NFKB1 -94 ins/del ATTG, NFKBIA -826C>T and NFKBIA -881A>G polymorphisms with risk of lung cancer in a Chinese population.To investigate the association of Genotyping of the polymorphisms were performed on 1,436 subjects (718 cases and 718 controls) by using PCR-RFLP technique, followed by DNA sequencing.NFKB1 polymorphism. In contrast, the heterozygous and variantgenotypes of theNFKBIA polymorphisms showed association with increased lung cancer risk . Several genotypic combinations of the three polymorphisms also showed significant association with lung cancer risk. The risk association of NFKB1 polymorphism remained significant when analyses were done according to gender and smoking status (P<0.05). The significance of NFKBIA risk association was not observed when gender-specific analyses were made (P>0.05), while only NFKBIA -881 GG genotype showed significant risk association among smokers when analyzed according to smoking status (P=0.032).We found a significant risk reduction associated with heterozygous ins/del  and variant del/del  genotypes of the NFKB1 and NFKBIAgenes were associated with risk of lung cancer.Polymorphisms in  Lung cancer is the most common type of cancer worldwide and represents a leading cause of cancer-related mortality.8NFKB1 and NFKBIA genes respectively, and it has been shown that disrupted expressions of these genes can result in carcinogenesis.13The nuclear factor-kappa B (NF-κB) family constitutes a group of transcription factors which serve as important mediators in inflammatory responses. In addition to inflammatory responses, a wide range of signal transduction processes converge on the NF-κB pathway, including cell proliferation, apoptosis, angiogenesis and many others.NFKB1 -94 ins/del ATTG polymorphism and NFKBIA -826C>T and -881A>G polymorphisms with the risk of lung cancer in a Chinese population.Functional polymorphisms within the promoter region of these genes could potentially influence the levels of the proteins encoded. These functional polymorphisms may therefore contribute to the inter individual differences in lung cancer risk. To test our hypothesis, we conducted a case-control study to investigate the association of 718 lung cancer patients and 718controls participated in this retrospective case-control study. The participants were recruited between September 2011 andAugust 2014 fromthe Zhengzhou Central Hospital Affiliated to Zhengzhou University. Controls were cancer-free individuals randomly selected from a cancer screening program, and were matched (in frequency) to cases by age, sex and smoking behavior. The study was approved by the Research Review and Ethics Board of Zhengzhou Central Hospital Affiliated to Zhengzhou University. Written informed consent was obtained from all the participants prior to the study.Genotyping was performed by using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method described15Differences in age, smoking status and gender between cases and controls were evaluated using the chi-square test. The association between the polymorphisms and lung cancer risk was determined using the logistic regression method to assess the odds ratio (OR). P values less than 0.05 were considered statistically significant.NFKB1 and NFKBIA polymorphisms and lung cancer risk is shown in NFKB1 polymorphism were found to be significantly associated with the reduced risk of lung cancer. On the other hand, the heterozygous and homozygous variant genotypes of the NFKBIA polymorphisms appeared to be significantly associated with increased lung cancer risk.The association between NFKB1 -94 ins/del ATTG polymorphism and NFKBIA -826C>T polymorphism with lung cancer risk is shown in The association of combinations of NFKB1 -94 ins/del ATTG polymorphism and NFKBIA -881A>G polymorphism and lung cancer risk is shown in Association between combinations of NFKBIA -826C>T polymorphism and NFKBIA -881A>G polymorphism and their association with lung cancer risk is shown in The combinations of NFKB1 polymorphism and lung cancer risk according to gender and smoking status is shown in The stratified association between the NFKBIA -826C>T polymorphism and lung cancer risk according to sex and smoking status is shown in The association between NFKBIA -881A>G polymorphism and the risk of lung cancer according to sex and smoking status is shown in NFKBIA -826C>T polymorphism, no association was observed for the heterozygous and variant genotypes and lung cancer risk in both males and females after stratification by sex. However, after stratification by smoking status, significant association was observed only for the variant genotype with lung cancer risk among smokers(P=0.032).The association between NFKB1 -94 ins/del ATTG, NFKBIA -826C>T and NFKBIA -881A>G polymorphisms have been described. These functional polymorphisms have been shown to affect the level of the proteins produced, with the deletion allele, T allele and G allele of the three polymorphisms are associated with the reduced production of the respective proteins products.17Given the important role of NF-κB and IκB in diverse biological pathways, genetic variations within key genes of the NF-κB pathway could obstruct the normal functioning of the protein products which in turn influences the risk of cancers. The NFKB1 -94 ins/del ATTG polymorphism could significantly reduce lung cancerrisk. Our results are in disagreement with those reported by Huang et al.NFKB1 polymorphism and lung cancer risk in a Chinese population. However, our results concur with Huo et al.We have showed that the variant del genotype of the NFKBIA -826C>T and -881A>G polymorphisms could increase lung cancerrisk. Similar to the NFKB1 -94 ins/del ATTG polymorphism, our results are contrary to the results of Huang et al.NFKBIA -881GG variant genotype among smokers. This was in partial agreement with Umar et al.NFKBIA -826C>T and -881A>G polymorphisms increased the risk of oral cancer among Chinese. The disagreements among the findings reported in the literature could be due to the different backgrounds  of the study subjects recruited, as well as the properness of subject matching.We have also showed in the current study that the variant genotypes of The strengths of this study are: (i) it is the first study which establishes an association between the polymorphism and lung cancer risk in China, and (ii) it was conducted in a relatively properly-matched population. However, the limitation is that only three SNPs were analyzed.NFKB1 -94 ins/del ATTG polymorphism, NFKBIA -826C>T polymorphism and NFKBIA -881A>G polymorphism could influence the risk of lung cancer in China.We have showed that the variant alleles of


This article contains data related to the research article Auzias et al. (2015)  Specifications Table•The sulcal pits, defined as the deepest points lying in each fold of the cortical surface, are gathering growing interest in the literature•The data presented here consist of well characterized subjects with high-quality cortical meshes, divided in two subgroups carefully matched according to several parameters, with the depth map and sulcal pits resulting from the extraction procedure proposed in•These data can be used for the benchmarking of new sulcal pit extraction algorithms, with a quantitative comparison with[1], and for the assessment of the consistency of the sulcal pits across two well matched populations.Value of the data1This data article contains data related to the research article 2www.oasis-brains.org) A particular attention was drawn to gather a large set of high-quality cortical surface tessellations from a well-controlled population of healthy individuals. For that purpose, 137 right-handed subjects were selected from the Open Access Series of Imaging Studies (OASIS) database (http://surfer.nmr.mgh.harvard.edu), in order to extract the inner cortical surface mesh (white), surface area and intracranial volume, and obtain spherical inter-individual correspondences through the Freesurfer sphere.reg mesh http://brainvisa.info/). We used this depth map to extract the sulcal pits of each fold using the technique detailed in For each cortical mesh, we then computed the Depth Potential Function (DPF) introduced in


Repetitive transcranial magnetic stimulation (rTMS) has been proposed as a tinnitus treatment option. Promising results have been obtained by consecutive stimulation of lateral frontal and auditory brain regions. We investigated a combined stimulation paradigm targeting the anterior cingulate cortex (ACC) with double cone coil rTMS, followed by stimulation of the temporo-parietal junction area with a figure-of-eight coil. The study was conducted as a randomized, double-blind pilot trial in 40 patients suffering from chronic tinnitus. We compared mediofrontal stimulation with double-cone-coil,  followed by left temporo-parietal stimulation with figure-of-eight-coil  to left dorsolateral-prefrontal-cortex stimulation with figure-of-eight-coil  followed by temporo-parietal stimulation with figure-of-eight-coil . The stimulation was feasible with comparable dropout rates in both study arms; no severe adverse events were registered. Responder rates did not differ in both study arms. There was a significant main effect of time for the change in the TQ score, but no significant time x group interaction. This pilot study demonstrated the feasibility of combined mediofrontal/temporoparietal-rTMS-stimulation with double cone coil in tinnitus patients but failed to show better outcome compared to an actively rTMS treated control group. Chronic Tinnitus Subjective tinnitus is characterized by the perception of sound in the absence of a corresponding sound source37Transcranial magnetic stimulation (TMS) is a non-invasive tool for inducing electric currents in the brain12131516171819202122However, rTMS treatment results are currently burdened by only moderate improvement and high inter-individual variability12132728293031333435361740Typically, the direct effect of the magnetic field produced by an rTMS coil remains limited to superficial brain areas and does not reach subcortical brain structures44451741The newly developed double cone coil (DCC) exerts an expanded penetration depth based on its angled geometry. A positron emission tomography study revealed that mediofrontal TMS using a double-cone coil is apt to modulate deeper located brain areas such as the dorsal and subgenual ACCanterior-cingulate-cortex-stimulation using a double cone coil (=ACDC-stimulation) has been investigated as an innovative approach for the treatment of affective disorders very recently and has given promising results so far55As the ACC has been shown to be critically involved in depression53et al. applying double-cone-coil-rTMS to the dorsal frontal cortex of 78 patientsFirst clinical experience in tinnitus patients has been gained by Vanneste anterior-cingulate-cortex-rTMS using a double-cone-coil in the treatment of patients with chronic tinnitus (TiCDC).Based on these previous results and the notion, that, in view of the involvement of both auditory and non-auditory brain regions in tinnitus27282930311740The study was conducted in a randomized, double-blind, parallel-group design with an active control group. Patients were assessed at screening, baseline, end of treatment (week 2), and two follow-up-visits taking place in week 4 and 12. We compared two combined study protocols consisting of medial frontal stimulation with double cone coil (study arm 1) vs. conventional prefrontal left DLPFC-stimulation (study arm 2/control group) both followed by stimulation of the left temporo-parietal junction area. Both study arms consisted of a total of 4000 stimuli per day. Detailed information on the study protocol and the applied stimulation parameters is provided in The study was approved by the Ethics Committee at the University of Regensburg. All participants gave written informed consent after a comprehensive explanation of the procedures. All study methods and procedures were carried out in accordance with the approved guidelines. The study had been previously registered with clinicaltrials.gov .Inclusion criterion was subjective tinnitus with duration of more than six months. Exclusion criteria comprise objective tinnitus (with a treatable cause), start of other treatments for tinnitus 3 months before study enrolment, presence of clinically relevant psychiatric comorbidities or unstable medical conditions, history or evidence of significant brain malformation or neoplasm, history of head injuries, cerebral vascular events, presence of irremovable metal objects in and around body, pregnancy, alcohol abuse or intake of illicit substances, and history of prior TMS treatment.Patients were recruited for participation in the study after presentation in the outpatient clinic of the Interdisciplinary Tinnitus Centre at the University of Regensburg, Regensburg, Germany.et al.65Primary objective was the responder rate at week 2 evaluation  of the left DLPFC was applied at 10 Hz frequency  with a figure-of-eight (=butterfly) coil  followed by 1 Hz stimulation of the left temporoparietal junction area6868All data are displayed as mean ± standard deviation. In case of missing data the last observation was carried forward, dropout participants were excluded from analysis. Responder rates were compared by chi-square tests of independence. For secondary outcome measures we calculated analyses of variance (ANOVA) with the within-subjects factor time  and the between-subjects factor group (“TiCDC” vs. “standard rTMS” protocol). CGI scores were compared using chi-square tests. All statistical tests were conducted two-tailed, unadjusted for multiple comparisons due to the pilot study character of the trial, and a value of p < 0.05 was assumed as statistically significant. The sample size was set in a range similar to prior actively controlled pilot trials regarding rTMS-based treatment of chronic tinnitus e.g.70, and wForty patients were randomized for participation in the study. One participant aborted the stimulation due to headache (TiCDC-participant after session 7). Three study subjects withdrew their consent during the stimulation period due to deterioration of their tinnitus . These four subjects were not included in the per-protocol analysis resulting in an analytical sample of 36 patients. The comparison of participants of both treatment groups revealed no differences in the clinical and demographic baseline characteristics. Detailed information is provided in No severe adverse events occurred during the course of the study. The incidence of adverse events was comparable in both study arms: six patients complained about headache (3 TiCDC/3 standard rTMS control group/1 dropout (TiCDC)), a deterioration of tinnitus was complained in one TiCDC-participant (leading to the second dropout of TiCDC-group) and two control-group-participants (2 dropouts), dizziness or a deterioration of pre-existing dizziness was reported by 1 patient of each group. Other adverse events included local pain (TiCDC: 1 report) and feelings of ear pressure on both sides (control group: 1 report).As a first step of the data analysis we aimed to compare the number of responders and non-responders in both treatment arms. The primary outcome criterion (responder rate week 2 vs. baseline) turned out non-significant see  leading 2(1) = 1.870, p = 0.171). With regard to the ANOVAs comparing secondary outcomes such as change in TQ, THI, TBF12, MDI, and WHOQoL-questionnaires, the main effect of time turned out significant for the change in the TQ score  and marginally significant for the THI with effect sizes being moderate to high (for details see left part of Response rates at week 12 (=end of study) turned out non-significant . Vanneste and colleagues reported the successful treatment of a patient suffering from a medication-resistant chronic depression by 10 sessions of ACDC-stimulation applying the same stimulation parameters as used in our pilot study 68Despite encouraging results of previous applications of mediofrontal double-cone-coil-rTMS in patients suffering from depression, the present study failed to confirm the alternative hypothesis of superiority as part of a combined treatment setting in subjects suffering from chronic tinnitus. This might be due to limited power of the present study, which would require comparisons to placebo-controlled study groups in larger samples. Apart from that, it is very likely that our understanding of stimulation settings is yet too mechanistic and in addition to the effect in the stimulated cortical area the impact of the stimulation on network activity and connectivity has to be taken into account in more detailHowever, despite these limitations of the present study and the negative results our data provide novel insights in the feasibility of double-cone-coil rTMS targeting the ACC in patients suffering from chronic tinnitus. Moreover, they allocate an estimation of clinical effects, thus allowing for further methodological adjustments and enabling power calculations for upcoming studies. In summary our pilot data confirm the potential of TiCDC-stimulation as a non-invasive, safe and well tolerated method of brain stimulation in the treatment of chronic tinnitus, but failed to demonstrate superiority compared to the clinical effectiveness of a combinatory rTMS study group addressing both the left DLPFC and the temporo-parietal junction area.How to cite this article: Kreuzer, P. M. et al. Combined rTMS treatment targeting the Anterior Cingulate and the Temporal Cortex for the Treatment of Chronic Tinnitus. Sci. Rep.5, 18028; doi: 10.1038/srep18028 (2015).


By blocking the stability and translation of protein-coding mRNAs, microRNAs impact most biological systems in health and disease, including the development and the function of the immune system . A recenT cells receive constant inputs from their surroundings - the challenge is that T cells must tolerate our own healthy cells and harmless commensals, yet mount vigorous immune responses against potentially harmful pathogens, transformed cells and other threats. If this discrimination fails, T cells may mistakenly attack healthy cells and cause autoimmunity, or they may fail to protect us from harm. A tried and tested strategy is that T cells tolerate signals that exclusively engage their T cell receptor (TCR), but go after signals that simultaneously engage the TCR and a set of co-stimulatory receptors that indicate the presence of tissue damage or inflammation . In isolTo probe the role of microRNAs in T cell activation, Marcais et al.  generateMtor allele and one defective Rictor allele into Dicer-deficient T cells. As a result, Mtor and Rictor proteins were derived from a single Mtor and a single Rictor allele. This approach compensated for the lack of post-transcriptional control, and restored near-normal levels of Mtor and Rictor proteins to microRNA-deficient T cells. Critically, reversing the overexpression of Mtor and Rictor also reduced the ability of microRNA-deficient T cells to produce IL-2 in the absence of co-stimulation [Detailed analysis of the signaling machinery in microRNA-deficient T cells revealed several lesions, including the abnormally strong and prolonged activation of mTORC1 and mTORC2 (‘mTOR’ stands for ‘mechanistic target of rapamycin’ and ‘C’ stands for ‘complex’). mTOR complexes integrate signaling pathways to regulate a broad range of cellular functions. A number of mTOR complex components were already known targets of microRNA regulation in normal  and in cIn this context it is of note that recent data suggest an additional role for microRNAs in developing thymocytes, the precursors of mature T cells . Here, m


A high P-wave duration and dispersion (Pd) have been reported to be a prognostic factor for the occurrence of paroxysmal atrial fibrillation (PAF), a condition linked to obstructive sleep apnea (OSA). We tested the hypothesis of whether a short-term increase of P-wave duration and Pd can be induced by respiratory manoeuvres simulating OSA in healthy subjects and in patients with PAF.12-lead-electrocardiography (ECG) was recorded continuously in 24 healthy subjects and 33 patients with PAF, while simulating obstructive apnea , obstructive hypopnea , central apnea (AP), and during normal breathing (BL) in randomized order. The P-wave duration and Pd was calculated by using dedicated software for ECG-analysis.P-wave duration and Pd significantly increased during MM and ITH compared to BL in all subjects . In MM, the increase was larger in healthy subjects when compared to patients with PAF (p<0.05).Intrathoracic pressure swings through simulated obstructive sleep apnea increase P-wave duration and Pd in healthy subjects and in patients with PAF. Our findings imply that intrathoracic pressure swings prolong the intra-atrial and inter-atrial conduction time and therefore may represent an independent trigger factor for the development for PAF. Obstructive sleep apnea (OSA) is highly prevalent, with approximately 20% of males and 10% of females in Western countries affected by asymptomatic OSA. Pathophysiologically, the disease is characterized by repetitive partial or complete obstruction of the pharynx during sleep. Despite increasing breathing efforts the upper airway collapse results in episodes of obstructive hypopneas or apneas resulting in negative intrathoracic pressure, intermittent hypoxia, autonomic nervous dysfunction, and consequently adverse effects for sleep-architecture and health.A large body of evidence suggests that OSA is associated with cardiac arrhythmias such as paroxysmal atrial fibrillation (PAF) with some studies proposing a causal relationship. ComparedInhomogeneous atrial depolarization secondary to insults such as atrial distension or autonomic nervous dysfunction can be quantified with a surface electrocardiography (ECG)-derived P-wave analysis. A high PCurrently, it is unclear whether hypopneas and apneas have an instant effect on P-wave duration and Pd and whether these pathophysiological events might subsequently alter atrial electrophysiology. Thus we aimed to study this important pathophysiological link and investigate the acute effects of simulated OSA on p-wave indices in healthy subjects and patients with PAF.The study was approved by the “Cantonal Ethics Committee Zurich”, Switzerland (KEK-ZH-Nr. 2012–0310 / EK 1672) and written informed consent was obtained from all participants.57 patients  aged between 18 and 75 years were recruited at the study site . The subjects overlap with two previously published studies and sample size estimation was adopted accordingly. In shortThe protocol for breathing manoeuvres simulating hypopneas and apneas (and therefore OSA) which was applied in this study has been described previously.,6,15,16 Digital 12-lead-ECG was recorded continuously in awake 24 healthy subjects and 33 patients with PAF in supine position . Room temperature and lighting were set at the same level for all measurements and all subjects remained in supine position five minutes before undergoing assessments. The beginning of the P-wave was defined as the point where the first atrial deflection crossed the isoelectric line and the end of the P-wave was defined as the point where the atrial deflection returned to the isoelectric line. The maximum and minimum P-wave durations in all 12 ECG-leads were retrieved and the most extreme ones were used to calculate the Pd (Pd = maximum P-wave duration − minimum P-wave duration). For compContinuous data were summarized as mean±SD and categorical data summarized using percentages. The primary outcomes of this study were the mean P-wave duration and Pd during each breathing manoeuvre. Mixed-effects linear regression with patient-level random-intercepts was used to compare the effect of each breathing manoeuvre on each outcome compared to BL. All analyses were adjusted for manoeuvre, heart rate, BMI, age, sex, and PAF status. An interaction between manoeuvre and PAF status was added to each model to compare the effects of each manoeuvre in patients with and without PAF. Mixed-effects linear regression was also used to assess the effect of each manoeuvre on Pd and P-wave duration measured in a single lead over time by including an interaction between manoeuvre and sector.In order to make sure our measurements were reproducible two blinded investigators each measured a total of 180 P-waves from 20 randomly chosen sectors in which all breathing manoeuvres were performed. Inter-observer agreement was assessed using the intra-class-correlation coefficient and a Bland-Altman plot. The validity of Pd  was checked by assessing its correlation with the standard deviation  of all corresponding P-wave measurements. A two-sided p-value less than 0.05 was considered to be statistically significant for all tests. All statistical analyses were performed with STATA version 14 .24 healthy subjects and 33 patients with PAF were included in this study. Clinical characteristics of the study population are presented in Overall results for Pd and P-wave duration are shown in None of the healthy subjects reported usage of cardio-active medication. Patients with PAF reported use of beta-blocker (82%), other blood pressure lowering agents (57%), and anti-arrhythmic agents (27%). During normal breathing (BL), patients with PAF did not differ from healthy subjects in P-wave duration (p = 1.00) and Pd (p = 0.55) after adjustment for heart rate, age, BMI, and sex. Despite no differences in BL, non-AF individuals were more susceptible to P-wave duration increases (p = 0.037) and Pd-increases (p = 0.009) during the MM than patients with PAF. P-wave durations and Pd for AF and non-AF participants during each manoeuvre are presented in The intra class correlation coefficient was 0.96 (95% CI 0.95–0.98) and the 95% limits of agreement were -4.49 ms / 5.10 ms showing very high inter-observer agreement in P-wave measurements. For further information see This study is, to our knowledge, the first to investigate the acute effects of