A running-in and starved lubrication experiment is designed to investigate the heavy-duty scuffing behavior of piston ring coatings against cast iron (Fe) cylinder liner using the piston ring reciprocating liner test rig. The scuffing resistance of the piston ring with the chromium-based ceramic composite coating (CKS), and that with the thermally sprayed nickel-chromium-molybdenum coating (NCM) is compared at different nominal pressures (40~100 MPa) and temperatures (180~250 °C). With the failure time as a criterion, the rank order is as follows: NCM/Fe > CKS/Fe. Before the scoring occurs at the interface of the piston ring and cylinder liner (PRCL), the cast iron liner enters into a “polish wear” stage, and iron-based adhesive materials begin to form on the piston ring surface. With the macroscopic adhesion formation, the plastic shearing cycle causes surface damages mainly due to abrasive effects for the CKS/Fe pairs and adhesive effects for the NCM/Fe pairs. As one of the main frictional pairs in the engine, the piston ring and cylinder liner have direct impacts on the engine’s mechanical efficiency and service life. With the development of the internal combustion engine in high power density, scuffing problems under severe working conditions have received more attention ,2.Scuffing is often recognized as an abrupt rise in the coefficient of friction accompanying by a sudden increase in noise and vibration. Many efforts have been made to investigate the scuffing phenomenon from different perspectives. Blok gave a criterion to judge the initiation of scuffing with the critical temperature, which was the sum of the bulk temperature and the flash temperature generated in the contact area . Czichos2O3 particles was improved remarkably [Boron phosphorus alloy cast iron with a high performance-price ratio, excellent processing properties, as well as excellent tribological properties has been widely applied in the manufacturing of the engine cylinder liners. A coating on the piston ring may offer advantages such as friction reduction, wear resistance, and better scuffing performance. Öner examined the structural changes on the CrN-coated surface due to thermal and mechanical shocks . Jishengmarkably . Lin premarkably . Wan dismarkably . With emThe purpose of this paper is to conduct and analyze a series of experiments to compare the heavy-duty scuffing behavior of a grey cast iron liner with typical NCM- and CKS-coated piston rings. It mainly presents the scuffing behavior and failure mechanism of the mating pairs under starved lubrication conditions. This provides a reference for the anti-scuffing design of piston ring coatings.The piston ring reciprocating liner test rig, as illustrated in 0.1. Its inner diameter is 110 mm, and its thickness is 10 mm. It was cut into 40 equal portions along the circumference. The piston ring has an inner diameter of 70 mm, an outer diameter of 110 mm, and a thickness of 3 mm. It was cut into 20 equal portions along the circumference. Liner samples utilize boron-phosphorus alloy cast iron + honing. Its roughness Ra  is 0.72 μm and its hardness is measured to be an average of 238 HV0.1. The scattered distribution of non-connected micro-cracks does not extend through the entire chromium coating (about 60 μm in thickness). The fine aluminum oxide particles exist in the micro-cracks. 0.1. The surface coating is a little rougher with the distribution pits of different size. The coating thickness is about 150 μm. It consists of pure Mo (light grey color) and NiCr alloy (dark grey color). NiCr alloys have self-lubricating properties at high temperatures. The basic material of the above two piston rings is gray cast iron.To guarantee experimental repeatability, the piston ring has no gap clearance to avoid the effects of the original shape. Mineral-based, fully-formulated engine oil (15W-40), which contains zinc dialkyl dithiophosphates (ZDDP) additives, serve as the lubricating oil in the experiments. To ensure the accuracy and repeatability of the experiment, the following procedure is used. First, the samples of piston ring and cylinder liner are cleaned with acetone. Second, after the liner sample is fixed, the ring sample is slowly lowered until