Spontaneous remission of epstein-barr virus-positive diffuse large B-cell lymphoma of the elderly.

A 94-year-old female patient presented with anorexia and left axillar lymphadenopathy on admission. Her past history was angina pectoris at 83 years of age and total gastrectomy due to gastric cancer at 87 years. The family history revealed that her son had had a malignant lymphoma, the histopathological diagnosis of which was diffuse large B-cell lymphoma. A physical examination showed both cervical, axillar, and inguinal lymphadenopathy without tenderness. She had elevated lactate dehydrogenase, ferritin, and soluble interleukin-2 receptor (sIL-2R). Whole-body computed tomography confirmed the cervical, axillary, and inguinal lymphadenopathy. Gallium-68 imaging revealed positive accumulation in these superficial lymph nodes. A right inguinal lymph node biopsy showed features of Epstein-Barr virus-associated lymphoproliferative disorder. Immunohistological studies on this lymph node biopsy showed CD20-positive large cells, CD3-positive small cells, and CD30-partly-positive large cells. In situ hybridization showed Epstein-Barr virus-positive, LMP-partly-positive, and EBNA2-negative cells. She refused chemotherapy as her son had died from hematemesis during chemotherapy. She received intravenous hyperalimentation for 1 month after admission. No palpable lymph nodes were identified by physical examination or computed tomography 3 months after admission, and regression of lactate dehydrogenase, ferritin, and sIL-2R was observed. She recovered from anorexia and was discharged. She died from pneumonia 10 months later after initial symptoms of anorexia. The autopsy showed no superficial lymphadenopathy.

A novel predictive method for assessing the quality of isolated pancreatic islets using scanning electrochemical microscopy.

INTRODUCTION: The current methods for evaluating islet potency are not useful in clinical transplantation. Therefore, we need reliable, rapid methods enabling accurate prediction of islet quality. MATERIALS AND METHODS: We evaluated respiratory activity using scanning electrochemical microscopy (SECM), glucose-stimulated respiratory activity, glucose-stimulated insulin release, ADP/ATP assays, insulin/DNA levels, and Trypan blue exclusion tests as predictive methods for the ability of isolated rat islets to cure syngeneic diabetic rats. RESULTS: Although glucose-stimulated respiratory activity, basal respiratory activity, ADP/ATP ratio, and glucose-stimulated insulin release were significantly correlated with the outcome of transplantation into diabetic rats, there was no correlation between outcomes, insulin/DNA ratios, and Trypan blue exclusion tests. The glucose-stimulated respiratory activity in islet preparations that could cure diabetic rats was significantly greater than those unable to cure diabetes. Rat islets with >1.5-fold glucose-stimulated respiratory activity consistently cured diabetic rats, whereas those with a value <1.5 hardly cured any rats. CONCLUSION: Measurement of the glucose-stimulated respiratory activity using SECM technique is a novel method that may be useful as a rapid, potent predictor of the outcome of clinical islet transplantation.

A microbial chip combined with scanning electrochemical microscopy.

A microbial chip for bioassay was fabricated and its performance was characterized by scanning electrochemical microscopy (SECM). The microbial chip was prepared by spotting a suspension of Escherichia coli on a polystyrene substrate by using a glass capillary pen. The respiration activity of the E. coli spot was imaged with SECM by mapping the oxygen concentration around the spot. The SECM images of the microbial chips clearly showed spots with lower reduction currents, indicating that E. coli in the spots uptake oxygen by respiration. The bactericidal effects of antibiotics (streptomycin and ampicillin) were measured using the E. coli-based microbial chip, and discussed in comparison with the minimum inhibitory concentration (MIC) determined by an agar plate dilution method.

Mapping peroxidase in plant tissues by scanning electrochemical microscopy.

