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Biomedical subjects

T Iwasaki

Publications and source records attributed to T Iwasaki.

At least 19 recordsLinked to original sources

Purification and characterization of a sialidase from Bacteroides fragilis SBT3182.

A sialidase from Bacteroides fragilis SBT3182 was purified 2,240-fold to apparent homogeneity by ammonium sulfate precipitation and sequential chromatographies on DEAE-Toyopearl 650M, Hydroxyapatite, MonoS and Superose6 columns. The N-terminal amino acid sequence of this sialidase, Ala-Asp-X-Ile-Phe-Val-Arg-Glu-Thr-Arg-Ile-Pro-, was determined. Substrate specificity of this enzyme using a variety of sialoglycoconjugates showed a 1.5- and 2.2-fold preference for sialyl alpha 2-8 linkages when compared with alpha 2-3 and alpha 2-6 bound sialic acids, respectively. The native sialidase had a molecular weight of 165kDa, as determined by Superose6 gel filtration chromatography and consisted of three subunits each of 55kDa by SDS-polyacrylamide gel electrophoresis. This enzyme had optimal activity at pH6.1 with colominic acid as substrate.

Amino Acid Sequence

Differential effect of self-stimulation on dopamine release and metabolism in the rat medial frontal cortex, nucleus accumbens and striatum studied by in vivo microdialysis.

Changes in the extracellular levels of dopamine (DA) and its metabolites in the dopaminergic terminal regions, the medial frontal cortex (MFC), nucleus accumbens (NAC), and striatum (STR), were measured by microdialysis during self-stimulation of the medial forebrain bundle (MFB) in rats pretreated with the DA uptake inhibitor, nomifensine (1 mg/kg, i.p.). Self-stimulation of the MFB in nomifensine-pretreated rats caused an increase in the extracellular DA level in the MFC and NAC but not in the STR. Self-stimulation also increased the extracellular concentrations of the main DA metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) to a similar extent in the MFC and NAC and to a lesser extent in the STR. Thus, there was a regional difference in the neurochemical changes following self-stimulation with either the MFC or the NAC showing larger extracellular levels of DA, DOPAC, and HVA than the STR. Furthermore, these changes were observed on both hemispheres ipsilateral and contralateral to the stimulation. The results indicate that self-stimulation of the MFB preferentially activates the mesocorticolimbic DA systems, thereby bilateral increases in the release of DA and its metabolism being produced in their terminal regions, the MFC and NAC.

3,4-Dihydroxyphenylacetic Acid

Herbimycin A blocks IL-1-induced NF-kappa B DNA-binding activity in lymphoid cell lines.

The mechanism by which interleukin-1 alpha (IL-1 alpha) activates NF-kappa B DNA-binding activity is not completely understood. While it is well established that protein kinase C can activate NF-kappa B, neither protein kinase C nor protein kinase A appears to be critical in the induction of NF-kappa B by IL-1 alpha. Since a number of growth factors signal via protein tyrosine kinase, in this study we examined a possible involvement of protein tyrosine kinase in the IL-1 alpha-induced NF-kappa B. The results showed that in the murine pre-B cell line 70Z/3 and in the murine T cell line EL-4 6.1 C10 IL-1 alpha-induced NF-kappa B was associated with transient increase in protein tyrosine kinase activity. Pre-treatment of these cell lines with herbimycin A, an inhibitor of tyrosine kinase activity, blocked the IL-1 alpha-enhanced protein tyrosine kinase activity and the IL-1 alpha-induced NF-kappa B DNA-binding activity. Herbimycin A at concentrations sufficient to block IL-1 alpha-induced NF-kappa B did not block the phorbol 12-myristate 13-acetate (PMA)-induced NF-kappa B. The data suggest that IL-1 alpha and PMA activate NF-kappa B by different pathways and that induction of NF-kappa B DNA-binding activity by IL-1 might be dependent on protein tyrosine phosphorylation.

Amino Acid Sequence

Direct interaction between an antigen-specific B cell clone and an MHC class II-reactive helper T cell clone.