simulated apnea and hypopnea on P-wave duration and Pd, markers of atrial conduction delay and inhomogeneous atrial depolarization. This study provides data to better understand the pathophysiological mechanisms that may be involved in patients with OSA and the onset of PAF. The presented data demonstrates how simulated obstructive hypopnea (ITH) and apnea (MM) increases P-wave duration and Pd and thus might have a direct effect on arrhythmogenesis. Our findings that P-wave duration and Pd are increased in patients with PAF are in line with several other studies in literature.The findings of this study may have some important implications for patients at risk for PAF. A large body of evidence suggests that OSA is linked to PAF via mechanisms like elevated blood pressure and intermittent hypoxia among others. This stuRecent research suggests that the success rate of PAF treatment (i.e. pulmonary vein isolation) is dependent on the diagnosis of OSA. Meta-analyses have shown that patients with OSA have significantly greater AF recurrence rates after pulmonary vein isolation and CPAP-treatment can reverse this excess risk. ECG-deriGenerally the mechanisms leading to the pathogenesis of PAF remain complex and a matter of debate. It is thought that atrial distortion and atrial enlargement are central to the development of PAF. In our pAccording to catheter measurements, approximately 85% of the airway pressure drop is transmitted into the atria. The physThe use of digital measurements, the high inter-observer agreement between blinded investigators, and the interchangeability between Pd and the SD of the measurements generate high confidence in the validity of the data. Compared with other studies in this field, the present one scores above average in these parametes.Given the design of this pathophysiological study it is important to emphasize that artificially induced changes in breathing patterns were evaluated which could limit the clinical relevance of the data. However, the physiology of the breathing manoeuvres in our study were validated using esophageal and aortic catheters the authIn conclusion this study provides evidence that breathing manoeuvres simulating obstructive sleep apnea promote an immediate increase of P-wave duration and Pd and provide a potential explanation for the increased risk of PAF in patients with OSA. Our findings imply that intrathoracic pressure swings prolong the intra-atrial conduction time and therefore could be an independent trigger factor for the development for PAF.S1 Appendix(PDF)Click here for additional data file.


Diverse ion channels and their dynamics endow single neurons with complex biophysical properties. These properties determine the heterogeneity of cell types that make up the brain, as constituents of neural circuits tuned to perform highly specific computations. How do biophysical properties of single neurons impact network function? We study a set of biophysical properties that emerge in cortical neurons during the first week of development, eventually allowing these neurons to adaptively scale the gain of their response to the amplitude of the fluctuations they encounter. During the same time period, these same neurons participate in large-scale waves of spontaneously generated electrical activity. We investigate the potential role of experimentally observed changes in intrinsic neuronal properties in determining the ability of cortical networks to propagate waves of activity. We show that such changes can strongly affect the ability of multi-layered feedforward networks to represent and transmit information on multiple timescales. With properties modeled on those observed at early stages of development, neurons are relatively insensitive to rapid fluctuations and tend to fire synchronously in response to wave-like events of large amplitude. Following developmental changes in voltage-dependent conductances, these same neurons become efficient encoders of fast input fluctuations over few layers, but lose the ability to transmit slower, population-wide input variations across many layers. Depending on the neurons' intrinsic properties, noise plays different roles in modulating neuronal input-output curves, which can dramatically impact network transmission. The developmental change in intrinsic properties supports a transformation of a networks function from the propagation of network-wide information to one in which computations are scaled to local activity. This work underscores the significance of simple changes in conductance parameters in governing how neurons represent and propagate information, and suggests a role for background synaptic noise in switching the mode of information transmission. Differences in ion channel composition endow different neuronal types with distinct computational properties. Understanding how these biophysical differences affect network-level computation is an important frontier. We focus on a set of biophysical properties, experimentally observed in developing cortical neurons, that allow these neurons to efficiently encode their inputs despite time-varying changes in the statistical context. Large-scale propagating waves are autonomously generated by the developing brain even before the onset of sensory experience. Using multi-layered feedforward networks, we examine how changes in intrinsic properties can lead to changes in the network's ability to represent and transmit information on multiple timescales. We demonstrate that measured changes in the computational properties of immature single neurons enable the propagation of slow-varying wave-like inputs. In contrast, neurons with more mature properties are more sensitive to fast fluctuations, which modulate the slow-varying information. While slow events are transmitted with high fidelity in initial network layers, noise degrades transmission in downstream network layers. Our results show how short-term adaptation and modulation of the neurons' input-output firing curves by background synaptic noise determine the ability of neural networks to transmit information on multiple timescales. Gain scaling refers to the ability of neurons to scale the gain of their responses when stimulated with currents of different amplitudes. A common property of neural systems, gain scaling adjusts the system's response to the size of the input relative to the input's standard deviation et al.Pacemaker neurons located in the ventrolateral (piriform) cortex initiate spontaneous waves that continue to propagate dorsally across the neocortex Along with the emergence of gain scaling during the first week of neural development, single neurons in the developing cortex participate in large-scale spontaneously generated activity which travels across different regions in the form of waves A simple model for propagating activity, like that observed during spontaneous waves, is a feedforward network in which activity is carried from one population, or layer, of neurons to the next without affecting previous layers We find that networks built from neuron types with different gain-scaling ability propagate information in strikingly different ways. Networks of gain-scaling (GS) neurons convey a large amount of fast-varying information from neuron to neuron, and transmit slow-varying information at the population level, but only across a few layers in the network; over multiple layers the slow-varying information disappears. In contrast, nongain-scaling (NGS) neurons are worse at processing fast-varying information at the single neuron level; however, subsequent network layers transmit slow-varying signals faithfully, reproducing wave-like behavior. We qualitatively explain these results in terms of the differences in the noise-modulated frequency-current curves of the neuron types through a mean field approach: this approach allows us to characterize how the mean firing rate of a neuronal population in a given layer depends on the firing rate of the neuronal population in the previous layer through the mean synaptic currents exchanged between the two layers. Our results suggest that the experimentally observed changes in intrinsic properties may contribute to the transition from spontaneous wave propagation in developing cortex to sensitivity to local input fluctuations in more mature networks, priming cortical networks to become capable of processing functionally relevant stimuli.linear/nonlinear (LN) models (see below). Before becoming efficient encoders of fast stimulus fluctuations, the neurons participate in network-wide activity events that propagate along stereotypical directions, known as spontaneous cortical waves Single cortical neurons acquire the ability to scale the gain of their responses in the first week of development, as shown in cortical slice experiments We have shown that in cortical neurons in brain slices, developmental increases in the maximal sodium  model by the stimulus standard deviation rapid fluctuations, while fixing the mean (DC) of the input current. The LN model We first characterized neuronal responses of conductance-based model neurons using methods previously applied to experimentally recorded neurons driven with white noise. The neuron's mature neurons have a common input-output curve with respect to the normalized stimulus  current input is quantified by the noise ,  . One canLth layer, line attractor: there appears an entire line of stable fixed points  . In thisnd 30 pA , at whicodulated . As the As the conductance ratio ecreases . Indeed,C values . This imThis analysis highlights the origins for the differential modulability of firing rate in NGS and GS neurons. Although the model reduction sacrifices some of the accuracy of the original model, it retains the essential features of action potential generation: the sudden rise of the action potential which turns on a positive inward sodium current, and its termination by a slower decrease in membrane potential which shuts off the sodium current and initiates a positive outward potassium current hyperpolarizing the cell. Although simpler neuron models  µm2 and µm2 for the maximal conductances of the GS neurons, so that their ratio was µm2 and µm2 for the maximal conductances of the NGS neurons, so that their ratio was where The rate coefficients, The equations were numerically solved using a first-order Euler method with an integration time step of For spike-triggered characterization we injected Gaussian noise current, where Intrinsic computation in these neuron types was previously characterized in experiments and model neurons where gain-scaling, because scaling the stimulus by i.e.nongain-scaling, because nonlinearities in the LN model vary with different values of the standard deviation when the stimulus is scaled by We refer to the neurons with aled by  , blue. TWe considered a feedforward network architecture with  neurons . We conswhere Instead of where where There were no recurrent connections among the neurons. Each neuron in the network also received an independent noise input with mean 0 and standard deviation lopment  , rather pment   . Since aFrom the When computing The input-output relationship plotted in Furthermore, we note that when predicting the propagation of firing rates across subsequent layers in this mean field analysis, the We first measured information transmission in the network about slow variations in the input . The mutwhere the signal-to-noise ratio can be written asAssuming Gaussian probability distributions, the noise isThis quantity computes the information between stimulus and response by taking into account how similar the response (reconstructed stimulus) is to the original stimulus. Due to the different firing rates evoked in the different networks, when computing the information we normalized the reconstructed stimulus (the PSTH) to have zero mean and unit variance.To quantify the information about fast fluctuations as a function of the mean and  neurons , we usedWhen examining the fidelity of firing rate transfer in networks composed of the two neuron types, we wanted a measure of how distinguishable is a discrete set of output firing rates in each layer given a set of input currents in the first layer see  layer 1.d noise  . We obtawhere To reduce the full conductance-based model (Eq. 2) that depends on four variables, whereand where To study the dynamics of this system in Figure S1µm2 and µm2).Information transmission through networks with gain-scaling neurons  A. PSTHs from each layer in two networks with different connectivity: left, weak synaptic strength (EPS)Click here for additional data file.Figure S2Firing rate transmission for different connection probability and synaptic strength. Top: The µm2 and µm2) and nongain-scaling networks with µm2 and µm2) with a noise level of (EPS)Click here for additional data file.Figure S3Firing rate transmission for a range of noise amplitudes. Top: The A. weak vs B. strong-connectivity, C. gain-scaling networks with D. nongain-scaling networks with (EPS)Click here for additional data file.


Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak (BLS) in rice, a serious bacterial disease of rice in Asia and parts of Africa. The virulence mechanisms of Xoc are not entirely clear and control measures for BLS are poorly developed. The solo LuxR proteins are widespread and shown to be involved in virulence in some plant associated bacteria (PAB). Here, we have cloned and characterized a PAB LuxR solo from Xoc, named as XocR. Mutation of xocR almost completely impaired the virulence ability of Xoc on host rice, but did not alter the ability to trigger HR  on non-host (plant) tobacco, suggesting the diversity of function of xocR in host and non-host plants. We also provide evidence to show that xocR is involved in the regulation of growth-independent cell motility in response to a yet-to-be-identified rice signal, as mutation of xocR impaired cell swimming motility of wild-type Rs105 in the presence but not absence of rice macerate. We further found that xocR regulated the transcription of two characterized virulence-associated genes (recN and trpE) in the presence of rice macerate. The promoter regions of recN and trpE possessed a potential binding motif (an imperfect pip box-like element) of XocR, raising the possibility that XocR might directly bind the promoter regions of these two genes to regulate their transcriptional activity. Our studies add a new member of PAB LuxR solos and also provide new insights into the role of PAB LuxR solo in the virulence of Xanthomonas species. N-acyl homoserine lactones (AHLs) are a major class of signal molecules. This signal is composed of a homoserine lactone ring carrying C4-C18 acyl chains and is thought to function as intraspecies communication  is a cell-density dependent cell-cell communication system that relies on small chemical signal molecules to control bacterial behavior and coordinate gene expression in a cell-density dependent manner  or to exogenous AHLs produced by neighboring bacteria, such as the SdiA from Salmonella enterica and E. coli to regulate target genes ; these PAB LuxR solos do not bind AHLs, but respond to yet unidentified plant signal(s)/compound(s)  is an important disease of rice in Asia and parts of Africa  caused by Xoc strains were grown at 28°C in nutrient broth (NB) medium  or on nutrient agar (NA). Medium containing macerated rice material was prepared as described previously  medium or on LB agar plates. When required, antibiotics were added to the medium at the following final concentrations: 50 μg/mL kanamycin (Km) for E. coli and Xoc, 10 μg/mL gentamicin (Gm) for E. coli and Xoc, and 100 μg/mL rifampicin (Rif) for Xoc.The bacterial strains and plasmids used in this study are listed in Table Xoc wild-type Rs105 was used as the parental strain to generate the in-frame deletion mutant via allelic homologous recombination. In-frame deletion of xocR was performed as described previously  using the primer pairs xocR-1F/xocR-1R and xocR-2F/xocR-2R (Table xocR-1 fragment (digested with BamHI and HindIII) and the xocR-2 fragment (digested with HindIII and XbaI) were ligated into BamHI/XbaI-digested pK18mobsacB  was selected on NA plates with Rif and Gm, and verified by PCR method. This final complemented strain, named as ΔxocR (xocR) was used for further study.The generation of complemented strains was performed as described previously . Then, cells were pelleted by centrifugation at 3099 × g and suspended in an equal volume of sterilized ddH2O. The final cell suspension was inoculated into the leaves of 2-week-old rice plants  using needless syringe. Water-soaking symptoms were measured 7 days after inoculation. Twenty-five leaves were inoculated for each Xoc strain in each treatment. The same experiment was performed three times.The pathogenicity testing and determination of bacterial load assays were performed as described previously . Cells were pelleted by centrifugation at 3099 × g and suspended in different volume of sterilized ddH2O resulting at three different concentrations of cell suspension . These three cell suspensions were used for HR tests. Briefly, Xoc strains were infiltrated into the leaves of greenhouse-grown tobacco (Nicotiana tabacum L. cv. Samsun), and the results were observed after 48 hours of infiltration. Four leaves were inoculated for each Xoc strain in each treatment. The same experiment was performed three times.The hypersensitive response assay was performed as described previously  and NB with rice macerate medium. In brief, Xoc strains were pre-incubated in NB broth at 28°C with shaking at 200 rpm, until the growth was reached to OD600 of 1.0. Cells were pelleted by centrifugation at 3099 × g and suspended in an equal volume of sterilized water. Then, 1 mL of cell suspension was inoculated into 100 mL of two testing media. All inoculation broths were grown at 28°C with shaking at 200 rpm and the OD600value was determined every 4 h until bacterial growth reached to the stationary stage. The experiments were performed three times, and each involves three replicates.Bacterial growth was monitored as described previously  broth at 28°C with shaking at 200 rpm, until the value of OD600 reached 1.0, and then 3 μl of each strain was inoculated onto the surface of the motility plates, including NA containing 0.3% soft agar with and without of rice macerate. Cell motility was examined at 72 h after incubation at 28°C. Five independent motility assays were performed for each strain.Swimming motility assay was performed as described previously  skim milk powder. After 48 h of incubation at 28°C, protease activity was assessed according to the hydrolytic zones around the bacterial colonies  following the manufacturer's instructions. To remove genomic DNA, the eluted RNA samples were treated with RNase inhibitors and DNaseI (TaKaRa Bio). RNA integrity was confirmed by electrophoresis on 1.2% agarose gels. Then, 2 μg of each RNA sample was used to synthesize cDNA with a cDNA Synthesis kit (TaKaRa). The same experiment was performed three times. The transcriptional levels of XOC_1737 and XOC_4211 in the wild-type Rs105 and the xocR-deletion mutant were also assessed and compared.The assays of RNA extraction, cDNA synthesis and real-time PCR were performed as described previously , was selected as a subject to perform a BlastP search in the genome of Xoc strain BLS256 (Accession number: NC_017267). This led to the identification of a corresponding ortholog, termed as XocR (XOC_1422), which shared 97% similarity/identity with OryR in sequence at amino-acid level. To determine whether XocR is a member of the PAB LuxR solos, the sequence features of XocR and its genomic organization in Xoc were further analyzed. XocR possessed a typical AHL-binding domain at the N terminus and a DNA-binding HTH domain at the C terminus  encoding a hypothetical protein located closely to the pip gene in Xoc, which was absent in Xoo. In Xoo on the other hand at the similar position, a locus (XOO1270) encoding a protein that putatively functions as an amino acid transporter was located adjacent to the pip gene  with that of well-characterized OryR from Xoo. These findings prompted us to test the potential role of xocR in the virulence of Xoc as well as other important pathogenesis related traits (see below).To examine whether the genome of R Figure . This oro Figure . Howevere Figure . Our resxocR in Xoc virulence, we generated the xocR in-frame deletion mutant as well as the corresponding complemented strain  in non-host (plant) tobacco. For this purpose, three different cell concentrations of wild-type Rs105 and the xocR mutant were infiltrated into the leaves of tobacco. We determined that mutation of xocR did not alter the ability to trigger HR in tobacco under all testing conditions and behaved as the wild-type  tobacco. Our results also provide a clue to reveal the diversity of function of xocR in host and non-host plants. The PAB LuxR solos have been shown to mediate virulence in diverse phytopathogenic bacteria by binding yet-to-be-identified plant signal/compound; these are XccR of X. campestris binding a signal present in cabbage  to regulate virulence in Xoc. This prompted us to examine whether any of the four well-studied virulence-associated traits, including cell motility, EPS production, biofilm formation and extracellular protease activity were impaired in the xocR mutant, resulting in virulence deficiency. We therefore determined the ability of the xocR mutant in regulating these phenotypes. As shown in Figure xocR mutant in the absence of rice macerate. As reported previously, the regulation of the PAB LuxR solos with regard to phenotypes occurs to the greatest extent in response to the presence of plant signal(s)  . Similarly to what occurs in Xoc, cell swimming motility is also regulated by OryR in Xoo  Figure . As shows Figure . Furtherm Figure , suggestXoo, OryR was found to positively control the transcriptional expression of a number of movement-related genes, including 30 flagella genes in response to rice macerate . Identification of the rice signal(s) that interacts with OryR or XocR will give more insight into this point. Nevertheless, our results suggest that response to host signal is important for XocR to regulate cell motility, which seems to facilitate the infection of Xoc on host rice.In oc Table . This fipip-box present in the promoter region of target genes to regulate their transcription. In view of this, an analysis of the promoter sequences of 29 reported virulence-associated genes of Xoc was made (Table pip box element)  and XOC_4211, encoding an anthranilate synthase component I (TrpE). To address whether the transcription of these two genes was controlled by xocR, we used qRT-PCR to determine their transcriptional levels in wild-type Rs105 and the xocR mutant. As shown in Figures recN (~1.7-fold) and trpE (~3.0-fold) in the wild-type background. However, the induced transcriptional effect of recN and trpE by rice macerate was reduced to ~1.4 and ~1.8-fold in the xocR