SUMMARYPurpose: The role of gap junctions in seizures is an area of intense research. Many groups have reported anticonvulsant effects of gap junction blockade, strengthening the case for a role for gap junctions in ictogenesis. The cerebral cortex is underrepresented in this body of research. We have investigated the effect of gap junction blockade on seizure-like activity in rat and mouse cerebral cortex slices. Methods: Seizure-like activity was induced by perfusing with low-magnesium artificial cerebrospinal fluid. The effect of three gap junction blockers was investigated in rat cortical slices; quinine (200 and 400 lM), quinidine (100 and 200 lM), and carbenoxolone (100 and 200 lM). In addition, the effect of mefloquine was investigated in wild-type mice and connexin36 knockout mice. The data were analyzed for the effect on frequency and amplitude of seizure-like events.Results: Paradoxical excitatory effects on seizurelike activity were observed for all three agents in rat cortical slices. Quinine (200 lM) and carbenoxolone (100 lM) increased both the frequency and amplitude of seizure-like events. Quinidine (100 lM) increased the frequency of events. Higher doses of quinine (400 lM) and carbenoxolone (200 lM) had biphasic excitatory-inhibitory effects. Similar excitatory effects were observed in adult wild-type mouse cortical slices perfused with mefloquine (5 lM or 10 lM), but were absent in slices from connexin36-deficient mice. Discussion: In conclusion, we have shown a paradoxical proseizure effect of pharmacologic gap junction blockade in a cortical model of seizurelike activity. We suggest that this effect is probably due to a disruption of inhibitory interneuron coupling secondary to connexin36 blockade.

Purpose: Specific markers of circulating tumor cells may be informative in managing lung cancer.Because the RE-1silencing transcription factor (REST/NRSF) is a transcriptional repressor that is inactivated in neuroendocrine lung cancer, we identified REST-regulated transcripts (CHGA, CHGB, SCG3,VGF, and PCSK1) for evaluation as biomarkers in peripheral blood. Experimental Design: Transcripts were screened across lung cancer and normal cell lines. Candidates were assessed by reverse transcription-PCR and hybridization of RNA extracted from the peripheral blood of 111 lung cancer patients obtained at clinical presentation and from 27 cancer-free individuals. Results: Expression profiling revealed multiple chromogranin transcripts were readily induced on RESTdepletion, most notably SCG3 was induced >500-fold.The SCG3 transcript was also overexpressed by 12,000-fold in neuroendocrine compared with nonneuroendocrine lung cancer cells. In peripheral blood of lung cancer patients and cancer-free individuals, we found that SCG3 was more tumor-specific and more sensitive than other chromogranin transcripts as a biomarker of circulating tumor cells. Overall, 36% of small cell lung cancer (SCLC) and 16% of non-SCLC patients scored positively for normalized SCG3 transcript. This correlated with worse survival among SCLC patients with limited disease (n = 33; P = 0.022) but not extensive disease (n = 29; P = 0.459). Interestingly, the subcohort of 6 SCLC patients with resistance to platinum/ etoposide chemotherapy all scored positively for peripheral blood SCG3 transcript (P = 0.022). Conclusions: SCG3 mRNA, a component of the REST-dependent neurosecretory transcriptional profile, provides a sensitive prognostic biomarker for noninvasive monitoring of neuroendocrine lung cancer.

Sleigh, Connexin36 knockout mice display increased sensitivity to pentylenetetrazolinduced seizure-like behaviors, Brain Research (2010),

IL-1R antagonist (IL-1Ra) is required for adequate host defense in invasive pneumococcal disease (IPD). The minor allele of an IL1RN gene (C/T) promoter polymorphism (rs4251961) has been shown to be associated with decreased IL-1Ra production in healthy adults. We genotyped 299 children with IPD, and examined 19 IL1RN haplotype-tagging single-nucleotide polymorphisms. Human embryonic kidney HEK293(T) cells were transfected with the promoter reporter plasmid pGL3p containing either allelic variant C (pGL3pCC) or T (pGL3pTT) with or without cotransfection with an expression construct overexpressing the globin transcription factor GATA-1. Plasma IL-1Ra concentrations were significantly higher in nonsurvivors compared with survivors (p < 0.0005), and the C allele of rs4251961 was associated with a significant increase in plasma IL-1Ra concentrations (p = 0.01) during the acute illness of IPD. These findings were validated in a cohort of 276 treatment-naive HIV-infected adults, with borderline significance (p = 0.058). Functional analyses demonstrated that the activity of the promoter constructs containing the T allele increased ~6-fold as compared with basal activity, and that containing the C allele by ~9-fold (p < 0.001) in the presence of GATA-1. Our findings suggest that the IL-1Ra single-nucleotide polymorphism rs4251961 plays a key role in the pathophysiology of IPD and in other human infections.

