Studies performed three decades ago in our laboratory supported the hypothesis that radiation efficacy may be augmented by bacterial extracts that stimulate non-specific systemic antitumor immune responses. Application to the clinic was halted by unacceptable side effects and toxicities resulting from exposure to whole bacterial pathogens. Later scientific advances demonstrated that DNA isolated from bacteria was immunostimulatory and could be reproduced with synthetic oligodeoxynucleotides (ODNs), thus fueling the transition from bugs to drugs. Unmethylated CpG motifs within bacterial DNA induce activation of Toll-like receptor 9 and subsequently activate antigen-specific cellular immune responses. CpG ODNs have demonstrated favorable toxicity profiles in phase I clinical trials. We showed that this potent immunoadjuvant can be used in combination with radiation therapy to enhance local and systemic responses of several murine tumors. Studies demonstrated that enhanced tumor response is mediated in part by the host immune system. Antitumor efficacy was diminished in immunocompromised mice. Animals cured by combination of radiation and CpG ODN were resistant to subsequent tumor rechallenge. This body of work contributes to our understanding of the dynamic interplay between tumor irradiation and the host immune system and may facilitate translation to clinical trials. Bacillus Calmette–Guérin and Corynebacterium parvum were used to stimulate antitumor immunity  of sarcomas ,b. The r disease .C. parvum was an effective adjunct to radiotherapy, since treatment response depends not only on radiobiological factors but also on the immune response of the tumor-bearing host (C. parvum increased radiosensitivity of well-established (8 mm diameter) immunogenic murine fibrosarcomas when local irradiation was given as a single-dose or in multiple fractions . In normal mice, 100% tumor take was achieved with inoculations as low as 2.5 × 105 tumor cells. At 100–120 days after treatment, mice cured by radiation alone required 2 × 105 tumor cells to produce 50% tumor take, whereas mice treated with CpG ODN 1826 plus irradiation were totally resistant to tumor rechallenge with cell numbers as high as 8 × 105. Like the animals rechallenged by the s.c. route, mice locally cured by CpG ODN 1826 plus irradiation were much more resistant to development of artificial metastases in the lung than were those cured by radiation alone. These results showed that the systemic antitumor rejection response generated by CpG ODN 1826 plus radiotherapy exerted antitumor effects long after exposure to the agents. Secondary tumor rejection was most likely due to development of a memory response and possibly specific T cell-mediated immunity  radiotherapy induced an immune-mediated abscopal effect when combined with anti-CTLA-4 antibody in two preclinical rodent tumor models  was well-tolerated and induced systemic antitumor responses even in patients with significant tumor burden (in situ vaccination strategy combined with radiation in patients with T cell lymphoma mycosis fungoides skin lesions (Early clinical reports showed CpG 7909 was an effective and well-tolerated adjuvant for improving vaccine responses ,b. Minorr burden . Encoura lesions .Treatment of mice bearing established immunogenic or non-immunogenic tumors with CpG ODN 1826 markedly enhanced response to single-dose and fractionated radiotherapy, likely through immune-mediated mechanisms. CpG ODN also induced a durable systemic immune memory response against subsequent rechallenge with tumor cells. These observations suggest CpG ODN could be used not only as an “immunosensitizer” in combination with radiotherapy but also as an adjuvant to prevent or reduce metastatic disease at sites distant from the primary irradiated tumor. These findings and others have demonstrated that CpG ODNs can be given in combination with conventional radiotherapy to improve therapeutic efficacy. Further studies are warranted to elucidate the dynamic interplay between tumor irradiation and the host immune system to facilitate translation to clinical trials. Our studies using CpG ODNs as radiation enhancing agents are being supplemented by new integrated approaches proposing a partnership between radiotherapy and immunotherapy designed to capitalize on radiation’s ability to enhance immunogenicity of the primary tumor and its microenvironment .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.


Helicobacter pylori (H. pylori) and the actual H. pylori treatments fail with increased regularity because of continuously rising antibiotic resistances. To meet this challenge, we focus on the development of a new anti-infective therapy against H. pylori by targeting a secreted enzyme, high temperature requirement A (HtrA). Release of the serine protease HtrA near the host’s gastric epithelial cells leads to loss of cellular adhesion due to E- cadherin cleavage [In vitro substrate cleavage assays as well as cellular infection assays fully support the biophysical data.More than 50% of the world population is infected with cleavage . We inve


Every day health care workers are exposed to dangerous and deadly blood borne pathogen like HBV. HBV is 100 times more likely to be transmitted after a percutaneous exposure through infected blood than HIV. Hepatitis B infection can be prevented by vaccination.To study the effect of vaccination program among the health care workers of Sri Ramachandra University (SRU), Porur, Chennai. It is mandatory for all health care workers of SRU to be vaccinated against HBV at the time of appointment, producing anti-HBs titer is mandatory. They are advised for re-vaccination according to their anti-HBs titer.This study was conducted among the health care workers of SRU who had needle stick injury for a period of 1 year (1st November 2010 to 31st October 2011).The blood samples were processed for serology screening by Micro Particle Enzyme Immuno assay (MEIA) followed by ELISA. The results were studied.85 samples from health care workers who had needle stick injury were processed .The samples were from doctors (38%), nurses (20%), technicians (7%), housekeeping staff (23 %), and students (10%). All the samples of injured persons were found seronegative by both methods, anti-HBs titer was found to be protective.Strict HBV vaccination program and monitoring the anti-HBs titer were effective in prevention of HBV transmission by needle stick injury to health care workers of SRU. This can be implemented in all health care organizations for the benefit of health care providers.


Genetically manipulated animals like mice or rabbits play an important role in the exploration of human cardiovascular diseases. It is therefore crucial to identify animal models that closely represent the human cardiac phenotype.Phase contrast MRI  was usedSimilar behavior in the global LV myocardial motion pattern was observed in all species, particularly for the radial and long-axis motion as can be seen in Figure The results revealed a significantly different myocardial motion pattern in mice and rabbits compared to humans. Especially systolic rotation of rabbits more closely resembled human left ventricular performance, a finding that should be considered when investigating myocardial performance using mouse models. These findings underline that in the assessment of myocardial motion animal models cannot be conferred one-to-one to human beings. This deeper knowledge of mammalian-related differences in myocardial motion can help to select animal models that best mimic the human phenotype when investigating myocardial diseases.DFG FO 507/3-1.


