Clostridium difficile toxins or infection induce upregulation of adenosine receptors and IL-6 with early pro-inflammatory and late anti-inflammatory pattern.

Clostridium difficile causes intestinal inflammation, which increases adenosine. We compared the expression of adenosine receptors (AR) subtypes A1, A2A, A2B, and A3 in HCT-8, IEC-6 cells, and isolated intestinal epithelial cells, challenged or not with Clostridium difficile toxin A and B (TcdA and TcdB) or infection (CDI). In HCT-8, TcdB induced an early A2BR expression at 6 h and a late A2AR expression at 6 and 24 h. In addition, both TcdA and TcdB increased IL-6 expression at all time-points (peak at 6 h) and PSB603, an A2BR antagonist, decreased IL-6 expression and production. In isolated cecum epithelial cells, TcdA induced an early expression of A2BR at 2s and 6 h, followed by a late expression of A2AR at 6 and 24 h and of A1R at 24 h. In CDI, A2AR and A2BR expressions were increased at day 3, but not at day 7. ARs play a role in regulating inflammation during CDI by inducing an early pro-inflammatory and a late anti-inflammatory response. The timing of interventions with AR antagonist or agonists may be of relevance in treatment of CDI.

Role of the haem oxygenase/carbon monoxide pathway in Clostridium difficile toxin A-induced enteritis in mice.

Clostridium difficile is the major cause of antibiotic-associated colitis, a disease with significant morbidity and mortality. This study investigated the role of the haem oxygenase-1 (HO-1)/carbon monoxide (CO) pathway in C. difficile toxin A-induced enteritis in mice. The HO substrate haemin, zinc protoporphyrin IX (ZnPP IX), a specific HO-1 inhibitor, dimanganese decacarbonyl (DMDC), a CO donor, or an equivalent volume of their respective vehicles were injected subcutaneously 30 min prior to local challenge with toxin A (25 or 50 µg per ileal loop) or PBS. Intestinal ileal loop weight/length ratios were calculated 3 h later. Ileal tissues were collected for histological analysis and measurement of myeloperoxidase (MPO) activity, tumour necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1ß) production by ELISA and immunohistochemistry for HO-1. Treatment of mice subjected to C. difficile toxin A (TcdA) with haemin or DMDC prevented oedema, mucosal disruption and neutrophil infiltration observed in histological analysis. It also decreased TcdA-induced MPO activity and TNF-α or IL-1ß production. In contrast, the specific HO-1 inhibitor (ZnPP IX) exacerbated all these evaluated parameters. TcdA increased HO-1 expression as seen by immunohistochemistry. These results suggest that the HO-1/CO pathway exerts a protective role in TcdA-induced enteritis and that its pharmacological modulation might be important for the management of C. difficile-associated disease.

Quercetin Suppresses Early Colon Carcinogenesis Partly through Inhibition of Inflammatory Mediators.

We have demonstrated that 0.45% quercetin added to a diet containing corn oil (15% w/w), as the lipid source, and cellulose (6% w/w), as the fiber source, was able to suppress the formation of high multiplicity aberrant crypt foci (ACF > 4 AC/focus), to lower proliferation and enhance apoptosis in a rat model of colon cancer. This experiment determined whether quercetin was acting as an antiinflammatory molecule in an in vivo model of colon cancer. We used weanling (21 d old) Sprague Dawley rats (n = 40) in a 2×2 factorial experiment to determine the influence of quercetin on iNOS, COX-1 and COX-2 expressions, all of which are elevated in colon cancer. Half of the rats received a diet containing either 0 or 0.45% quercetin, and within each diet group, half of the rats were injected with saline or azoxymethane (AOM, 15 mg/kg BW, sc, 2× during wk 3 and 4). The colon was resected 4 wk after the last AOM injection, and the mucosa scraped and processed for RNA isolation. Data from this experiment were analyzed using a mixed model in SAS for main effects and their interaction. AOM injection stimulated (P < 0.0001) iNOS expression. However there was an interaction such that, relative to rats injected with saline, AOM-injected rats consuming diets without quercetin had significantly elevated iNOS expression (5.29-fold), but the expression in AOM-injected rats consuming the diet with quercetin was not significantly elevated (1.68-fold). COX-1 expression was 20.2% lower (P < 0.06) in rats consuming diets containing quercetin. COX-2 expression was 24.3% higher (P < 0.058) in rats consuming diets without quercetin. These data suggest inflammatory processes are elevated in this early stage of colon carcinogenesis, yet quercetin may protect against colon carcinogenesis by down-regulating the expressions of COX-1 and COX-2.

