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Biomedical subjects

N Ishiguro

Publications and source records attributed to N Ishiguro.

At least 73 records · Page 4Linked to original sources
PubMed1998

Synovial fluid cartilage metabolism marker concentrations in osteonecrosis of the femoral head compared with osteoarthrosis of the hip.

OBJECTIVE: Biochemical aspects of the etiology of articular cartilage degeneration in osteonecrosis of the femoral head have not been investigated extensively. We analyzed biochemical conditions in the hip joint cavity, regarding articular cartilage turnover in osteonecrosis of the femoral head (ONFH). METHODS: Cartilage metabolism markers in synovial fluid were measured. Synovial fluid (SF) samples were collected from 19 ONFH cases and 17 control hips with osteoarthrosis (OA). Concentrations of carboxy-terminal type II procollagen peptide (pCOL-II-C), matrix metalloproteinase-3 (MMP-3), and tissue inhibitor of metalloproteinase-1 (TIMP-1) were measured. In addition, concentrations of unsaturated disaccharides of hyaluronic acid (delta di-HA), chondroitin 4-sulfate (delta di-4S), and chondroitin 6-sulfate (delta di-6S) in SF were measured by high performance liquid chromatography. RESULTS: The mean SF concentration of pCOL-II-C was higher on ONFH than in OA. The mean SF MMP-3 level was higher in ONFH than in OA, while the mean SF TIMP-1 level was the same in the 2 groups. The SF concentration of delta di-HA and the delta di-6S/delta di-4S ratio were higher in ONFH than in OA. CONCLUSION: Higher concentrations of cartilage metabolism markers in ONFH compared with OA SF suggest elevated cartilage turnover in the former disease.

Adolescent↗
PubMed1997

Activation transcription factor 1 involvement in the regulation of murine H-2Dd expression.

Resistance to radiation leukemia virus-induced leukemia is correlated with an increase in H-2D expression on the thymocyte surface. Recently, it has been shown that elevated H-2Dd expression on the infected thymocyte is a result of elevated mRNA transcription and that the transcriptional increase is correlated with elevated levels of a DNA binding activity, H-2 binding factor 1 (H-2 BF1), which recognizes the 5'-flanking sequences (5'-TGACGCG-3') of the H-2Dd gene. This target for transcription factor binding has been found to be identical in the 5'-regulatory region of 12 rodent class I genes, nine of which have been shown to be functional genes. Furthermore, this cis-element is found 5' of 20 primate class I genes (15 human genes), seven of which are known to be functional. Here, we demonstrate that activation transcription factor 1 (ATF-1) is one component of H-2 BF1. In addition, the levels of ATF-1 mRNA in uninfected and radiation leukemia virus-infected thymocytes parallel those of H-2Dd mRNA, and therefore, it is suggested that ATF-1 up-regulates the transcription of the H-2Dd gene after radiation leukemia virus infection of thymocytes. Transfection experiments also demonstrate that ATF-1 activates a reporter plasmid that contains the H-2 BF1 motif, but not a reporter lacking this motif. This is the first demonstration of the interaction of ATF-1 with 5'-regulatory sequences of major histocompatibility complex class I genes.

Activating Transcription Factor 1↗
PubMed1997

Alteration of the kinetics of type I procollagen synthesis in human osteosarcoma cells by 1,25-dihydroxyvitamin D3.

