyanin (13-HPODE-KLH) as immunogen. The antibody recognized 13-HPODE-modified bovine serum albumin (BSA), but not aldehyde-modified proteins, such as malondialdehyde-modified BSA. The antibody also recognized adducts derived from 13-HPODE and 13S-hydroperoxy-9Z, 11E, 15Z-octadecatrienoic acid (13-HPOTRE(alpha)). The oxidized alpha-linolenic acid- and linoleate-protein adducts were recognized by the antibody. Oxidized phospholipid-protein adducts were scarcely recognized by the antibody. However, when ester bonds of phospholipids containing linoleic acid were hydrolyzed by alkaline treatment, the cross-reactivities appeared. The result suggests that a phospholipid hydroperoxide can react with a protein directly or indirectly, and a carboxyl terminal (COOH) of the lipid in an adduct was needed as an epitope. Oxidized LDL (ox-LDL) was prepared by the incubation of LDL with copper ion or 2,2'-azobis(2-amidinopropane)dihydrochloride (AAPH), and the formation of lipid hydroperoxide-modified apolipoprotein was confirmed using the antibody. A slight immunoreactivity was observed in ox-LDL without alkaline treatment. When the ox-LDL was treated with alkali to hydrolyze the ester bonds of the lipid, enhanced antigenicity appeared with time-dependency. The results suggest that lipid hydroperoxide-modified apolipoprotein was formed during the oxidation of LDL.

Antibodies↗
PubMed1996

Inhibition of lipid peroxidation and the active oxygen radical scavenging effect of anthocyanin pigments isolated from Phaseolus vulgaris L.

No attention has been paid to anthocyanin pigments from the viewpoint of inhibitors of lipid peroxidation and scavengers of active oxygen radicals; therefore, we investigated the antioxidative, radical scavenging, and inhibitory effects on lipid peroxidation by UV light irradiation of three anthocyanin pigments, pelargonidin 3-O-beta-D-glucoside (P3G), cyanidin 3-O-beta-D-glucoside (C3G), and delphinidin 3-O-beta-D-glucoside (D3G), isolated from the Phaseolus vulgaris L. seed coat, and their aglycons, pelargonidin chloride (Pel), cyanidin chloride (Cy), and delphinidin chloride (Del). All pigments had strong antioxidative activity in a liposomal system and reduced the formation of malondialdehyde by UVB irradiation. On the other hand, the extent of antioxidative activity in a rat liver microsomal system and the scavenging effect of hydroxyl radicals (-OH) and superoxide anion radicals (O2-) were influenced by their own structures.

Animals↗