Biomedical subjects
E Tanaka
Publications and source records attributed to E Tanaka.
Inhibition of in vivo tumor growth by the beta chemokine, TCA3.
TCA3 is a proinflammatory murine glycoprotein that shares structural features with cytokines of the beta chemokine family and is a chemoattractant for neutrophils and monocytes. To assess the in vivo functions of TCA3, the cDNA was expressed in two mouse myeloma cell lines. Although the transfected and control cells had similar growth rates in vitro, TCA3-expressing tumors demonstrated impaired growth in both normal and immunodeficient mice. Histologic evaluation of the injection sites demonstrated that TCA3 expression resulted in an early neutrophil and monocyte infiltrate accompanied by tumor necrosis. There was complete regression of the TCA3-transfected tumor in some immunocompetent syngeneic mice. The TCA3-transfected cells induced specific and long-lasting immunity in mice that showed complete tumor regression; these animals were resistant to challenge with nontransfected tumor cells. In contrast, priming with irradiated tumor cells provided little protection against challenge with nontransfected tumor, which indicates that TCA3 specifically augments tumor immunogenicity. Mixing TCA3-transfected cells with normal tumor cells causes retarded growth of the normal tumor cells provided the latter are injected into the same site. Furthermore, direct in situ injection of soluble rTCA3 early during the course of tumor implantation also inhibits tumor growth. The data suggest that TCA3 may perform two roles in tumor protection: it induces lymphocyte-independent antitumor activity and stimulates tumor-specific immunity. We speculate that TCA3 has natural adjuvant activities that result in augmented immune responses.
Induction of cytochrome P450 isozymes in rat renal microsomes by cyclosporin A.
To examine the effects of cyclosporin A (CsA) on renal cytochrome P450 forms, CsA was administered to rats, and the renal levels of P450 were determined by immunoblotting. CsA treatment for 17 days increased total renal P450 content by 40% with a concomitant elevation of the omega- and (omega-1)-hydroxylation activities of lauric acid. Arachidonic acid omega-hydroxylation activity was also induced 2-fold by treatment with CsA for 17 days. Among the P450 forms, CYP4A2 was induced significantly, whereas CYP2C23, CYP4A1 and CYP4A8 were unaffected. These changes were accompanied by slight but significant increases in blood urea nitrogen and systolic blood pressure. These data suggest that CsA increased arachidonic acid omega-hydroxylation activity by the induction of CYP4A2. The specific induction of CYP4A2 may be related to CsA-induced nephrotoxicity and elevated blood pressure, because omega-hydroxyarachidonic acid is a potent vasoconstrictor.
Trimethadione metabolism, a useful indicator for assessing hepatic drug-oxidizing capacity.
The metabolism of trimethadione (TMO), a useful indicator of hepatic drug-oxidizing capacity in rats and humans, was studied using 14 different forms of rat cytochrome P450 (CYP1A1, 1A2, 2A1, 2A2, 2B1, 2B2, 2C6, 2C7, 2C11, 2C12, 2C13, 2E1, 3A2 and 4A2) and three forms of human cytochrome P450 (CYP1A2, 2C and 3A4). TMO N-demethylation was increased by treating rats with phenobarbital. CYP2C11 and 2B1 had high TMO N-demethylase activity, but 1A1 and 1A2 had low activity. Antibodies raised to CYP2C11 and 2B1/2 inhibited TMO N-demethylation in hepatic microsomes of untreated and phenobarbital-treated rats, respectively. In a reconstituted system, human CYP3A4 and 2C produced efficiently dimethadione (DMO), but CYP1A2 did not catalyse TMO N-demethylation. Antibodies raised to CYP3A2 and 2C11 inhibited TMO N-demethylation in human hepatic microsomes. These results indicated that the N-demethylation of TMO is catalysed mainly by CYP2C11 and 2B1 in rat hepatic microsomes, and that human CYP3A4 and an unspecified isoform of the 2C subfamilies contribute to TMO N-demethylation in human liver.
Changes in antibody titers to hepatitis C virus following interferon therapy for chronic infection.