The kinetics of type I procollagen synthesis in a human osteosarcoma cell line, MG 63, were investigated after treatment with 1,25-dihydroxyvitamin D3 (1,25-(OH)2 D3), a hormonal inducer of phenotypic differentiation. Pulse label and chase experiments demonstrated greatly enhanced production and more rapid reduction of intracellular procollagen molecules in the 1,25-(OH)2 D3-treated cells as compared to the nontreated case. After a chase for 1 h, labeled procollagen was reduced by nine-tenths in 1,25-(OH)2 D3-treated cells, while half of the radioactivity still remained in nontreated cells. The expression rate of type I collagen, which was examined by pulse label experiment, was elevated in association with an increase in the mRNA coding for the type I collagen alpha 1 chain by 1,25-(OH)2 D3 treatment. However, the amount of intracellular procollagen present after 4 h continuous labeling was almost the same, independent of the 1,25-(OH)2 D3 treatment. Thus, we conclude that strage of the molecule was not affected. The results therefore suggest an increase in both the synthesis and secretion of type I collagen. The 1,25-(OH)2 D3 treatment was also found to induce the alpha subunit of prolyl 4-hydroxylase and to be associated with an elevated level of hydroxyproline in the procollagen. Moreover, gelatinase B-resistant procollagen molecules, indicative of intracellular procollagen molecules in the stable triple helical form, were detected only in the 1,25-(OH)2 D3-treated cells. These data suggest more efficient proline hydroxylation is involved in rapid secretion of procollagen after hormone administration. The present evidence points to posttranslational control of procollagen synthesis.

Animals↗
PubMed1997

Deletion of c-myb exon 9 induced by insertion of repeats.

A simple repeat was found to be inserted into exon 9 of the c-myb gene in three out of 20 bovine T lymphomas. The repeat was composed of multiple copies of a 12-nucleotide motif and had no significant homology to the sequences reported so far. Tumor cells containing the repeat expressed two kinds of c-myb mRNA: (1) are that included the repetitive sequence in exon 9, and (2) are that lacked the whole sequence of exon 9. Transfection of an expression vector containing exon and intron sequences and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of the mRNA demonstrated that the insertion of the repeat enhanced exon skipping of the transfected minigene. These observations imply that the insertion of the repeat may enhance exon skipping of the c-myb pre-mRNA. Although the transcription-activating activity by the c-Myb with the repeat was low, that by the c-Myb without exon 9 was three- to eightfold higher than the wild-type c-Myb. These data suggest that insertion of the 12-nucleotide repeat in codon 359 may result in c-Myb proteins having high- and low-transcription-activating activity.

Animals↗
PubMed1997

Macrophage activation and migration in interface tissue around loosening total hip arthroplasty components.

The bone-cement interface tissue of failed total hip arthroplasty (THA) has inflammatory characteristics, such as the presence of prostaglandin E2 and interleukin 1 (IL-1). We considered that the bone-cement interface tissue could be the site of granulomatous inflammation caused by a foreign-body reaction. It has been demonstrated that inflammatory cytokines and chemokines have an important role in granulomatous inflammation. Bone-cement interface tissue was obtained at revision from nine patients with failed cemented THA, and the role of macrophages was assessed by immunohistochemistry, electron microscopy, and molecular biological techniques. We used the reverse-transcriptional polymerase chain reaction to examine the expression of mRNA for IL-1 alpha, IL-1 beta, tumor necrosis factor alpha (TNF alpha), macrophage inflammatory protein (MIP)-1 alpha, MIP-1 beta, IL-8, and monocyte chemoattractant protein. Polyethylene debris surrounded by macrophages and phagocytosis of debris by macrophages was frequently observed in the interface tissue. Macrophage activation and the production of inflammatory cytokines such as IL-1 and TNF alpha might induce the development of interface tissue. Expression of chemokine mRNAs was also commonly seen, suggesting that this led to recruitment of macrophages into the bone-cement interface tissue. Debris released from implants appears to cause activation of macrophages and the production of inflammatory cytokines and chemokines that induce cellular recruitment into interface tissue. This mechanism might form a vicious cycle that aggravates THA loosening.

Aged↗
PubMed1997

Alternative usage of exon 1 of bovine PrP mRNA.

Here we report two types of bovine prion protein (PrP) mRNA that possessed different lengths of the 5'-untranslated region and were expressed in various bovine tissues. The two mRNA species were transcribed from identical positions but differed in the usage of the splice site for exon 1/intron. One mRNA possessed exon 1 consisting of 53 nucleotides and the other possessed exon 1 consisting of 168 nucleotides. Usage of exons 2 and 3 was identical for the two mRNA species. The two mRNA species were detected in all but spleen tissue; the mRNA possessing 168-nt exon 1 was not detected in bovine spleen. This is the first report on the tissue-specific alternative splicing of PrPc mRNA in any other species. Only a low level of PrPc appeared to be present in bovine spleen. These results suggested the possibility that the mRNA possessing 53-nt exon 1 was inefficiently translated into Prp; however, in vitro translation analysis showed no marked difference in translational efficiency between the two mRNA species.

