Photocurrent generation of a single-gate graphene p-n junction fabricated by interfacial modification.

A back-gate graphene p-n junction was achieved by selective interfacial modification of a chemical vapor deposition (CVD)-grown graphene field effect transistor (FET). Silane self-assembled monolayer (SAM) patterns were used to fabricate uniform p- and n-doped regions and a sharp p-n junction in the graphene FET channel. A gate-dependent photocurrent response was observed at the graphene p-n junction, and exhibited a maximum signal between two Dirac point voltages of SAM-doped graphene regions. A spatial photocurrent map shows that the photocurrent generated at the junction region was much larger than that from graphene/electrode junctions under the same incident laser power. This single-peak characteristic photocurrent in CVD graphene is dominated by the photothermoelectric contribution, and is highly sensitive to the power of incident laser. The SAM interfacial modification method provides a feasible route for the fabrication of efficient graphene-based photodetectors.

Remineralizing agents: effects on acid-softened enamel.

This study sought to evaluate whether remineralizing toothpastes can protect acid-softened enamel against further erosive episodes. Fifty enamel slabs of bovine teeth with preformed erosion-like lesions were randomly assigned to 1 control and 4 experimental groups (n = 10): group 1, nanohydroxyapatite (nanoHAp) dentifrice; group 2, arginine and calcium carbonate (CaCO3) dentifrice; group 3, potassium nitrate (KNO3) and high-fluoride (F) availability dentifrice; group 4, ordinary fluoridated dentifrice (OFD); and group 5, control (deionized water). Initial hardness measurements were taken after the different treatments were applied. Statistically significant mineral gains of 8.0% and 10.0% were exhibited in groups 1 and 4, respectively. Groups 2 and 3 showed mineral gains of 4.5% and 2.1%, respectively; these were not statistically significant. Group 5 showed mineral loss (-11.8%). A 1-way analysis of variance showed no statistically significant differences in the mean microhardness values among groups. However, there are indications that the nanoHAp and OFD toothpastes may decrease erosive lesions after treatment, while the arginine + CaCO3 and KNO3 + F pastes may prevent the progression of erosive lesions.

Potential agents to control enamel caries-like lesions.

OBJECTIVES: To assess whether pastes containing casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) and calcium sodium phosphosilicate (CSP) control artificial caries lesion progression. METHODS: Enamel slabs of bovine teeth were embedded in polyester resin and had their enamel surfaces serially polished, and tested for surface microhardness (SMH, 25 g, 5 s). Incipient caries-like lesions were pre-formed and specimens were evaluated by microhardness test (SMH(post-lesion)) and randomly assigned to five treatment groups (n=15): (1) regular dentifrice (RE, 1,100 ppm F); (2) dentifrice with calcium sodium phosphosilicate (CSP); (3) amorphous calcium phosphate stabilized by casein phosphopeptide (CPP-ACP); (4) CPP-ACP with 900 ppm F (CPP-ACP+F) and (5) control group-unexposed to any remineralizing agent. Treatments were applied five times, after the de-remineralization period in the cariogenic challenges. Post-treatments SMH measurements were conducted (SMH(post-treatment)). RESULTS: ANOVA was applied for data evaluation and revealed a significant difference among the treatments (p=0.0161). Tukey's test was conducted and the percentage of mineral loss was calculated. Specimens exposed to CSP (7.1%), RE (6.7%) and CPP-ACP+F (3.8%) showed lower mineral loss than those that in the control group (-11.0%). CPP-ACP group (3.2%) differed from the control, CSP and RE groups. CONCLUSIONS: Depending on the agent used, a remineralizing effect may be expected, which reflects in caries lesions progression.

Brain MRI in mucopolysaccharidosis: effect of aging and correlation with biochemical findings.

