Neuronal NADPH oxidase 2 regulates growth cone guidance downstream of slit2/robo2

NADPH oxidases (Nox) are membrane‐bound multi‐subunit protein complexes producing reactive oxygen species (ROS) that regulate many cellular processes. Emerging evidence suggests that Nox‐derived ROS also control neuronal development and axonal outgrowth. However, whether Nox act downstream of receptors for axonal growth and guidance cues is presently unknown. To answer this question, we cultured retinal ganglion cells (RGCs) derived from zebrafish embryos and exposed these neurons to netrin‐1, slit2, and brain‐derived neurotrophic factor (BDNF). To test the role of Nox in cue‐mediated growth and guidance, we either pharmacologically inhibited Nox or investigated neurons from mutant fish that are deficient in Nox2. We found that slit2‐mediated growth cone collapse, and axonal retraction were eliminated by Nox inhibition. Though we did not see an effect of either BDNF or netrin‐1 on growth rates, growth in the presence of netrin‐1 was reduced by Nox inhibition. Furthermore, attractive and repulsive growth cone turning in response to gradients of BDNF, netrin‐1, and slit2, respectively, were eliminated when Nox was inhibited in vitro. ROS biosensor imaging showed that slit2 treatment increased growth cone hydrogen peroxide levels via mechanisms involving Nox2 activation. We also investigated the possible relationship between Nox2 and slit2/Robo2 signaling in vivo. astray/nox2 double heterozygote larvae exhibited decreased area of tectal innervation as compared to individual heterozygotes, suggesting both Nox2 and Robo2 are required for establishment of retinotectal connections. Our results provide evidence that Nox2 acts downstream of slit2/Robo2 by mediating growth and guidance of developing zebrafish RGC neurons.     

Oxidant and antioxidant status of Turkish marasmic children: A single center study

The aim of this study was to investigate plasma oxidant and antioxidant status in Turkish marasmic children. The study population consisted of 38 marasmic children (group I) and 28 age-matched children (group II) who were apparently well, with weight-forage >80% of the standards in the same region. After overnight fasting, venous blood samples were drawn and immediately transferred to heparinized and normal tubes. Plasma antioxidant potential (AOP), and malondialdehyde (MDA) levels were measured in both groups. The plasma MDA levels were found to be higher in group I than in group II. However, plasma AOP values were lower in group I than in group II. The present study suggests that AOP is reduced due to an impaired antioxidant system in the plasma of malnourished patients. This oxidant stress causes significant peroxidation. Also, the antioxidant defense system of the patients is deteriorated in marasmus.     

The effects of Nigella sativa (Ns), Anthemis hyalina (Ah) and Citrus sinensis (Cs) extracts on the replication of coronavirus and the expression of TRP genes family

Extracts of Anthemis hyalina (Ah), Nigella sativa (Ns) and peels of Citrus sinensis (Cs) have been used as folk medicine to fight antimicrobial diseases. To evaluate the effect of extracts of Ah, Ns and Cs on the replication of coronavirus (CoV) and on the expression of TRP genes during coronavirus infection, HeLa-CEACAM1a (HeLa-epithelial carcinoembryonic antigen-related cell adhesion molecule 1a) cells were inoculated with MHV-A59 (mouse hepatitis virus–A59) at moi of 30. 1/50 dilution of the extracts was found to be the safe active dose. ELISA kits were used to detect the human IL-8 levels. Total RNA was isolated from the infected cells and cDNA was synthesized. Fluidigm Dynamic Array nanofluidic chip 96.96 was used to analyze the mRNA expression of 21 TRP genes and two control genes. Data was analyzed using the BioMark digital array software. Determinations of relative gene expression values were carried out by using the 2^???Ct method (normalized threshold cycle (Ct) value of sample minus normalized Ct value of control). TCID₅₀/ml (tissue culture infectious dose that will produce cytopathic effect in 50 % of the inoculated tissue culture cells) was found for treatments to determine the viral loads. The inflammatory cytokine IL-8 level was found to increase for both 24 and 48 h time points following Ns extract treatment. TRPA1, TRPC4, TRPM6, TRPM7, TRPM8 and TRPV4 were the genes which expression levels changed significantly after Ah, Ns or Cs extract treatments. The virus load decreased when any of the Ah, Ns or Cs extracts was added to the CoV infected cells with Ah extract treatment leading to undetectable virus load for both 6 and 8hpi. Although all the extract treatments had an effect on IL-8 secretion, TRP gene expression and virus load after CoV infection, it was the Ah extract treatment that showed the biggest difference in virus load. Therefore Ah extract is the best candidate in our hands that contains potential treatment molecule(s).     