Scanning electrochemical microscopy has been firstly used to map the enzymatic activity in natural plant tissues. The peroxidase (POD) was maintained in its original state in the celery (Apium graveolens L.) tissues and electrochemically visualized under its native environment. Ferrocenemethanol (FMA) was selected as a mediator to probe the POD in celery tissues based on the fact that POD catalyzed the oxidation of FMA by H(2)O(2) to increase FMA(+) concentration. Two-dimensional reduction current profiles for FMA(+) produced images indicating the distribution and activity of the POD at the surface of the celery tissues. These images showed that the POD was widely distributed in the celery tissues, and larger amounts were found in some special regions such as the center of celery stem and around some vascular bundles.

Oxygen consumption of single bovine embryos probed by scanning electrochemical microscopy.

Oxygen consumption of individual bovine embryos was noninvasively quantified by scanning electrochemical microscopy (SECM). A probe microelectrode was used to scan near a single embryo surface in a culture medium to monitor the oxygen reduction current at 37 degrees C, under a water-saturated atmosphere of 5% CO2 and 95% air. The oxygen concentration profiles near the embryos were in good agreement with the theoretical spherical diffusion. When an embryo reached the stage of a morula with a 74-microm radius on day 6 after in vitro fertilization, the oxygen concentration difference (deltaC) between the bulk solution and the morula surface was 6.90 +/- 1.35 microM. The oxygen consumption rate (F) of the single morula was estimated to be (1.40 +/- 0.27) x 10(-14) mol s(-1). After the SECM measurement, the embryo was continuously cultured for another 2 days and grew to the stage of a blastocyst with a 100-microm radius. For the blastocyst, the deltaC values for the inner cell mass side and the trophoblast side were 16.40 +/- 1.83 and 9.14 +/- 1.68 microM, respectively. The oxygen consumption rate of the blastocyst was found to be in the range of (2.50 +/- 0.46) x 10(-14) mol s(-1) < F < (4.49 +/- 0.50) x 10(-14) mol s(-1). We have carried out SECM measurements for 19 embryos, and the results were compared in detail with these from an optical microscopic observation. The deltaC values for the morulae on day 6 after in vitro fertilization were strongly related to the morphological embryo quality. The morulae showing a larger deltaC value developed into blastocysts of a larger size, and the deltaC value after the subsequent 2 days of cultivation was found to be increased.

Dielectrophoretic manipulation of a single chlorella cell with dual-microdisk electrode.

Dielectrophoretic manipulation of a single chlorella cell was performed using a dual-microdisk electrode, which consists of two Pt-Rh ultrafine wires (ca. 1-microm radius) sealed in a glass capillary. An attractive or repulsive force was induced on the chlorella depending on the frequency of the ac voltage applied between the two disk electrodes. To avoid the direct contact of a chlorella with the metal, a dual electrode with retracted disks was fabricated and used for forming a micropattern of chlorellas at a solid substrate. The effect of both the frequency and ion concentration of the solutions on the dielectrophoretic force exerted on a chlorella cell was investigated in detail based on the theories of dielectrophoresis.

[Epidemiological survey for hemolytic streptococci isolated from children in Tokyo].

During the 20-year period between 1979-1998, a total of 4,176 strains of hemolytic streptococci have been isolated from 20,118 healthy primary school children and little children in Tokyo Metropolitan (Tokubetsuku, Tama and Tosho). Culture of throat swabs every November and the following February during the 20-year period were made and serological grouping and typing for isolates were done by T agglutination method. The results were as follows. 1) Serological group of hemolytic streptococci isolated from children were 3,188 strains (76.3%) for isolates of group A out of total strains of 4,176, 569 strains (13.6%) for isolates group B, 63 strains (1.5%) for isolates of group C and 356 strains (8.5%) for isolates of group G. 2) The most dominant was T12 during 1979-1998, and other relatively frequent serotypes were T28, T1, T4, T6 in that order. These ranks of and the main epidemic serotypes showed a similar trend in the 3 areas. 3) The isolation rates of group A streptococci were 15.9% in Tokubetsuku, 17.1% in Tama and 14.9% in Tosho. The average of 3 areas were 15.8%. 4) The epidemic cases seemed to be caused by group A streptococci were 20 cases, their isolated serotype were 7 cases by T28, 5 cases by T12, 4 cases by T6, 2 cases by T4, each 1 case by T1 and T25. 5) A total of 2,927 strains of group A streptococci were examined for drug sensitivity. All strains were sensitive to beta-lactam group of antibiotics (benzylpenicillin and cephaloridine). Resistant (MIC > or = 25 micrograms/ml) to TC, CP and EM etc. were 740 strains (25.3%) in this study. The incidence of resistant strains were to TC 493 strains (66.6%) out of 740 strains, 81 strains (10.9%) for TC.CP, 72 strains (9.7%) for EM and 66 strains (8.9%) for TC.CP.EM.OL.LCM. TC resistant strains have not varied much through the whole period, but CP and EM resistant strains were very variable by year. Many resistant strains to TC were T4, to EM and multiple drug resistant were T12. 6) The rates of isolates of the same type of group A streptococci in school child individual during for the tests taken twice a year were 12.3%, indicating group A streptococci, according to the duration of the carrier state, seems to be a short period.