The use of quantitative assays for hepatitis C virus specific antibodies (anti-HCV) as a prognostic marker was evaluated in 31 patients with chronic hepatitis C treated with interferon (IFN). Changes in titers of serum HCV-RNA and anti-HCV antibodies; anti-C11 (anti-core), anti-C100 (anti-NS3), and anti-C7 (anti-NS3) were investigated. Recombinant IFN-alpha 2a was administered and the patients were followed for more than 1 year. The patients were classified into three groups according to their responses to IFN: 11 sustained responders with continuous normalizations of serum alanine aminotransferase (ALT) levels; 14 transient responders with transient decreases in ALT; and six nonresponders who had no changes in ALT levels. Ten of 11 sustained responders had a continuous decrease in anti-C11 titers after completion of treatment, decreasing to less than half of pretreatment titers. No patients in the other two groups had a continuous decrease in anti-C11 titers. Although sustained responders had decreases in anti-C100 and anti-C7 titers after IFN therapy, these titers also decreased in some patients in the other two groups. HCV-RNA was not detected in the sera of 10 of 11 sustained responders following IFN therapy. In contrast, while 9 of 10 transient and non-responders had a decrease or disappearance of HCV-RNA at the completion of therapy, they had increased levels thereafter. These results indicate that anti-HCV-core (anti-C11) titers most closely reflect the status of HCV replication. A quantitative assay for anti-HCV-core antibody can be used as a predictive marker of remission in IFN-treated patients with chronic hepatitis C.
Immunoperoxidase staining of hepatitis C virus in formalin-fixed, paraffin-embedded needle liver biopsies.
The localization of hepatitis C virus (HCV) in the liver has not been well clarified. We report successful indirect immunoperoxidase staining of the HCV core antigen using polyclonal antibodies raised in rabbits and conventional formalin-fixed, paraffin-embedded needle biopsy sections of liver. The core antigen was distributed in a fine granular pattern diffusely, perisinusoidally, or focally within the hepatocellular cytoplasm of livers from patients with HCV infection. The staining tended to show a more heterogeneous pattern in terms of intensity and distribution in cases of more advanced disease. Hepatocellular carcinoma cells were also frequently stained. HCV immunostaining will provide important information on the pathogenesis and treatment of HCV-related liver diseases.
Viral and host factors that contribute to efficacy of interferon-alpha 2a therapy in patients with chronic hepatitis C.
Using conventional statistical analysis and multiple regression analysis, we investigated the viral and host factors that influence the response to recombinant interferon-alpha 2a therapy in patients with chronic hepatitis C. A total of 36 patients was randomly assigned to three administration schedules, 12 patients in each. Response to treatment was set as the criterion variable. Four variables were statistically significant in the conventional method in predicting a good therapeutic outcome: HCV genotype III and IV, lower histology activity index (HAI) score for liver, higher total dose of interferon administration, and lower serum HCV RNA concentration. In multiple regression analysis, a combination of the above four variables resulted in a higher multiple correlation coefficient (R = 0.84, P < 0.0001) using a stepwise method. Of those four, HCV genotype had the highest absolute value of standard partial regression coefficient (0.51). The HCV RNA concentration was correlated with HCV genotype and HAI score, whereas HCV genotype and HAI score showed no correlation. Thus, HCV RNA concentration was not statistically significant in multiple regression analysis. These findings indicate that HCV genotype, HAI score, and schedule of administration can be important predictors of the response to interferon therapy.
Adaptation of hepatitis C virus for persistent infection in patients with acute hepatitis.
Nucleotide sequencing was used to analyze amino acid substitutions in the putative envelope 1 (E1) and envelope 2/nonstructural 1 (E2/NS1) regions of hepatitis C virus (HCV) to clarify a viral mechanism of persistent infection in three patients with acute hepatitis C who developed chronic hepatitis and two patients with chronic hepatitis. The HCV RNA titer in serum decreased markedly after the onset of acute hepatitis and then re-elevated. During this period, the substitution rate in the E2/NS1 region (especially in the hypervariable region located in the N-terminus) was significantly higher in patients with acute hepatitis than in patients with chronic hepatitis (P < 0.05). When patients with acute hepatitis C became persistent HCV carriers, the substitution rate decreased to the level seen in patients with chronic hepatitis. The amino acid substitution rate in the E1 region in the acute phase was similar to that found in the chronic HCV carrier state. These observations suggest that rapid substitution of the amino acid sequence in the hypervariable region of the E2/NS1 region may be one of the mechanisms of persistent HCV infection.
Transmission of hepatitis C in an isolated area in Japan: community-acquired infection. The South Kiso Hepatitis Study Group.
