Biomedical subjects
Publications and source records attributed to S Watanabe.
A urea broth microdilution method to assay the susceptibilities of seven Malassezia species was developed. This method indicated the same sensitivities as the agar plate dilution method for isolates of Malassezia furfur, M. pachydermatis, M. slooffiae, and M. sympodialis.
The nucleotide sequences of CAP59 genes from five serotypes of Cryptococcus neoformans were analyzed for their phylogenetic relationships. Approximately 600-bp genomic DNA fragments of the CAP59 gene were amplified from each isolate by PCR and sequenced. The CAP59 nucleotide sequences of C. neoformans showed more than 90% similarity among the five serotypes. By phylogenetic analysis, their sequences were divided into three clusters: serotypes A and AD, serotypes B and C, and serotype D. In addition, the results of reduced amino acid sequences were similar to the nucleotide sequence data. These data revealed that serotype AD was genetically close to serotype A rather than serotype D, although it had been considered to be a mixed type of serotype A and D by serological analysis. Furthermore, the nucleotide sequences of the serotype B and C isolates of C. neoformans were very similar to each other. These results indicated that serotype B and C isolates belonging to C. neoformans var. gattii were genetically homogeneous and closely related. The molecular analysis of the CAP59 gene will provide useful information for the differentiation of serotypes of C. neoformans and for an understanding of their phylogenetic relationships.
Ebola virus contains a single glycoprotein (GP) that is responsible for receptor binding and membrane fusion and is proteolytically cleaved into disulfide-linked GP1 and GP2 subunits. The GP2 subunit possesses a coiled-coil motif, which plays an important role in the oligomerization and fusion activity of other viral GPs. To determine the functional significance of the coiled-coil motif of GP2, we examined the effects of peptides corresponding to the coiled-coil motif of GP2 on the infectivity of a mutant vesicular stomatitis virus (lacking the receptor-binding/fusion protein) pseudotyped with the Ebola virus GP. A peptide corresponding to the C-terminal helix reduced the infectivity of the pseudotyped virus. We next introduced alanine substitutions into hydrophobic residues in the coiled-coil motif to identify residues important for GP function. None of the substitutions affected GP oligomerization, but some mutations, two in the N-terminal helix and all in the C-terminal helix, reduced the ability of GP to confer infectivity to the mutant vesicular stomatitis virus without affecting the transport of GP to the cell surface, its incorporation into virions, and the production of virus particles. These results indicate that the coiled-coil motif of GP2 plays an important role in facilitating the entry of Ebola virus into host cells and that peptides corresponding to this region could act as efficient antiviral agents.
BACKGROUND: Measles virus is implicated in the aetiology of Crohn's disease. This measles hypothesis is mainly supported by immunohistochemical findings that the measles related antigen is present in the intestine of patients with Crohn's disease. Recently we isolated this antigen from the intestine of a patient with Crohn's disease using a molecular cloning technique and produced the monoclonal antibody against it (designated 4F12). AIM: To discover whether the measles related antigen is uniquely present in Crohn's disease. SUBJECTS/METHODS: Colonic mucosa samples from 20 patients with Crohn's disease, 20 with ulcerative colitis, 11 with non-inflammatory bowel disease (IBD) colitis, and nine controls were immunohistochemically stained with the anti-measles monoclonal antibody 4F12. The numbers of positive cells, the ratio of positive cells to nucleated cells, and the staining intensity of the positive cells were compared. Furthermore, the distribution of the measles antigen in other human organs was examined. RESULTS: Both the number of positive cells and the ratio of positive cells to nucleated cells were significantly increased in Crohn's disease, ulcerative colitis, and non-IBD colitis compared with controls (p<0.05) but were similar among the three disease groups. The staining intensity of the positive cells was also similar among the three disease groups. Small numbers of positive cells were observed in the oesophagus, stomach, duodenum, jejunum, and lung. CONCLUSIONS: The presence of the measles related antigen in the colonic mucosa was not unique to Crohn's disease. These results, together with the observation that such a measles related antigen was derived from host protein, do not support the hypothesis that measles virus causes Crohn's disease.