mutant, respectively, compared to the control medium. Meanwhile, the transcription of recN or trpE in the wild-type strain and the xocR mutant was similar in the absence of rice macerate, whereas in the presence of rice macerate, mission of xocR significantly alter the expression pattern of both genes. In details, the expression level of recN and trpE was reduced to (~1.5-fold) and (~2.0-fold), respectively, compared to the wild-type strain in the presence of rice macerate. Although the complemented strain was able to restore the deficiency of the xocR mutant in cell motility  was highly similar with their cases in the xocR mutant under all testing conditions (data not shown). It is possible that the transcriptional level of both genes in the pBBR1-MCS5 vector was relatively high compared to their native levels in the genome, further suggesting the transcriptional regulation of xocR on both genes was probably precise in Xoc. Nevertheless, our results collectively suggest that the rice macerate may activate the positive regulation effect of xocR on the transcription of two known virulence-associated genes in Xoc. It is currently not known whether XocR directly binds the promoter region of recN or trpE to regulate their transcription in the presence of rice macerate or whether this regulation is indirect. RecN is a highly conserved, SMC -like protein in bacteria , which is a member of tryptophan biosynthetic pathway. AS catalyzes the formation of anthranilate from chorismate and ammonia (or glutamine), which is the first committed step branching from shikimate pathway toward the biosynthesis of l-tryptophan  to mediate virulence. As described above, XocR most likely binds to a DNA sequence called de Table ; this le) Figure . They arZeigler, . It playZeigler, . MoreoveXoc. XocR shares high sequence similarity at amino-acid level with OryR from Xoo. XocR, responds to rice-derived unidentified compound(s) and regulates virulence on host rice and cell motility similar to OryR. We further provided evidence that XocR mediated Xoc virulence through the control of the transcription of two known virulence-associated genes (recN or trpE) in the presence of rice signal(s). Future studies will focus on the identification of the rice signal(s) that interacts with XocR, and the regulatory mechanisms of XocR on cell motility as well as the transcription of recN or trpE in Xoc.This study reports for the first time the presence of a PAB LuxR solo designated as XocR in The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


RNA interference (RNAi) is a mechanism by which small double-stranded RNAs called micro(mi) or small interfering(si) RNAs bind messenger RNAs (mRNAs) to inhibit their expression. The mechanism involves the RNA-induced silencing complex (RISC) composed of Dicer, TRBP and Ago2 proteins in which TRBP loads miRNAs into the active complex . SeveralWe used a model based on miRNA Let7 activity on a reporter gene (RL or EGFP) linked to a complementary sequence (cLet7) to measure RNAi activity or its suppression. We used RNA-immunoprecipitation (IP) and gel mobility shift assays to compare TRBP binding to RRE or siRNAs. We studied RRE activity on RNAi in the context of the entire HIV-1, a lentiviral or an adenoviral vector.We observed that RRE, acts as an RNAi suppressor with no modification of the endogenous RISC . In contrast, RRE RNA displaces siARNs from TRBP, which suggests a change in miRNA incorporation into the RISC. RNAi remains functional in HIV-1 infected cells, whereas a lentiviral vector expressing RRE has a suppressive activity. The suppression is alleviated when Rev or GagPol is expressed. Adenovirus is known to be suppressed by RNAi and RRE reverses this inhibition as seen by increased viral replication.RRE is a new RNAi suppressor, which acts by competition with siRNA and miRNA for binding to TRBP and therefore incorporation into the RISC. This could explain in part the alteration of certain gene expression and modifications of the cell metabolism in patients with long-term HIV-1 infection.


Sinonasal lymphoma is a non-Hodgkin lymphoma (NHL) representing 1.5% of all lymphomas. It presents as an unremitting ulceration with progressive destruction of midline sinonasal and surrounding structures. Poor prognosis warrants early treatment although diagnosis is challenging and frequently delayed. It is usually primary in origin and to our knowledge the sinonasal region has never been reported as a sanctuary site in leukaemia/lymphoma relapse. We present a unique case of B-cell ALL (acute lymphoblastic leukaemia) with late relapse to the nasal septum as a sinonasal lymphoblastic lymphoma and with genetic support for this as a sanctuary site. Sinonasal lymphoma presents as an unremitting ulceration affecting midline sinonasal region and surrounding structures , 2. It iA case of rapid destruction of the nose and face was first described by McBride in 1897 . In 1933Sinonasal lymphoma is rare in Western countries with higher prevalence in South America and Asia . PatientWe report a unique case of B-lymphoblastic lymphoma presenting as a late relapse of B-acute lymphoblastic leukaemia (B-ALL) in the sinonasal space which we believe represents a newly described sanctuary site.A 44-year-old Caucasian woman presented with a three-month history of nasal obstruction, yellow nasal discharge, and forehead pain. She was initially treated for allergic rhinitis and referred to the ENT department when symptoms persisted. Examination revealed a mass involving the nasal septum and the lower part of the nasal cavity and extending into the nasopharynx. Histology showed a monomorphic infiltrate of immature cells positive for CD19, PAX-5, HLADR, CD34, Tdt, and CD56 consistent with a diagnosis of B-cell ALL .9/L, CNS disease at presentation, and presence of residual leukemic cells on flow cytometry after first cycle of induction, she had been treated with eight cycles of Hyper-CVAD followed by a cyclophosphamide/total body irradiation conditioned allogeneic stem cell transplant from a sex-mismatched, unrelated donor. She had suffered from hyperacute graft-versus-host disease (GVHD) which was steroid responsive and chronic GVHD of the liver treated with cyclosporine. This resolved and she had not been on any immunosuppression for 20 months at the time of presentation.There was a history of B-ALL, with normal female karyotype at diagnosis, six years prior to current presentation. Due to poor risk features, including a high presenting white cell count of 122 × 10Pretreatment staging revealed an isolated sinonasal B-ALL, Ann Arbor stage 1 . The LDHChimerism analysis by FISH revealed 100% XY cells in the bone marrow and peripheral blood. FISH analysis of the nasal biopsy showed most cells (191 out of 200) to have three copies of chromosome X with only 9 having chromosomes X and Y, consistent with relapse from initial female leukaemia cells. Further cytogenetic analysis of the sample was unsuccessful.Induction chemotherapy was initiated according to the UKALL12 protocol. This was complicated by neutropenic fever, fatty infiltration of liver, hyponatraemia secondary to antibiotic induced SIADH, quite marked weight loss, and peripheral neuropathy secondary to vincristine. The L-asparaginase lowered her antithrombin levels and antithrombin concentrates were given to counteract her hypercoagulable state. Due to the significant toxicity chemotherapy was discontinued after Phase 1 induction and the planned CNS prophylaxis with high dose methotrexate was cancelled as she was too unwell with an ECOG performance status of three. She then received 3600 cGy consolidative radiotherapy in 20 fractions to the nasal cavity over four weeks. There was no evidence of systemic relapse at diagnosis or after completion of treatment.Two months later she complained of hard crusts within her nose. Examination revealed a septal perforation with some necrotic bone at the posterior half of the nasal septum and associated crusting of the nasal cavity, colonised by pseudomonas. A persistent nonspecific opacity within the ethmoid and maxillary sinuses was also identified on CT imaging. The crusts were removed under local anaesthetic and necrotic bone was debrided. She was treated medically for suspected sinusitis secondary to local irradiation. Examination at her one-month and eleven-month follow-up revealed a septal perforation with healthy mucosal edges. There was no suggestion of recurrence of her sinonasal lymphoblastic lymphoma or leukaemia eighteen months after completion of chemoradiotherapy.B-cell lymphoblastic leukaemia/lymphoma (B-LBL/ALL) is a form of Precursor Lymphoid Neoplasm under the World Health Organization (WHO) classification of lymphoid neoplasm  constituLBL and ALL have similar clinical, cytogenetic, and histological features and are frequently treated as the same disease. Clinical distinction of LBL from ALL relies on the fact that LBL shows minimal involvement of blood or bone marrow . They arExtramedullary relapse in ALL is not uncommon but usually involves the lymph nodes, CNS, spleen, liver, and testis . The devThe CNS and testes are well-recognised sanctuary sites for extramedullary relapse, protected from antileukemic T-lymphocytes and a graft-versus-leukemic effect possibly through biological barriers or increased blast tropism for these regions , 21. ThePrognosis of adult ALL is poor with only 30–40% of patients achieving long-term disease remission . A high We report a sinonasal lymphoma as an isolated extramedullary relapse of B-ALL. This is an unusual site for B-ALL relapse and the female lineage of the tumour cells strongly supports our contention that the nasal septum represents a newly described sanctuary site for lymphoma.Disease relapse is frequently diagnosed late further worsening an already grave prognosis of the disease. Early diagnosis improves clinical outcome highlighting the need for an awareness that patients presenting with sinonasal symptoms refractory to initial medical treatment are at risk of relapse.