OBJECTIVE: To investigate the influence of aging on conventional MRI and magnetic resonance spectroscopy (MRS) findings of mucopolysaccharidosis (MPS) patients and to test the correlation of enzyme levels, urinary glycosaminoglycans (GAG), and neuroimaging findings. METHODS: Sixty patients with MPS types I (n = 8), II (n = 31), IV-A (n = 4), and VI (n = 17) underwent T2, fluid-attenuated inversion recovery (FLAIR), and MRS of the brain. For analysis of MRI variables, we measured the normalized cerebral volume (NCV), CSF volume (NCSFV), ventricular volume (NVV), and lesion load (NLL) on FLAIR using semiautomated and automated segmentation techniques. For MRS, a point-resolved spectroscopy technique was used. Voxels were positioned at the white and gray matter. Statistical analysis involved Pearson or Spearman tests for correlation between neuroimaging, age, enzyme levels, and urinary GAG. RESULTS: The median age at onset of the disease was 20 months. Patients with longer disease duration had more NLL in the white matter (r = 0.28, p = 0.03), and this difference was more pronounced in MPS II patients (r = 0.44, p = 0.02). Metabolites ratios in MRS, NCV, NCSFV, and NVV did not correlate with disease duration or age of the patients (p > 0.05). MRI and MRS variables in either the white or the gray matter did not correlate with enzymatic activity or GAG levels. Patients with MPS II had a lower mean NCV (p < 0.001). CONCLUSIONS: Our data showed that white matter lesion is more extensive as disease duration increases, especially in mucopolysaccharidosis type II patients. MRI and magnetic resonance spectroscopy findings did not correlate with either enzymatic or glycosaminoglycan levels.

Production and characterization of recombinant human anti-HBs Fab antibodies.

Recombinant human Fab antibodies were generated with different reactivities against the hepatitis B virus surface (HBs) antigen. To isolate the antibodies, a method was used that combined transformation of human B cells by Epstein-Barr virus (EBV) infection with a primer-vector system developed for isolating DNA fragments of human Ig Fab portions. With this method, monoclonal and oligoclonal cell lines producing anti-HBs antibodies were established and three anti-HBs Fab antibodies were isolated from two of these cell lines. From analysis of affinity characteristics, immunohistochemical activity, and cytolysis activity, these three Fab antibodies were classified into three different groups. The first group had high affinity for HBs, the second had the ability to kill HBV-infected cells, and the third was applicable to immunohistochemical staining with HBV-infected cells. The combined effect of these antibodies was also investigated by complement-dependent cytotoxicity assay.

Experience-dependent modulation of motor corticospinal excitability during action observation.

The primate premotor cortex is endowed with an "action observation/execution matching system", that is, the same premotor neurons discharge when actions are performed and when actions are observed. Hence, this system predicts a strong visual input to the motor system. Whether this input is dependent on visual experience or not has not been previously investigated. To address this issue we compared corticospinal excitability while subjects viewed frequently observed and less frequently observed hand actions of others and of themselves. Motor corticospinal excitability was larger when the action orientations were as they are frequently observed (Self-away, subject's own hand facing out from the subject, or Other-toward, an unknown hand facing toward the subject) compared with less frequently observed actions (Self-toward, subject's own hand facing "toward" the subject, or Other-away, an unknown hand facing out from the subject). This finding suggests that the modulation of motor corticospinal excitability during action observation and hence the "action observation/execution matching system" is largely dependent upon visual experience.

Cloning and expression of human anti-tumor necrosis factor-alpha monoclonal antibodies from Epstein-Barr virus transformed oligoclonal libraries.

Peripheral blood was obtained from a healthy human volunteer and transformed with Epstein-Barr virus (EBV). This produced an oligoclonal cell library in culture medium that was screened by ELISA for anti-human tumor necrosis factor-alpha (TNFalpha) activity. RNA from two positive clones was applied to RT-PCR using antibody-specific primers, and the light (kappa and lambda) and heavy chain genes (gamma and mu) were cloned into the plasmid vector pFab1-His2. The antibodies produced in Escherichia coli as Fab fragments were assayed for anti-TNFalpha activity utilizing ELISA. Two IgG1/kappa anti-TNFalpha antibodies and two IgM/kappa anti-TNFalpha antibodies were isolated. DNA sequence analysis showed that the VL and VH gene families of IgM and IgG were the same. Both the antibodies showed almost the same activity on ELISA-testing. Ten clones randomly selected from light (kappa and lambda) and heavy (gamma and mu) chain genes in the oligoclonal cell library 1D5 were sequenced, and each gene (kappa, lambda, gamma, and mu) was found to be composed of one to three different genes. These data support the conclusion that the cell clone is oligoclonal at the molecular level.