With electrophysiological techniques, we found phase-dependent modification of the efficacy of signal transmission in the procerebrum (PC), the oscillatory olfactory center, of the terrestrial mollusk Limax marginatus and elucidated its neuronal mechanism. Previous studies have indicated that about 10(5) PC neurons can be classified into only two types: bursting (B) neurons and nonbursting (NB) neurons, and both types of neurons have ongoing and phase-locked periodic oscillation of their membrane potentials. On olfactory nerve stimulation, excitatory postsynaptic potentials (EPSPs) were evoked with a constant latency in NB neurons, while EPSPs with a variable latency were evoked in B neurons. These findings suggest a monosynaptic connection from the olfactory nerve to NB neurons, but a polysynaptic connection between the olfactory nerve and B neurons. This polysynaptic transmission is most likely mediated by NB neurons because the olfactory nerve makes synaptic connection only with NB neurons in the PC. The latency of the evoked EPSPs in B neurons depended on the phase of the PC oscillatory activity, presumably because of the oscillation of the intervening NB neurons. These results suggest that the efficacy of olfactory nerve-B neuron polysynaptic transmission is regulated by the activity level of the phasically oscillating NB neurons. Thus, the intrinsic oscillation in the PC can serve as a filter for olfactory information conveyed from the olfactory nerve as a train of neuronal spikes. This filtering system may also produce a phase-dependent modification by the olfactory input of the PC oscillation frequency.
Retinal amacrine cells have abundant dendro-dendritic synapses between neighboring amacrine cells. Therefore an amacrine cell has both presynaptic and postsynaptic aspects. To understand these synaptic interactions in the amacrine cell, we recorded from amacrine cells in the goldfish retinal slice preparation with perforated- and whole cell-patch clamp techniques. As the presynaptic element, 19% of the cells recorded (15 of 78 cells) showed spontaneous tetrodotoxin (TTX)-sensitive action potentials. As the postsynaptic element, all amacrine cells (n = 9) were found to have GABA-evoked responses and, under perforated patch clamp, 50 microM GABA hyperpolarized amacrine cells by activating a Cl(-) conductance. Bicuculline-sensitive spontaneous postsynaptic currents, carried by Cl(-), were observed in 82% of the cells (64 of 78 cells). Since the source of GABA in the inner plexiform layer is amacrine cells alone, these events are likely to be inhibitory postsynaptic currents (IPSCs) caused by GABA spontaneously released from neighboring amacrine cells. IPSCs were classified into three groups. Large amplitude IPSCs were suppressed by TTX (1 microM), indicating that presynaptic action potentials triggered GABA release. Medium amplitude IPSCs were also TTX sensitive. Small amplitude IPSCs were TTX insensitive (miniature IPSCs; n = 26). All of spike-induced, medium amplitude, and miniature IPSCs were Ca(2+) dependent and blocked by Co(2+). Blocking of glutamatergic inputs by DL-2-amino-phosphonoheptanoate (AP7; 30 microM) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 2 microM) had almost no effect on spontaneous GABA release from presynaptic amacrine cells. We suggest that these dendro-dendrotic inhibitory networks contribute to receptive field spatiotemporal properties.