More than thirty years after the discovery of HIV, the HIV/AIDS epidemic continues to expand worldwide. Undiagnosed HIV infections and late diagnoses remain major public health challenges in all HIV-affected countries. According to recent estimates, the proportion of persons living with HIV unaware of their infection varies from >50% in sub-Saharan Africa to 20-50% in European countries and 20-30% in North America. In high-income countries, around 50% of new HIV diagnoses are made late ; mean CD4 cell count at first presentation to medical care in high-income countries was slightly above 300 cells per μL in 2011, and has not increased significantly over the past 20 years. In low- and middle-income countries, most patients start HIV treatment with CD4 counts well below the recommended thresholds; the main reason for late initiation of HIV treatment is late diagnosis.Undiagnosed HIV infections and late diagnoses have serious implications for both the individual and public health. Patients diagnosed late cannot benefit fully from timely initiation of HIV treatment and are more likely to experience HIV-related morbidity and premature mortality. Undiagnosed HIV-infected individuals have a higher risk of transmitting HIV than individuals aware of their HIV infection, because they are not engaged in HIV treatment (which greatly reduces the risk of transmission) and cannot make informed decisions about their sexual behavior. As a consequence, the majority of new HIV transmissions may originate from individuals who are unaware of their HIV infection.Improving prevention and early diagnosis are thus key objectives to reduce mortality and turning the tide on HIV transmission. In this presentation, we will describe the non-biomedical HIV prevention methods, including early diagnosis, review and summarize the evidence supporting (or not) their effectiveness and discuss how these methods could be improved.


The hippocampal circuitry undergoes attentional modulation by the cholinergic medial septum. However, it is unclear how septal activation regulates the spatial properties of hippocampal neurons. We investigated here what is the functional effect of selective-cholinergic and non-selective septal stimulation on septo-hippocampal system. We show for the first time selective activation of cholinergic cells and their differential network effect in medial septum of freely-behaving transgenic rats. Our data show that depolarization of cholinergic septal neurons evokes frequency-dependent response from the non-cholinergic septal neurons and hippocampal interneurons. Our findings provide vital evidence that cholinergic effect on septo-hippocampal axis is behavior-dependent. During the active behavioral state the activation of septal cholinergic projections is insufficient to evoke significant change in the spiking of the hippocampal neurons. The efficiency of septo-hippocampal processing during active exploration relates to the firing patterns of the non-cholinergic theta-bursting cells. Non-selective septal theta-burst stimulation resets the spiking of hippocampal theta cells, increases theta synchronization, entrains the spiking of hippocampal place cells, and tunes the spatial properties in a timing-dependent manner. The spatial properties are augmented only when the stimulation is applied in the periphery of the place field or 400–650 ms before the animals approached the center of the field. In summary, our data show that selective cholinergic activation triggers a robust network effect in the septo-hippocampal system during inactive behavioral state, whereas the non-cholinergic septal activation regulates hippocampal functional properties during explorative behavior. Together, our findings uncover fast septal modulation on hippocampal network and reveal how septal inputs up-regulate and down-regulate the encoding of spatial representation. The projections from the medial septum to the hippocampus are proposed to have important roles in cognition by modulating the activity of episodic memory circuits Winson, . Beyond in vitro , to a recording system, which also allowed for animal position tracking. The recordings took place in rectangle-shaped linear track (80 × 80 × 9.5 cm and wide) situated in the center of a room with multiple background cues available. Signals were amplified  and band-pass filtered between 380 Hz and 6 kHz for single-unit detection. To maximize cell separation, only waveforms of sufficient amplitude (at least three times noise threshold) were acquired. Candidate waveforms were discriminated off-line using graphical cluster-cutting software (Axona Ltd.), which allows waveform separation based on multiple features including spike amplitude, spike duration, maximum and minimum spike voltage, and the time of occurrence of maximum and minimum spike voltages. Autocorrelation histograms were calculated for each unit, and the unit was removed from further analysis if the histogram presented spiking within the first 2 ms (refractory period), inconsistent with good unit isolation.For the inactive (immobile) recording session pellets were positioned in the middle of the square arena where the rats consumed the pellets throughout the recordings session. For the active (mobile) recording session the rats were placed in the open field and 20 mg food pellets  were thrown in every 20 s to random locations within the open field (pellet-chasing task); in this way, animals locomoted continuously, allowing for complete sampling of the environment. A recording session was considered as inactive if the animal locomoted less than 14 m per 12 min (with average velocity of <2 cm/s) and active if the animal locomoted more than 72 m per 12 min (average velocity of >10 cm/s). The chosen duration of 12 min allowed the rats to explore evenly the arena in two subsequent recordings (baseline and stimulation sessions). Baseline recordings >15 min result in insufficient exploration of the subsequent TBS session, while recordings < 10 min reduced the sampling of the explored environment. The rats were habituated to the square arena before the recordings.p-value is calculated, indicating whether the correlation is significant or not. Place field activity is not considered to be spatially coherent if the p-value is greater than 0.001. The spatial information content  is expressed in bits per spike  recordings were performed as previously described  between local field potentials  and perfused transcardially with ice-cold 0.9% saline followed by 4% paraformaldehyde . Brains were removed, post-fixed in paraformaldehyde for up to 24 h and cryoprotected in 25% sucrose for >48 h. Brains were sectioned coronally at 40μm on a freezing microtome. Primary antibody incubations were performed overnight at 4°C in PBS with BSA and Triton X-100 (each 0.2%). The concentration for primary antibodies was anti-ChAT 1:200 (Millipore). Sections were then washed and incubated in PBS for 10 min and secondary antibodies were added (1:200) conjugated to Alexa Fluor 594 dye (Invitrogen) for 2 h at room temperature. For visualization, the sections were mounted onto microscope slides in phosphate-buffered water and cover-slipped with Vectashield mounting medium (Fisher Scientific). The YFP fluorescence was evaluated within a selected region that was placed below the fiber tip in area of 1.5 × 1.5 mm. Fluorescence was quantified based on the average pixel intensity within the selected region  paired with post-hoc Newman–Keuls test. The probability level interpreted as significant was p < 0.05. All data points are plotted ± sem.All data were analyzed using Prism software . Statistical significance was estimated by using two-tailed n = 7 rats, n = 85 cells). We applied laser stimulation of septal cholinergic neurons in the animals implanted with optic fiber and recording tetrodes and evaluated the local network effect in medial septum. Trains of square pulses of blue light were applied continuously every 6 s. From 215 cells recorded in medial septum (n = 7 rats) we identified 70 cells that changed their firing frequency as a result of the stimulation  of the affected cells, are defined as non-cholinergic neurons. The ChAT neurons group is characterized with tonic, regular slow-frequency firing 3.7 ± 1.4 Hz . The firing frequency of the place cells was significantly affected only for the inactive state, where the stimulation/baseline spiking ratio decreased during the 50 Hz stimulation, compared to the ratio after the stimulation protocol  the spiking of hippocampal neurons and (2) theta band (5–12 Hz) of hippocampal local field oscillations. The optic fiber was located in medial septum, while the recording tetrodes were located in dorsal hippocampal CA1 area  in four rats. We used the phase-locking value , which measures the degree of local field synchrony between all stimulation epochs in the theta frequency range. We analyzed the values for the first 100 ms of the stimulation protocols. The phase-locking values were higher for 10 Hz  for both inactive  in active behavioral state during pellet-chasing task. Similarly to the optogenetic septal stimulation we observed three types of responses from the hippocampal interneurons: (1) group of neurons that responded with initial inhibition, followed by subsequent potentiation  = 1.730, n = 120 trials p < 0.05), which occurred at the end of TBS protocol and shortly after TBS . We also analyzed the firing properties of the targeted unit with the firing properties of the units with place fields in a distal location (controls). We investigated the mean and center firing rate, the spatial coherence, the spatial specificity , field size and centroid difference between clockwise and counter-clockwise directions. We represented the CA1 place field properties as ratios of the measured values from baseline session over the values of the TBS session. Our data show significant difference of the place field center rate between the periphery-stimulated fields and the center-stimulated fields , the center firing frequency increases. Importantly, the timing of increased place cell spiking coincides with intrinsically-entrained synchronization of hippocampal theta cells and concurrent theta power increase. The center-stimulated fields reflect spiking decrease that parallels the timing of extrinsically-induced entrainment of septo-hippocampal circuitry. This result suggests that stimulation of the medial septum activates the hippocampal neural circuitry involved in the generation of theta field activity in a non-physiological manner, dissociated from the normal behavioral correlates (Bland et al., Our data show that the modulatory role of septal cholinergic control over the hippcampal place field representation is mediated by the rhythmic entrainment of hippocampal neurons and the ability of septo-hippocampal circuits to resonate in theta frequency. We used specific optogenetic stimulation of the septo-hippocampal cholinergic neurons in transgenic rats and non-specific theta-burst stimulation protocols to demonstrate that medial septum is a major player not only in the transition between behavioral states but also enables rapid regulation of the hippocampal spatial encoding.