Brain tissue changes in autism spectrum disorders seem to be rather subtle and widespread than anatomically distinct. Therefore a multimodal, whole brain imaging technique appears to be an appropriate approach to investigate whether alterations in white and gray matter integrity relate to consistent changes in functional resting state connectivity in individuals with high functioning autism (HFA). We applied diffusion tensor imaging (DTI), voxel-based morphometry (VBM) and resting state functional connectivity magnetic resonance imaging (fcMRI) to assess differences in brain structure and function between 12 individuals with HFA  and 12 healthy controls . Psychological measures of empathy and emotionality were obtained and correlated with the most significant DTI, VBM and fcMRI findings. We found three regions of convergent structural and functional differences between HFA participants and controls. The right temporo-parietal junction area and the left frontal lobe showed decreased fractional anisotropy (FA) values along with decreased functional connectivity and a trend towards decreased gray matter volume. The bilateral superior temporal gyrus displayed significantly decreased functional connectivity that was accompanied by the strongest trend of gray matter volume decrease in the temporal lobe of HFA individuals. FA decrease in the right temporo-parietal region was correlated with psychological measurements of decreased emotionality. In conclusion, our results indicate common sites of structural and functional alterations in higher order association cortex areas and may therefore provide multimodal imaging support to the long-standing hypothesis of autism as a disorder of impaired higher-order multisensory integration. Autism spectrum disorders (ASD) are neurodevelopmental and behaviorally defined disorders characterized by deficits in social reciprocity, impaired communication and repetitive patterns of behavior and interests The precise pathophysiology of ASD remains unclear apart from ASD that occur secondary to clearly defined neurological or genetic disorders like tuberous sclerosis or Rett syndrome. So far, genetic These altered neural tissue properties have been proposed to be exponentially distributed meaning that early brain abnormalities in ASD increasingly affect additional regions and functional systems throughout development A data-driven, multimodal structural  and functional (fcMRI) imaging approach might be able to detect superregional, system-level alterations in HFA as it does not limit the analysis to predefined regions, tissue types or functional systems.Diffusion tensor imaging (DTI) is based on the measurement of diffusion properties of water molecules Voxel based morphometry (VBM) is a complementary approach to investigate structural brain abnormalities by assessing macroscopic volumetric changes of gray and white matter brain tissue. Structural neuroimaging studies in HFA have reported decreased white matter Task dependent functional magnetic resonance imaging (fMRI) studies in HFA have shown various alterations Functional connectivity magnetic resonance imaging (FcMRI) is an approach that allows for the detection of functional brain networks without task constraints. Functional connectivity has been defined as the temporal correlation of a neurophysiologic index measured in different brain areas In summary the reported VBM, DTI and fcMRI findings indicate that individuals with HFA display alterations of gray matter volume, fractional anisotropy and functional connection strength, and that these alterations predominantly affect frontal and temporal association cortices.We hypothesized that structural and functional alterations in HFA, although complex and widely distributed, affect similar or overlapping brain regions. We furthermore expected that these convergent structural and functional alterations are more likely to be located in multimodal association cortex areas rather than in unimodal primary sensory or motor cortex regions. To test these hypotheses, this study combined structural  and functional (fcMRI) data-driven imaging approaches.All participants gave written consent before participating. The study was approved by the institutional review board of Ludwig-Maximilians University Munich and complies with the declaration of Helsinki. HFA participants were recruited from the outpatient clinic of the Department of Psychiatry and Psychotherapy, Ludwig-Maximilians University Munich. Healthy controls were recruited by advertisement and word of mouth. For subject characteristics see All participants had a full scale IQ >85. The language-based IQ of HFA and HC individuals did not differ significantly (p = 0.70). The language based IQ was estimated using a vocabulary test Two additional psychological tests were applied to the HFA group exclusively. The first one was the Freiburg Personality Inventory (FPI) which is a psychological test to assess personality traits http://fcon_1000.projects.nitrc.org/indi/abide/.Data acquisition was performed at a magnetic field strength of 3.0 Tesla . DTI was performed using a singleshot spin-echo sequence and SENSE parallel imaging (undersampling factor of 3). Diffusion-weighted images were acquired in 20 non-collinear diffusion-encoding directions with multiple diffusion weighting b-values (b = 1000 s/mm2 and b = 0). Data acquisition parameters included the following: 25 contiguous (no-gap) 4.0 mm-thick axial slices with an acquisition matrix of 128×128 over a FOV of 230 mm, repetition time (TR)  =  9300 ms, echo time (TE)  =  102 ms, and spatial resolution  =  1.8×1.8×4.0 mm. The DTI sequence was repeated three times and averaged to obtain stable diffusion parameters. For an anatomical reference and the VBM analysis a high-resolution isotropic Magnetization Prepared Rapid Gradient Echo (MPRAGE) sequence was acquired . During the 6-minute resting state functional experiment participants were instructed to minimize head movement and stay awake with their eyes closed. Data was acquired using T2*-weighted echo planar imaging . All high-resolution T1 MPRAGE images were quality controlled by a board certified radiologist to detect potential artifacts such as ringing due to motion, spiking or signal loss. The imaging raw data illustrated in this study has been made publically available through the ABIDE (Autism Brain Imaging Data Exchange) consortium and can be downloaded here: To investigate between group differences in FA values, we analyzed DTI data using the tract-based spatial statistics (TBSS) approach implemented in FSL 4.16  Software Library, Oxford, UK). Images were corrected for eddy currents due to changing gradient fields and head motions Structural data analysis was performed using the VBM tool implemented in FSL 4.16. Structural images were brain-extracted using BET http://afni.nimh.nih.gov/afni). The first 5 functional scans of each session were discarded. We used the FMRI Expert Analysis Tool (FEAT) All data analysis steps were performed using FSL 4.16 and Analyses of Functional Images . Previously preprocessed DTI and VBM files of HFA participants and controls served as the input. The algorithm was set to decompose the multimodal input into eight co-varying components. The resulting pseudo-z-scored component maps were normalized to values between -8 and 8, analogous to Groves and colleagues To test the relationship between measures of different imaging modalities an additional analysis called „linked independent component analysis for multimodal data fusion“ was performed. Linked ICA is a robust data fusion model that takes multi-modal data and characterizes inter-subject variability in terms of a set of multimodal components Psychological test results were correlated with FA, VBM and functional connectivity values within regions showing the most significant differences between HFA and HC. In order to maintain statistical power, we limited the correlation analysis to two out of twelve subscales of the FPI , namely emotionality and social interaction. These test items were chosen because alterations in these domains represent core features of HFA, rather than other subscales like aggressive behavior or health sorrows. The QEAS (questionnaire to estimate autistic symptoms) only consisted of 2 subscales, namely empathy and adequate social behavior, both of which were tested for correlation with imaging measures. In addition, we tested all subscales  and the sum of the Autism Quotient Questionnaire. Within each modality the significance of the Pearson product moment correlation results was Bonferroni-corrected for multiple testing . To correlate functional connectivity results we defined one region of interest (ROI) in the left STG based on the statistical group difference map obtained during fcMRI analysis and calculated the connection strength to the DMN. To correlate VBM measures, individual gray matter volume was summed up in a ROI in the left MTG, based on the VBM group difference map obtained previously. To correlate DTI measures we defined two regions of interest (ROI) based on the statistical group difference map obtained during DTI analysis. ROIs had a diameter of 16 mm around the peak voxel on the FA skeleton. FA values for each HFA subject were averaged within each of these two ROIs and correlated to psychological test results.corr≤0.01, DTI analysis revealed three main clusters of significantly reduced FA values in HFA when compared to HC: HFA individuals displayed lower FA values in a right-sided cluster reaching from the splenium of the corpus callosum into the superior longitudinal fasciculus within the parietal and temporal lobe and into the lateral occipital cortex (At pcorr≤0.1). At an uncorrected p-value (puncorr≤0.001) clusters of reduced gray matter volume of HFA were detected within the bilateral temporo-parietal junction area  and functional (fcMRI) data-driven imaging approaches to detect subtle, complex and widely distributed brain alterations in individuals with HFA and to investigate whether these alterations affect similar brain regions across modalities. We found three regions of convergent structural and functional alterations in HFA, all of which were located in higher order association cortex. The two regions displaying the most prominent FA value decrease, namely the right temporo-parietal junction area and the left frontal lobe showed concomitant functional connectivity decrease to the DAN and the LFPN respectively and a trend towards decreased gray matter volume. Interestingly, DTI alterations in these two main clusters seemed to be located down the neuronal stream to VBM changes. A formal testing of the relation between these structural properties revealed that DTI and VBM measures in these two clusters co-varied across participants, meaning that high FA values coincided with high gray matter volume and vice versa. In a third cluster of convergent structural and functional alterations decreased functional DMN connectivity in the bilateral superior temporal gyrus was accompanied by a trend towards gray matter volume decrease in the temporal lobe of individuals with HFA.The functional and structural differences between HFA and HC individuals in the right temporo-parietal cortex found in this study are in line with previously reported abnormalities of parietal lobe anatomy in terms of alterations in FA-properties The temporal lobe is another region known to be affected by autism pathology Convergent structural and functional alterations in HFA were also revealed in a mainly left-sided cluster including the prefrontal cortex, the ACC and the anterior corpus callosum. Differences in frontal lobe anatomy and function have previously been described in Asperger’s syndrome The main sites of functional and structural pathology reported in the present study can be attributed to higher order association cortex while primary sensory and motor cortices show almost no significant differences in functional connectivity strength or structural integrity. Higher order association areas are located in the prefrontal, the superior temporal and in the inferior parietal cortex and are known to display similar cytoarchitectonic properties One limitation to our study is that no formal analysis of the relationship between functional connectivity and structural changes was performed. While the relation between DTI and VBM measurements had been successfully explored before In conclusion, the results of the present study indicate that brain alterations in HFA, although subtle and widely distributed, converge into common sites of structural and functional differences in higher order association cortex areas such as the temporo-parietal junction and the prefrontal cortex in individuals with high functioning autism. These findings support the long-standing hypothesis of autism as a disorder of impaired higher-order multisensory integration.Figure S1Relation between DTI and VBM parameters. To investigate the relationship between DTI and VBM variance, “linked independent component analysis for multimodal data fusion”, a data fusion model that characterizes inter-subject variability of multi-modal data in terms of a set of multi-modal components, was performed. Here we show one component where DTI and VBM parameters co-varied across participants. This component comprised multiple brain regions, including the left frontal cortex and right temporo-parietal cortex. The images are color-coded according to significance (z-statistic), with red-yellow showing larger values for subjects with positive weights . To show the effects within regions that show a disease effect in HFA, DTI and VBM results shown in (TIFF)Click here for additional data file.


Gene silencing of O6-methylguanine–DNA methyltransferase (MGMT) by promoter methylation improves the outcome of glioblastoma patients after combined therapy of alkylating chemotherapeutic agents and radiation. The purpose of this study was to assess the frequency of MGMT promoter methylation in soft tissue sarcoma to identify patients eligible for alkylating agent chemotherapy such as temozolomide.Paraffin tumor blocks of 75 patients with representative STS subtypes were evaluated. The methylation status of the MGMT promoter was assessed by methylation-specific polymerase-chain-reaction analysis (PCR). Furthermore, immunohistochemistry was applied to verify expression of MGMT. MGMT gene silencing was assumed if MGMT promoter methylation was present and the fraction of tumor cells expressing MGMT was 20% or less. Methylation specific PCR detected methylated MGMT promoter in 10/75 cases. Immunohistochemical staining of nuclear MGMT was negative in 15/75 cases. 6/75 tumor samples showed MGMT promoter methylation and negative immunohistochemical nuclear staining of MGMT. In none of the tested STS subtypes we found a fraction of tumors with MGMT silencing exceeding 22%.MGMT gene silencing is a rare event in soft tissue sarcoma and cannot be recommended as a selection criterion for the therapy of STS patients with alkylating agents such as temozolomide. Soft tissue sarcoma (STS) form a heterogeneous group of malignant tumors arising in the mesenchymal tissues. A considerable fraction of patients presents with locally advanced disease and requires multimodal therapy including chemotherapy or chemoradiation. DoxorubParaffin blocks of tumor biopsie or resections of 75 patients with representative histological sarcoma subtypes were obtained from the local tumor tissue bank Table . These tFor methylation specific PCR, genomic DNA was isolated from paraffin sections of the selected tissue blocks and bisulfite converted with a commercial kit  according to the manufacturer’s instructions. The bisulfite treated DNAs were amplified by a two-step PCR approach to improve the sensitivity of detecting methylated alleles. For theImmunohistochemical analysis of formalin-fixed, paraffin-embedded sections of the selected tumor samples was performed according to standard protocols. The primary antibody used was MGMT Monoclonal Antibody MT 23.2  after heat antigen retrieval in pH 9 buffer . Only nuclear staining was considered positive. MGMT expression was considered lost when there were 20% or less stained tumor cell nuclei.MGMT gene silencing was assumed if methylation specific PCR detected MGMT promoter methylation AND immunohistochemistry demonstrated MGMT expression in less or equal than 20% of tumor cells. Results of these analyses are presented descriptively. Median and range are given if appropriate.Methylation specific PCR detected unmethylated MGMT promoter in 65/75 cases. In 10/75 tumor samples we found MGMT promoter methylation. Yet, in each of these cases also unmethylated MGMT promoter was detected. An example of the methylation specific PCR is given in Figure Immunohistochemical staining showed inhomogeneous expression of MGMT in most patients. Immunohistochemistry was regarded negative in 15/75 cases. An example of negative and positive staining for MGMT is depicted in Figure In 6/75 tumor samples we detected both MGMT promoter methylation and negative staining of MGMT. In none of the tested STS subtypes we found a fraction of tumors with MGMT silencing (MGMT promoter methylation and negative immunohistochemistry for MGMT) that exceeded 22% , malignant fibrous histiocytoma (MFH) and synovial sarcoma and reported MGMT promoter methylation in 21/62 cases. KawagucA selection criterion for a personalized therapy in soft tissue sarcoma should be easy and reliable to assess, discriminate between tumors that are targeted well and positive in a relevant number of patients. AlthougAll authors declared that they have no competing interest.JJ, FW, PS and PH participated in the design of the study. PH took responsibility for the funding. MH, CS and PS carried out the methylation specific PCR and immunohistochemistry. JJ and MH performed the statistical analysis. JJ drafted the manuscript. All authors read and approved the final manuscript.