Alternative Splicing↗
PubMed1997

The immunosuppressive effect of fresh allogeneic bone graft in mice.

We investigated the immunosuppressive effect of fresh bone allografts after donor specific spleen cell transfusion in mice. Allografts from major histocompatibility complex incompatible mice were performed one week after the transfusion. Cellular and humoral immune responses were recorded by measuring alloreactive cytotoxic T-lymphocyte activity and antibody activity. Survival in days of a second set skin graft was measured for determining clinical immunity. Alloreactive cytotoxic T-lymphocyte activity was stimulated at 17 days (10 days after bone grafting) and then suppressed at 31 days (24 days after bone grafting). The survival of the second set skin graft was not diminished at this time. This outcome indicates that the spleen cell stimulated cellular immune response was suppressed after fresh bone allografts, suggesting that these allografts have an immunosuppressive effect and induce immunosuppressive actions.

Animals↗
PubMed1997

Characterization of differentiation antigens expressed in bovine lymphosarcomas.

To characterize the cell-surface antigens expressed in tumour cells derived from bovine leukosis and to determine their cell lineages, the immunophenotypes of the tumour cells from 13 bovine lymphosarcomas were examined with 13 monoclonal antibodies (mAbs). Of 13 cattle with lymphosarcomas, four were identified clinically as having the thymic-type sporadic bovine leukosis (SBL) and one as having the skin-type; two had enzootic bovine leukosis (EBL) and six were untypable. Flow cytometric analysis revealed that the tumour cells from nine cases were of T-cell lineage (BoCD5+, BoCD2+ or BoCD2-) and two were of B-cell lineage (MHC-II+, BoCD5+, IgM+); there were two bovine leukaemia virus-infected cattle (EBL). T-cell tumours appeared to originate from immature (BoCD4-, BoCD8-) T cells, but there was no significant relationship between clinical type (EBL, calf-, skin- and thymic-type) and tumour-cell immunophenotype.

Animals↗
PubMed1997

p53 mutation as a potential cellular factor for tumor development in enzootic bovine leukosis.

Mutations of p53 in the lymphocytes from peripheral blood and from tumoral lymph nodes in six naturally occurring bovine leukemia virus (BLV)-infected cows were examined. A point mutation of the p53 gene was found in three of six (50%) BLV-infected cows. These p53 gene mutations resulted in amino acid substitutions of codons 144, 167 and 241. The BLV-infected cow in the tumor stage had abnormally proliferating monoclonal B-lymphocytes having the p53 mutation. However, the mutation was not found in somatic cells, except for tumor cells. These results show that p53 mutation plays an important role in the pathogenesis of BLV-induced neoplasms, and that the B-lymphocyte bearing p53 mutations may be a target cell for tumor formation of enzootic bovine leukosis.

Animals↗
PubMed1997

Sensitive enzyme-linked immunosorbent assay for detection of PrP(Sc) in crude tissue extracts from scrapie-affected mice.

An enzyme-linked immunosorbent assay (ELISA) was developed that detects PrP(Sc) in crude extracts from brain and spleen tissue of scrapie-affected mice with high sensitivity and specificity. Brain tissue was homogenized in 8% Zwittergent 3-12 and 0.5% Sarkosyl. The homogenate was treated with collagenase and DNase I and then subjected to proteinase K digestion. Precipitates containing PrP(Sc) were obtained by ultracentrifugation. Spleen tissue was homogenized in 4% Triton X-100 and 0.5% Sarkosyl, and the homogenate was treated firstly with collagenase and DNase I, and secondly with proteinase K. PrP(Sc) was then extracted with 6.25% Sarkosyl and precipitated through salting-out with NaCl and by ultracentrifugation. When PrP(Sc) was dissolved in 3-4 M guanidine thiocyanate and adsorbed to microtiter plates, strong and specific reactions to the formation of antigen-antibody complexes could be detected by ELISA. The sensitivity of PrP(Sc)-detection for this ELISA, as measured by serial dilution of scrapie material in tissue homogenates from uninfected animals, was equal or higher than that attained by Western blot. This ELISA is more rapid than Western blot and seems to be more suitable for screening large numbers of animals. It also has potential application for the diagnosis of the transmissible spongiform encephalopathies.