The role of the dorsolateral prefrontal cortex during sequence learning is specific for spatial information.

Many studies have implicated the dorsolateral prefrontal cortex in the acquisition of skill, including procedural sequence learning. However, the specific role it performs in sequence learning has remained uncertain. This type of skill has been intensively studied using the serial reaction time task. We used three versions of this task: a standard task where the position of the stimulus cued the response; a non-standard task where the color of the stimulus was related to the correct response; and a combined task where both the color and position simultaneously cued the response. We refer to each of these tasks based upon the cues available for guiding learning as position, color and combined tasks. The combined task usually shows an enhancement of skill acquisition, a result of being driven by two simultaneous and congruent cues. Prior to the performance of each of these tasks the function of the dorsolateral prefrontal cortex was disrupted using repetitive transcranial magnetic stimulation. This completely prevented learning within the position task, while sequence learning occurred to a similar extent in both the color and combined tasks. So, following prefrontal stimulation the expected learning enhancement in the combined task was lost, consistent with only a color cue being available to guide sequence learning in the combined task. Neither of these effects was observed following stimulation at the parietal cortex. Hence the critical role played by the dorsolateral prefrontal cortex in sequence learning is related exclusively to spatial cues. We suggest that the dorsolateral prefrontal cortex operates over the short term to retain and manipulate spatial information to allow cortical and subcortical structures to learn a predictable sequence of actions. Such functions may emerge from the broader role the dorsolateral prefrontal cortex has in spatial working memory. These results argue against the dorsolateral prefrontal cortex constituting part of the neuronal substrate responsible for general aspects of implicit or explicit sequence learning.

Four cases of bronchiolitis obliterans organizing pneumonia associated with thyroid disease.

We present 4 cases of bronchiolitis obliterans organizing pneumonia (BOOP) associated with thyroid diseases. Case 1 had previously undergone surgery for thyroid cancer, and had a secondary hypothyroidism at the onset of BOOP. Case 2 developed Basedow's disease 3 months after the onset of BOOP and BOOP relapsed 21 months after the onset of Basedow's disease. Case 3 developed subacute thyroiditis 33 months after the onset of BOOP, and has had no relapse of BOOP for 7 years. Case 4 had hypothyroidism at the time of diagnosis of BOOP, and her BOOP relapsed 3 months after the initial onset of BOOP. Two of these 4 cases of BOOP with thyroid diseases relapsed, and thyroid dysfunction could modify the pathophysiology of BOOP.

Interhemispheric asymmetry of motor cortical excitability in major depression as measured by transcranial magnetic stimulation.

BACKGROUND: Neuroimaging studies of major depressive disorder (MDD) indicate interhemispheric differences in prefrontal cortical activity (right greater than left). AIMS: To investigate whether there are any interhemispheric differences of motor cortical excitability in MDD. METHOD: Eight patients with treatment-refractory MDD off medication were assessed for the severity of their depression, and transcranial magnetic stimulation studies (bilateral motor threshold and paired-pulse studies) were conducted. Eight normal controls were also studied. RESULTS: MDD patients showed significant interhemispheric differences in motor threshold and paired-pulse curves, both of which showed lower excitability on the left hemisphere. Such differences were absent in controls. CONCLUSIONS: Our findings may aid the further understanding of the neurophysiology underlying MDD.

Interindividual variability of the modulatory effects of repetitive transcranial magnetic stimulation on cortical excitability.