Effects of 12-h tobacco deprivation on event-related potentials elicited by visual smoking cues.

RATIONALE: A cigarette smoker's reactivity to smoking cues, or cue-reactivity, traditionally has been indexed by self-report and/or measures of autonomic nervous system activity. Recent evidence suggests that measures of central nervous system activity in the form of event-related brain potentials (ERPs) may also index cue-reactivity in smokers. OBJECTIVE: The present study sought to confirm the sensitivity of ERPs to smoking cues and to investigate the question of whether 12 h of smoking deprivation would enhance ERP cue-reactivity to such stimuli. METHODS: Scalp ERPs were recorded to 80 smoking-related pictures and 80 neutral pictures, i.e., similar pictures with a nonsmoking theme, in 19 tobacco-deprived smokers, 17 nondeprived smokers, and 19 nonsmokers. RESULTS: Smokers' N300 amplitudes over fronto-central scalp were larger to neutral than to smoking-related stimuli, thus reflecting N300 smoking cue-reactivity. N300 cue-reactivity was greater for deprived than for nondeprived smokers. Smokers' P300 values were greater to smoking-related than to neutral stimuli, particularly over the centro-parietal area; however, nonsmokers also showed a P300 main effect to smoking cues. Smoking deprivation did not affect P300 cue-reactivity, nor did deprivation affect self-reported urges to smoking relative to neutral cues. CONCLUSIONS: These data confirm the sensitivity of ERPs to tobacco cues in smokers and suggest, additionally, that the cue-reactivity of the N300 component is modulated by smoking deprivation. N300 cue-reactivity may reflect an internally generated priming of the semantic network related to the smokers' need states. Stimulus-category differences in P300 may reflect cue-reactivity in smokers and/or nonaddiction-specific factors in both smokers and nonsmokers.

Event-related brain potentials as indicators of smoking cue-reactivity.

OBJECTIVE: Reactivity to smoking cues, shown previously by autonomic and self-report variables, was investigated in smokers and nonsmokers using event-related brain potentials (ERPs). METHODS: Average ERPs to 20 color pictures of people smoking and 20 neutral pictures depicting nonsmoking themes, randomly mixed with 4 repetitions/stimulus, were measured from 20 light-moderate smokers and 18 nonsmokers, following a stressor. Smoker status and stimulus type effects on ERPs and principal components factor scores (FS) were tested by repeated-measures ANOVAs. RESULTS: Smokers' N268 showed significant medial and midline smoking cue-reactivity (ERP to smoking-related minus neutral stimuli); while a P300-like, P412, showed significant smoking cue-reactivity over medial and left hemisphere scalp. FS analyses confirmed most of the foregoing. P412 smoking cue-reactivity was correlated with unpleasantness-pleasantness cue-reactivity but not with urge-to-smoke cue-reactivity. Nonsmokers' N268 stimulus differences were not significant, but significant P412 stimulus effects (unconfirmed by FS analyses) were found in central-to-posterior and in left parietal-temporal areas. CONCLUSIONS: Smokers' N268 is identified with a process detecting stimuli incongruent with tobacco-addicted states; and P412 smoking cue-reactivity is discussed in terms of an automatic, perceptual-categorization system, consistent with Tiffany's drug-use and Johnson's P300 models. Implications of ERP smoking cue-reactivity for study of tobacco and other addictions are discussed.