Real-time multisite observation of glutamate release in rat hippocampal slices.

A multichannel glutamate sensor was fabricated that consists of enzyme modified electrodes and has a high sensitivity and selectivity to glutamate. We placed a rat hippocampal slice on the sensor and monitored the current at four electrodes resulting from the stimulation with muscimol, a gamma-aminobutyric acid(A) (GABA(A)) receptor agonist. We obtained different glutamate concentration increases at the different positions, suppressed by bicuculline, a GABA(A) receptor antagonist. This demonstrated that the sensor can monitor the glutamate released via GABA(A) receptors pathways, and the difference in the concentrations may indicate differences in the distribution of GABA(A) receptor as well as diverse receptor functions. This multichannel sensor may be useful for non-invasive, real-time monitoring of glutamate distribution, which would make it a valuable tool for pharmacological analysis.

Reduced-dose chop therapy for elderly patients with non-Hodgkin's lymphoma.

While CHOP therapy is effective for malignant lymphoma, the optimum schedule for elderly patients remains controversial. The present study investigated the usefulness of reduced-dose CHOP therapy for elderly patients. Previously untreated patients aged 65 years or older with intermediate to high-grade non-Hodgkin's lymphoma were given up to 6 courses of reduced-dose CHOP therapy at 3-week intervals. Group A patients were given (5/6) of the standard dose and Group B received 7/12 of the standard dose. Filgrastim was administered when the white blood cell count fell below 2,000/microL. Fifty-seven patients were evaluable and the scheduled therapy was completed in 37. For patients aged from 65 to 79 years and for patients older than 80 years, the complete response rate was 79.5% and 46.2%, overall 3-year survival was 58.2% and 30.4%, and event-free 3-year survival was 49.3% and 44.4%, respectively. Major toxicities (> or = grade 3) included leukopenia in 42 patients and documented infection in 7 patients. Grade 3 cardiac plus renal failure, grade 3 peritonitis due to small bowel perforation, and grade 3 liver dysfunction occurred in 1 patient each. One patient died of toxicity (grade 4 hematological toxicity and pneumonia). In conclusion, it seems that in the elderly patients with non-Hodgkin's lymphoma, response to reduced-dose ((5/6) dose) CHOP therapy is comparable to that for standard CHOP in younger adults, mainly because of improved dose-intensity.

Infectivity and oocyst excretion patterns of Cryptosporidium muris in slightly infected mice.

To determine the infectivity of Cryptosporidium to hosts in slight infections, we examined the infectivity and oocyst output patterns of Cryptosporidium muris in mice inoculated with small numbers of oocysts. One of the 25 ICR mice inoculated with 2.4 x 10(1) oocysts and 19 of the 25 mice inoculated with 2.4 x 10(2) oocysts shed oocysts in the feces after inoculation. Four of the 50 mice inoculated with 2.4 x 10(1) oocysts for 10 consecutive days also shed oocysts and their OPG values were similar to that of the mice which received 2.4 x 10(2) oocysts. Consequently, it is clear that less than 10% of the mice which received 2.4 x 10(1) C. muris oocysts for 10 consecutive days.

Imaging localized horseradish peroxidase on a glass surface with scanning electrochemical/ chemiluminescence microscopy.

