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Biomedical subjects

N Inoue

Publications and source records attributed to N Inoue.

At least 55 records · Page 3Linked to original sources
PubMed1996

Development of a canine chemotherapeutic model with ifosfamide.

The purpose of this study was to develop a canine experimental model for neoadjuvant chemotherapy of primary bone tumors with ifosfamide, which is safe and clinically relevant for use in human beings with bone tumors. Our study was divided into two steps, each with four dogs. In the first step ifosfamide was administered for 4 consecutive days in three cycles with 3-week intervals between each cycle. For this first step a daily dosage of 300 mg/m2 of body surface resulted in only moderate leukopenia, whereas a daily dosage of 450 mg/m2 caused severe leukopenia. Therefore, to determine the maximal dose tolerable and to verify the results from step 1, we administered the higher daily dosage of 450 mg/m2 in step 2 for four successive cycles with 3-week intervals. In each step one dog died acutely after the first cycle of chemotherapy. In addition during step 2 one dog died of overwhelming sepsis after the second cycle of ifosfamide. The remaining five dogs survived without other appreciable laboratory abnormalities. Neither hematuria nor proteinuria was observed throughout the course of study, and relevant findings were not observed at autopsy. We determined that 450 mg/m2 was the maximal tolerated dosage of ifosfamide for our regimen, with the dose-limiting factor being myelosuppression, specifically leukopenia. Using this canine model, we can estimate the effect of ifosfamide on bone graft incorporations and the fixation of biologic prostheses that is clinically the most important aspect of limb salvage surgery.

Animals↗
PubMed1996

[Immunoglobulin heavy chain variable region(VH) genes in B cell clones producing anti-colon antibodies in ulcerative colitis].

We have established anti-colon antibodies producing B cell clones from the patient with ulcerative colitis, and analyzed the immunoglobulin heavy chain variable region (VH) gene segments of these clones using RT-PCR with VH family specific primers. VH3 family was used by all ten clones from ulcerative colitis that produced anti-colon antibodies, while various VH gene families were used by eight clones from normal controls. By nucleotide sequence analysis, two LPL clones were thought to be derived from a germ line gene segment, VH26. However, other three PBL clones were derived from 22-2B, 9-1 and 1.9-III, respectively. These results suggest the heterogeneity of the colonic antigens that autoantibodies in ulcerative colitis recognize.

Antibody-Producing Cells↗
PubMed1996

[Intraventricular arachnoid cyst--on the origin of intraventricular arachnoid cysts].

Arachnoid cysts very rarely occur within the ventricular system, where no arachnoid tissue exists. We present three cases of intraventricular arachnoid cyst with special reference to its origin. The first patient was a 5-year-old boy who complained of headaches and enlargement of his head. A CT scan revealed obstructive hydrocephalus and a large cystic lesion in the right lateral ventricle. The symptoms resolved after fenestration of the cyst and cystoperitoneal shunt. The second patient was a 49-year-old woman who complained of headache and numbness in her left upper extremity. A CT scan and MRI revealed a large cyst in the trigone of the right lateral ventricle. Fenestration of the cyst wall and cystoperitoneal shunt were performed relieving her complaints. The third patient was a 42-year-old man who complained of frequent seizures and dizziness. A CT scan and MRI demonstrated a moderate size cystic mass in the inferior horn of the right lateral ventricle. The symptoms were improved by partial resection of the cyst wall. Immunohistochemical studies and light microscopy confirmed that the cyst walls were composed of arachnoid membrane, implying that the cysts were arachnoid cysts. CT and MRI in these three cases showed widening of the choroidal fissure bordering the cyst wall, occasionally involving a part of the protruding cyst wall. On enhanced CT and MRI, the choroid plexus in the trigone of ipsilateral lateral ventricle was displaced anterolaterally, implying that the cysts had grown from outside the choroid plexus. Postoperative MRI demonstrated the shrunken cyst wall attached to the choroidal fissure. The surgical findings in case 3 also showed that the cyst wall was attached firmly to the choroid plexus. These findings appeared to indicate that the intraventricular arachnoid cyst originated from the arachnoid layer drawn into the choroidal fissure with choroidal vascular mesenchyme.

Adult↗
PubMed1995

Analysis of cis-acting regions upstream of the rat Na+/K(+)-ATPase alpha 1 subunit gene by in vivo footprinting.