Transcriptionally active MuDR, the regulatory element of the mutator transposable element family of Zea mays, is present in some accessions of the Mexican land race Zapalote chico.

To date, mobile Mu transposons and their autonomous regulator MuDR have been found only in the two known Mutator lines of maize and their immediate descendants. To gain insight into the origin, organization, and regulation of Mutator elements, we surveyed exotic maize and related species for cross-hybridization to MuDR. Some accessions of the mexican land race Zapalote chico contain one to several copies of full-length, unmethylated, and transcriptionally active MuDR-like elements plus non-autonomous Mu elements. The sequenced 5.0-kb MuDR-Zc element is 94.6% identical to MuDR, with only 20 amino acid changes in the 93-kD predicted protein encoded by mudrA and ten amino acid changes in the 23-kD predicted protein of mudrB. The terminal inverted repeat (TIR) A of MuDR-Zc is identical to standard MuDR; TIRB is 11.2% divergent from TIRA. In Zapalote chico, mudrA transcripts are very rare, while mudrB transcripts are as abundant as in Mutator lines with a few copies of MuDR. Zapalote chico lines with MuDR-like elements can trans-activate reporter alleles in inactive Mutator backgrounds; they match the characteristic increased forward mutation frequency of standard Mutator lines, but only after outcrossing to another line. Zapalote chico accessions that lack MuDR-like elements and the single copy MuDR a1-mum2 line produce few mutations. New mutants recovered from Zapalote chico are somatically stable.

In vitro elution of tobramycin and vancomycin polymethylmethacrylate beads and spacers from Simplex and Palacos.

In vitro studies of antibiotic elution from polymethylmethacrylate cement comparing Simplex (Howmedica, Rutherford, NJ) with Palacos brands (Richards, Memphis, TN) have shown variable results. This study compares the elution of tobramycin and vancomycin from Simplex and Palacos beads and spacers. Six-millimeter beads and spacers were incubated in phosphate-buffered saline, and the solution was sampled and changed daily until the concentration of antibiotic fell below the minimum inhibitory concentration (MIC) for Staphylococcus aureus. In all groups, the Palacos PMMA beads and spacers showed elution at higher levels and remained above the MIC longer than did the Simplex PMMA beads. Tobramycin had superior elution to vancomycin in all groups. This in vitro study shows that Palacos PMMA has superior elution properties to Simplex PMMA in tobramycin and vancomycin beads and spacers.

Relation of surface area to in vitro elution characteristics of vancomycin-impregnated polymethylmethacrylate spacers.

To study the relationship between surface area and antibiotic elution from antibiotic-impregnated polymethylmethacrylate (PMMA) spacers, a standard block spacer made of vancomycin (4 g) and 40 g of PMMA was compared with two unique spacer designs, the "donut" and "fenestrated." The spacers were incubated in phosphate-buffered saline, which was changed daily, and a microbiologic assay was used to measure the antibiotic activity of the eluates. The donut and fenestrated spacers had 12% and 40% more surface area than the standard spacer, respectively. There was no significant difference, however, in daily elution levels of antibiotic between the donut spacer and the standard spacer. The fenestrated spacer displayed significantly better elution than either the standard or donut spacers, with an average of 20% more antibiotic eluted on any given day.

Effects of Ayahuasca on the human EEG.