Scanning electrochemical/chemiluminescence microscopy (SECM/SCLM) was developed for simultaneous imaging of chemiluminescence and topography of immobilized horseradish peroxidase (HRP) on a glass substrate. When the SECM tip for electrochemical generation of H2O2 scanned over the immobilized spot in an aqueous luminol solution, the immobilized enzymes catalyzed the oxidation of luminol to emit chemiluminescence, which was detected by a photon counter. The photon-counting intensity was plotted against the position of the tip to give a two-dimensional image, indicating the activity of immobilized enzymes. Since the current was sensitive to the topography of substrate, the topography image was also obtained by mapping the oxygen reduction current.

Immunoassay of the MRSA-related toxic protein, leukocidin, with scanning electrochemical microscopy.

Scanning electrochemical microscopy (SECM) was applied to the immunoassay of leukocidin, which is a toxic protein produced by methicillin-resistant Staphylococcus aureus (MRSA), with the intention of developing and early diagnostic for MRSA infection. An antibody-chip for leukocidin was prepared by self-assembling of anti-leukocidin on a protein A-coated glass substrate. A sample solution containing leukocidin was spotted onto the antibody-chip, followed by labeling with horseradish peroxidase (HRP) via a sandwich method. The reduction current of the oxidized form of ferrocenylmethanol generated by the HRP reaction was monitored to view SECM images of the spot of captured leukocidin. The amplitude of reduction current depended on the concentrations of sample solutions used for making spots. This SECM-based immunoassay detects as low as 5.25 pg mL(-1) leukocidin.

Patterning and characterization of surfaces with organic and biological molecules by the scanning electrochemical microscope.

A novel approach for micropatterning of surfaces with organic and biological microstructures using the scanning electrochemical microscope (SECM) is described. The approach is based on the introduction of the spatial resolution by local deposition of gold particles followed by monolayer formation and functionalization. Specifically, gold patterns were deposited locally on silicon wafers with the SECM as a result of the controlled anodic dissolution of a gold microelectrode. The gold patterns were further used as microsubstrates for assembling cystamine monolayers to which either fluoresceine isothiocyanate (FIT) or glucose oxidase (GOD) were covalently attached. Characterization of the organic monolayers, as well as the biological activity of the enzyme patterns, was carried out by fluorescence microscopy and the SECM, respectively.

Microamperometric measurements of photosynthetic activity in a single algal protoplast.

The effects of p-benzoquinone (BQ) on photosynthetic and respiratory electron transport in a single algal protoplast (radius, 100 microm) was investigated quantitatively by amperometric measurements using microelectrodes. Under light irradiation (25 kLx) in the presence of 1.00 mM BQ, a single protoplast consumed BQ by (2.9 +/- 0.2) x 10(-13) mol/s and generated p-hydroquinone (QH2) by (2.7 +/- 0.3) x 10(-13) mol/s, suggesting that BQ was quantitatively reduced to QH2 via the intracellular photosynthetic electron-transport chain. The generation of QH2 increased with light intensity and with concentration of BQ added to the outside solution but became saturated when the light intensity was above 15 kLx or the BQ concentration was higher than 0.75 mM. The addition of 3-(3, 4-dichlorophenyl)-1,1-dimethylurea, a photosynthetic electron-transport inhibitor, decreased the generation of QH2 upon light irradiation, suggesting that BQ accepts electrons from a site in the photosynthetic electron-transport chain after the photosystem II site. The presence of 1.00 mM BQ increased the generation of photosynthetic oxygen by approximately (2.6 +/- 1.0) x 10(-13) mol/s, which was approximately 1.5-2 times larger than that expected from the consumption of BQ. The electrons produced by the additional generation of oxygen is used to reduce intracellular species as well as to reduce BQ.

In vivo analysis of hydrogen peroxide and lipid radicals in the striatum of rats under long-term administration of a neuroleptic.