By means of in vivo footprinting, we examined the putative cis-acting DNA elements located between -50 and -122 of rat Na+/K(+)-ATPase alpha 1 subunit gene ATP1A1. Proximal and distal GC box sequences and a consensus sequence for the active transcription factor (ATF) were protected for all the tissues examined (kidney, brain and liver). Putative cooperation between two binding factors on the ATF site and the proximal GC box was observed. The overall in vivo footprinting profiles of the three tissues did not exhibit any marked differences that could account for the variation in the extent of tissue-specific transcription. The alpha 1 regulatory element (ARE) found by Suzuki-Yagawa et al. does not appear to be an element responsible for tissue-specific regulation of the gene.

Animals↗
PubMed1995

Structure and chromosomal localization of the GPI-anchor synthesis gene PIGF and its pseudogene psi PIGF.

Posttranslational modification by the GPI glycolipid anchor is essential for the surface expression of many membrane proteins. Defect of GPI biosynthesis due to somatic mutation in the hematopoietic stem cell is the basis for an acquired genetic disease, paroxysmal nocturnal hemoglobinuria (PNH). Previously, an X-linked gene PIGA (phosphatidylinositol glycan class A), which participates in the first step of the biosynthesis, was shown to be mutated in abnormal cells from all 60 patients with PNH. The cDNA of another GPI synthesis gene PIGF was previously cloned, but it is not involved in pathogenesis of PNH. In the present study, we have analyzed PIGF genomic clones. The PIGF gene contained six exons spanning about 40 kb and was located to the short arm of chromosome 2 at 2p16-p21. The frequency of mutations on both alleles of PIGF should be much lower than that of mutation in the X-linked PIGA, accounting for a lack of involvement of PIGF in PNH. We also identified the processed pseudogene of PIGF (psi PIGF) and mapped it to 5q35.

Base Sequence↗
PubMed1995

Reduction of masseter muscle activity in bottle-fed babies.

Our previous studies suggested that there are significant differences in the growth of the jaw and in muscle activity between breast- and bottle-fed infants. To confirm these differences quantitatively, myoelectric activities of the masseter muscles of bottle-fed babies were studied. Twelve bottle-fed babies, as well as 12 breast-fed babies as a control group, were examined electromyographically during bottle or breast feeds. The duration time of sucking bursts, interval time, cycle time, 0-to-peak amplitude, integrated amplitude of bursts, and integrated amplitude/duration time were measured and the number of bursts over 30 microV was counted. All differences of means were significant by t-test. The masseter muscle activity in bottle-fed babies is significantly reduced. Our results are contrary to previous papers in which almost the same sucking actions in both breast and bottle feeding were reported. The reason why previous researchers thought that the sucking patterns in breast- and bottle-fed babies are essentially the same is considered, and the implications of the differences for dental health are discussed.

Bottle Feeding↗
PubMed1995

An assay method for nitric oxide synthase in crude samples by determining product NADP.

An assay method for nitric oxide synthase (NOS) was developed based on fluorometric or enzymatic determination of NADP+. An aliquot (< or = 2 microliters) of crude enzyme sample, homogenate or supernatant of rat cerebellum, was added to a reaction mixture containing arginine, NADPH, and O2 and incubated at 25 degrees C for 30 min. A strongly fluorescent substance was formed from a product, NADP+, and measured (fluorometric assay). When rat cerebellar layers were assayed, freeze-dried sections of layers (0.2 to 2 micrograms dry wt) were added directly into 1.24 microliters of NOS reaction mixture and the total NADP+ formed in picomole amounts was specifically amplified up to 4000-fold and determined, using an enzymatic NADP cycling amplification reaction (enzymatic-cycling assay). NOS activity was calculated as the difference in NADP+-forming activity in the absence and presence in the reaction mixture of NG-nitro-L-arginine, a specific inhibitor of NOS. Enzymatic activity was analyzed in rat cerebellar supernatants by two procedures. With supernatants (and purified macrophage NOS), the ratio between the specific activities on a protein basis using the present NADP+ formation assay and using [3H]citrulline formation from [3H]arginine as substrate was 2. The distribution of NOS activity was shown between the particulate and supernatant fractions of rat cerebellum. The molecular and granular layers of rat cerebellum contained similar NOS activities, while the activity in the white matter was negligibly low. NOS distribution is also reported among rat organs.

Animals↗
PubMed1995

Effects of creatine and beta-guanidinopropionic acid on the growth of Ehrlich ascites tumor cells: i.p. injection and culture study.