While optimal cytoreduction is the standard of care for advanced ovarian cancer, the related post-operative morbidity has not been clearly documented outside pioneering centers. Indeed most of the studies are monocentric with inclusions over several years inducing heterogeneity in techniques and goals of surgery. We assessed the morbidity of optimal cytoreduction surgery for advanced ovarian cancer within a short inclusion period in 6 referral centers dedicated to achieve complete cytoreduction.The 30 last optimal debulking surgeries of 6 cancer centers were included. Inclusion criteria included: stage IIIc- IV ovarian cancer and optimal surgery performed at the site of inclusion. All post-operative complications within 30 days of surgery were recorded and graded using the Memorial secondary events grading system. Student-t, Chi2 and non-parametric statistical tests were performed.180 patients were included. There was no demographic differences between the centers. 63 patients underwent surgery including intestinal resections (58 recto-sigmoid resection), 24 diaphragmatic resections, 17 splenectomies. 61 patients presented complications; One patient died post-operatively. Major (grade 3–5) complications requiring subsequent surgeries occurred in 21 patients (11.5%). 76% of patients with a major complication had undergone an ultraradical surgery (P = 0.004).While ultraradical surgery may result in complete resection of peritoneal disease in advanced ovarian cancer, the associated complication rate is not negligible. Patients should be carefully evaluated and the timing of their surgery optimized in order to avoid major complications. Residual disease after surgery is the main prognostic factor in advanced ovarian cancer Extensive debulking surgery may increase morbidity and delay initiation of chemotherapy The following specialized and high-volume care centers were included in this longitudinal retrospective study.Institut Claudius Regaud, Toulouse; Institut Gustave Roussy, Villejuif; Institut Paoli Calmettes, Marseille; Hôpital Européen Georges Pompidou, Paris; Centre Oscar Lambret, Lille; Institut Bergonié, Bordeaux.In order to avoid any bias due to case selection we included the last 30 patients who underwent complete cytoreductive surgery for an advanced ovarian cancer in each center.All surgeries were performed by/or under the supervision of a senior surgeon (more than 5 years of experience in the management of advanced ovarian cancers).Inclusion criteria: we considered patients with primary epithelial ovarian carcinoma stages IIIC and IV operated in the participating centers. Stage IIIC disease included patients with bulky peritoneal disease, but not those with lymph node involvement only. All surgical procedures had to be performed in the inclusion center, achieving complete  cytoreductive surgery. In case of neo-adjuvant chemotherapy, diagnosis and tumor extent was assessed by an initial laparoscopy.Primary debulking surgery (PDS) was defined as patients undergoing a debulking surgery before any chemotherapeutic treatment. Interval debulking surgeries (IDS) was defined as patient undergoing a debulking surgery after cycles of neo-adjuvant treatment. The numbers of cycles were at the discretion of the treating physician.All data were recorded without identifiers therefore our study did not require informed consent from patients. The need for written informed consent from the participants was waived as this was an audit with no identifiers accessible to the external observers.2, liver resection.All patients’ charts were collected and analyzed by external observers  according to a predefined checklist. Demographic, per and postoperative data were recorded. Sugarbaker scoring system The Memorial Sloan-Kettering Cancer Center surgical secondary events grading system We considered all events occurring within 30 days after debulking surgery.All patients were staged according to the International Federation of Gynecology and Obstetrics (FIGO) system Categorical variables were compared using Fisher exact test and continuous variables were compared using the Student t-test. All statistical tests were two-sided and differences were considered significant at a level of p<0.05.180 patients operated between 2005 and 2008 were included. Demographic characteristics of the population are presented in The mean Sugarbaker score was 14.8, ranging from 10 to 33. Sixty (33.3%) patients had an extensive disease with Sugarbaker score above 10. 128 (71%) patients underwent a laparoscopy before the cytoreductive surgery. Seventy one patients (40%) underwent PDS followed by adjuvant chemotherapy. One hundred and nine patients (60%) received neo-adjuvant chemotherapy and underwent IDS after a mean of 3.2 cycles of neoadjuvant chemotherapy. There were no demographic differences between these two groups. The procedures performed are depicted in There were 61 (33%) complications comprising 40 (22%) minor and 21 (11%) major complications . One hunWe analyzed the different factors involved in the occurrence of complications. ASA score, presence of ascitis and previous surgeries were not associated to the occurrence of complications  in univariate analysis.As demonstrated in Diaphragmatic and small bowel resections were not associated with increased complications. Operative parameters such as duration of surgery, results of the surgery  and use of drains were not associated with post-operative complications on univariate analysis.In this multicentric analysis of surgical morbidity during cytoreductive surgery for advanced ovarian cancer we identified the following variables being predictive for complications: timing of surgery (PDS vs. IDS), the extent of surgery  and realization of a colic or rectal resections. This is concordant with a recent study published by Aletti et al. We used the MSKCC scoring system to classify complications occurring in our patients. Our overall rate of complications was 33% (61 patients/180 patients), with 11% (22 patients/180 patients) having severe complications. The complication rate in a meta-analysis using population-based reports varied between 2.5% to 4.8% We chose a different approach and established an independent audit. We selected 6 different reference centers for the treatment of advanced ovarian carcinomas. In order to avoid selection bias we performed a longitudinal study including the 30 last patients who underwent optimal debulking surgery for ovarian cancer. Two external reviewers independently reviewed the records. The patients were homogenous with no significant demographic differences across the six centers. We included only the last 30 patients per center to reduce as much as possible the bias induced by heterogeneity of care induced by long inclusion periods. Therefore we have limited most of the bias induced by retrospective studies. Several studies have documented that gynecologist-oncologist usually gives optimal level of care in advanced ovarian cancers The inclusion criteria of this study was complete surgery achieved we did not record the number of patients in which complete surgery was attempted. However in multicentric studies performed in France including many of the centers which participated in this study, the rate of complete surgery was around 70%–85% As demonstrated by the univariate and multivariate analysis, most of the major complications were related to bowel resections (14/22).Other radical procedures such as extensive peritoneal resections or diaphragmatic resections were not associated to significant increase of major complications. This is in accordance with Chereau et al., who found an acceptable complication rate of diaphragmatic surgery for stage III/IV ovarian cancer surgeries All patients in our study had immediate re-anastomosis. 13 patients (7.2%) had diverting protective stomas, two were performed for the management of a complication during reoperation.. Most of the anastomosis were performed mechanically (49/61).Mourton et al. published their experience of 70 patients with recto-sigmoid resection and primary anastomosis Our overall leakage related complications were slightly above the range of the literature with 14/180 patients (7.7%) with a lower rate of protective stomas. The rate of complications due to anastomotic leak was 22% (14/61) if we only consider patients with bowel resection. Several factors can explain higher complication rate related to bowel resection in our study: (i) most of the patients had advanced disease with important tumor burden (ii) we only included patients with optimal surgeries and maximal surgical efforts. (iii) there is no reporting bias as external auditors longitudinally included all cases. (iv) finally the low rate of protective stoma might explain why a significant number of the anastomotic leaks directly lead to complications requiring radiological or surgical intervention. Whether we should perform diverting protective stoma in case of bowel (recto-sigmoid) resection in advanced ovarian cancer surgery cannot be answered by our study.We were able to identify some risk factors of anastomotic leak. First Sugarbaker scores of patients with bowel resections who presented major complications were significantly higher than peritoneal carcinosis index of patients with bowel resection without major complications (19+/−5.03 versus 10+/−6.32). It seems that increased morbidity is not due to a unique surgical procedure but to accumulation of multiple bowel resections associated to extensive peritoneal resections that might act as a protective barrier. We also observed a trend for patients who had neo-adjuvant chemotherapy to develop less major complications following bowel resections (13% for patient with neo-adjuvant chemotherapy versus 32% for patients without neo-adjuvant chemotherapy). We had recorded characteristics for bowel anastomosis. While most of them were made automatically with few protective stomas the numbers of complications did not allow us to carry an insightful statistical analysis. In the participating centers the bowel resection and anastomosis was directly made by the gynecologist-oncologists .A meta-analysis by Bristow et al. demonstrated that each cycle of neo-adjuvant chemotherapy will have a negative incidence on the overall survival. However a recent meta-analysis demonstrated than neoadjuvant chemotherapy helped the gynecologic oncologist achieve an increased rate of optimal cytoreduction Our study demonstrates a lower number of complications in patients with neoadjuvant chemotherapy, in relation to the reduction of the need for radical surgery. The recently EORTC randomized trial has demonstrated that neo-adjuvant chemotherapy might be as good as upfront surgery in advanced ovarian cancer This is a multicentric study of post-operative complications in optimal surgery of advanced ovarian carcinomas performed as an independent audit. While the overall complication rate is acceptable and justifies active surgical approach we have been able to point out bowel resection as the main cause of major complications and therefore suggest that patients requiring such procedure to be clearly identified and optimal preventive procedures applied to prevent occurrence of major complications.