BACKGROUND/AIMS: The spread of hepatitis C virus (HCV) infection not due to drug needle sharing or transfusion is largely unknown in communities. A search for risk factors for HCV infection in an endemic area might elucidate inapparent modes of transmission. METHODS: We conducted screening for hepatitis virus markers and parenteral exposures to blood among 435 inhabitants in an isolated area known for its endemicity for non-A, non-B hepatitis and in a nonendemic area with 1542 inhabitants. RESULTS: The prevalence of hepatitis B surface antigen was the same in both areas. The prevalence of antibody to HCV verified by the recombinant immunoblot assay was 32.4% in the highly endemic area and 2.3% in the nonendemic area (P < 0.001). Risk factors for HCV infection in the highly endemic area were complex but included folk remedies such as acupuncture and "vacuuming" for congested blood in muscle by the use of a warm glass bottle. CONCLUSIONS: Folk remedies such as acupuncture and cutting of the skin using nonsterilized knives should be considered as possible routes of HCV transmission not associated with blood transfusion or sharing of drug paraphernalia.
A three-dimensional finite element model of the mandible including the TMJ and its application to stress analysis in the TMJ during clenching.
The purpose of this study was to develop a three-dimensional finite element model of the mandible including the temporomandibular joint (TMJ) and further to investigate stress distributions in the TMJ during clenching. The model consisted of 2088 nodes and 1105 elements, comprising cortical and cancellous bones, articular disc, articular cartilage layer and periodontal ligament. For loading condition, a resultant force of 500 N derived from the cross-sectional areas of the muscles was applied to the model. Compressive stresses were induced in the anterior, middle and lateral areas of the TMJ during clenching, whereas the tensile stresses were induced in the posterior and medial areas. The mean principal stresses on the surface of condyle were -1.642, -0.543, 0.664, -1.017, 0.521 MPa in the anterior, middle, posterior, lateral and medial areas, respectively. The mean stresses on the surface of glenoid fossa were approximately 1/5 to 4/5 those on the surface of condyle, although the patterns of stress distributions were almost similar. In both the articular disc and cartilage layer, the stress distributions were very approximate in qualitative and quantitative aspects. Thus, it is shown that stresses during clenching vary substantially in different structures and areas of the TMJ and that the stresses are reduced by the articular disc and cartilage layer in comparison with actual biting forces exerted by the masticatory muscles, if anatomic relation between various TMJ components is well maintained.
Simulation studies on a pseudo-three-dimensional reconstruction algorithm for volume imaging in multi-ring PET.
The imaging performance of a pseudo-three-dimensional (3D) reconstruction algorithm has been investigated. The algorithm is based on ID filtering followed by 3D backprojection. High- and low-frequency components of the image are reconstructed separately, and oblique projections are used only for reconstructing the high-frequency component. The cross talk between different slices due to oblique backprojection is eliminated by high-pass filtering. 'Variable filters', the cut-off frequencies of which are proportional to the tilt angles of projections, provide higher signal-to-noise ratio than 'fixed filters' with similar capability for cross-talk rejection. The maximum ring difference used in the reconstruction of each slice is extended as far as possible, while still satisfying the requirement that the total density of backprojection beams deposited in the slice is approximately uniform in the field of view. This allows the use of almost all available projection data to improve signal-to-noise ratio in peripheral slices. The root mean square noise of the reconstructed images evaluated with simulation studies is in good agreement with numerical calculations, and it is shown that the loss in signal-to-noise ratio caused by discarding the low-frequency component of oblique projections is negligibly small as long as suitable variable filters are used.
Inhibitory effects of nicorandil on sympathetic coronary vasoconstriction.
OBJECTIVE: The aim was to investigate whether nicorandil suppresses the rise in coronary vascular resistance that occurs during stimulation of the sympathetic nerve supply to the heart and, if so, what are the mechanisms of action. METHODS: The effects of nicorandil on coronary vascular resistance during ventrolateral cardiac nerve stimulation and on the reactivity of the coronary vasculature to intracoronary infusion of noradrenaline or neuropeptide Y (NPY) were examined under beta receptor blockade. The effects of nicorandil on the overflow of noradrenaline and NPY during ansae subclaviae stimulation were compared with those in a control group and in a group treated with glyceryl trinitrate and N omega-nitro-L-arginine (L-NNA) under both alpha and beta receptor blockade with vagotomy. RESULTS: Intracoronary infusion of nicorandil decreased coronary vascular resistance prior to cardiac nerve stimulation, and during stimulation it suppressed the percentage increase in resistance from the prestimulation value. Nicorandil suppressed the reactivity of the coronary vasculature to exogenous noradrenaline and NPY. Intra-atrial infusion of nicorandil significantly reduced the overflow of NPY but not of noradrenaline during stimulation of the ansae subclaviae at 20 Hz. This suppressive effect was not observed in the glyceryl trinitrate + L-NNA group. CONCLUSIONS: Nicorandil reduces sympathetic coronary vasoconstriction by decreasing the reactivity of the vasculature to sympathetic neurotransmitters and by suppressing NPY overflow during cardiac sympathetic nerve stimulation. The suppressive action on NPY overflow is thought to be due to the opening of ATP sensitive potassium channels in the sympathetic nerve endings rather than to a glyceryl trinitrate-like action of nicorandil.