Stress hormone and sleep differences in a competition versus training setting are yet to be evaluated in elite female team-sport athletes. The aim of the current study was to evaluate salivary cortisol and perceptual stress markers during competition and training and to determine the subsequent effects on sleep indices in elite female athletes. Ten elite female netball athletes (mean ± SD; age: 23 ± 6 years) had their sleep monitored on three occasions; following one netball competition match (MATCH), one netball match simulation session (TRAIN), and one rest day (CONTROL). Perceived stress values and salivary cortisol were collected immediately pre- (17:15 pm) and post-session (19:30 pm), and at 22:00 pm. Sleep monitoring was performed using wrist actigraphy assessing total time in bed, total sleep time (TST), efficiency (SE%), latency, sleep onset time and wake time. Cortisol levels were significantly higher (p < .01) immediately post MATCH compared with TRAIN and CONTROL (mean ± SD; 0.700 ± 0.165, 0.178 ± 0.127 and 0.157 ± 0.178 μg/dL, respectively) and at 22:00 pm (0.155 ± 0.062, 0.077 ± 0.063, and 0.089 ± 0.083 μg/dL, respectively). There was a significant reduction in TST (-118 ± 112 min, p < .01) and SE (-7.7 ± 8.5%, p < .05) following MATCH vs. TRAIN. Salivary cortisol levels were significantly higher, and sleep quantity and quality were significantly reduced, following competition when compared to training and rest days.

Selenium (Se) compounds have demonstrated anticancer properties in both preclinical and clinical studies, with particular promise in combination therapy where the optimal form and dose of selenium has yet to be established. In a phase I randomised double-blinded study, the safety, tolerability and pharmacokinetic (PK) profiles of sodium selenite (SS), Se-methylselenocysteine (MSC) and seleno-l-methionine (SLM) were compared in patients with chronic lymphocytic leukaemia and a cohort of patients with solid malignancies. Twenty-four patients received 400 μg of elemental Se as either SS, MSC or SLM for 8 weeks. None of the Se compounds were associated with any significant toxicities, and the total plasma Se AUC of SLM was markedly raised in comparison to MSC and SS. DNA damage assessment revealed negligible genotoxicity, and some minor reductions in lymphocyte counts were observed. At the dose level used, all three Se compounds are well-tolerated and non-genotoxic. Further analyses of the pharmacodynamic effects of Se on healthy and malignant peripheral blood mononuclear cells will inform the future evaluation of higher doses of these Se compounds. The study is registered under the Australian and New Zealand Clinical Trials Registry No: ACTRN12613000118707.

Aim-To examine mesotheliomas for a possible relation between p53 immunostaining, p53 gene mutation, simian virus 40 (SV40), and asbestos exposure. Methods-ParaYn sections from 11 mesotheliomas were used for p53 immunostaining and also to extract DNA. This was analysed for the presence of mutations in exons 5 to 8 of the p53 gene using a "cold" single strand conformational polymorphism method, together with sequencing. The DNA from the paraYn sections was also used to search for SV40 sequences. A 105 base pair segment at the 3' of the SV40 large T antigen (Tag) was targeted and any PCR amplification products were sequenced to confirm that they were of SV40 origin. EDAX electron microscopic diVerential mineral fibre counts were performed on dried lung tissue at a specialist referral centre. Results-The fibre counts showed that seven of the mesotheliomas were associated with abnormally high asbestos exposure. Of these, two showed p53 immunostaining, none showed p53 gene mutation, and five showed SV40. Of the four other mesotheliomas, three showed p53 immunostaining, one showed a (silent) p53 mutation, and none showed SV40. The diVerence in frequency of SV40 detection was significant at the p < 0.05 level. Conclusions-Immunostaining for the p53 gene was relatively common but p53 mutations were rare in this series. SV40 virus sequence was detected in five of seven asbestos associated mesotheliomas but in none of the non-asbestos-associated mesotheliomas. This suggests there may be a synergistic interaction between asbestos and SV40 in human mesotheliomas. A study with a larger number of cases is needed to investigate these observations further. (J Clin Pathol 1999;52:291-293)

Selenium, a trace element with anticancer properties, can reduce harmful toxicities of chemotherapy and radiotherapy without compromising efficacy. However, the dose-response relationship in normal versus malignant human cells is unclear. We evaluated how methylseleninic acid (MSA) modulates the toxicity and efficacy of chemotherapy and radiation on malignant and non-malignant human mononuclear blood cells in vitro. We specifically investigated its effects on endoplasmic reticulum stress induction, intracellular glutathione concentration, DNA damage and viability of peripheral blood mononuclear cells and THP1 monocytic leukaemia cells in response to radiation, cytosine arabinoside or doxorubicin chemotherapy. MSA, at lower concentrations, induced protective responses in normal cells but cytotoxic effects in malignant cells, alone and in conjunction with chemotherapy or radiation. However, in normal cells higher concentrations of MSA were directly toxic and increased the cytotoxicity of radiation but not chemotherapy. In malignant cells higher MSA concentrations were generally more effective in combination with cancer treatments. Thus, optimal MSA concentrations differed between normal and malignant cells and treatments. This work supports clinical reports that selenium can significantly reduce dose-limiting toxicities of anticancer therapies and potentially improve efficacy of anticancer treatments. The optimal selenium compound and dose is not yet determined.