We reported previously that stimulation of glycoprotein 130 (gp130) by a combination of human IL-6 and soluble IL-6 receptor (sIL-6R) could support proliferation, differentiation, and terminal maturation of erythroid cells in the absence of erythropoietin (EPO) from human CD34(+) cells in culture with stem cell factor (SCF). This observation suggested that differentiation of hematopoietic stem/progenitor cells to erythroid cells progressed according to an intrinsic program and that EPO receptor (EPOR) could be replaced by other cytokine receptors. In other words, EPOR appeared to be dispensable for erythropoiesis. Here we examined the role of EPOR in erythropoiesis stimulated by SCF, sIL-6R, and IL-6. Surprisingly, reduction of EPOR expression using antisense oligodeoxynucleotides suppressed erythropoiesis stimulated not only by SCF and EPO, but also by SCF, sIL-6R, and IL-6. EPO mRNA was detected in erythroid cells but not myeloid cells cultured in the presence of SCF, sIL-6R, and IL-6. Furthermore, high concentrations of anti-EPO-neutralizing antibody abrogated erythropoiesis in cultures without exogenous EPO. Based on these results, we suggest that erythroid progenitors themselves secrete EPO and that they have the potential to differentiate and mature in response to this endogenous EPO.
We tested the hypothesis that haemodynamic changes to intubation and postoperative pharyngolaryngeal morbidity are similar for blind intubating laryngeal mask (ILM)-guided compared with laryngoscope-guided tracheal intubation in adults with normal airways. We also compared intubation success rates and airway complications. One-hundred and fifty paralysed, anaesthetized adult patients undergoing elective surgery were randomly assigned to one of three equal-sized groups: 1. blind intubation via the ILM using a straight, silicone tube; 2. intubation with a Macintosh laryngoscope using a straight silicone tube and 3. intubation with a Macintosh laryngoscope using a polyvinyl chloride tube (controls). A standard sequence of adjusting manoeuvres was followed if intubation was difficult. The number of adjusting manoeuvres and intubation attempts, time to intubation, intubation success rate (first attempt and within 3 min), haemodynamic changes (pre-induction, post-induction, post-intubation), oesophageal intubation, mucosal trauma (blood detected), hypoxia (SpO2 < 95%) and postoperative pharyngolaryngeal morbidity (double-blinded) were documented. Time to successful intubation was longer (57 vs 35 s), and more intubation attempts were required in the ILM group (P < 0.0001). The intubation success rate was 100% (all first attempt) for the laryngoscope groups and 94% (56% first attempt) for the ILM group. There were no significant differences in heart rate or blood pressure among groups. Oesophageal intubation (26 v 0%) and mucosal trauma (19 v 2%) were more common in the ILM group. Hypoxia and postoperative pharyngolaryngeal morbidity were similar among groups. Blind intubation through the ILM offers no advantages over the Macintosh laryngoscope for adult patients requiring intubation for elective surgery with normal airways, but it is a feasible alternative.
To obtain information on the effects of Ca2+ on the membrane properties of the lipid A analogue E5531, we have determined the aggregate size, zeta potential, membrane fluidity, micropolarity and permeability of the E5531 membrane as a function of Ca2+ levels. Within the molar ratios of [Ca2+]/[E5531] = 1 and 3, Ca2+ increased the zeta potential of the E5531 membrane but had no effect on aggregate size (approximately 20 nm). Within the above ratios, Ca2+ decreased the membrane fluidity, as measured by micropolarity of E5531 and increased the phase transition temperature. The pharmacokinetics in rats for these samples with different membrane fluidity, prepared by changing the pre-dose formulation concentration of Ca2+, was determined and a correlation between membrane fluidity and pharmacokinetics was clearly observed. It thus appears that Ca2+ effects the membrane fluidity of E5531 as well as its pharmacokinetics in rats.
A 58-year-old male patient had apical hypertrophic cardiomyopathy (HCM) associated with a life-threatening tachycardia due to atrial flutter. Following palpitation and dyspnea for 2-3 h, he became unconscious because of circulatory catastrophe, but was fully resuscitated. An electrocardiogram recorded just before the loss of consciousness revealed atrial flutter at a rate of 260 beats/min with a 2:1 ventricular response. He was diagnosed as having apical HCM based on the echocardiographic and left ventriculographic findings. Atrial stimulation at a rate of 150 pacings/min for 1 min caused a marked drop in systemic systolic blood pressure from 170 to 120 mmHg. The patient was treated with 150 mg of cibenzoline per day to prevent supraventricular tachyarrhythmias and to improve left ventricular diastolic function. At the time of the recent follow-up at 2 and a half years, he felt quite well.