Screening of common and novel familial mediterranean fever mutations in south-east part of Turkey

Familial mediterranean fever (FMF) is an autosomal recessive autoinflammatory disorder (MIM# 249100), particularly common in populations of Mediterranean extraction. MEFV gene, responsible for FMF, encoding pyrin has recently been mapped to chromosome 16p13.3. In the present study, 3,341 unrelated patients with the suspicion of FMF in south-east part of Turkey between the years 2009 and 2013 were enrolled and genomic sequences of exon 2 and exon 10 of the MEFV gene were scanned for mutations by direct sequencing. We identified 43 different type of mutations and 9 of them were novel. DNA was amplified by PCR and subjected to direct sequencing for the detection of MEFV gene mutations. Among the 3,341 patients, 1,598 (47.8 %) were males and 1,743 (52.1 %) were females. The mutations were heterozygous in 806 (62.3 %), compound heterozygous in 188 (14.5 %), homozygous in 281 (21.8 %) and mutations had complex genotype in 17 (1.32 %) patients. No mutation was detected in 2,051 (61.4 %) patients. The most frequent mutations were M694V, E148Q, M680I(G/C) and V726A. We could not find any significant differences between the two common mutations according to the gender. Molecular diagnosis of MEFV is a useful tool in clinical practice, thus a future study relating to genotype/phenotype correlation of FMF in more and larger group in Turkish population involving the whole MEFV gene mutations is necessary.     

Informed Consent Prior to Coronary Angiography in a Real World Scenario: What Do Patients Remember?

Background

Patients'' informed consent is legally essential before elective invasive cardiac angiography (CA) and successive intervention can be done. It is unknown to what extent patients can remember previous detailed information given by a specially trained doctor in an optimal scenario as compared to standard care.

Methodology/Principal Findings

In this prospective cohort study 150 consecutive in-patients and 50 out-patients were included before elective CA was initiated. The informed consent was provided and documented in in-patients by trained and instructed physicians the day before CA. In contrast, out-patients received standard information by different not trained physicians, who did not know about this investigation. All patients had to sign a form stating that enough information had been given and all questions had been answered sufficiently. One hour before CA an assessment of the patients'' knowledge about CA was performed using a standard point-by-point questionnaire by another independent physician. The supplied information was composed of 12 potential complications, 3 general, 4 periprocedural and 4 procedural aspects.95% of the patients felt that they had been well and sufficiently informed. Less than half of the potential complications could be remembered by the patients and more patients could remember less serious than life-threatening complications (27.9±8.8% vs. 47.1±11.0%; p<0.001). Even obvious complications like local bleeding could not be remembered by 35% of in-patients and 36% of out-patients (p = 0.87). Surprisingly, there were only a few knowledge differences between in- and out-patients.

Conclusions

The knowledge about CA of patients is vague when they give their informed consent. Even structured information given by a specially trained physician did not increase this knowledge.     