The effect of erythromycin on Pseudomonas aeruginosa and neutrophil mediated epithelial damage.
Erythromycin therapy for long periods may benefit patients with chronic bronchial sepsis colonized by Pseudomonas aeruginosa despite the lack of antibacterial activity. We have investigated the effect of filtrates of 24 h P. aeruginosa cultures (CF) with or without erythromycin 0.5, 5, 20 mg/L on human nasal epithelium in the absence or presence of polymorphonuclear leucocytes (PMN). Ciliary beat frequency (CBF) and epithelium integrity were examined for 4 h. Erythromycin (20 mg/L) alone did not affect epithelium. CF without erythromycin slowed CBF by 63.5% of control at 4 h, and caused disruption of surface integrity in 80% of the epithelium. Addition of erythromycin to CF did not inhibit these effects. Erythromycin did not affect growth of P. aeruginosa. Filtrates of P. aeruginosa cultured with erythromycin (5 and 20 mg/L) caused less CBF slowing (37.2% and 19.2% of control, respectively) and epithelial disruption (4.2% and 6.7%, respectively). Unstimulated PMN (10(7)/mL) slowed CBF by 13% of control at 4 h but did not cause epithelial disruption. PMN and CF together slowed CBF (95.4% of control) and damaged epithelium (93.3% of epithelium disrupted) synergically. Pre-incubation of PMN with erythromycin did not inhibit these effects. PMN and filtrates of P. aeruginosa cultured with erythromycin (5 and 20 mg/L) caused less CBF slowing (58.0% and 33.6% of control, respectively) and epithelial disruption (40.0% and 13.3%, respectively). Erythromycin may benefit patients by reducing P. aeruginosa production of factors which damage epithelium and stimulate neutrophil mediated cytotoxicity.
"Silent" hepatitis B virus mutants are responsible for non-A, non-B, non-C, non-D, non-E hepatitis.
Since the recent introduction of diagnostic kits for hepatitis C and E, some cases of non-A, non-B, non-C, non-D, non-E hepatitis (so-called hepatitis F) have been revealed. We attempted to demonstrate that so-called hepatitis F is caused by hepatitis B virus (HBV) variants. Polymerase chain reaction (PCR) was used to amplify serum HBV DNAs from 20 patients with acute hepatitis and 20 patients with chronic hepatitis who had been diagnosed as having so-called hepatitis F on the basis of conventional serological markers. The PCR technique successfully amplified HBV DNAs in 18 (90%) cases of acute hepatitis and 17 (85%) cases of chronic hepatitis. Sequencing of HBV DNAs of six patients (acute 3, chronic 3) revealed equally a T-to-C mutation of DR2 and an 8-nucleotide deletion of the 3'-terminus of the X gene coding region, giving rise to the generation of a C-terminally truncated X protein and probable damage to the enhancer II/core promoter elements. These mutations of the X gene coding region may lead to suppression of replication and expression of HBV DNAs. Thus virtually all cases of so-called hepatitis F appear to be caused by "silent" HBV mutants, at least in Japan.
Recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) or tumour necrosis factor-alpha (TNF-alpha) activate human alveolar macrophages to inhibit growth of Mycobacterium avium complex.
We investigated the effects of certain macrophage-active cytokines on the phagocytosis and growth inhibition of Mycobacterium avium complex (MAC) by human alveolar macrophages (AM). We also evaluated the effects of pretreatment with each cytokine on the superoxide anion release (O2-) from AM. The cytokines that we used were recombinant GM-CSF, natural type TNF-alpha, recombinant interferon-gamma (IFN-gamma), and recombinant IL-2. We found that phagocytosis by the various cytokine-stimulated AM was similar to that of unstimulated AM. On the other hand, significant growth inhibition of MAC was observed in the macrophages treated with GM-CSF or TNF-alpha, while no growth inhibition of MAC was observed in the macrophages treated with IFN-gamma or IL-2. Pretreatment with all cytokines tested enhanced the O2- release from AM, but there was no correlation between the enhancement of O2- release and the growth inhibition of MAC. Thus, we concluded that GM-CSF or TNF-alpha could activate AM to inhibit growth of MAC, probably not through the enhanced production of reactive oxygen intermediates.