EEG data were recorded under field conditions from 11 members of the Santo Daime Doctrine, a Brazilian shamanistic religion, before and after ingesting the psychoactive alkaloid preparation, ayahuasca, or daime, as they term it. Post-ingestion, we observed increases in power in the 36-44 Hz frequency band ("40 Hz") from the left occipital-temporal-parietal scalp electrodes in the eyes-closed condition, which extended to most of the posterior scalp in the eyes-open condition. The results are consistent with many reports that ayahuasca intensifies visual imagery. These results are discussed in terms of a thalamocortical model of the role of 40 Hz activity in brain function and conscious experience. We also noted tendencies toward decreases in the power of slow (theta and alpha) brain rhythms, and increases in the 14-30 Hz beta band, in accord with studies reported 30 years ago with other consciousness-altering compounds. Analysis of four ayahuasca samples yielded an average composition per ingested dose (75 ml) of 55.6 mg harmine, 43.9 mg tetrahydroharmine, 41.3 mg N,N-dimethyltryptamine (DMT), 4.6 mg harmaline, and 3.1 mg harmol. The DMT appeared to be of sufficient concentration to promote psychoactive effects, while the ß-carbolines functioned to supply MAO inhibitors necessary to prevent degradation of DMT and to maintain its oral activity.

Fluconazole disc diffusion testing for the routine laboratory.

The increasing incidence of resistance to the antifungal agent, fluconazole, prompted the need for a rapid, reliable and easy-to-use susceptibility test. We have developed a disc diffusion test for fluconazole against Candida spp. suitable for a clinical laboratory. Disc diffusion tests on six different media were compared with MIC values. On the basis of correlation coefficient with MICs (r = -0.95), quality of growth and zone edge definition, Yeast Nitrogen Base agar with glucose (YNBG) produced the best results. Further studies on YNBG showed that the method is reliable for Candida albicans and for resistant isolates with no zone of inhibition, but results for the slower growing and uncommon species must be interpreted with some caution. Implementation of this test in the clinical laboratory has provided a much needed therapeutic service for clinicians within the hospital. It has also reduced the reliance on the reference laboratory for susceptibility results and the consequent costs involved.

P3B and positive slow wave following real and dummy feedback on arithmetic rule-learning and perceptuomotor tasks.

Event-related Potentials (ERPs) were recorded to feedback during a cognitively demanding, arithmetic rule-learning task and a relatively simple, skill-oriented, perceptuomotor task. For both tasks, a compound feedback display was employed. It consisted of numeric feedback information presented simultaneously with a red or green light (50% each) which indicated whether the numeric information was real (valid) or dummy (invalid). The task and feedback-validity manipulations showed a functional dissociation between the P3b (350-450 msec.) and a Positive Slow Wave (600-900-msec.). P3b was larger for real than for dummy feedback; Positive Slow Wave was larger for rule-learning than for perceptuomotor tasks.

Positive event-related potentials to real and dummy rule-learning feedback and to perceptuomotor feedback.

Amplitudes of 5 event-related potentials (ERPs) were recorded at 5 sites of 10 males to real (R-) and dummy (D-) feedback (FB) over two difficulty levels of a rule-learning task, with interspersed perceptuomotor (PM) task trials. Rule-learning R-FB for positive slow wave (PSW) and P3b was greater than for D-FB for both mean time-window and principal component factor score measures. FB effects varied by site for P2/P3a (mainly Fz-C4-Pz) and for a late PSW (LPSW; mainly C4-C3-Fz). A new ERP, P508, showed the greatest topographic differentiation, but no FB main effect. The following ERPs may reflect different sources: PSW versus P3b; P2/P3a versus LPSW; R-FB versus D-FB P2/P3a; R-FB versus D-FB LPSW; and P508 versus all others. LPSW was greater to simple than complex task difficulty; while the "P508" factor score trended towards being greater for complex than simple. ERP interpretation is in terms of stimulus recognition classification, comparative evaluation and development elaboration of mental models. Rule-learning D-FB exceeded PM accuracy R-FB for all ERPs but P2/P3a. Strongly implicated in these differences are preparatory acts in the former task as reflected by the PSW and LPSW.

Event-related potentials in a guessing task: the gleam in the eye effect.