Serotonin Receptor 6 Mediates Defective Brain Development in Monoamine Oxidase A-deficient Mouse Embryos

Monoamine oxidases A and B (MAO-A and MAO-B) are enzymes of the outer mitochondrial membrane that metabolize biogenic amines. In the adult central nervous system, MAOs have important functions for neurotransmitter homeostasis. Expression of MAO isoforms has been detected in the developing embryo. However, suppression of MAO-B does not induce developmental alterations. In contrast, targeted inhibition and knockdown of MAO-A expression (E7.5–E10.5) caused structural abnormalities in the brain. Here we explored the molecular mechanisms underlying defective brain development induced by MAO-A knockdown during in vitro embryogenesis. The developmental alterations were paralleled by diminished apoptotic activity in the affected neuronal structures. Moreover, dysfunctional MAO-A expression led to elevated levels of embryonic serotonin (5-hydroxytryptamine (5-HT)), and we found that knockdown of serotonin receptor-6 (5-Htr6) expression or pharmacologic inhibition of 5-Htr6 activity rescued the MAO-A knockdown phenotype and restored apoptotic activity in the developing brain. Our data suggest that excessive 5-Htr6 activation reduces activation of caspase-3 and -9 of the intrinsic apoptotic pathway and enhances expression of antiapoptotic proteins Bcl-2 and Bcl-XL. Moreover, we found that elevated 5-HT levels in MAO-A knockdown embryos coincided with an enhanced activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and a reduction of proliferating cell numbers. In summary, our findings suggest that excessive 5-HT in MAO-A-deficient mouse embryos triggers cellular signaling cascades via 5-Htr6, which suppresses developmental apoptosis in the brain and thus induces developmental retardations.     

Cometabolic Degradation of Trichloroethylene by Single- and Two-Phase Systems

The cometabolic degradation of trichloroethylene (TCE) by Pseudomonas putida F1 (strain ATCC 700007) at different concentrations was studied in single- and two-phase systems using 2-undecanone as the second organic phase. Toluene vapors were used as the primary growth substrate for Pseudomonas putida F1. The effects of the biomass concentration and the phase ratio on the biodegradation process were investigated. The best biomass concentration and the most suitable phase ratio were found to be 0.462 and 0.025 g/L (v_org/v_aq), respectively. In the single-phase system, 36.5 mg/L TCE was degraded completely in 15 hours and only 78% of 55 mg/L TCE was degraded in 27 hours, while in the two-phase system 55 mg/L TCE was degraded completely in 14 hours. The use of the two-phase system not only decreased the biodegradation time of TCE but also prevented the inhibition effect of high concentrations of TCE on the microbial biomass.     

Serum proinflammatory mediators at different periods of therapy in patients with multiple myeloma

Multiple myeloma (MM) is a malignant disease characterized by the clonal proliferation of plasma cells within the bone marrow. Several cytokines have been demonstrated to be involved in the control of growth, progression, and dissemination of MM. We determined serum levels of interleukin-1beta (IL-1beta), soluble interleukin-2 receptor (sIL-2R), interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-alpha), and C-reactive protein (CRP) in 14 newly diagnosed MM patients. The median age of the patients was 63.4 +/- 10.8 years and all of the patients were stage III (classified according to the Durie-Salmon classification). The same parameters were measured in 15 healthy controls. In addition, we also examined the effects of vincristine-adriamycin-dexamethasone (VAD) therapy on the same parameters and mediators as well as the relationship among the parameters in the same patient groups. The serum concentrations of TNF-alpha, IL-1beta, sIL-2R, IL-6, IL-8, and CRP (18.6 +/- 3.7 pg/mL, 10.1 +/- 2.8 pg/mL, 730 +/- 220 U/mL, 11.4 +/- 3.3 pg/mL, 23.9 +/- 8.3 pg/mL, and 49.9 +/- 19.5 mg/dL, resp) were significantly higher in newly diagnosed MM patients than in healthy controls (P < .0001). All of the parameters were found to be significantly reduced after chemotherapy. In conclusion, we found that after the VAD therapy, the level of these cytokines which are thought to play an important role in the pathogenesis of MM was significantly suppressed. This is the first study demonstrating strong impact of VAD treatment on circulating mediators of sIL-2R and IL-8 levels parameters.