Animals↗
PubMed1997

Differential display analysis of murine collagen-induced arthritis: cloning of the cDNA-encoding murine ATPase inhibitor.

We used the differential display technique in order to detect a new gene involved in murine type II collagen-induced arthritis (CIA). In this study, we have identified a novel gene, IF1, whose expression level is increased during the natural course of CIA. Northern blot analyses suggest that IF1 is involved in the natural course of CIA but is not involved as a trigger of CIA. IF1 is considered to be the murine ATPase inhibitor gene for several reasons. First, IF1 shows an extremely high homology to the rat ATPase inhibitor; the highly conserved region between rat and bovine amino acid residues 22-45, which is the minimum sequence showing ATPase inhibitory activities, is also highly conserved in IF1. Second, IF1 possesses a histidine-rich region in the same area, which is thought to be important for regulation of mammalian inhibitors. Third, the tissue distribution of IF1 is very suggestive. The expression of IF1 was very strong in energetic organs such as the heart, brain and kidney, and the development of arthritis requires great amounts of ATP. As arthritis develops rapidly, the cellular ATP pool may be decreased. Before the ATP pool is exhausted, the ATPase inhibitor may serve as a brake for ATP hydrolysis. If the supply of free energy can be reduced, the inflammation of arthritis may in turn be restored. Our hypothesis is that the ATPase inhibitor is involved in regulating the inflammatory responses.

Amino Acid Sequence↗
PubMed1997

Effect of dibutyryl cyclic adenosine monophosphate on skeletal muscle reperfusion injury in the rat.

To clarify the action of dibutyryl cyclic adenosine monophosphate (DBcAMP) on reperfused ischemic muscle, experiments were conducted using the hindlimbs of 18 male Lewis rats. At the midportion of the thigh all tissues except for the femoral artery and vein were transected, and a route for continuous intravenous infusion was secured in a contralateral limb vein. After inducing total ischemia by clamping the femoral artery and vein with a vascular clamp for 4 h, the limb was reperfused for 1 h. Blood flow was then compared using the hydrogen gas clearance method in a group in which DBcAMP 10 mg was continuously infused from a vein in the contralateral hindlimb from 1 h prior to the induction of ischemia to 1 h after the completion of ischemia (DBcAMP group), a group in which saline was infused in the same manner (control group), and a group which was subjected to biopsy alone (biopsy group). The percent change in blood flow was significantly higher in the DBcAMP group than in the control group at 15 and 30 min after the release of the clamp. Adenosine triphosphate (ATP), phosphocreatine (PCr), and lipid peroxide (LPO) were measured in tissue samples obtained 1 h after reperfusion. Serum LPO was measured in blood samples collected at the same time. ATP values were higher in the DBcAMP group than in the control group. PCr was significantly higher in the DBcAMP group than in the control group. LPO levels in skeletal muscle tissue did not differ significantly between the DBcAMP and control groups. In contrast, serum LPO levels were significantly lower in the DBcAMP group than the control group. On morphologic analysis the control and DBcAMP groups showed normal vascular endothelial cells and absence of the 'no-reflow' phenomenon. These data confirm that in this reperfusion model the administration of DBcAMP enhances the viability of skeletal muscle cells. Moreover, mediated by an effect on vascular endothelial cells this agent is thought to be of help in mitigating the vascular endothelial cell injury occurring in acute ischemic injury. DBcAMP may be a useful agent in mitigating skeletal muscle ischemia-reperfusion injury.

Adenine Nucleotides↗
PubMed1997

Instability of the 12-nucleotide repeat in c-myb gene of bovine T-lymphoma cells.