Gene regulatory networks play essential roles in living organisms to control growth, keep internal metabolism running and respond to external environmental changes. Understanding the connections and the activity levels of regulators is important for the research of gene regulatory networks. While relevance score based algorithms that reconstruct gene regulatory networks from transcriptome data can infer genome-wide gene regulatory networks, they are unfortunately prone to false positive results. Transcription factor activities (TFAs) quantitatively reflect the ability of the transcription factor to regulate target genes. However, classic relevance score based gene regulatory network reconstruction algorithms use models do not include the TFA layer, thus missing a key regulatory element.Gene expression and Transcription factor activity based Relevance Network (GTRNetwork). Different combinations of TFA prediction algorithms and relevance score functions have been applied to find the most efficient combination. When the integrated GTRNetwork method was applied to E. coli data, the reconstructed genome-wide gene regulatory network predicted 381 new regulatory links. This reconstructed gene regulatory network including the predicted new regulatory links show promising biological significances. Many of the new links are verified by known TF binding site information, and many other links can be verified from the literature and databases such as EcoCyc. The reconstructed gene regulatory network is applied to a recent transcriptome analysis of E. coli during isobutanol stress. In addition to the 16 significantly changed TFAs detected in the original paper, another 7 significantly changed TFAs have been detected by using our reconstructed network.This work integrates TFA prediction algorithms with relevance score based network reconstruction algorithms to reconstruct gene regulatory networks with improved accuracy over classic relevance score based algorithms. This method is called E. coli gene transcriptome data gives a set of potential regulatory links with promising biological significance for isobutanol stress and other conditions.The GTRNetwork algorithm introduces the hidden layer TFA into classic relevance score-based gene regulatory network reconstruction processes. Integrating the TFA biological information with regulatory network reconstruction algorithms significantly improves both detection of new links and reduces that rate of false positives. The application of GTRNetwork on  Gene regulatory networks play an essential role in controlling gene expression and ensuring that the right genes are expressed or silenced at the right time in the right place to make the organism function appropriately. Better understanding of gene regulatory structure aids biological researchers and biochemical engineers in obtaining more complete views of the complex gene expression and regulatory mechanisms in organisms.In the gene regulation process, an active transcription factor (TF) can bind DNA and control gene expression. However, many TFs are not inherently active. Complex mechanisms, such as forming dimers, interacting with signal metabolites or binding specific micro-RNAs, are needed in order to control the activities of these TFs . The actSince TFA is governed by various complex molecular interactions, it is difficult to determine directly from experiments, especially if the activation mechanism is unknown. However, it is possible to computationally predict the change of TFAs relative to a reference state using transcriptome data and a known TF-gene network architecture ,2. Netwoin vivo  is the relative gene expression level matrix and L × M matrix [TFAr] is the relative transcription factor activities, the elements ijEr(t) = ijE(t)/ijE(0) and kjTFAr(t)/kjTFAr(0), N × L matrix [CS] is the control strength matrix of transcription factors and genes. The gene expression model in Eq. (4) can be decomposed into matrix [CS] and matrix log([TFAr]) using different algorithms.where Er] can be obtained from transcriptome experiments such as DNA microarrays or RNAseq, and the control strength information must be initialized from the literature e.g. RegulonDB  and [CS], transcription factor activities log([TFAr]) can be estimated. There are three major approaches to estimate log ([TFAr]) expectation maximization (EM) approach (e.g. gNCA-r) ) matrix, log ( and log ([TFAr]) need to fit three criteria given below to ensure the uniqueness of the decomposition  must have full-column rank.(i) The connectivity matrix  must have at least L-1 zeros.(ii) When a node in the regulatory layer is removed along with all of the output nodes TFAr], must have full row rank. In other words, each regulatory signal cannot be expressed as a linear combination of the other regulatory signals.(iii) The matrix, log  with elements ij Zgene regulatory networks with different sensitivities can be reconstructed by searching for gene regulatory links containing TF genes with the Z score larger than the threshold. The information of TF genes (which genes encode TFs) can be found from database such as RegulonDB Inputs: a) Log 2 ratio transcriptome data in matrix [C]b) Initial TF-gene network topology in adjacency matrix  and matrix [C]. 1. Match the genes between the matrix [Er] and store the reduce matrix as [Er0]. Remove unmatched genes in [C] and store the reduced matrix in [C0].Remove unmatched TFs and genes in [Er0] and [C0] to fit the criteria.2. If the TFA prediction algorithm is gNCA-r or FastNCA, check the three criteria described in TFA prediction section and reduce the matrix [2 ratio TFA matrix [TFA] from matrix [Er0] and [C0].3. Apply TFA prediction algorithm to predict the logM] between TFAs and all expression levels of all genes from matrix [TFA] and [Er].4. Calculate the relevance score matrix [Z] of relevance score matrix [M] using CLR algorithm.5. Calculate the joint statistical likelihood matrix [Z] so that there are S elements in [Z] greater than T. For all the TF-gene pairs having a Z score greater than T, construct a regulatory link.6. Set a threshold T for matrix [7. If the operon list is available, check and add all genes in the same operon of TF target genes to the regulatory target set of the TF.YF developed and implemented the GTRNetwork algorithm and drafted this manuscript. LRJ and JD developed the initial concept and suggested ways to improve the algorithm and testing methods. All authors read and approved the final version of the manuscript.Test results of GTRNetwork Algorithm combinations, GTRNetwork algorithm using TF-gene network topologies providing different level of information as input, the input initial TF-gene network topologies are obtained by randomly deleting 70%, 50%, 30% or 10% links of the TF-gene links data from RegulonDB 7.0 [onDB 7.0 .Click here for fileE. coli predicted using GTRNetworkPotential new regulatory links of . Gene expression data is obtained from M3D database [database  and contdatabase  is used database Click here for file