It has been hypothesized that free radicals play a causative role in tardive dyskinesia, which is an inveterate movement disorder caused by chronic administration of neuroleptics. To verify this hypothesis, rats were reared while being regularly treated with water containing a neuroleptic, haloperidol (HPD), for 1 year (HPD group). The changes in the striatal hydrogen peroxide content of the rats in the HPD and control groups were measured by using a Pt-disk microelectrode while the animals were in a freely moving state following intraperitoneal administration of HPD (HPD challenge). We also performed electron spin resonance (ESR) detection of lipid radicals in the striatum before the HPD challenge. HPD challenge led to significant elevation of the intrastriatal hydrogen peroxide in all animals, but the elevation in the HPD group was smaller than that in the control group. However, in the HPD group, marked ESR signals of intrastriatal lipid radicals were observed. We think that these results support the hypothesis on the role of free radicals in tardive dyskinesia.

Permeation of redox species through a cell membrane of a single, living algal protoplast studied by microamperometry.

Permeation of several redox species through a cell membrane of a single algal protoplast (radius 100 microns) was investigated by amperometry with a Pt microdisk electrode (disk radius, 6.5 microns) located near the membrane. The redox current observed at the microelectrode decreased as the microelectrode approached the cell membrane since the membrane acted as a barrier for diffusion of redox species from bulk to the microelectrode. Permeability coefficient (Pm) of the protoplast membrane was determined by the quantitative analysis of the variation of the redox current with microelectrode-membrane distance using digital simulation. The Pm values for Fe(CN)6(4-), Fe(CN)6(3-), Co(phen)3(2+), ferrocenyl methanol(FMA) and p-hydroquinone(QH2) were < or = 1.0 x 10(-4), < or = 1.0 x 10(-4), 1.0 x 10(-3), 5.0 x 10(-3) and 2.0 x 10(-2) cm/s, respectively. Using these Pm values, the concentration changes inside a model cell and chloroplast were theoretically calculated.

Hydrogen peroxide augmentation in a rat striatum after methamphetamine injection as monitored in vivo by a Pt-disk microelectrode.

We fabricated a Pt-disk microelectrode (diameter 30 microns) to conduct differential double-pulse amperometry (first step: 750 mV, 1 s; second step: 1,100 mV, 1 s) to detect hydrogen peroxide in the brain of a freely moving animal. This measurement determined hydrogen peroxide (detection limit, 0.03 microM) without any observable influence from other oxidizable species, such as dopamine (DA), ascorbic acid, or uric acid. The electrode was implanted into the right striatum of a rat. After intraperitoneal injection of methamphetamine (MAP), hydrogen peroxide concentrations were directly detected while the behavioral changes were monitored. MAP injection led to significant augmentation of hydrogen peroxide, the elevation of which depended on the dose of MAP. This is consistent with a previous report on the increase of DA-release caused by amphetamines and indirect evidence of the production of hydrogen peroxide via DA-metabolism.

Detection of microspotted carcinoembryonic antigen on a glass substrate by scanning electrochemical microscopy.

Microspots of carbinoembryonic antigen (CEA) on glass substrates were characterized by scanning electrochemical microscopy (SECM). CEA was immobilized via a sandiwch method using horseradish peroxidase (HRP)-labeled anti-CEA. The reduction current of the oxidized form of ferrocenylmethanol generated by the HRP reaction was monitored to view SECM images. This method detects as low as ∼10(4) CEA molecules in a single 20-µm-radius spot.

Microamperometric estimation of photosynthesis inhibition in a single algal protoplast.

The single-cell level inhibition of photosynthesis by some chemicals was investigated from the light-irradiation induced response of intracellular oxygen reduction current at an ultramicroelectrode inserted into a single algal protoplast. The addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an electron-transport inhibitor, decreased a current peak appeared immediately after the light irradiation. The pI50 value (the negative decadic logarithm of the concentration needed for 50% inhibition) was determined from the change in response patterns and found to be 6.9 for DCMU. 2,4-Dichlorophenoxyacetic acid (2,4-D), an auxin herbicide, showed no effect on photosynthesis in the effective concentration range. 2,4-Dinitrophenol (DNP), a proton carrier, accelerated the oxygen production upon light irradiation.