it comes into contact with the liner sample. The interface between the samples must contact evenly along their arc directions.The experiment was divided into three stages: the running-in stage with the light load (RLL), the running-in stage with the heavy load (RHL), and the oil starvation stage (OS). The running-in stage was supplied with adequate lubricating oil at the speed of 0.1 mL/min. The oil was evenly distributed along the entire 30-mm stroke. The RLL and RHL stages were provided to eliminate machining defects, such as large burrs, and provide a stable contact state before starvation. At the end of the RHL period, the lubricating oil supply was stopped. Then, starved lubrication of the OS stage occurred at the PRCL interface. The experiment essentially simulates the starvation at the top dead center in a diesel engine. For the cast iron cylinder liner sliding against the CKS and NCM ring coatings, the scuffing resistance test results on the anti-scuffing time duration as a function of nominal pressure and temperature are shown in Some transferred cast iron patches result in adhesion on the NCM ring surfaces, as shown in The worn patches are transferred to the mating surface, as shown in (a)The surface honing textures of the cast iron cylinder liner gradually becomes shallow and its friction condition is in the “polishing wear” stage.(b)The formation of microscopic and macroscopic adhesions causes cast iron plastic shearing, resulting in different damage modes of the surface materials transfer. Meanwhile, the subsurface accompanied with the deformation contributes to the formation of abrasive particles. (c)Most transferred materials adhere to the piston ring surface with different particle aggregation patterns.Based on the analysis of the scuffed PRCL, the scuffing sequence of the damage to the final failure seems to involve the following events for the two piston ring coatings:(1)When mated with a cast iron cylinder liner, the NCM coating has better scuffing resistance properties than the CKS coating with the failure time criterion.(2)At the heavy-duty sliding conditions, when the nominal pressure and temperature exceed more than 60 MPa and 220 °C, respectively, the anti-scuffing performance exhibits an abrupt downward trend. Exceeding these harsh conditions, the anti-scuffing performances are maintained at a weak level.(3)Before the scoring occurs at the PRCL interface, the cast iron liner enters into a “polish wear” stage, and iron-based adhesive materials begin to form on the piston ring surface.(4)The plastic shearing cycle causes surface damages mainly due to adhesive effects for the NCM/Fe pairs, and abrasive effects for the CKS/Fe pairs.Heavy-duty scuffing behavior of the PRCL interface is obtained using the piston ring reciprocating liner test rig. The interface contacts are starved of lubrication after the running-in period. The effects of nominal pressures and temperatures on the scuffing resistance of the piston ring coatings are evaluated. The experimental results can be summarized as follows:


Tumor-associated inflammatory cells in classical Hodgkin lymphoma (CHL) typically outnumber the neoplastic Hodgkin/Reed-Sternberg (H/RS) cells. The composition of the inflammatory infiltrate, particularly the fraction of macrophages, has been associated with clinical behavior. Emerging work from animal models demonstrates that most tissue macrophages are maintained by a process of self-renewal under physiologic circumstances and certain inflammatory states, but the contribution from circulating monocytes may be increased in some disease states. This raises the question of the source of macrophages involved in human disease, particularly that of CHL. Patients with relapsed CHL following allogeneic bone marrow transplant (BMT) provide a unique opportunity to begin to address this issue. We identified 4 such patients in our archives. Through molecular chimerism and/or XY FISH studies, we demonstrated the DNA content in the post-BMT recurrent CHL was predominantly donor-derived, while the H/RS cells were derived from the patient. Where possible to evaluate, the cellular composition of the inflammatory infiltrate, including the percentage of macrophages, was similar