Campylobacter jejuni transcriptional response under iron replete and iron limited conditions was characterized using RNA-seq. We have identified 111 novel C. jejuni 5’UTRs and mapped 377 co-transcribed genes into 230 transcriptional operons. In contrast to previous microarray results, the C. jejuni iron stimulon is less extensive than previously believed and consists of 77 iron activated genes and 50 iron repressed genes. As anticipated, the iron repressed genes are primarily those involved in iron acquisition or oxidative stress defense. Interestingly, these experiments have revealed that iron is an important modulator of flagellar biogenesis with almost all the components of the flagella found to be iron activated. Given that motility is a well-known C. jejuni colonization factor, this suggests that there is an important regulatory coupling of flagellar biogenesis and iron level in C. jejuni. In addition we have identified several consensus mutations in the C. jejuni NCTC11168 strain that are widespread in the Campylobacter research community and which may explain conflicting phenotypic reports for this strain. Comparative analysis of iron responsive genes with the known Fur regulon indicates that many iron responsive genes are not Fur responsive; suggesting that additional iron regulatory factors remain to be characterized in C. jejuni. Further analysis of the RNA-seq data identified multiple novel transcripts including 19 potential ncRNAs. The expression of selected ncRNAs was confirmed and quantified with qRT-PCR. The qRT-PCR results indicate that several of these novel transcripts are either Fur and/or iron responsive. The fact that several of these ncRNAs are iron responsive or Fur regulated suggests that they may perform regulatory roles in iron homeostasis.The genome-wide  Campylobacter jejuni face additional challenges in acquiring sufficient iron as their hosts typically restrict free iron to prevent unwanted microbial growth  were plotted according to Log2(fold change). Genes identified as being iron repressed by both techniques are either dark purple (iron repressed) or dark green (iron activated). Genes only identified in the iron seq analysis are shown as light purple or green squares (repressed and activated respectively). Genes in blue were iron activated in the RNA-seq and iron repressed in the microarray. Genes in grey were not significantly differentially expressed in the RNA-seq analysis. See Table S2 for specific gene details.(TIF)Click here for additional data file.Figure S3Comparison between Fur regulation and iron responsive genes.The RNA-seq iron regulon was compared to the direct and indirect Fur regulons determined using ChIP-chip and microarrays respectively . While the majority of the iron repressed genes were regulated by Fur, most iron activated genes were not Fur responsive. This suggests the presence of additional iron responsive regulatory elements in C. jejuni. The length of the bars/ribbons is proportional to the number of genes in each group. Figure made using Circos v0.54 [52].(TIF)Click here for additional data file.Figure S4Average insert size of Illumina sequencing libraries.The average insert size for each of the Illumina sequencing libraries constructed was calculated based on the paired-read alignment to the C. jejuni NCTC 11168 genome.(TIF)Click here for additional data file.Figure S5Additional ncRNA validation experiments.Panel A: Northern blot showing expression of ncRNA_Z across different conditions. The 5S RNA is also shown as a loading control. Densitometry analysis did not reveal differential expression of ncRNA_Z when normalized to the loading control. Panels BCD: qRT-PCR experiments of ncRNAs AA, Z and P with varying levels of template RNA for each RT reaction. Differentially expressed conditions  are indicated with an asterisk. (TIF)Click here for additional data file.Table S1Statistics for high throughput sequencing runs.Listed are the read statistics for the high throughput sequencing runs performed in this study as well as runs that were reanalyzed from other studies .(XLSX)Click here for additional data file.Table S2Genes differentially expressed between iron limited and iron replete conditions.Listed are the genes that were found to be significantly differentially expressed under iron limitation .(XLSX)Click here for additional data file.Table S3Fold changes in gene expression as determined by RNA-seq and microarray analysis.Listed are the fold changes in gene expression as determined by either RNA-seq or previous microarray analysis [4].(XLSX)Click here for additional data file.Table S4RPKM values for each gene and sequencing run.Listed are the RPKM values for each C. jejuni gene in each sequencing reaction.(XLSX)Click here for additional data file.Table S5C. jejuni.Percent contribution of COG categories to the total transcriptome of Listed are the overall contributions of each COG category to the total transcriptome.(XLSX)Click here for additional data file.Table S6C. jejuni as determined by RNA-seq.5'UTRs in Listed are the locations and lengths of the 5’UTR regions that were identified by manually inspecting reads that extended beyond the C. jejuni NCTC11168 predicted start codon. Included are those identified using previous a previous RNA-seq study [16] and those identified using dRNA-seq [17].(XLSX)Click here for additional data file.Table S7C. jejuni as determined by RNA-seq.Novel transcripts identified in Listed are the locations and strand origin of the novel C. jejuni NCTC11168 transcripts identified in this study.(XLSX)Click here for additional data file.Table S8Genes Identified in Operonic Structure.Listed are the genes identified as being in operonic structure by using the aligned RNA-seq paired-end reads to infer co-transcription. (XLSX)Click here for additional data file.Table S9C.Operonic Transcripts in jejuni. Listed are genes present in operonic structure that may form potential operonic transcripts.(XLSX)Click here for additional data file.


A 58 year-old male was referred to our institute for recurrent palpitations followed by ICD shocks. The patient had a history of ischemic cardiomyopathy (EF 25%) and had received a dual-chamber ICD two years earlier for primary prevention of sudden cardiac death. The device interrogation showed multiple episodes of tachycardia and anti-tachycardia pacing (ATP), which frequently terminated the tachycardia. However, occasional failure to terminate the tachycardia or immediate re-initiation within the redetection window led to ICD shocks. A typical episode of tachycardia with its initiation is shown in Device interrogation showed the onset of tachycardia with a near-simultaneous atrial and ventricular activity (fourth beats). Such an onset of tachycardia with less atrial beats than ventricular beats rules out atrial tachycardia. Device interrogation also revealed multiple sequences of ATP that entrained the tachycardia with a VAV response further ruling out atrial tachycardia. The ventricular electrogram morphology during sinus rhythm was similar to that during tachycardia, and the 12-lead EKG demonstrated identical wide QRS morphology during sinus rhythm and during tachycardia . The similarity in QRS morphology (and ventricular electrogram) during sinus rhythm and tachycardia could be consistent with bundle branche reentry VT as those arrhythmias are typically seen in patients with severe cardiomyopathy (like our patient). However, the tachycardia initiation with simultaneous V and A makes this diagnosis unlikely.  Based on these observations, the two differential diagnoses are:-junctional ectopic tachycardia (JET) that starts with V (4th beat) and conducts retrogradely to A (4th beat).-typical slow fast atrioventricular nodal reentrant tachycardia (AVNRT) that initiates with A (3rd beat) leads to V (3rd beat) through the fast pathway and gives a second V (4th beat) through the slow pathway (1:2 response).The patient underwent an electrophysiology study after discontinuing the beta blockers for 5 days. The baseline intracardiac intervals were unremarkable. The mode of tachycardia initiation   was either spontaneous see  or afterFurther EP maneuvers were performed to differentiate JET and AVNRT as recently reported . APD delBased on these findings, the tachycardia was thus diagnosed as AVNRT. The slow pathway area was successfully targeted with one RF application to render the tachycardia non-inducible.  No arrhythmia has been observed on the ICD telemetry for more than a year.In the case presented here, a sinus beat led to 1:2 response with conduction via the fast pathway generating the first QRS* followed by conduction over the slow pathway leading to the second QRS** and induction of AVNRT .An atrial extrasystole causing 1:2 response is an uncommon phenomenon that has been previously described . However


CRT is an effective treatment for patients with heart failure and intraventricular conduction delay. However, clinical non-response rates remain unacceptably high; emphasizing the need for improved selection criteria. Central to the underlying mechanism of resynchronization is the relationship between mechanical and electrical activation. The purpose of this study was to demonstrate that mechanical and electrical activation can be quantified in patients with CRT devices combining cardiovascular magnetic resonance (CMR) and electrical myocardial imaging (EMI).Non-invasively characterize mechanical and electrical activation patterns in patients undergoing cardiac resynchronization therapy (CRT)Patients implanted with biventricular CRT for greater than 9 months (N=10) were mapped with 120 body surface electrodes during both intrinsic activation (device off) and CRT pacing. CMR tagging was also acquired with pacing on/off to perform strain analysis. The circumferential uniformity ratio (CURE) was computed to characterize temporal-spatial mechanical dyssynchrony . CMR images of the torso and heart were then used to reconstruct electrical epicardial activation maps using a novel EMI technique. Electrical synchrony was indexed by the mean total activation difference time . Only 2 patients underwent full analysis.CURE decreased from 0.73 to 0.51 when the device was turned off, indicating an increase in mechanical dyssynchrony. The corresponding LVEF decreased as well in on/off modes from 42% to 29%. Figure CMR and non-invasive EMI are feasible in patients with CRT devices. This ability to quantify mechanical and electrical synchrony before and after CRT may provide insights into mechanisms of resynchronization and lead to robust dyssynchrony metrics that will identify appropriate candidates for CRT.


Radioembolization offers a novel way to treat the nonresectable, liver predominant hepatic malignancies with better tumor response and overall progression-free survival rates. Transarterial catheter-based radioembolization procedure involves the hepatic arterial administration of glass- or resin-based beta emitting Yttirum-90 microspheres. Safe delivery of the tumoricidal radiation dose requires careful angiogram planning and coil embolization to quantify lung shunting and prevent systemic toxicity, respectively. Diagnostic pretreatment angiogram also serves to identify the hepatic arterial variant anatomy and other coexisting pathologies that might require a different or alternative approach. We describe a complex case of celiac artery stenosis with tortuous pancreaticoduodenal arterial arcade precluding access to the right hepatic artery for performing radioembolization. Celiac artery stenting of the stenosis was performed to facilitate subsequent safe and successful Yttrium-90 microsphere radioembolization. She was referred for Yttrium-90 (90Y) radioembolization procedure for treatment of unresectable stable right hepatic lobe metastatic lesions  within the right hepatic artery (A planning or mapping angiogram was performed. The aortogram demonstrated a tight (approximately >90%) stenosis of the proximal celiac axis with mild poststenotic dilatation . A superstenosis . Subsequstenosis . Absencec artery . 90Y microspheres  was administered within the right hepatic artery through the stented celiac artery approach during the treatment phase utilizing the standard protocol till the end point. Bremsstrahlung images obtained after the 90Y radioembolization confirm proper delivery of microspheres to the targeted region without any abnormal extratumoral distribution (Approximately 27.8 mCi (1029 MBq) of ribution . Patient90Y microspheres are permanent biocompatible and non-biodegradable medical devices with a 90Y beta emitter within glass spheres or adsorbed onto resin spheres. Tumor cells receive the blood supply predominantly through the arterial system compared to the normal liver tissue which receives supply from the portal venous system. This preferential flow facilitates selective delivery of the radioactive microspheres to the tumor during transcatheter arterial therapies [90Y microsphere treatment, multiple studies reported improved tumor response and delayed time to progression compared to other treatment modalities [Microsphere radioembolization therapy is currently approved for unresectable hepatic metastatic disease from colorectal cancer  and unresectable hepatocellular carcinoma as a humanitarian device exemption  within the United States. herapies . The selherapies . Given tdalities –7. 90Y microspheres are administered selectively up to the calculated dose or until flow stasis is achieved. A posttreatment Bremsstrahlung scan confirms dose delivery to tumor and absence of extrahepatic activity. These technical details and guidelines have been meticulously described previously [The first step in the multidisciplinary approach is the planning or mapping angiogram phase that serves to confirm the normal or variant hepatic arterial anatomy and to coil embolize the GDA. Subsequently, Tc-99m MAA is injected to confirm lack of extrahepatic distribution and quantify the intrinsic tumoral lung shunting, which is used in conjunction with anatomical imaging volume and laboratory data for dose calculation. The treatment phase angiogram serves to reconfirm the arterial anatomy of the tumor, after which the eviously –10. Posteviously , 12. Arterial variants and abnormalities in the mesenteric and hepatic circulations might require alternate techniques for safe and effective radioembolization . Celiac 90Y microsphere therapy in cases of celiac artery stenosis [90Y radioembolization treatment in English literature.Up to date, there have been a few case reports demonstrating the utilization of the SMA-inferior PDA approach for stenosis , 13. The90Y microspheres radioembolization offers a unique opportunity to treat nonresectable, liver predominant hepatic malignancies with good tumor response, and overall progression free survival rates. Hepatic arterial variants and abnormalities such as arterial stenosis might limit access to the hepatic arterial branches supplying the tumor, and therefore, require alternate approaches or additional minimally invasive endovascular interventions. We report the use of stenting procedure in a case of celiac artery stenosis to facilitate hepatic 90Y microsphere radioembolization. This technique can be performed as a first line approach or in cases where the hepatic branches are not technically accessible through the conventional SMA-PDA arcade approach. Although the use of celiac axis stenting for 90Y microspheres radioembolization has been successful in our case, this approach should be evaluated further in a larger study population.