TP67.14 established by somatic hybridization is a 2,4,6-trinitrobenzenesulfonic acid (trinitrophenyl, TNP)-specific B cell clone with a receptor molecule for TNP on the cell membrane, and MS202 is an interleukin-2 (IL-2)-dependent T helper (Th) cell clone reactive to auto-MHC class II antigens (IAk and IEk) as previously reported. In the present study it was shown that MS202 considerably induced the maturation of TP67.14 into anti-TNP plaque-forming cells (PFCs), and this response was markedly augmented by the addition of TNP-keyhole limpet hemocyanin (KLH). Recombinant cytokines and the culture supernatant of MS202 with TP67.14 did not affect the generation of anti-TNP antibodies by TP67.14. Also, neither anti-IL-4 nor anti-IL-5 monoclonal antibody (mAb) inhibited the maturation of TP67.14 mediated by MS202. The differentiative effect of MS202 on TP67.14 was completely lost when each cell was separately cultured using a semipermeable membrane. Monoclonal antibodies against LFA-1 beta molecules significantly blocked the development of anti-TNP PFCs induced by MS202, as well as anti-IAk and anti-IEk mAbs. Interestingly, the plasma membrane-enriched fraction (PM) derived from MS202 exhibited much more differentiative effects on TP67.14 treated with TNP-KLH than PM from other T cell lines and concanavalin A-induced T lymphoblasts. In addition, TNP-conjugated PM from MS202 by itself induced a great number of anti-TNP PFCs. The present findings indicate that MS202 is capable of inducing the maturation of TP67.14, which is considered to represent a population of B cells with antigen specificity in a late lineage of B cell maturation, through direct cell contact but not soluble factors. This suggests that B cells with antigen specificity, in the presence of antigen, can be induced to mature into antibody-secreting cells through direct contact with Th cells; in this process surface major histocompatibility complex class II and lymphocyte function-associated antigen 1 (LFA-1) molecules are directly involved and the cell membrane derived from Th cells provides a transductional signal for maturation of B cells with antigen specificity in the presence of antigen.

Animals

Detection of capsid antigen of human papillomavirus (HPV) in benign lesions of female genital tract using anti-HPV monoclonal antibody.

We established a murine monoclonal antibody (K1H8) to human papillomavirus (HPV) using alkaline-disrupted virions of HPV type 1 (HPV-1) as the immunogen. K1H8 recognized a 57 kD capsid protein of HPV-1 and detected the antigen in paraffin sections of formalin-fixed tissue. With K1H8, we examined immunohistochemically 68 biopsy specimens obtained from the female genital tract. The specimens were histologically condyloma acuminatum or koilocytotic lesions with or without dysplasia and each specimen was found to harbour a single type of genital HPV, such as types 6, 11, 16, 18, 31, 33, 42, 51, 52, 56, and 58, by Southern blot hybridization analysis. The antigen was localized in the nuclei and occasionally in the cytoplasm of squamous cells showing koilocytotic changes. Eighty-four per cent of the specimens (57 cases) showed positivity for the antigen, indicating that K1H8 is a broadly-reactive antibody to various genital HPVs. The results suggest that benign mucosal lesions of the female genital tract are more frequently associated with viral production and are a potential source of transmission.

Animals

Immunohistochemical study of skin lesions in herpes zoster.

Thirty-seven biopsy skin tissues of herpes zoster taken from 27 patients were analysed immunohistochemically using two monoclonal antibodies detecting either nucleocapsid or glycoproteins of varicella-zoster virus (VZV) on paraffin sections of formalin fixed tissues. Skin lesions of herpes zoster were divided clinically into four stages: erythematous, vesicular, pustular and ulcerative. In the erythematous stage, VZV antigens, if detected, were found only within ballooning cells in the lower epidermis or follicular epithelium. In the vesicular stage, antigens were detected in the cells around and within the intraepidermal vesicles and in histiocytes or fibrocytes of the dermis in all cases and in the endothelial or perineural cells in 10 of 14 cases. In the pustular stage, the antigens were observed in degenerated or necrotic keratinocytes and multinucleated giant cells within pustules and some necrotic cells in the dermis. In the ulcerative stage, the viral antigens were detected only at the ulcer margin and around the hair shaft in 2 of 7 cases. These results suggest that VZV initially involves the epidermis in the erythematous stage, subsequently invades the dermis in the vesicular stage, and disappears in the early ulcerative stage.

Antigens, Viral

Myocardial uptake of antimyosin antibody compared with serum myosin light chain I levels in patients with myocardial infarction.

Myocardial accumulation of In-111-antimyosin (InAM) was evaluated in comparison with circulating serum myosin light chain I (LCI) level at the time of InAM injection. Seventeen consecutive patients were studied at various stages ranging from 6 days to 34 days after myocardial infarction (MI). The infarct area was positive for InAM uptake in all patients (100%), and significant myocardial uptake was observed in 14 patients (82.4%). The intensity of InAM uptake correlated with the infarct location shown by ECG and CAG. In contrast, 12 patients (70.6%) had normal or undetectable serum myosin LCI levels, with 5 being normal (0.42-2.5 ng/ml) and 7 undetectable (0.42 ng/ml or less). Only 5 patients (29.4%) had elevated serum myosin LCI levels at the time of InAM injection, and this elevation was slight, ranging from 3.4 to 4.5 ng/ml (mean: 3.75 ng/ml). Among patients with undetectable, normal, and elevated serum myosin LCI levels, there was no significant correlation between InAM uptake and the serum myosin LCI level. Thus, even after the serum myosin LCI level has decreased to normal, InAM can still bind to cardiac myosin in patients with MI, presumably until there is complete recovery from the hibernating myocardium due to ischemic damage.