Repetitive transcranial magnetic stimulation (rTMS) appears to have effects on cortical excitability that extend beyond the train of rTMS itself. These effects may be inhibitory or facilitatory and appear to depend on the frequency, intensity, duration and intertrain interval of the rTMS. Many studies assume facilitatory effects of high-frequency rTMS and inhibitory effects of low-frequency rTMS. Nevertheless, the interindividual variability of this modulation of cortical excitability by rTMS has not been systematically investigated. In this study, we applied 240 pulses of rTMS at 90% of the subjects' motor threshold to their motor cortex at different frequencies (1, 10, 15 and 20 Hz) and examined the effects on motor evoked potentials (frequency tuning curve). Although the averaged group data showed a frequency-dependent increase in cortical excitability, each subject had a different pattern of frequency tuning curve, i.e. a different modulatory effect on cortical excitability at different rTMS frequencies. The interindividual variability of these modulatory effects was still high, though less so, when the number of rTMS pulses was increased to 1,600. These findings illustrate the degree of variability of the rTMS effects in the human brain.

Modulation of corticospinal excitability by repetitive transcranial magnetic stimulation.

OBJECTIVE: Repetitive transcranial magnetic stimulation (rTMS) is able to modulate the corticospinal excitability and the effects appear to last beyond the duration of the rTMS itself. Different studies, employing different rTMS parameters, report different modulation of corticospinal excitability ranging from inhibition to facilitation. Intraindividual variability of these effects and their reproducibility are unclear. METHODS: We examined the modulatory effects of rTMS to the motor cortex at various frequencies (1, 10, 20 Hz) and at different time-points in twenty healthy volunteers. RESULTS: We observed significant inhibition of MEPs following 1 Hz rTMS and significant facilitation of MEPs following 20 Hz rTMS for both day1 and day 2. Interestingly, at 1 Hz and 20 Hz rTMS, the modulatory effect produced by rTMS was greater on day 2. However, there was no significant change in corticospinal excitability following 10 Hz rTMS neither on day 1 nor day 2. CONCLUSION: Our findings raise questions as to how stimulation parameters should be determined when conducting studies applying rTMS on multiple days, and in particular, studies exploring rTMS as a treatment modality in neuropsychiatric disorders.

Understanding taijin kyofusho through its treatment, Morita therapy.

This article presents a brief review of the definition, nosology, history, clinical features, and etiology of taijin kyofusho. Special attention is also given to Morita therapy for taijin kyofusho. The term taijin kyofusho literally means the disorder (sho) of fear (kyofu) of interpersonal relations (taijin). Morita therapy was developed by Masatake Morita in the 1910s to treat the Japanese mental disorders called shinkeishitsu and taijin kyofusho. It is suggested that taijin kyofusho is an excellent example of a mental disorder in which understanding its treatment is an integral part of its conceptualization.

Bacterial expression of a human recombinant monoclonal antibody fab fragment against hepatitis B surface antigen.

The Fab fragment was cloned from the monoclonal cell line TAPC301-CL4, which was produced using the Epstein-Barr virus (EBV) transformation method. This cell line produces a human monoclonal antibody (CL4MAb) against the hepatitis B surface antigen (HBsAg). This MAb was shown to have hepatitis B virus (HBV) neutralizing activity in chimpanzees. The Fab fragment was produced by subjecting the heavy and light chain antibody genes of the TAPC301-CL4 cell line to reverse transcription-polymerase chain reaction, cloning the products in the plasmid vector pFab1-His2 and introducing the plasmid into bacteria. Sequence analyses of the CL4Fab fragment revealed that the light and heavy chains belong to the Vk3a and VH3 groups of the immunoglobulin (Ig) family, respectively. An enzyme-linked immunosorbent assay confirmed that specificity of the recombinant CL4Fab antibody against HBsAg was the same as that of the parental MAb. Flow cytometric analysis using PLC/PRF/5 (Alexander) cells, which express HBsAg, showed the reactivities of the CL4MAb and CL4Fab antibody were the same. These results suggest that the recombinant CL4Fab antibody produced by Escherichia coli using the new vector-primer system developed for human IgG Fab fragments has a very high affinity for the HBsAg and may be useful clinically. A source for generation of human MAb for human therapy with very stable and specific expression was thus produced by isolating antibodies from EBV-transformed cell lines.