Chronic myelogenous leukemia (CML) is a leukemic stem cell (LSC)-driven myeloproliferative disorder and is associated with a characteristic chromosomal translocation which generates a constitutively active tyrosine kinase, the BCR-ABL oncoprotein. The standard treatment therapy for CML patients is the BCR-ABL tyrosine kinase inhibitor (TKI) imatinib. It is a life-long treatment due to the fact that LSCs are resistant to TKIs. Since its introduction, imatinib has improved the 5-year survival rate up to 90%. An emerging problem is resistance to imatinib, which is mainly caused by mutations inside the BCR-ABL kinase domain, and its increasing incidence during disease progression. It has been reported that BCR-ABL drives its own mutation via upregulation of reactive oxygen species (ROS) causing oxidative DNA damage. Among the several dozens of intensively characterized mediators of BCR-ABL action, the transcription factor STAT5 is among the few ones that is critical for leukemia initiation and maintenance and it has been shown that STAT5 becomes upregulated during disease progression.qPCR analysis of primary CML patient samples reveal a positive correlation of STAT5 mRNA levels and BCR-ABL mutations. Using BCR-ABL transformed murine cell lines retrovirally overexpressing STAT5A or STAT5B, we can show that STAT5 triggers ROS production leading to an increase in DNA double-strand breaks.We hypothesize that STAT5 is an important mediator of imatinib resistance in CML due to its ability to drive ROS production consequently leading to BCR-ABL mutations.


Neurons co-express a multitude of different ion channels that determine the electrophysiological behaviour of the cell. Recent experimental observations have revealed that the expression of different ion channels varies dramatically within neurons of a defined type, even when these neurons exhibit stereotyped electrical properties -5. This


Approved drugs serve as functional probes for analyzing disease-relevant biochemical pathways or are screened against proteins to identify novel medical applications resulting in a repurposing of known drugs. Despite these benefits of approved drugs, it is almost impossible to physically obtain a complete collection. Commercial vendors cover about 50-65% of drugs. Academic efforts such as the Johns Hopkins Clinical Compound Library (JHCCL)  and the


Random or stochastic monoallelic expressed genes (StMA genes) represent a unique form of monoallelic expression where allelic choice is made at random early in development. The consequential clonal diversity provides opportunity for functional heterozygosity in tissues such as the brain, and can impact on both development and disease. We investigate the relationship of StMA expressed genes previously identified in clonal neural stem cells with the neurodevelopmental disorders autism and schizophrenia. We found that StMA genes show an overrepresentation of schizophrenia risk candidates identified by genome wide association studies from the genetic association database. Similar suggestive enrichment was also found for genes from the NHGRI genome-wide association study catalog and a psychiatric genetics consortium schizophrenia dataset although these latter more robust gene lists did not achieve statistical significance. We also examined multiple sources of copy number variation (CNV) datasets from autism and schizophrenia cohorts. After taking into account total gene numbers and CNV size, both autism and schizophrenia associated CNVs appeared to show an enrichment of StMA genes relative to the control CNV datasets. Since the StMA genes were originally identified in neural stem cells, bias due to the neural transcriptome is possible. To address this, we randomly sampled neural stem cell expressed genes and repeated the tests. After a significant number of iterations, neural stem cell expressed genes did not show an overrepresentation in autism or schizophrenia CNV datasets. Therefore, irrespective of the neural derived transcriptome, StMA genes originally identified in neural stem cells show an overrepresentation in CNVs associated with autism and schizophrenia. If this association is functional, then the regulation (or dysregulation) of this form of allelic expression status within tissues such as the brain may be a contributory risk factor for neurodevelopmental disorders and may also influence disease discordance sometimes observed in monozygotic twins.  Several forms of monoallelic gene expression have been identified. Imprinting, a form of monoallelic gene expression based on parent-of-origin effects, is highly represented in the brain transcriptome , and sevWe utilized allelic expression measures from a previous study of clonal neural stem cells to construct a list of stochastic monoallelic (StMA) expressed genes . A clonehttp://geneticassociationdb.nih.gov), NHGRI GWAS catalog (http://www.genome.gov/gwastudies) and 81 associated SNPs from the psychiatric genetics consortium [P < 2 × 10−5 . Genes mapping close to or containing the risk SNPs in the latter study were identified with Galaxy analysis server [Enrichment analysis was carried out on both association studies  and CNV datasets (using StMA gene lists only). Candidate genes from association studies were taken from the Genetic Association Database (s server . Associahttp://bbgre-dev.iop.kcl.ac.uk), 120 autism patients with de novo CNVs from the DECIPHER consortium repository (http://decipher.sanger.ac.uk), 843 autism spectrum disorder patients from a developmental disability CNV study [http://www.ncbi.nlm.nih.gov/dbvar ) [Disease associated CNV locations were obtained from a number of sources: 75 autism patients from the Brain Body and Genetic Resource Exchange (BBGRE) CNV database  –17. For We used publically available neuropsychiatric datasets to look for enrichment of StMA genes relative to control data. First, occurrence of StMA genes and BA genes within datasets from the genetic association database  were exaSince allelic status of StMA genes correlates with overall transcript levels ,6,19, suDisease datasets were obtained from CNV databases and publications. Autism CNVs from BBGRE and DECIPHER databases show high enrichment of StMA gene occurrence relative to the total number of genes (3.2-5.0 fold). The number of CNV regions containing StMA genes also showed a higher proportion of autism and schizophrenia CNV regions containing StMA genes (14 to 25%) compared with dbVAR control CNVs (2%). When the CNV size is taken into account to calculate gene density, the frequency of StMA genes per megabase of CNV was 2 to 4 fold higher than the dbVAR control CNV regions. The remaining two autism publication derived datasets represent a comprehensive CNV morbidity map of neurodevelopmental delay, with some of the patients having an autism spectrum disorder diagnosis  and 5 geThe schizophrenia datasets were obtained from two publications. We used 12 replicated schizophrenia CNVs documented in a recent meta-analysis review by Since our identified StMA genes were originally identified in neural stem cells , it is cWe have demonstrated an associative link of neural stem cell identified StMA genes with autism and schizophrenia GWAS and CNV identified genes. Pathological neuropsychiatric-associated CNVs are likely to alter the transcript levels of specific risk genes . SimilarConsistent with our disease enrichment findings, allelic imbalances identified within induced pluripotent stem cell derived differentiated human neurons also show a modest association for schizophrenia and autism candidate genes . Allele In summary, we have previously identified genes showing stochastic monoallelic expression patterns in clonal neural stem cells, together with a number of genetic and epigenomic associations . Here weTable S1Control CNV datasets used to construct the dbVAR control dataset.Total gene content and StMA gene occurrence are shown for each data set. The Conrad et al (2010) control study consisted of two CNV lists – a CNV discovery dataset and HapMap based dataset as shown in the table.(DOCX)Click here for additional data file.Table S2Summary of CNV data sets based on genes mapped to minimal overlap CNV coordinates.Total gene content and StMA gene occurrence are shown for autism (BBGRE and DECIPHER) and schizophrenia  CNV datasets based on genes mapped to minimal overlap CNV coordinates. Enrichment tests were carried out relative to dbVAR controls, a merger of CNV loci as shown in table S2. (DOCX)Click here for additional data file.File S1Allelic expression gene lists and CNV coordinates and their associated genes used in the study.(XLSX)Click here for additional data file.File S2R code used for CNV enrichment analysis and permutation test.(TXT)Click here for additional data file.


Urosepsis associated with urinary tract calculi is a critical disease. Patients with this condition occasionally require drainage, mostly ureteric stent placement and these patients need longer hospitalization. However, indications for timely ureteric stenting for urosepsis associated with urinary tract calculi have not been clearly determined. The objective of the study was to evaluate whether earlier stent placement might lead to shorter length of stay (LOS) in hospitals.For patients who required ureteric stent procedures for urosepsis associated with urinary tract calculi in our hospital from July 2008 to July 2012, we compared the LOS in our hospital between patients with earlier stenting and those without it. A linear regression model was used for multivariable-adjusted comparison.In a total of 30 patients , the mean days from emergency room admission to ureteric stenting was 3.53days  and the overall mean LOS was 36 days . The early-stenting group  had significantly shorter LOS than the delayed-stenting group  with adjusted β coefficient of -26 days .In patients with urosepsis associated with urinary tract calculi, earlier stenting within 2 days of admission may reduce the hospital LOS.