Adult

Preoperative evaluation of myocardial viability by thallium-201 imaging in patients with old myocardial infarction who underwent coronary revascularization.

The myocardial uptake and redistribution in thallium scintigraphy and the regional wall motion by echocardiography were evaluated by a semi-quantitative method in 42 patients who previously had myocardial infarction (50 target vessels) and underwent coronary revascularization. The aim of this study was to elucidate the significance of the initial image, delayed image and redistribution on thallium-201 scintigraphy for clinical diagnosis of the myocardial viability. As a semi-quantitative analysis, we used a bull's-eye display for thallium image and centerline method for echocardiographic wall motion, and compared the results before and after revascularization. As a result, the thallium grade improved postoperatively in all 17 areas which preoperative had showed redistribution, and also in 11 of the 32 areas without preoperative redistribution. The sensitivity, specificity and accuracy of preoperative thallium redistribution for predicting myocardial viability were 61%, 100% and 78%, respectively, when the postoperative improvement in the thallium grade was used as the standard. The postoperative probability of improvement in the thallium grade increased in proportion to the preoperative thallium grade (delayed image) (p less than 0.01). There was no correlation between the preoperative thallium delayed image and postoperative improvement in wall motion. Postoperative improvement in thallium image and wall motion could not be predicted from the preoperative wall motion. Thus, postoperative improvement in thallium images can be anticipated if redistribution is present on the preoperative thallium image, and the preoperative thallium delayed image is useful for predicting myocardial viability. Improvement in wall motion could not be predicted preoperatively by these methods.

Adult

Functional role of self IA molecules in polyclonal B cell activation using an autoreactive B cell clone derived from (NZB X NZW) F1 mice.

The mechanism of polyclonal B cell activation in autoimmune diseases was investigated by using an autoreactive B cell clone established by somatic hybridization with B cells derived from NZB X NZW (B/W) F1 mice. Briefly, splenic B cells from B/W F1 mice were fused with M12.4.1, a mutant of a B cell line, in the presence of polyethylene glycol and DMSO. NW47.7, a subclone of a resulting hybridoma, expresses B cell surface antigens on the cell membrane, namely IAd, IgM, B220, the receptors for the C3 fragment of complement (C3R), and the Fc portion of IgG (Fc gamma R). It also possesses a receptor molecule for mouse red blood cells treated with bromelain (Br-MRBC) on its surface, by rosette-forming assay with Br-MRBC. In contrast, parental M12.4.1 does not express IAd and IgM on the cell membrane, and does not bind to Br-MRBC under the same conditions. Thus, it is likely that NW47.7 may be an autoreactive B cell clone specific for Br-MRBC. Interestingly, NW47.7 was induced to generate a significant number of IgM-secreting cells when treated with Br-MRBC and rIL-5. Furthermore, mAb against IAd molecules, but not IAk and KdDd, markedly inhibited the differentiative effect of polyclonal activators such as LPS and rIL-5. Also, when MHC identical irradiated B cells were added to the culture of NW47.7 as a stimulator, the induction of IgM-producing cells was greatly augmented, but this augmenting effect was lost by interfering with direct contact of NW47.7 cells with stimulator B cells using a semipermeable membrane, as well as by the addition of mAb against IAd molecules. In addition, irradiated NW47.7, but not M12.4.1, by itself could enhance the secretion of IgM by NW47.7 as a stimulator, but this enhancing effect markedly decreased in the presence of anti-IAd mAb. The present results suggest that surface IA molecules on B cells are involved during the differentiative response to polyclonal activators, and may directly provide a differentiative signal for maturation of B cells into IgM-secreting cells.

Animals

Molecular cloning of a novel human papillomavirus (type 60) from a plantar cyst with characteristic pathological changes.

In a plantar cyst composed of the wall of the squamous cell layer and the horny inner substance in the lower dermis, we found characteristic pathological changes, such as cytoplasmic eosinophilic inclusions and vacuolated structure, and, immunohistochemically, the papillomavirus capsid antigen. The human papillomavirus (HPV) DNA cloned from the cyst showed no homology with other known prototypes of HPV (HPV 1 through HPV 59) by Southern blot analysis under stringent conditions and was named as HPV 60. HPV 60 DNA was found in three other cases of plantar cyst with the identical pathological changes, but not in a plantar cyst without such changes. The results suggest that HPV 60 has unique biological properties to induce a plantar cyst as a distinct type of cutaneous HPV.