Insertion of a 12-nucleotide repeat in c-myb gene exon 9 was observed in about 15% of sporadic bovine T-lymphomas. The 12-nucleotide repeat in the T-lymphoma cells showed deletion and insertion of the repeat units during cultivation of the cells. To know whether deficiency in DNA loop repair is involved in the instability of the repeat, abilities to bind and correct the loop structure in nuclear extracts were examined. The nuclear extracts of all examined cells had ability to bind and correct the loop structure. These data suggest that instability of the 12-nucleotide repeat in bovine T-lymphoma cells might be independent of deficiency of DNA loop repair function.

Animals↗
PubMed1997

Epitope mapping of a monoclonal antibody specific to feline panleukopenia virus and mink enteritis virus.

To obtain monoclonal antibodies (MAbs) specific to feline panleukopenia virus (FPLV) and mink enteritis virus (MEV), 15 hybridomas secreting MAbs against MEV-Abashiri were established and the properties of the MAbs were analyzed. The cross-reactivity of MAbs revealed that one MAb, P2-215 was specific for FPLV and MEV, whereas the remaining fourteen MAbs reacted with canine parvovirus (CPV), FPLV, and MEV. Epitope analyses using various CPV/MEV chimeric viruses revealed that the MAb P2-215 recognized the epitope comprised of amino acid 93-Lys in VP2, which is known to be FPLV and MEV-specific.

Amino Acid Sequence↗
PubMed1997

Characterization of the bovine prion protein gene: the expression requires interaction between the promoter and intron.

We cloned the part of the bovine PrP gene which contains the 5'-flanking region, exon 1, exon 2 and intron 1 to analyze its promoter region. The 5' non-coding region of the bovine PrP gene consisted of three exons and two introns, and its organization was similar to that of the mouse, rat and sheep PrP genes. The 5'-flanking region of the bovine PrP gene from the transcription start site to nucleotide position -88 was (G + C)-rich (78%) and contained three potential binding sites for the transcription factor Sp1, but no CCAAT-box or TATA-box. This region showed high homology (89%) with that of the sheep PrP gene, but relatively low homology (approximately 46-62%) with the same region of the mouse, rat, hamster and human PrP genes. The position from -88 to -30 within the 5'-flanking region of the bovine PrP gene showed major promoter activity. However, this region was able to function properly only in collaboration with the region at +123 to +891 of intron 1 of the bovine PrP gene.

Animals↗
PubMed1996

A perfluorochemical prevents ischemia-reperfusion injury of muscle.

This investigation evaluated the effect of oxygenated perfluorochemical (PFC) perfusion on the viability of ischemic skeletal muscle compared to hypothermic preservation. Twenty-five hindlimbs of 13 white rabbits were divided into five groups: a PFC group (7 hr of ischemia with 6 hr of PFC perfusion), a PFC/ reperfusion group (8 hr of ischemia with 6 hr of PFC perfusion and 1 hr reperfusion), a hypothermia group (7 hr of ischemia with 6 hr of 4 degrees C cold preservation), a hypothermia/reperfusion group (8 hr of ischemia with 6 hr of 4 degrees C cold preservation and 1 hr of reperfusion), and a control group. The levels of adenine nucleotides, creatine phosphate, hypoxanthine, xanthine, and lipid peroxide were determined in each group. In the PFC and PFC/reperfusion groups, the ATP level remained at 90% of that in the control group. The hypoxanthine level in the hypothermia/reperfusion group was decreased to 70% of that in the hypothermia group and the xanthine level was increased to 130%. In the PFC and PFC/reperfusion groups, hypoxanthine and xanthine levels were much lower and were similar to those in the control group. Lipid peroxide levels were also lower in the PFC and PFC/reperfusion groups than in the hypothermia/reperfusion group. Electron microscopy showed that endothelial cells from the PFC/reperfusion groups were not swollen. These results suggest that PFC perfusion is superior to hypothermia in inhibiting the generation of free radicals and in preventing ischemia-reperfusion injury of skeletal muscle.

Adenine Nucleotides↗