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Altered function of the medial temporal lobe (MTL) is a valuable indicator of conversion from amnestic mild cognitive impairment (aMCI) to Alzheimer’s disease. This study is to delineate the functional circuitry of multiple subdivisions of parahippocampal gyrus and hippocampus (HIP) and to examine how this knowledge contributes to a more principled understanding of the contributions of its subregions to memory in aMCI. The functional connectivity (FC) analysis was performed in 85 aMCI and 129 healthy controls. The aMCI demonstrated the distinct disruptive patterns of the MTL subregional connectivity with the whole-brain. The right entorhinal cortex (ERC) and perirhinal cortex (PRC) showed increased connectivity with the left inferior and middle occipital gyrus, respectively, which potentially indicated a compensatory mechanism. Furthermore, the right altered MTL subregional FC was associated with episodic memory performance in aMCI. These results provide novel insights into the heterogeneous nature of its large-scale connectivity in MTL subregions in memory system underlying the memory deficits in aMCI. It further suggests that altered FC of MTL subregions is associated with the impairment of the differential encoding stages of memories and the functional changes in the specific right HIP-ERC-PRC-temporal circuitry may contribute to the impairment of episodic memory in aMCI. Amnestic mild cognitive impairment (aMCI), which is characterized by the impairment of episodic memory, is a high rate of conversion to Alzheimer’s disease (AD)1Growing evidence suggests a more differentiated picture, one of functional diversity in the MTL subregions581075151617Recently, several neuroimaging evidences indicate that aMCI can be characterized by abnormalities in resting-state FC of MTL subregions202122The objective of the present study was therefore to investigate the large-scale functional organization and functional circuitry of MTL subregional networks in aMCI. We hypothesized that aMCI would present differentially abnormal connectivity patterns in the MTL subregional networks compared to healthy controls (HC). And we further predicted that the altered FC in a specific pathway along the MTL cortical processing hierarchy system would link to the impairment of episodic memory in aMCI. To answer this issue, the present study evaluated the connectivity patterns based on two parallel arrays of seeds along the longitudinal axis of the parahippocampal gyrus (PHG) and the HIP in each hemisphere (p > 0.05). Compared with HC, aMCI showed significantly lower MMSE and MDRS scores, and significant deficits in performance in multiple-domains of cognitive functions, including episodic memory, information processing speed, executive function, and visuospatial cognition . Detailed raw scores and the corresponding Z scores from individual neuropsychological tests are provided in Demographic and neuropsychological characteristics are shown in SI Results). Furthermore, schematic polar plots illustrated that there were different FC patterns among four PHG and three HIP seeds, which showed similar patterns between aMCI and HC by visual inspection  and age were quadratic, namely inverse U-shaped curve in aMCI see . FurtherThe present study identified a distinct altered FC pattern associated with cognitive impairment and a selective topography of functional changes in the MTL subregional networks, indicated that the disruption of the right HIP–ERC–PRC–temporal pathway was responsible for the impairment of episodic memory, and provided new insights to understand the pathophysiology underlying the memory deficits in aMCI.The present study indicated that the MTL subregions showed different patterns of connectivity with cortical and subcortical areas, which corroborated and extended findings of functional heterogeneity along the longitudinal axis of the HIP and PHG in both animals1918171827285619Our results showed that there were differentially abnormal connectivity patterns along the anterior to posterior axis of PHG and HIP and the asymmetry of damaged degree between left and right hemispheres in aMCI compared to HC see , Fig. S1233031Based on our findings, as well as these findings from structural connectivity, as indicated by tract-tracing in animals71617524735571718373839As shown in 7177427Our results showed that the pPHC network in aMCI exhibited decreased connectivity with parietal lobe  and middle temporal gyrus, which locate in the parieto-temporal visuospatial pathway351745474919The present study showed that the ERC network mainly exhibited decreased connectivity with hippocampus and cerebellum. Numerous nonhuman primate studies have demonstrated that the PRC and PHC provide nearly two-thirds of the cortical input to the ERC. The anterolateral aspects of the ERC receive highly integrated visual object information from occipito-temporal visual object processing pathway via the PRC7525457Our findings complement findings of functional heterogeneity along the longitudinal axis of the HIP in both animals and humans. The present study showed the decreased connectivities between right aHIP and visual cortex (right fusiform gyrus), between mHIP and parietal cortex (precuneus), middle temporal gyrus, cerebellum, and visual cortex, between right pHIP and visual cortex in aMCI. These results suggest that aMCI present differentially abnormal connectivity patterns along the anterior to posterior axis of HIP. The previous studies have indicated that the fusiform gyrus, which is located on the ventromedial surface of the temporal and occipital lobes60614950The relationship between the decreased FC and the neuropsychological performance supported the notion that the disrupted functional neurocircuitry was the basis of cognitive impairment in aMCI , suggest197Interestingly, the present study demonstrated that the relationships between the increased FC strength and age were quadratic, namely reverse U-shaped curve . This fi3, so each ROI would have included mixed signals from adjacent regions. To assess the effect degree of the resting connectivity patterns by the choice of smoothing, we compared the differences of these results from the different smoothing sizes  and no significant differences were found.The resting data of this study were smoothed with a Gaussian kernel of 4 × 4 × 4 mmThe current study provides novel insights into the heterogeneous nature of its large-scale connectivity in MTL subregions underlying the memory deficits in aMCI. It further suggests that the functional changes in the right HIP-ERC-PRC-temporal circuitry may be an early indicator for early stage changes and progression of this disease.This study was approved by the responsible Human Participants Ethics Committee of the Affiliated ZhongDa Hospital, Southeast University. All participants were assessed at the Affiliated ZhongDa Hospital, Southeast University, and written informed consent was obtained from all of the participants and the methods were carried out in accordance with the approved guidelines.SI Methods) used to choose subjects can be found in our previously published studiesThe present study initially recruited 222 elderly individuals , including 87 aMCI, and 135 HC, through normal community health screening, newspaper advertisement, and hospital outpatient service. Due to motion artifacts, of these, 6 HC subjects and 2 aMCI subjects were excluded. The remaining 129 HC and 85 aMCI subjects were eligible and entered further analysis. The inclusion and exclusion criteria . The details regarding the neuropsychological assessments are provided in SI Methods.MRI images were acquired using a 3.0 Tesla Trio Siemens scanner  with a 12-channel head-coil at ZhongDa Hospital Affiliated to Southeast University. Details on image acquisition parameters are provided in http://www.fil.ion.ucl.ac.uk/spm). Briefly, this preprocessing included removal of the first ten volumes, slice timing correction and head motion correction723 voxels and spatially smoothed using a 4-mm full-width half-maximum (FWHM) Gaussian kernel. Linear detrending and temporal band-pass filtering (0.01–0.08 Hz) were applied to reduce the effect of low-frequency drifts and high-frequency physiological noise. Finally, several nuisance variables, including six head motion parameters, global mean signal76Data analyses were conducted with Matlab  and SPM8 . Then, a two-sample t test was performed to determine whether the GM was atrophied in aMCI  head displacement or rotation (in mm or °) derived from the motion-correction procedure477273This study created two parallel arrays of four-millimeter spherical seeds along the longitudinal axis of the PHG and HIP respectively . Specifi111718858611171826The average time courses for all voxels within each MTL subregion were extracted as the reference time course, separately, and voxelwise cross-correlation analysis was then carried out between the averaged time courses of all voxels within the seed region and the whole brain within the GM mask. The GM mask was created by thresholding (a probability threshold of 0.2) the GM probability map obtained from all subjects in this study. A Fisher’s z-transform was then applied to improve the normality of the correlation coefficients.t test and chi-square tests (applied only in the comparisons according to gender) were used to test the differences in demographic data and neuropsychological performances between aMCI and HC (p < 0.05).The statistical analyses were performed using SPSS 20.0 software . The two-independent samples t-tests with a stringently threshold of p < 0.01 using family-wise error (FWE) correction together with a cluster extent k > 100 voxels (800 mm3) to reveal regions most robustly correlated with each seed. Only clusters within the GM mask were retained. To avoid ambiguous biological interpretations related to apparently negative connectivity resulting from correction for global signal changesTo highlight the FC patterns of MTL subregions at a group level, the spatial maps of FC in each group were submitted to a random-effect analysis using one-sample p (uncorrected) <0.005 and cluster extent k > 80 voxels (640 mm3) was used to achieve a corrected statistical significance of p < 0.05, determined by Monte-Carlo simulation (see program AlphaSim by D. Ward).To assess the between-group differences of the FCs of each MTL subregion, we used GLM analysis  with age, gender, years of education, and GM volume treated as covariates. A statistical threshold of To assess the potential overlap between the between-group difference connectivity maps derived from the seven seeds of interest, these between-group difference maps from GLM analysis were thresholded as indicated above, binarized and summed, generating a so-called “overlapping” map.SI Methods. Analysis of variance (ANOVA) was used to test the difference of FC patterns of MTL seeds between aMCI and HC (p < 0.05).To characterize abnormal FC patterns of MTL seeds to specific target brain ROIs, schematic polar plots were used to describe FC patterns of each seed with target regions throughout the whole-brain. Details on definition of target ROIs are provided in SI Methods) .To assess the links between altered FC and neuropsychological performance, we extracted the averaged FC strengths of the regions showing altered MTL subregional connectivity. The multiple linear regression model analysis was used to examine the relationships between the extracted FC strengths and the neuropsychological performance in each group. To increase statistical power by reducing random variability, as previously described82p < 0.05, Bonferroni-corrected).To investigate the links between the age and increased FC in the whole-brain in aMCI due to HC, the curve estimation was used to examine the relationships between FC strength  and age in each group .