C), which acquires altered conformation, form aggregates and is resistant to proteolysis. PrPC is a cell surface glycosyphosphatidylinositol (GPI)-anchored protein of 208-209 aminoacids which is codified by a single exon and whose expression is developmentally regulated. In adult animals PrPC is highly expressed in the central nervous system but peripheral nervous system and other organs or tissues also express this protein.Prions are proteinaceus infectious agents associated to invariable fatal neurodegenerative disorders named transmissible spongiform encephalopathies or prion diseases. The mechanism associated with disease propagation lays on post-translation modifications of the cellular prion protein  or Hop (its human homologue). Comparative experiments using neurons derived from wild type and PrPC-null mice demonstrated that PrPC binding to vitronectin promoted axonal growth in dorsal root ganglion neurons (DRG). In addition, PrPC interaction with laminin activated metabotropic glutamate receptors, Ca2+ mobilization and PKC activation promoting neuronal plasticity and memory formation in rats. STI1 was the most explored PrPC ligand. The PrPC-STI1 engagement induces PKA an ERK1/2 activation, which increase neuronal survival and differentiation. The signaling pathways modulated by PrPC-STI1 also included Ca2+ influx by activation of α-7 nicotinic acetylcholine receptor, induction of PI3K and mTOR. These last two pathways were associated to increased protein synthesis. An increment on the self-renewed of neuronal progenitors is also modulated by PrPC-STI1 engagement.The use of cellular models led to the characterization of PrPC at the neuronal membrane and recently we demonstrated that this protein is abundantly secreted by astrocytes. Secreted STI1 plays an autocrine activity upon astrocytes modulating their proliferation and differentiation. The mechanism associated with STI1 secretion is under evaluation but preliminary results indicated that this protein is secreted by microvesicles derived from multivesicular bodies.The co-chaperone STI1 is known to interact with PrPC and STI1/Hop in cellular survival and differentiation raised questions regarding their involvement in tumoral processes. In fact, the literature has been pointed that PrPC expression contributes to cancer progression and resistance to various cancer therapies. STI1/Hop expression was also associated with the proliferation of tumor cells. Our data pointed that STI1 secreted by glioblastoma cells has an autocrine function mediating proliferation of these cells by binding to PrPC. We are presently evaluating how to interfere with the PrPC-STI1 binding in order to modulate tumor growth.The involvement of PrPC-STI1 engagement is an interesting therapeutic target for both neurodegeneration and cancer and investments should be done to address these issues.Together our results point that PrPThere are no competing interests in this presentation.


Dear Editor,I was interested to read the paper entitled "Effects of clonidine premedication upon postoperative shivering and recovery time in opium addicted and non-addicted patients by elective leg fracture surgeries" . I note


ΔMye mice, which lack IKKβ in myeloid cells. In summary, the oral or systemic route of allergic sensitization and IKKβ signaling in myeloid cells regulate how the gender influences allergen-specific responses and lung allergic inflammation.Gender influences the incidence and/or the severity of several diseases and evidence suggests a higher rate of allergy and asthma among women. Most experimental models of allergy use mice sensitized via the parenteral route despite the fact that the mucosal tissues of the gastrointestinal and respiratory tracts are major sites of allergic sensitization and/or allergic responses. We analyzed allergen-specific Ab responses in mice sensitized either by gavage or intraperitoneal injection of ovalbumin together with cholera toxin as adjuvant, as well as allergic inflammation and lung functions following subsequent nasal challenge with the allergen. Female mice sensitized intraperitoneally exhibited higher levels of serum IgE than their male counterparts. After nasal allergen challenge, these female mice expressed higher Th2 responses and associated inflammation in the lung than males. On the other hand, male and female mice sensitized orally developed the same levels of allergen-specific Ab responses and similar levels of lung inflammation after allergen challenge. Interestingly, the difference in allergen-specific Ab responses between male and female mice sensitized by the intraperitoneal route was abolished in IKKβ Thus, the magnitude and quality of the allergen-specific immune response are controlled by the dose and frequency of exposure to allergen, the type of adjuvant, the genetic background, the route of sensitization and the gender. Previous studies showed that the subcutaneous route was more effective than the intraperitoneal (ip) route at inducing IgE antibodies Cytokines produced by myeloid cell subsets, including macrophages and dendritic cells, influence differentiation of naive T lymphocytes into Th2 cells capable of supporting induction of IgE Abs and effector cells involved in allergic responses. The nuclear factor κB (NF-κB) pathway plays an important role in cytokine responses to a variety of stimuli Experimental murine models of allergy and asthma use mostly female mice and the possible influence of gender on allergen-induced lung inflammation has not been thoroughly investigated. To address this question, we compared allergen-specific Ab responses and subsequent responses to nasal allergen challenge in male and female mice sensitized either orally or via ip injection with the model food antigen ovalbumin (OVA) and cholera toxin as adjuvant. We show that ip sensitization of adult mice results in a significant gender-disparity with female mice developing higher IgE and Th2-associated response and allergic airway inflammatory responses upon allergen challenge. Such differences were not seen in male and female mice sensitized via the gastrointestinal tract, suggesting that gender differentially influences allergic sensitization depending on the route of exposure.ΔMye mice, in which IKKβ-dependent NFκB signaling was eliminated in myeloid cells Wild-type C57BL/6 mice were obtained from the Jackson Laboratories  or NCI  and acclimated in our facility before use. The IKKβImmunohistochemistry was performed on 5 μm sections of formalin-fixed and paraffin-embedded tissues. Lung tissues were inflated before fixation and tissue sections were stained with hematoxylin and eosin according to standard protocols. Quantification of lung inflammation was performed in a blinded fashion by two independent investigators as previously described Oral sensitization to the food antigen was performed on days 0 and 7 by intragastric administration of 250 μl of PBS containing 1 mg of ovalbumin (OVA) and 10 μg of cholera toxin as adjuvant. Parenteral sensitization was performed on days 0 and 7 by intraperitoneal injection of 100 μl of PBS containing 100 μg of OVA and 1 μg of cholera toxin. Nasal antigen challenges were performed on days 15, 16 and 19 with 100 μg of OVA as previously described Plasma samples were collected one week after each sensitization, on days 7 and 14, and plasma levels of OVA-specific antibodies were measured by enzyme-linked immunosorbent assay (ELISA) as previously described Mechanical properties of the mouse lung were assessed on anesthetized mice using the forced-oscillation technique as previously described Cytokine mRNA responses antibodies were analyzed by Real-Time RT-PCR as previously described 2 below the detection limit. All tests were considered significant at a probability of p<0.05.Results are expressed as the mean ±1 SD. Unless otherwise indicated, statistical significance was determined by Student’s T test or by ANOVA followed by the Fisher Least Significant Difference Test . For statistical analysis, antibody levels below the detection limit were recorded as two logWe first examined the antibody responses after oral or parenteral  sensitization. In mice sensitized by the ip route, the IgE, IgG2a/c levels were higher in females . In contWe next examined whether the gender-induced difference in allergen-specific Ab responses would affect responses to nasal antigen challenge . HistoloAllergen-induced airway hyper-responsiveness (AHR) is an important outcome of inflammation and airway remodeling in asthma. Twenty-four hours after the last challenge with OVA , there wAirway hyper-responsiveness also is controlled by cytokines secreted in response to antigen challenge; we and others have previously shown that differences in cytokine responses can lead to discrepancies between lung inflammation and AHR ΔMye mice). Interestingly, no difference was found in the level of serum IgG and IgE antibodies between male and female IKKβΔMye mice sensitized by the parenteral route, except for IgA, which was higher in females (ΔMye mice sensitized by the oral route exhibited no difference in antibody responses (Myeloid cells are major contributors of pro-inflammatory cytokines. As mentioned above, it has been suggested that an increased expression of interleukin-6 by monocytes in women could contribute to the higher risks of inflammatory disorders in this population when compared to men  females . As with females , male anesponses .ΔMye mice sensitized orally failed to develop airway reactivity to methacholine (ΔMye mice sensitized via the parenteral route also failed to show increased lung resistance to methacholine challenge. Surprisingly, male IKKβΔMye mice sensitized via the parenteral route retained the ability to develop allergen-induced airway hyper-responsiveness, suggesting that the IKKβ pathway plays a major role in allergen-induced airway hyper-responsiveness in female mice. This discrepancy in lung responses between male and female IKKβΔMye mice sensitized via the parenteral route was further confirmed by the analysis of lung inflammation (ΔMye mice showed higher levels of inflammation than those of female IKKβΔMye mice sensitized via the same parenteral route (We next examined whether the changes in allergen-specific responses induced by IKKβ deficiency in mice sensitized by the parental route also affect allergen-induced airway hyper-responsiveness. Twenty-four hours after the last antigen challenge, both male and female IKKβacholine . Female ammation . Thus, lal route .Experimental and clinical data indicate a difference in gender susceptibility to airway hyper-responsiveness and allergic inflammation. However, these differences appear to vary with the experimental system used and the end point analyzed. There are known differences in lung airway size, alveolar number and volume between males and females Several mouse strains have been studied to characterize pulmonary functions Airway hyper-responsiveness to cholinergic agents like methacholine is a defining feature in experimental asthma. Furthermore, airway hyper-responsiveness in the context of allergic asthma generally is associated with allergic inflammation characterized by eosinophilia and mucus production Most murine models of asthma evaluate responses after intraperitoneal sensitization followed by several nasal challenges. The oral route of sensitization we used in this study more likely recapitulates events that occur during common sensitization to food allergens. The importance of the route of sensitization has been already demonstrated by Hansen et al. who compared ip versus nasal sensitization ΔMye mice further supports the notion that myeloid cells and other innate cells at the site of allergic sensitization control the profile of allergen-specific B and T cell responses and subsequent pathologic responses to allergen challenge.The immune system in mucosal tissues of the GI tract has evolved alongside, but separate from the general bloodstream to adapt to the unique environment in this site. Mine et al. In summary, we have shown than allergic sensitization via the gastrointestinal tract leads to similar patterns of allergen-specific immune responses in male and female mice. We also showed that IKKβ in myeloid cells plays a key role in the gender disparity with regard to responses to intraperitoneal sensitization and further studies are warranted to better understand underlying mechanisms. Finally, our findings add support to the notion that airway hyper-responsiveness to methacholine challenge do not always reflect differences in airway inflammation. Thus, careful selection of experimental models and assays are crucial to study mechanisms of gender disparity for allergic diseases.