Growth of Ehrlich ascites tumor (EAT) cells in the abdominal space of mice or in cell culture was studied in response to i.p. injection or addition, respectively, of creatine or creatine analogue beta-guanidinopropionic acid (beta-GPA). The increase in body weight of the mice due to cancer growth was less in the beta-GPA-injected than in the creatine- or sham-injected group. The volume of abdominal ascites and total cell counts at 11th day after implantation of EAT cells was significantly less in the beta-GPA than in the other groups. The proliferation rate of EAT cells in the beta-GPA group was 27% and 35% of the creatine- and sham-injected groups, respectively. Supplementation of creatine tended to enhance the growth of EAT cells. The creatine concentration in ascites fluid was approximately 4-times greater than in blood plasma of sham-injected control mice. But the creatine content in EAT cells was significantly reduced to approximately 50% in response to beta-GPA injection. Cell culture without creatine caused a significant decrease in viability. The viability was improved, however, by addition of either creatine or serum into the medium. By contrast, it was not significantly increased by addition of serum alone which caused only a minor elevation of the creatine level (23 microM). It is suggested that EAT cell growth is inhibited by lowering the availability of creatine in association with some unknown factors in serum or ascites fluid.

Abdomen↗
PubMed1995

Change of phosphotyrosine immunoreactivity on microglia in the rat substantia nigra following striatal ischemic injury.

Using immunohistochemistry, we investigated changes in phosphotyrosine (P-Tyr) immunoreactivity on the microglia of the rat substantia nigra (SN) following striatal ischemic injury produced by transient middle cerebral artery (MCA) occlusion. Anterograde axonal degeneration in the SN due to striatal ischemic injury was detected by depletion of calcineurin immunoreactivity in that region from 1 day after operation. From 3 days to 1 month (the longest period examined in this study) after MCA occlusion, there was a significant increase in P-Tyr immunoreactivity in the SN ipsilateral to the MCA occlusion. Also, light microscopic observation showed that the microglia exhibited an increased immunoreactivity for P-Tyr and characteristic morphological changes in the ipsilateral SN. The present results indicate that a signal transducing cascade(s) associated with tyrosine phosphorylation may be involved in the activation of the microglia in the SN responding to anterograde degeneration of the striatonigral pathway.

Animals↗
PubMed1995

The endothelial cell nitric oxide synthase: is it really constitutively expressed?

During the past two years, the enzyme responsible for production of endothelium-derived nitric oxide, the endothelial cell NO synthase (ecNOS) has been cloned and the gene encoding this enzyme isolated, cloned and its structure characterized. This research has provided direction for a variety of studies of regulation of the ecNOS. Several features of the ecNOS are compatible with a constitutively expressed, poorly regulated gene, including absence of a TATA box and numerous SP-1 sites. The promoter also contains a number of putative binding domains which suggest that it may be regulated by a variety of transcription factor mediated signals. In this review we will discuss evidence to support the concept that the ecNOS is a constitutively expressed gene subject to a modest degree of regulation by important physiological influences.

Amino Acid Oxidoreductases↗
PubMed1995

Experimental study on the optimum flow rate and pressure for selective cerebral perfusion.

The optimum flow rate and pressure for selective cerebral perfusion during moderate hypothermia (25 degrees C) were investigated in 36 mongrel dogs. Cerebral perfusion was performed for 90 minutes at a flow rate of 100% (the physiologic flow rate), 50%, 25%, and 0%, or no flow (cerebrocirculatory arrest). Somatosensory evoked potentials were monitored to assess brain function. An excess lactate level was considered an index of anaerobic cerebral metabolism, and histopathologic evaluation was performed. Somatosensory evoked potentials showed no abnormalities at flow rates of 100% and 50%, but became abnormal in some dogs at 25% and in all dogs under no-flow conditions. The excess lactate level only increased at a no-flow rate, but not significantly. Histopathologic evaluation showed no ischemic changes at flow rates of 100% and 50%, but there were slight ischemic changes at 25% and severe ischemic damage at no flow. The mean carotid arterial pressure was 63.1 +/- 5.9, 39.8 +/- 6.2, 24.9 +/- 6.0, and 11.3 +/- 3.5 mm Hg at a flow rate of 100%, 50%, 25%, and no flow, respectively. These results suggest that the safe range of flow rates for cerebral perfusion during moderate hypothermia is more than 50% of the physiologic level with a carotid arterial pressure of about 30 mm Hg or more.