Natural history of hepatitis C.
With advances in tools for the diagnosis of hepatitis C virus (HCV) infection, it has become easier to evaluate the natural course of hepatitis C. Although HCV infection initially occurs in adult individuals, in most patients with acute hepatitis C (68%) it develops into chronic hepatitis. Once chronic hepatitis is established, the rate of spontaneous cure of the liver disease is very rare (below 2%). The duration from the onset of acute hepatitis until the time of diagnosis of cirrhosis of the liver and of hepatocellular carcinoma is about 20 and 30 years, respectively. The long-term clinical course of hepatitis C is divided into the three phases of acute, silent, and reactivated. The acute phase lasts from the onset of disease until 2-3 years thereafter, and the silent phase which follows lasts for 10-15 years. In the silent phase, the serum transferase level remains relatively low, below 100 IU/l, and is sometimes within the normal range. In the reactivated phase, the level of serum aminotransferase increases and remains at a high or moderate level until hepatocellular carcinoma develops. The mechanism of the chronicity of hepatitis C is unknown. However, recent advances in molecular analysis may soon elucidate this. Successive antigenic change of the HCV E2/NS1 hypervariable domain as a result of mutations may represent a mechanism by which this virus escapes the host immunosurveillance system, as well as a mechanism of its chronicity.
Inhibition by dexamethasone of retinoic acid-induced enhancement of leukotriene C4 synthesis in rat basophilic leukemia-1 cells.
We investigated inhibitory actions of dexamethasone (DEX) on retinoic acid (RA)-induced enhancement of leukotriene C4 (LTC4) synthesis in rat basophilic leukemia-1 (RBL-1) cells. Cultured cells were preincubated with RA for 16 h with or without DEX, and generation of LTC4 was measured by high performance liquid chromatography in cell-free and intact cell systems. RA (0.1 microgram/ml) significantly potentiated calcium ionophore-stimulated production of LTC4 synthesis. DEX inhibited the RA-induced enhancement of LTC4 synthesis by up to approximately 95% in intact cells when stimulated with calcium ionophore. RA-induced LTC4 synthase activity, which was determined by enzyme assay, was also inhibited by DEX by 65% in a cell-free system. This discrepancy of inhibition between the intact and cell-free systems was due to a partial inhibition of phospholipase A2 activity by DEX in the intact cells. These results indicate that the production of LTC4 is predominantly regulated at a level of LTC4 synthase. The induction of new LTC4 synthase activity by RA and inhibition of the RA-induced activity by DEX are important regulatory mechanisms of LTC4 synthesis.
Interaction of Pseudomonas aeruginosa with human respiratory mucosa in vitro.
Pseudomonas aeruginosa commonly infects the airways of patients with cystic fibrosis and bronchiectasis. It produces several toxins that slow ciliary beat, stimulate mucus production and damage epithelium. It adheres to epithelial cells, damaged mucosa (in animal models), and mucus. However, little is known of the interaction of P. aeruginosa with intact human respiratory mucosa. We have studied the interactions of a nonmucoid clinical isolate of P. aeruginosa with adenoid tissue in a novel organ culture model with an air-mucosal interphase P. aeruginosa (5.9 +/- 0.9 x 10(6) colony-forming units (cfu)) was pipetted onto the organ culture surface, and incubated for 15 min, 1, 2, 4, 8, 12, 16, and 24 h, at 37 degrees C in 5% CO2 in a humidified atmosphere. Assessment has been made by transmission and scanning electron microscopy. Transmission electron microscopy (TEM) showed that uninfected organ cultures had normal ultrastructure. TEM of infected organ cultures at 8 h showed significant epithelial damage: 43.9 +/- 10% of cells extruding from the epithelial surface, 17.7 +/- 3% of cells with loss of cilia, 32.9 +/- 10.2% of cells with mitochondrial damage, and 11.6 +/- 3% of cells with cytoplasmic blebbing. P. aeruginosa only infrequently adhered to normal epithelium, but adhered to areas of epithelial damage and to basement membrane. Scanning electron microscopy (SEM) of organ cultures up to 2 h found P. aeruginosa only infrequently associated with mucus. SEM at 4 h revealed P. aeruginosa predominantly associated with mucus and extruded damaged epithelial cells, but also occasionally associated with cilia, and very occasionally with unciliated cells.(ABSTRACT TRUNCATED AT 250 WORDS)