Preparation of recombinant human monoclonal antibody Fab fragments specific for Entamoeba histolytica.

Genes coding for human antibody Fab fragments specific for Entamoeba histolytica were cloned and expressed in Escherichia coli. Lymphocytes were separated from the peripheral blood of a patient with an amebic liver abscess. Poly(A)+ RNA was isolated from the lymphocytes, and then genes coding for the light chain and Fd region of the heavy chain were amplified by a reverse transcriptase PCR. The amplified DNA fragments were ligated with a plasmid vector and were introduced into Escherichia coli. Three thousand colonies were screened for the production of antibodies to E. histolytica HM-1:IMSS by an indirect fluorescence-antibody (IFA) test. Lysates from five Escherichia coli clones were positive. Analysis of the DNA sequences of the five clones showed that three of the five heavy-chain sequences and four of the five light-chain sequences differed from each other. When the reactivities of the Escherichia coli lysates to nine reference strains of E. histolytica were examined by the IFA test, three Fab fragments with different DNA sequences were found to react with all nine strains and another Fab fragment was found to react with seven strains. None of the four human monoclonal antibody Fab fragments reacted with Entamoeba dispar reference strains or with other enteric protozoan parasites. These results indicate that the bacterial expression system reported here is effective for the production of human monoclonal antibodies specific for E. histolytica. The recombinant human monoclonal antibody Fab fragments may be applicable for distinguishing E. histolytica from E. dispar and for use in the serodiagnosis of amebiasis.

Bacterial expression of a neutralizing mouse monoclonal antibody Fab fragment to a 150-kilodalton surface antigen of Entamoeba histolytica.

A mouse monoclonal antibody (MAb) (EH3015, IgG1 with a K light chain) prepared by hybridoma technology recognizes a 150-kD surface antigen of Entamoeba histolytica and inhibits adherence and cytotoxicity of the ameba to mammalian cells. The genes encoding the light chain and the Fd region of the heavy chain of the MAb were cloned and expressed in Escherichia coli. The plasmid used was designed for the expression of Fab with a hexa-histidine tag in the periplasmic space. Recombinant Fab fragments were purified and analyzed by an indirect immunofluorescence antibody test and Western immunoblot. The specificity of the recombinant Fab fragment was comparable with the parent whole IgG. In addition, the Fab fragments significantly inhibited the adherence of E. histolytica to erythrocytes. These results suggest that the production of a neutralizing MAb in Escherichia coli is practical and efficient with this expression system.

The development of a sensitive and specific enzyme immunoassay for FK480, a novel cholecystokinin type-A receptor antagonist, in human plasma.

A sensitive and specific enzyme immunoassay for FK480, a novel cholecystokinin type-A (CCK-A) receptor antagonist, was developed to study the pharmacokinetics of the drug at low-dose administration using a specific monoclonal antibody. The high performance liquid chromatography (HPLC) method had been used for studying toxicokinetics, but its determination limit (2.5 ng ml-1) was too high for use in clinical studies. Subsequently we developed an enzyme immunoassay (EIA) using rabbit anti-FK480 serum (polyclonal antibody). It had higher sensitivity (0.1 ng ml-1) when 0.5 ml of plasma was used but its specificity was low because of the cross-reactivity of the metabolites of FK480. Therefore we produced several monoclonal antibodies for FK480 by cell fusion, and selected the antibody which was least cross-reactive for the isolated metabolites of FK480. Finally we developed a sensitive and specific EIA using this monoclonal antibody. The lower limit of quantification of this method was 0.2 ng ml-1 when 0.2 ml of human plasma was used. The coefficient of variation over the calibration range (0.2-10 ng ml-1) was less than 15%. We used this method for clinical studies, and it showed a good correlation to the HPLC method when plasma concentration was 2.5 ng ml-1 or more.

Human monoclonal anti-HCMV neutralizing antibody from phage display libraries.