Animals↗
PubMed1995

Principle of the bark of Phellodendron amurense to suppress the cellular immune response: effect of phellodendrine on cellular and humoral immune responses.

Previously we have isolated the quaternary base alkaloids, magnoflorine and phellodendrine, from Phellodendri Cortex (cortex of Phellodendron amurense Rupr., Rutaceae) as the biologically active principles to suppress local graft-versus-host (GvH) reactions in mice. In this paper, we focus on phellodendrine. Phellodendrine suppressed local semisyngeneic GvH reactions and systemic allogeneic GvH reactions in X-ray irradiated recipient mice. Phellodendrine also suppressed the induction phase of sheep red blood cell (SRBC)-induced delayed type hypersensitivity in mice and tuberculin-induced delayed type hypersensitivity in guinea pigs, but did not suppress the effector phase of these reactions. Surprisingly, phellodendrine, unlike prednisolone and cyclophosphamide, did not affect antibody production in mice to SRBC. Phellodendrine was expected to be a valuable new type of immunosuppressor against the cellular immune response.

Animals↗
PubMed1995

Human herpesvirus 6B origin-binding protein: DNA-binding domain and consensus binding sequence.

We previously demonstrated by a DNA-binding assay that the human herpesvirus 6B (HHV-6B) replication origin has a structure similar to those of alphaherpesviruses, although the HHV-6B and herpes simplex virus type 1 (HSV-1) origin-binding proteins (OBPs) and origins are not interchangeable. Here we describe additional properties of the interaction between HHV-6B OBP and the HHV-6B origin. Competitive electrophoretic mobility shift assays (EMSAs) with DNA duplexes containing single-base alterations allowed deduction of a consensus DNA sequence for HHV-6B-specific OBP binding, YGWYCWCCY, where Y is T or C and W is T or A, while that for HSV-1-specific binding was reported to be YGYTCGCACT. By EMSA, the HHV-6B OBP DNA-binding domain was mapped to a segment containing amino acids 482 to 770. However, in Southwestern (protein-DNA) blotting, the region sufficient for the DNA binding encompassed only amino acids 657 to 770. Similarly, Southwestern blotting showed that amino acids 689 to 851 of HSV-1 OBP had HSV-1 origin-binding activity, although this region was insufficient for origin binding in the EMSA. Although the longer DNA-binding domains identified by EMSA have marginal overall homology among HHV-6B and alphaherpesvirus OBP homologs, the smaller regions sufficient for the binding observed by Southwestern blotting have significant similarity. From these results, we propose a hypothesis that the DNA-binding domain of herpesvirus OBPs consists of two subdomains, one containing a conserved motif that contacts DNA directly, and another, less well conserved, that may modulate either the conformation or accessibility of the binding domain.

Amino Acid Sequence↗
PubMed1995

Molecular regulation of the bovine endothelial cell nitric oxide synthase by transforming growth factor-beta 1.

The promoter region of the endothelial cell nitric oxide synthase (ecNOS) gene contains potential response elements for transforming growth factor-beta 1 (TGF beta 1). TGF beta 1 plays an important role in the pathogenesis of atherosclerosis, vascular hypertrophy, and angiogenesis. We therefore sought to determine whether TGF beta 1 might modulate ecNOS expression in bovine aortic endothelial cells (BAEC). TGF beta 1 increased ecNOS mRNA in a dose-dependent manner. TGF beta 1 also increased ecNOS protein content. The production of nitrogen oxides (NOx), assessed by chemiluminescence, and nitric oxide synthase activity, assessed by arginine/citrulline conversion were increased in TGF beta 1-treated cells. Transcriptional activity of the 5'-flanking promoter region of the ecNOS gene was increased by TGF beta 1, as assessed by transfection with promoter/luciferase constructs. Deletion analysis suggested that the TGF beta 1-response element was present between nucleotides -1269 and -935 from the first transcription start site, in which a putative nuclear factor-1 (NF-1) binding site existed. Gel shift assays showed that nuclear protein(s), immunologically similar to CCAAT transcription factor/NF-1, bound to the putative NF-1 binding site in a sequence-specific manner. Mutation of the putative NF-1 binding site in the promoter/luciferase construct significantly decreased the responsiveness to TGF beta 1. In conclusion, TGF beta 1 increases ecNOS expression associated with an increase in production of NO in BAEC. This response is probably mediated by transcriptional activation of the ecNOS gene promoter.

Amino Acid Oxidoreductases↗