The aim of the present investigation was to compare the hemodynamic and thermal responses to a 30-min aerobic exercise between middle- or old-aged patients with normal left ventricular function and those with left ventricular dysfunction. Constant-load sitting ergometer exercise of approximately 90% of the subject's oxygen uptake (VO2) at the anaerobic threshold for 30 min was conducted in 21 patients with left ventricular dysfunction (61+/-10 years, left ventricular ejection fraction (LVEF) 35+/-7%) and 24 patients with normal left ventricular function (59+/-9 years, LVEF 71+/-7%). Heart rate (HR), blood pressure, deep temperatures in the forehead and thigh, and forearm skin blood flow (SkBF) were measured every minute, and cardiac output (CO) and stroke volume (SV) were determined every 10 min with the dye-dilution technique during the exercise. Patients of both groups exhibited a progressive elevation in each temperature and an increase in SkBF during the exercise. Although the VO2 and CO remained stable, almost the same magnitude of decrease in SV as increase in HR was seen after the 10th min of exercise in both groups. The magnitude of the decrease in SV was greater in old-aged than middle-aged patients with left ventricular dysfunction. Thus, the downward drift in SV during a 30-min constant-load aerobic exercise might not be influenced by left ventricular function, but intensified by aging in patients with left ventricular dysfunction.
The present study attempted to detect the viability of myocardium by quantitative automatic 3-dimensional analysis of the improvement of regional wall motion using an magnetic resonance imaging (MRI) tagging method. Twenty-two subjects with ischemic heart disease who had abnormal wall motion on echocardiography at rest were enrolled. All patients underwent dobutamine stress echocardiography (DSE), coronary arteriography and left ventriculography. The results were compared with those of 7 normal volunteers. MRI studies were done with myocardial tagging using the spatial modulation of magnetization technique. Automatic tracing with an original program was performed, and wall motion was compared before and during dobutamine infusion. The evaluation of myocardial viability with MRI and echocardiography had similar results in 19 (86.4%) of the 22 patients; 20 were studied by positron emission tomography or thallium-201 single photon emission computed tomography for myocardial viability, or studied for improvement of wall motion following coronary intervention. The sensitivity of dobutamine stress MRI (DSMRI) with tagging was 75.9% whereas that of DSE was 65.5%. The specificity of DSMRI was 85.7% (6/7) and that of DSE was 100% (7/7). The accuracy of DSMRI was 77.8% (28/36) and that of DSE 72.2% (26/36). DSMRI was shown to be superior to DSE in terms of evaluation of myocardial viability.
Three-dimensional images of the coronary arteries using electron-beam computed tomography (EBCT) data with shaded surface rendering makes it possible to achieve images easily with a short reconstruction time. However, a lower threshold is required to estimate vessel diameters and there is a quantitative problem compared with conventional coronary arteriography. In combination with volume rendering, EBCT may be useful to detect the normal coronary artery wall, the major components of the atherosclerotic plaque (lipid, fibrous connective tissue and calcium). EBCT scans offer a new, non-invasive alternative to conventional coronary arteriography for diagnosis of coronary artery disease.
The present study investigated the relationship between aortic atherosclerosis and carotid atherosclerosis, and studied the effects of coronary risk factors for these arteries. The subjects consisted of 78 patients with coronary artery disease (CAD) and 69 patients without CAD. All subjects underwent enhanced computed tomography and B-mode ultrasonography within a short time period to determine the extent of aorta and carotid atherosclerosis. Significant correlations between maximal aortic wall thickness (MAWT) and aortic wall volume (AWV) with carotid intima-media thickness (IMT) were demonstrated. MAWT, AWV and IMT were significantly higher in patients with CAD compared with controls (p=0.009, p=0.024, p=0.001, respectively). Furthermore, there were significant differences in MAWT, AWV and IMT among groups classified by the number of coronary artery stenoses, and no significant differences among groups classified by risk factors, but it was shown that MAWT, AWV and IMT increased gradually as the risk factors increased in number. MAWT, AWV and IMT had positive correlations with age, systolic blood pressure and triglyceride, and a negative correlation with high density lipoprotein-cholesterol. This study demonstrated that both aortic atherosclerosis and carotid atherosclerosis are closely correlated with coronary atherosclerosis, and that the atherosclerosis indices are independently associated with age and hyperlipidemia.