L-Gulonolactone oxidase-deficient mice (Gulo(−/−)) were used to study the effects of ascorbate deficiency on aortic relaxation and lowering of blood pressure by nitroglycerin (GTN). Special emphasis was given to changes in expression and activity of vascular aldehyde dehydrogenase-2 (ALDH2), which has been recently characterized as key enzyme of vascular GTN bioactivation.Ascorbate deficiency was induced in Gulo(−/−) mice by ascorbate deprivation for 4 weeks. Some of the animals were concomitantly treated with the proteasome inhibitor bortezomib and effects were compared with ascorbate-supplemented Gulo(−/−), untreated or nitrate-tolerant wild-type mice. Aortic relaxation of the experimental groups to GTN, acetylcholine and the NO donor diethylamine NONOate was studied. Changes in mRNA and protein expression of vascular ALDH2 were quantified by real-time PCR and immunoblotting, respectively, and aortic GTN denitration rates were determined by thin layer chromatography. Ubiquitination of proteins was analyzed by immunoblotting.Like GTN treatment, ascorbate deprivation induced vascular tolerance to GTN that was associated with markedly decreased rates of GTN denitration. Ascorbate deficiency did not affect ALDH2 mRNA levels, but reduced ALDH2 protein expression and the total amount of ubiquitinated proteins to about 40% of wild-type controls. These effects were largely prevented by ascorbate supplementation or treating Gulo(−/−) mice with the 26S proteasome inhibitor bortezomib.Our data indicate that ascorbate deficiency results in vascular tolerance to GTN via proteasomal degradation of ALDH2. The results support the view that impaired ALDH2-catalyzed metabolism of GTN contributes significantly to the development of vascular nitrate tolerance and reveal a hitherto unrecognized protective effect of ascorbate in the vasculature.


Buthus scorpions using mtDNA sequences. We sampled 91 individuals of the genus Buthus from 51 locations scattered around the Atlas Mountains . We sequenced 452 bp of the Cytochrome Oxidase I gene which proved to be highly variable within and among Buthus species. Our phylogenetic analysis yielded 12 distinct genetic groups one of which comprised three subgroups mostly in accordance with the orographic structure of the mountain systems. Main clades overlap with each other, while subclades are distributed parapatrically. Geographic structures likely acted as long-term barriers among populations causing restriction of gene flow and allowing for strong genetic differentiation. Thus, genetic structure and geographical distribution of genetic (sub)clusters follow the classical theory of allopatric differentiation where distinct groups evolve without range overlap until reproductive isolation and ecological differentiation has built up. Philopatry and low dispersal ability of Buthus scorpions are the likely causes for the observed strong genetic differentiation at this small geographic scale.The immense biodiversity of the Atlas Mountains in North Africa might be the result of high rates of microallopatry caused by mountain barriers surpassing 4000 meters leading to patchy habitat distributions. We test the influence of geographic structures on the phylogenetic patterns among  The Mediterranean region comprises a highly diverse geographic area characterized by a variety of peninsulas and islands. This underlying geographic structure is reflected in biogeographical patterns of the region already described by de Lattin (1949) De Lattin (1949) Buthus, which has undergone extensive speciation within NW Africa Buthus scorpions are widely distributed over large parts of Africa and show a circum-Mediterranean stronghold including the Mediterranean islands, with extraordinarily high diversity in North Africa In order to test for the influence of orographic structures on differentiation and speciation processes, we selected the scorpion genus Buthus species from 51 locations over major parts of the Antiatlas, High Atlas, Middle Atlas and Jebel Sahro during spring 2008 and 2009. Three specimens of the related genus Androctonus from two locations served as outgroup. We analysed a fragment of the mitochondrial Cytochrome Oxidase I (COI) gene, which has been shown to be highly informative at species level and consequently is one of the most commonly used markers for phylogenetic and phylogeographic studies. Due to its high potential for species identification it is often chosen as the barcoding gene in zoological studies. Here we use its properties to address the following questions:For this study we collected a total of 91 individuals belonging to 4 described Buthus exhibit geographic structuring?Does genetic variation within Do genetic structures follow any geographical and/or evolutionary pattern?Are genetic lineages in accordance with current taxonomy?Androctonus mauritanicus (JF820097) was obtained from genbank as an additional outgroup. Details about analyzed individuals, including genbank accession numbers, are given in We generated 94 sequences (including three outgroup specimens) with a total length of 452 base pairs. One sequence from The second dataset included 84 additional sequences obtained from genbank . In totaButhus lineages, which were recovered in all analyses  Several lineages are composed of different species: four of the five lineages with ten or more individuals analysed contain two to three species; even lineage H  includes two different morpho-types.The 12 lineages A through L do not match with current taxonomy (as reflected by morphology based identification keys): (i) The two tree hypotheses (ML and BI) constructed from our original dataset distinguish 12 genetic lineages. One of these clusters (lineage K) is further structured into three sub-clades. The genetic clusters exhibit a clear geographic pattern with all the lineages being confined to relatively restricted areas. None of the lineages is widely distributed. Most lineages are separated from each other by geographic barriers such as high mountain ranges or steep valleys. Only some basal lineages show range overlap (e.g. K) while all younger groups are strictly allopatric or parapatric (e.g. the three sub-groups of K). Four groups (A–D) are situated around the western and north-western parts of the High Atlas. One group is located in the westernmost Antiatlas (E), three others at the northern foothills of these mountains (F–H) and one at the southern foothills of the Antiatlas (J). The largest cluster (K) is distributed south of the High Atlas, east of the Draa valley, and splits further into three subclades, which form distinct strictly parapatric groups following the orographic structures of this region . Another cluster (I) extends from the Draa valley to the south western edge of the Antiatlas, probably including parts of the distribution ranges of clade G and H.Buthus scorpions mostly does not exceed elevations higher than 2500 m asl Buthus individuals. A similar pattern was observed for the same taxonomic group using various molecular markers such as allozymes, nuclear and mitochondrial DNA all of which showed significant genetic differentiation patterns even among local populations Buthus lineages also include their last centre of differentiation. Thus, our study area may comprise 12 allopatric refugia during the last ice age revealing a much more fine grained biogeographical sub-structure than previously observed in the Maghreb e.g. Buthus in Morocco is much more complex than the structures observed in other Mediterranean sub-centres Buthus individuals  gene was amplified using standard PCR procedures with the following primers: forward 5′ GGT CAA CAA ATC ATA AAG ATA TTG G 3′, reverse 5′ TAA ACT TCA GGG TGA CCA AAA AAT CA 3′For all individuals, DNA was isolated from leg, caudal segment or telson muscle tissue using the Qiagen DNeasy kit with standard protocols for tissue samples. A 452 bp fragment of the mitochondrial Cytochrome Oxidase I ). Sequences were inspected and aligned using Geneious 5.0.3 We included 91 sequences of General alignment statistics  were calculated using DNAsp v. 5.10 The best fitting model of sequence evolution for phylogenetic tree reconstruction was chosen using MrModeltest 2.3 Buthus scorpions obtained from genbank  In a second analysis, we included 84 sequences from bank see  resultinbank see . When thAppendix SIDNA sequences obtained from NCBI genbank.(DOC)Click here for additional data file.


The mechanisms mediating this effect are still unclear. The activity of several ion channels is sensitive to changes in temperature, leading to cytosolic calcium influx. Calcium is universally involved in cellular processes, its effects depending on the cellular context. Although not much is known about the role of calcium in immune cells, it has been shown that calcium is necessary for inflammasome assembly. Conventional methods of measuring calcium are not ideal to study the relation between temperature and intracellular calcium concentrations  is highly dependent on temperature. This is not established in detail because the indicator is mostly used in temperature independent assays. Here, we determined KdFluo-4 for temperatures in a range of 5-55oC, enabling calculation of exact intracellular calcium concentrations. We applied the assay in different cell lines and primary human innate immune cell populations.[Ca2+]i to basal levels. This [Ca2+]i increase can be blocked by the aspecific ion channel antagonist Ruthenium Red.Exposing cells to decreasing temperature induces elevated intracellular calcium concentrations. Subsequently rising temperature restores [Ca2+]i measurements in innate immune cell populations under tight temperature control.We show that the intracellular calcium concentration in immune cells increases significantly upon decreasing temperature. Blockade by Ruthenium Red indicates that this effect is mediated by thermosensitive ion channels. We successfully optimized a 96-well format fluorescense-based assay using a qPCR machine, enabling [CaNone declared