Blotting, Southern

Magnetic resonance microscopy of rabbit eyes.

Magnetic resonance (MR) micro-imaging was performed on enucleated eyes from rabbits previously injected with perfluoropropane gas (C3F8), with or without the surgical creation of retinal detachment. Condensed vitreous, which exhibited shortened longitudinal relaxation time (T1), could be differentiated with proton-density and T1-weighted imaging. Gradient-echo imaging could in addition detect vitreo-retinal tractions. The detached retina itself was also seen. Further, proton-density but not T1-weighted imaging showed lens opacities appearing as high-intensity regions. MR microscopy is a convenient method for gross morphological examination of intact eyes.

Animals

Reconstitution of the active form from the amino- and carboxyl-terminal fragments of a reactive site-modified subtilisin inhibitor of adzuki beans (Vigna angularis).

The complex of an adzuki bean subtilisin inhibitor (ASI-II) with its target enzyme, prepared at pH 7.6, was subjected to reversed-phase HPLC in a trifluoroacetic acid-acetonitrile system. Two peptide fragments derived from the reactive site-modified ASI-II were obtained. The analyses of the amino acid composition and sequence of these two fragments revealed that one corresponded to the region from the amino-terminal Lys to the reactive site P1 Ala and the other, to the region from the reactive site P1' Asp to the carboxyl-terminal Gly of the inhibitor. Although neither fragment alone showed inhibitory activity against subtilisin, an equimolar mixture of both fragments was found to inhibit strongly the target enzyme, as did the intact inhibitor. Thus, it was suggested that the two fragments have strong specific affinity with each other, regenerating the reactive site-modified ASI-II, to inhibit the target enzyme.

Amino Acids

Suprahepatic vena cava anastomosis of the donor liver to the recipient retrohepatic vena cava in canine liver transplantation.

A revised method of the upper caval anastomosis in canine orthotopic liver transplantation is described. It uses the recipient retrohepatic inferior vena cava below the suprahepatic veins for vascular suturing. Ten consecutive operations were performed to assess the feasibility of this method, with special reference to the outflow obstruction at the level of the suprahepatic inferior vena cava. Seven of 10 dogs survived more than 6 days. The cause of death was not related to the outflow obstruction in any instance. Regardless of the duration of survival, free hepatic vein pressures as well as portal vein pressures of all dogs remained in the physiological range even after the skin closures. Based on these observations, we conclude that this approach is safe and reproducible in experimental transplantation of the canine liver.

Anastomosis, Surgical

Syntheses and antiulcer activities of 2-aminonorbornene derivatives.

2,2,-Disubstituted norbornenes (1, 2), 2,2-Disubstituted norbornane (3), 2,2,3-trisubstituted norbornenes (4, 5), oxanorbornenes (6) and azanorbornenes (7) were synthesized by the Diels-Alder reaction using alpha, beta-dehydroamino acids as a key step, and their antiulcer activities were examined. The oxazolidine derivative (1h) exhibited the most potent activities against several ulcer-models in rat.

Animals

A synthesis of 2-substituted 2-aminoethanol derivatives having inhibitory activity against protein kinase C.

A series of 2-aminoethanol derivatives was synthesized and their inhibitory activities against protein kinase C were investigated. Among these compounds, 2-endo-hexadecylamino-5-norbornene-2- exo-methanol (4h) and 2-endo-hexadecylamino-5-norbornene-2,3-exo-dimethanol (4i) inhibited protein kinase C at the IC50 values of 2 x 10(-5) and 1 x 10(-5) M, respectively, but not protein kinase A at a concentration of 1 x 10(-3) M. The structure-activity relationships are discussed.

Animals

[Protective effects of the fractions extracted from the callus of Acer nikoense Maxim. on alpha-naphthylisothiocyanate induced liver injury].

It has been reported that the methanol extract from the bark of Acer nikoense MAXIM. (AN) and the fractions separated from this extract showed the protection for liver injuries induced by carbontetrachloride (CCl4) in rat. On the other hand, the evidence manifested that the fractions of the callus of young branches from AN were recognized to have protective effects on the CCl4-induced liver injuries in rat. In this study, the protective activities of the fractions extracted from the callus of AN were investigated on the alpha-naphthylisothiocyanate (ANIT)-induced liver injuries in rat. The protectively active principles on the ANIT-induced liver injuries were found in the ether soluble fraction. The components of the protective effect were purified by repeated silica gel chromatographies. One of the protective substances in this fraction was identified to be beta-sitosterol. The protective fractions on the ANIT-induced liver injuries showed the protective effects on ANIT-induced cholestasis, but did not show cholagogic effects.

1-Naphthylisothiocyanate