to that of the original tumor. Our findings suggest that the H/RS cells themselves define the inflammatory environment. In addition, our results demonstrate that tumor-associated macrophages in CHL are predominantly derived from circulating monocytes rather than resident tissue macrophages. Given the association between tumor microenvironment and disease progression, a better understanding of macrophage recruitment to CHL may open new strategies for therapeutic intervention. Lymph nodes involved by classical Hodgkin lymphoma (CHL) typically contain relatively few neoplastic Hodgkin/Reed-Sternberg (H/RS) cells, which are scattered throughout a mixed inflammatory background composed of lymphocytes, macrophages, neutrophils, eosinophils and plasma cells. The H/RS cells can comprise as little as 1% of the cellularity –10. It iOf particular interest are the tumor-associated macrophages and their source as increased numbers of tumor-associated macrophages correlate with a poor prognosis in patients with CHL –16. WhilIn addition, while self-renewal of resident macrophages may enable recovery in response to various forms of tissue injury 18), lethal radiation prior to bone marrow transplant (BMT) results in gradual replacement of tissue macrophages (with the exception of microglia) by donor monocytes in both mouse models , lethal  and humaThe complex interplay between residual/recovering tissue macrophages and colonizing donor monocytes following BMT is of particular interest in recurrent CHL. Recurrent CHL often involves tissues with a large resident macrophage population such as lung and liver. Given the association with macrophage frequency and disease prognosis, we sought to further understand this process, including whether the inflammatory environment may change post-transplant. We characterize the inflammatory infiltrate in pre- and post-BMT tumor specimens in patients who had received an allogeneic BMT and investigate the origin of the inflammatory cells (donor or recipient) in the post-BMT specimens.Prior to commencing the study, approval was obtained from the Johns Hopkins Hospital Institutional Review Board (JHH IRB) under the IRB-approved protocol IRB NA_00083069. Per the JHH IRB, specific oral or written consent was not required for inclusion of specimens in this retrospective study, as the findings do not affect either diagnosis or treatment and both the diagnosis and treatment plans were established prior to and independent of the study findings. The pathology database was then searched to identify patients with CHL who were treated with an allogeneic BMT at our institution and subsequently developed relapsed disease. Clinical data were reviewed by one of the authors (A.S.D.) in accordance with IRB-approved protocol. Decalcified bone marrow biopsy specimens were excluded from the study.Formalin-fixed and paraffin-embedded (FFPE) tissue was used to perform XY chromosome studies using fluorescence in situ hybridization  in patients who received a sex mismatched BMT. Representative fields were chosen based on the corresponding H&E stained sections, and XY FISH images were captured with a fluorescence microscope. XY FISH was also performed subsequent to standard immunohistochemical staining for CD68 on the same slide.Molecular chimerism studies  were performed on the post-transplant FFPE tissue in 3 of the 4 patients. This system uses PCR and capillary electrophoresis  to determine the length of 15 tetranucleotide short tandem repeats (STRs) in order to generate a molecular “signature” for the patient and donor. The relative percentages of donor and patient DNA in the post-BMT CHL samples were determined using the patient and donor STR profiles that were performed prior to the BMT.The FFPE tissue was also subjected to hematoxylin and eosin (H&E) staining as well as immunohistochemical staining using routine methods for clinical diagnosis. All antibodies were from Ventana  with the exception of CD3 .Four patients were identified that had relapsed CHL after an allogeneic BMT and also had post-BMT FFPE tissue involved by CHL available in our archives . Two of Three patients had undergone myeloablative transplants , and one patient (Patient A) had a non-myeloablative (reduced intensity) transplant. Three of the four patients had opposite sex donors: patients (Patients A and C) were male but had a female donor and patient (D) was a female but had a male donor. Two of the patients (B and D) had bone marrow chimerism studies available near the time of relapse. In patient B, studies performed both one month before relapse and one month after relapse (but before the initiation of treatment) showed approximately 10% patient DNA and 90% donor DNA. Patient D had 100% donor DNA with no patient DNA detected both 8 months prior to relapse and 2 months after relapse.To assess the origin of the inflammatory infiltrate in the recurrent lymphoma we performed XY FISH on post-BMT tumor from patients A, C and D. Samples from all three patients demonstrated that the inflammatory infiltrate was largely composed of donor derived inflammatory cells . ScatterTo confirm the FISH findings as well as to investigate the composition of the tumor in the patient with a same-sex donor (Patient B), molecular chimerism studies were performed on 3 of the 4 patients (excluding patient D). For all three, the molecular identity testing showed that the majority of the cells in the tumor were of donor origin  .th relapse, though the patient had been treated with the anti-PD-1 monoclonal antibody nivolumab prior to the biopsy (For the two patients where pre- and post-BMT tissue was available (Patients A and B), IHC stains were performed to visualize the neoplastic cells and the composition of the inflammatory infiltrate . The pere biopsy .In this study we utilized post-BMT tumor specimens in order to more fully characterize the relationship between H/RS cells and the tumor-associated inflammatory cells, in particular the origin of the tumor-associated macrophages. These findings support the idea that H/RS cells, perhaps through cytokines and chemokines, actively recruit and define the inflammatory microenvironment –35. In pAdditionally, we found that the composition of the inflammatory infiltrate was very similar in the pre- and post-BMT samples where evaluable. The percentage of macrophages for the two patients examined in this manner fell into the “intermediate risk” category as defined by Steidl, et al. with between 5 and 25% macrophages  and simiOne potential limitation on the generalizability of this study for understanding tumor-infiltrating macrophage recruitment to the CHL microenvironment is the demonstration in prior studies that total body irradiation is associated with gradual replacement of tissue macrophages by bone marrow-derived monocytes , 25–27. While the number of cases is small, our findings raise the possibility of novel treatment strategies for recurrent CHL, including the potential to pharmacologically inhibit monocyte recruitment to tumor lesions to slow or alter disease progression ,40.S1 FigA separate area of recurrent CHL involving the lung from patient D is shown (compared to (TIF)Click here for additional data file.S2 FigThe same image shown in (TIF)Click here for additional data file.S3 FigEven in the reduced intensity BMT setting, the majority of cells in the recurrent tumor are donor-derived. XY FISH studies of the pre- and post-BMT tumor from Patient A are shown. Nuclei are stained with DAPI (blue). Red and green signals correspond to the probes on the X and Y chromosomes, respectively. The pre-BMT tissue  demonstrates both X and Y chromosomes in all cells whereas the post-BMT tissue  shows that most cells are XX  with only scattered large XY  cells. In 21% of nuclei, the X, Y FISH probes could not be definitively evaluated due to the plane of section.(TIF)Click here for additional data file.S4 Figth relapse; 200x). There is a suggestion of a decrease in the percentage of CD8+ cells as compared to both the initial pre and post-BMT specimens, but the findings are otherwise similar. All antibodies were from Ventana , with the exception of lymphocyte-activation protein 3  and indolemine 2,3-dioxegenase . IDO is visualized as red staining.Representative high power fields from an axillary lymph node involved by CHL from Patient A post-BMT and post-treatment with the anti-PD-1 monoclonal antibody nivolumab (4(TIF)Click here for additional data file.