Event-related potentials (ERPs) were recorded from a single subject performing a forced-choice guessing task. On each trial, ERPs were elicited by four, sequential, graphic images; 2 1/2 seconds after the last stimulus was delivered, the subject guessed which of the four images was experimentally (randomly) designated as the target. P200 had greater amplitude over the posterior scalp for stimuli which were guessed by the subject to be targets than for not-guessed stimuli. The amplitude of the P100, N100, and P300 components was unrelated to the subject's guess. A positive displacement evident in the waveforms from about 150-500 ms post-stimulus onset suggested that Slow Wave may have been partially responsible for the observed differences. These results suggest that ERPs may contain predictive information about a subject's subsequent responses in forced-choice guessing tasks. We termed this the "gleam in the eye" effect.

Mutator activity in maize correlates with the presence and expression of the Mu transposable element Mu9.

Mutator is a powerful system for generating new mutants in maize. Mutator activity is attributable to a family of transposable, multicopy Mu elements, but none of the known elements is an autonomous (regulatory) element. This paper reports the discovery of Mu9, a 4942-base-pair Mu element that was cloned after it transposed into the Bronze-2 locus. Like other Mu elements, Mu9 has approximately 215-base-pair terminal inverted repeats and creates a 9-base-pair host sequence duplication upon insertion. A small gene family of elements that cross-hybridize to Mu9 has been found in all maize lines, and one of the other known Mu elements, Mu5, probably arose as a deletion of Mu9. Mu9 has several of the properties expected for the proposed regulator of Mutator activity. (i) The presence of Mu9 parallels the presence of Mutator activity in individuals from a line that genetically segregates for the Mu regulator. (ii) Lines that transmit Mutator to greater than 90% of their progeny have multiple copies of Mu9. (iii) Most maize lines that lack Mutator activity and that are not descended from Mutator lines lack the Mu9 element. (iv) Transcripts that hybridize to Mu9 are abundant in active Mutator lines, but they are absent from lines that have epigenetically lost Mutator activity. These correlations suggest that Mu9 is a candidate for the autonomous Mutator element.

Movement-related potentials during development: a replication and extension of relationships to age, motor control, mental status and IQ.

Three age groups of normals (children, preadolescents, and adults) and a group of mentally retarded adults performed a noncued button press task from which averaged movement-related potentials (MRPs) were derived. MRP wave shapes replicated our previously reported modal waveform types, except for the preadolescents, who showed no modal MRP. The normal groups showed a developmental shift in the prevalence of waveforms with the retarded differing from normal adults. The modal child waveform had a large amplitude positive-negative-positive form; the retarded had a uniphasic positive form; while normal adults showed the usual negative form. Major MRP types among children could not be attributed to differences in sampling or number of the trials averaged, or to trial-to-trial MRP variability. Background EEG activity did not differ in different modal MRP types. Modal child MRPs showed an initial peak positivity at Fz, and a late peak at Cz. The retarded had a sustaining central positivity developed by midepoch. MRP positivity and negativity were related to age, inhibition of extraneous eye movement (EM), and IQ. In children, greater cognitive proficiency is associated with adult-like MRP. The results suggest that positivity, in part, reflects a subject's efforts at inhibiting movement extraneous to the instructed task.

In-vitro antigonococcal activity of rosoxacin (WIN 35213).

The in-vitro activity of rosoxacin against 173 isolates of Neisseria gonorrhoeae, including 17 beta-lactamase-producers, was tested by an agar dilution method. Of the isolates, 167 (including 16 of the beta-lactamase-producers) were inhibited by 0 . 06 mg/l of rosoxacin. The remaining six isolates, one of which produced beta-lactamase and the others were moderately resistant to penicillin, were inhibited by 0 . 12-0 25 mg/l of the compound. There was little correlation between the minimum inhibitory concentrations (MICs) of rosoxacin and penicillin, except for isolates with MICs of penicillin of 0 . 06-1 mg/l, for which correlation was good.