A wealth of genome sequences has provided thousands of genes of unknown function, but identification of functions for the large numbers of hypothetical genes in phytopathogens remains a challenge that impacts all research on plant-microbe interactions. Decades of research on the molecular basis of pathogenesis focused on a limited number of factors associated with long-known host-microbe interaction systems, providing limited direction into this challenge. Computational approaches to identify virulence genes often rely on two strategies: searching for sequence similarity to known host-microbe interaction factors from other organisms, and identifying islands of genes that discriminate between pathogens of one type and closely related non-pathogens or pathogens of a different type. The former is limited to known genes, excluding vast collections of genes of unknown function found in every genome. The latter lacks specificity, since many genes in genomic islands have little to do with host-interaction.Dickeya dadantii 3937 and Pectobacterium carotovorum WPP14 were used for development of this tool, because these pathogens are important on multiple high value crops in agriculture worldwide and more genomic and functional data is available for the Enterobacteriaceae than any other microbial family. Our approach achieved greater than 90% precision and a recall rate over 80% in 10-fold cross validation tests.In this study, we developed a supervised machine learning approach that was designed to recognize patterns from large and disparate data types, in order to identify candidate host-microbe interaction factors. The soft rot Enterobacteriaceae strains Application of the learning scheme to the complete genome of these two organisms generated a list of roughly 200 candidates, many of which were previously not implicated in plant-microbe interaction and many of which are of completely unknown function. These lists provide new targets for experimental validation and further characterization, and our approach presents a promising pattern-learning scheme that can be generalized to create a resource to study host-microbe interactions in other bacterial phytopathogens.The online version of this article (doi:10.1186/1471-2164-15-508) contains supplementary material, which is available to authorized users. Interactions between plant-associated microbes and their eukaryotic hosts are complex biological processes involving hundreds, if not thousands, of genes from each organism. Understanding the molecular mechanisms of such complex processes at the systems-scale is seriously hampered by the lack of a comprehensive list of gene products that contribute for even a single bacterial or fungal pathogen. Variation in lifestyles and pathogenic potential between organisms makes the challenge all the greater. Genome sequencing has dramatically increased the potential for large-scale screens to identify genes involved with host-microbe interactions. Direct experimental evidence is the obvious gold standard, but not all significant pathogens are experimentally tractable, and selection of experimental conditions and convenient hosts for high-throughput screens can limit discovery. More targeted experiments can be designed to probe function more completely, but these are time consuming and generally limited to a smaller number of candidate genes. Further, it is unclear what experiments to conduct if a candidate gene is of completely unknown function. Importantly, genes of unknown function make up a substantial fraction of each sequenced genome, and it is likely that among these lie some of the greatest potential for discovery of truly novel aspects of host-microbe interaction .Computational approaches to identify potential host-microbe interaction factors and predict their specific functions can be a valuable way to guide experimentation, and may be the only option for some recalcitrant organisms. Typical bioinformatics strategies include searching for sequence similarity to gene products known to contribute to host-microbe interaction in other organisms, and comparing genomes to identify gene islands that discriminate between pathogens of one type and closely related non-pathogens or pathogens of a different type. Both strategies are useful, but the former is limited to known genes and detectable levels of sequence similarity, and thus excludes the vast collections of genes of unknown function. The latter lacks specificity, since many genes in genomic islands may have little to do with host interactions, and the definition of rules for the distribution across organisms can be arbitrary. There are no simple rules to define the relevant distribution for the set of orthologous genes across genomes, especially when there are a large number of genomes being compared. Further, it is preferable in many situations to factor in other features such as genome context or gene expression data as additional evidence sources to predict whether a gene is associated with host-microbe interaction processes.Dickeya dadantii (aka. Erwinia chrysanthemi) 3937 .The data sets supporting the results of this article are available in the LabArchives repository, [List of individual protein-coding genes from Dd3937 (a) and WPP14 (b) assigned host-microbe interaction GO annotations corresponding to Table .(ZIP 33 KB)Additional file 1:(a) Strain information of all gamma-proteobacteria with complete or nearly complete genomes used in this study and their associations with various phenotypes of interest . (b) List of all non gamma-proteobacteria with complete or nearly complete genomes used in this study. (c) List of attribute categories used to generate summary taxonomic and phenotypic attributes based on sequence homology, the count of genomes in each category used as BLASTP search databases, and the number of proteins in Dickeya dadantii (Dd3937) and Pectobacterium carotovorum (WPP14) with hits to target organisms in those categories. (d) List of attributes used in the category of phenotypes of interest, including taxonomic groups and lifestyle statistics. (ZIP 223 KB)Additional file 2:Functional genomics data sets used in this study. (a) Data sets generated from microarray or proteomics studies. (b) Binding sites prediction of 32 putative transcriptional factors that may be related with host-microbe interactions. (ZIP 34 KB)Additional file 3:Description of selected supervised machine learning schemes.(DOCX 150 KB)Additional file 4:(a) List of selected attributes with importance measurement score and subsets of attributes defined to be used in data decay analysis forDickeya dadantii3937 andPectobacterium carotovorumWPP14, (b) probability score plot used to define a compact set of attributes for both strains.(ZIP 1 MB)Additional file 5:(a) RP curve to compare strategies of data transformation for boosting classifier performance. (b) PR curve to compare classifier performance using five data sets partitioned according to GO terms for different aspects of host-microbe interaction  List of 211 genes predicted to be host-microbe interaction factors for Dickeya dadantii 3937, (b) list of 216 genes predicted to be host-microbe interaction factors for Pectobacterium carotovorum WPP14. (c) list of top 300 genes that could not be confidently classified as host-microbe interaction factors for both Dickeya dadantii 3937 and Pectobacterium carotovorum WPP14. (ZIP 17 KB)Additional file 7:


Escherichia coli and human cells. Naturally occurring viral variants with single mutations within the loxLTRu sequence are also shown to be efficiently targeted by uTre, further increasing the range of applicability of the recombinase. Potential off-target sites in the human genome are not recombined by uTre. Furthermore, uTre expression in primary human T cells shows no obvious Tre-related cytopathic or genotoxic effects. Finally, uTre expressing mice show no undesired phenotypes during their normal lifespan. We have developed a broad-range HIV-1 LTR specific recombinase that has the potential to be effective against the vast majority of HIV-1 strains and to cure HIV-1 infected cells from the infection. These results strongly encouraged us in our confidence that a Tre recombinase-mediated HIV eradication strategy may become a valuable component of a future therapy for HIV-infected patients.Current drugs against HIV can suppress the progression to AIDS but cannot clear the patient from the virus. Because of potential side effects of these drugs and the possible development of drug resistance, finding a cure for HIV infection remains a high priority of HIV/AIDS research. We recently generated a recombinase (termed Tre) tailored to efficiently eradicate the provirus from the host genome of HIV-1 infected cells by specifically targeting a sequence that is present in the long terminal repeats (LTRs) of the viral DNA . However


A Norfolk terrier was referred to the Animal Health Trust neurology department with suspected dystrophin-deficient muscular dystrophy (DD-MD), which was confirmed by clinical workup and immunohistochemistry.Exon resequencing of the canine Duchenne Muscular Dystrophy (DMD) gene was undertaken to screen for potential disease causing mutations. The sequence data generated from all coding DMD exons revealed a 1 bp deletion in exon 22, causing a frameshift and premature termination of the coding sequence. Gene expression analysis indicated reduced levels of dystrophin transcript in the DD-MD case and western blot confirmed the absence of full length protein.The finding represents a novel mutation causing DD-MD in the dog.The online version of this article (doi:10.1186/s40575-015-0019-4) contains supplementary material, which is available to authorized users. In healthy skeletal muscle the dystrophin protein protects cells against damage when contracting during movement. In dystrophin deficient muscular dystrophy (DD-MD) the absence of dystrophin results in damage to the muscle. This leads to loss of mobility and reduced life expectancy.A six month old male Norfolk terrier was referred to the Animal Health Trust neurology department, and was confirmed as having DD-MD. This was the first time the disease had been described in this breed. The aim of this study was to identify the genetic cause of the disease in this dog.DMD) were sequenced, in both the affected dog and its mother. A variant in exon 22 was identified which was predicted to result in the production of a truncated dystrophin protein. The lack of functional dystrophin in the affected dog was confirmed through expression analysis.The coding exons of the dystrophin gene  is an X-linked disease in humans  and dogs . In DD-MDMD) of dogs. Examples include a 4 base pair (bp) deletion in exon 65 in a Cocker spaniel [A number of pathogenic mutations have been discovered in the dystrophin gene . Sequence read alignments were visualised in IGV [Due to the large size of the dystrophin gene we focused our investigation on the 79 coding exons of DMD gene expression, relative quantification by quantitative reverse transcription PCR (qRT-PCR) was performed. Assays were designed for the DMD gene and for the ubiquitously expressed TATA box binding protein (TBP) gene. RNA was extracted from skeletal muscle samples obtained from one case and five control dogs. Muscle samples were not matched by breed or the muscle from which they were biopsied. qRT-PCR was carried out in triplicate, and reaction efficiencies calculated by generating a seven point, doubling dilution standard curve for both assays [DMD expression in the DD-MD case  targeted a region close to the N-terminus of the dystrophin protein. The immunoblot showed bands for full length dystrophin in both of the controls, but not in the case (Figure 1315 targIn the clinical investigation by Beltran and colleagues, immunohistochemical analysis using antibodies targeting the rod-domain and the c-terminus showed an absence of dystrophin in the skeletal muscle of the Norfolk terrier DD-MD case . These fIn this study we have successfully identified a novel frameshift mutation causing a case of DD-MD in a Norfolk terrier dog.Muscle tissue from the case was taken as part of a veterinary diagnostic procedure, and a small subsection of this tissue used for the RNA and protein analyses. Muscle samples from controls were obtained post-mortem. Samples of DNA were collected by buccal swabbing which is a non-invasive procedure and does not require a U.K. home office licence. All samples were taken with full owner consent.The data set supporting the results of this article is included within the article (and Additional file