Human cytomegalovirus (HCMV) infection in immunocompromised patients causes considerable morbidity and mortality. Although ganciclovir prophylaxis reduces the incidence of HCMV disease, severe side effects raise serious problems. Thus, the development of new strategies for prophylaxis are clearly needed, and human monoclonal antibodies offer a potential alternative. We describe the cloning, using the phage display system, of a recombinant human Fab fragment against HCMV. A phage display library with 4 x 10(6) clones was panned three times against lysates of HCMV-infected cells, and screened by ELISA. Of six antigen-binding clones, one monoclonal antibody reacted strongly to HCMV. In immunostaining analysis, this Fab was able to stain HCMV-infected cells from 24 h post-infection (pi) through to 96 h pi, but not at 6 h pi. In the presence of cytosine arabinoside, HCMV-infected cells were not stained, even at 24 h pi. These results indicate that an HCMV protein that was recognized by the Fab was synthesized in the late phase of infection. In addition, this Fab exhibited neutralizing activity: at 1 microg/ml it reduced HCMV plaque formation by 50%. The Fab was able to neutralize three HCMV strains, but it did not neutralize HSV-1 or -2 infection.

[Neutralizing human antibody specific for human cytomegalovirus in E. coli expression system].

We isolated neutralizing human Fab fragment specific for human cytomegalovirus (HCMV) by phage display system. Fab libraries were constructed from peripheral lymphocyte of healthy individual. In several clones reacted for HCMV-infected HEL cells, one clone, designated 13-3, stained HCMV infected cells at 96 hrs post infection. It didn't react to cells at 6 hrs post infection or infected cells in the presence of AraC, meaning that 13-3 recognized protein synthesized at late times of infection. It also neutralized HCMV, Towne strain, efficiently. This neutralizing activity was specific for HCMV and no effect for HSV-1 and 2.

Factors related to the relapse of bronchiolitis obliterans organizing pneumonia.

STUDY OBJECTIVE: The purpose of this study is to determine factors, including laboratory data, related to the relapse of bronchiolitis obliterans organizing pneumonia (BOOP). DESIGN: Retrospective study. SETTING AND PATIENTS: The medical files of Fukuoka University Hospital and Nishi Fukuoka Hospital patients from 1984 to 1996 were reviewed, and 18 cases of BOOP that had been diagnosed using transbronchial or open lung biopsy were selected for evaluation. MEASUREMENTS: The 18 cases were put into two groups composed of 7 patients who relapsed and 11 who did not relapse. Their clinical symptoms and laboratory data at first admission, including hemograms, blood chemistry tests, and pulmonary function tests were compared. Patients with or without associated diseases, such as collagen vascular diseases, were compared using the same parameters in order to examine the relationship between the associated diseases and BOOP relapse. RESULTS: The serum levels of total protein and albumin in patients who relapsed were significantly lower than in patients who did not relapse, respectively: 5.8 (range, 4.4 to 6.2) vs 6.3 (range, 4.5 to 6.8) g/dL, p < 0.05; and 2.9 (range, 2.5 to 3.4) vs 3.7 (range, 2.8 to 4.3) g/dL, p < 0.01. Levels of serum albumin in BOOP patients with associated diseases, however, were significantly lower than in those without associated diseases, respectively: 2.95 (range, 2.5 to 3.9) vs 3.65 (range, 2.8 to 4.3) mg/dL, p < 0.05. The fall in serum albumin levels in patients who relapsed, therefore, was probably due to associated diseases. The fact that 5 of 8 patients with associated diseases relapsed but only 2 of 10 without associated diseases relapsed suggests that a relationship exists between associated diseases and the prognosis of BOOP, although this finding was not statistically significant because of the small number of cases and the heterogeneity of the associated diseases. The most striking observation was that PaO2 levels in patients who relapsed were significantly lower than in those who did not, respectively: 55.4 (range, 39.9 to 73.2) vs 78.0 (range, 48.4 to 89.4) mm Hg, p < 0.05. However, PaO2 levels were not statistically different between patients with and without associated diseases, respectively: 66.0 (range, 45.4 to 78.8) vs 71.4 (range, 39.9 to 89.4) mm Hg. CONCLUSIONS: The severity of hypoxemia at first medical examination may be an important determinant for the subsequent BOOP relapse.