In 21 patients with chronic aortic dissections and proven patent false lumens, the flow volume and flow patterns in the patent false lumens was evaluated using velocity-encoded cine magnetic resonance imaging (VENC-MRI) and the relationship between the flow characteristics and aortic enlargement was retrospectively examined. Flow patterns in the false lumen were divided into 3 groups: pattern A with primarily antegrade flow (n=6), pattern R with primarily retrograde flow (n=3), and pattern B with bidirectional flow (n=12). In group A, the rate of flow volume in the false lumen compared to the total flow volume in true and false lumens (%TFV) and the average rate of enlargement of the maximum diameter of the dissected aorta per year (deltaD) were significantly greater than in groups R and B (%TFV: 74.1+/-0.07 vs 15.2+/-0.03 vs 11.8+/-0.04, p<0.01; deltaD: 3.62+/-0.82 vs 0 vs 0.58+/-0.15 mm/year, p<0.05, respectively). There was a significant correlation between %TFV and deltaD (r=0.79, p<0.0001). Evaluation of flow volume and flow patterns in the patent false lumen using VENC-MRI may be useful for predicting enlargement of the dissected aorta.
The study examined the association between aortic wall volume (AWV) detected by enhanced computed tomography and coronary artery atherosclerosis observed on angiography. In 180 cases, AWV was measured as the total wall volume of a 7-cm portion of the descending thoracic aorta distal from the tracheal bifurcation. Coronary artery atherosclerosis was angiographically quantified by both Gensini score, in terms of the severity of coronary artery stenosis, and Extent score, in terms of the severity of coronary artery involvement. Mean AWV values between the patients with significant coronary artery stenosis and those without significant stenosis were 9.83+/-4.04 cm3 and 8.09+/-2.39 cm3, respectively (p<0.001). AWV was a significantly independent variable for significant coronary artery disease (p=0.0097) and an Extent score > or = 60 (p=0.0092). Calcification of AWV, however, was not associated with coronary atherosclerosis. The quantification of aortic atherosclerosis was useful for diagnosing coronary artery disease.
The term for re-entrainment to a new light-dark cycle in streptozotocin (STZ)-induced diabetic rats was significantly longer than that in control rats. In STZ-induced diabetic rats, the same level of phase delay in the suprachiasmatic nucleus neuronal firing rhythm was observed in control rats after glutamate application. Therefore, 5-HT function in the hypothalamus is thought to be decreased in STZ-induced diabetic rats. These results suggest that postsynaptic neuronal function is still maintained in the suprachiasmatic nucleus of STZ-induced diabetic rats. Therefore, 5-HT mechanisms may play an important role in the maintenance of this function.
Adenosine was intraperitoneally (i.p.) injected to Wistar rats every 3 days with a total of 5 administrations. After a 7-day withdrawal, the animals were challenged with methamphetamine (0.5 mg/kg, i.p.). The effect of methamphetamine on locomotor activity was significantly potentiated by repeated adenosine pretreatment. Moreover, methamphetamine-induced dopamine release was also increased in the striatum. Methamphetamine-induced hyperactivity and dopamine release were significantly potentiated by repeated pretreatment of an adenosine A1 agonist, N6-cyclohexyladenosine (0.5 mg/kg, i.p.). These results suggest that the acute effect of methamphetamine is potentiated by repeated pre-treatment of adenosine via adenosine A1 receptors.