A randomized controlled study (RCT) recently showed that short-term heart rate variability (HRV) biofeedback in addition to standard rehabilitation care for alcohol dependence can reduce craving, anxiety and improve cardiovascular autonomic function. In this one-year follow-up study we aimed to explore whether completion of 2-week HRV-Biofeedback training is associated with long-term abstinence. Furthermore, we sought to identify potential predictors of post-treatment abstinence.We conducted a survey on abstinence in patients with alcohol dependence 1 year after completion of an RCT comparing HRV-biofeedback in addition to inpatient rehabilitation treatment alone (controls). Abstinence rates were compared and analysed for association with demographic data as well as psychometric and autonomic cardiac assessment before and after completion of the biofeedback training using bivariate and multivariate regression analyses.Out of 48 patients who participated in the RCT, 27 patients  completed our one-year follow-up. When including in the analysis only patients who completed follow-up, the rate of abstinence tended to be higher in patients who underwent HRV-biofeedback 1 year earlier compared to those who received rehabilitative treatment alone . This non-significant trend was also observed in the intention-to-treat analysis where patients who did not participate in the follow-up were assumed to have relapsed . Neither cardiac autonomic function nor psychometric variables were associated with abstinence 1 year after HRV-biofeedback.Our follow-up study provide a first indication of possible increase in long-term abstinence after HRV-biofeedback for alcohol dependence in addition to rehabilitation.DRKS00004618). This one-year follow-up survey has not been registered.The original randomized controlled trial was registered in the German Clinical Trials Register ( Alcohol dependence is a major global public health problem which affects 5.1% of the global population and causes up to 3.3 million deaths per year, the majority of those being related to cardiovascular diseases , 3. MoreWe recently showed in a randomized controlled study that heart rate variability (HRV) biofeedback in addition to standard rehabilitation care for alcohol dependence can reduce craving and anxiety more effectively than rehabilitative treatment alone . In thisIn this follow-up study we aimed to determine the relapse rate 1 year after HRV-biofeedback training and integrative inpatient rehabilitative treatment in order to explore potential treatment effects and acquire first long-term data that could form a basis for confirmatory research in large study populations. Furthermore, we sought to identify predictors of abstinence following combined rehabilitative and HRV-biofeedback treatment.This is a one-year follow-up survey study after a randomized controlled trial on the effects of HRV-biofeedback on cardiac autonomic function assessed via time and frequency domain parameters of HRV, autonomic neurovascular function assessed via laser Doppler flowmetry of cutaneous blood flow after sympathetic stimulation as well as craving, anxiety and depressive symptoms evaluated using psychometric tests. These techniques were reported in detail elsewhere . BrieflyTo perform a one-year follow-up assessment of abstinence, study participants were contacted by mail 1 year after discharge from rehabilitative therapy after having given written permission to be contacted during inpatient treatment. They were asked to answer a standard questionnaire about their social situation, alcohol and drugs consumption, as well as need for further institutional treatment in the period of 12 months after discharge. Abstinence from alcohol and drugs was considered no consumption during 12 months after discharge from our clinic. We extracted demographic characteristics in subjects who had undergone HRV-biofeedback in addition to rehabilitation 1 year earlier and those who had received rehabilitative treatment alone from the original dataset.Abstinence rates 1 year after discharge from the rehabilitative therapy were evaluated applying standard criteria of the German Society for Addiction Research and Addiction Treatment . IndividIn order to define individual factors related to abstinence we compared patients who reported being abstinent 1 year post-intervention and those who reported relapse in terms of demographic characteristics as well as psychometric measurements and parameters of the autonomic cardiac function. These analyses were performed in patients who completed follow-up. We included in our analyses psychometric scores at baseline and those obtained immediately following the final HRV-biofeedback training session (post-biofeedback or post-observation period for patients in the control group). Psychometric scores were obtained using the Obsessive Compulsive Drinking Scale, as measure of craving as well as the subscales Anxiety and Depression from the Symptom Checklist-90, as measures of anxiety and depression. Measures of the cardiac autonomic function comprised the time-domain parameter coefficient of variation of R-R intervals (CVNN) and frequency-domain parameters high frequency (HF), low frequency (LF) and total power (TP) at baseline and immediately post-biofeedback or observation period. In order to improve our understanding of the association between applied treatments, and sustained abstinence, this analysis was undertaken in both the biofeedback and the control group.We then went on to identify specific predictors of abstinence after rehabilitative inpatient care by relating baseline data extracted from the original dataset of the randomized controlled trial to data from the present survey. We conducted these analyses separately for control patients and those who had received HRV-biofeedback. In order to identify any specific changes in biological or psychological characteristics due to HRV-biofeedback which relate to sustained abstinence, analyses were repeated using data on patient characteristics and outcomes obtained immediately after completion of the biofeedback intervention, the duration of the control period, respectively.2 test in patients who completed follow-up. Continuous data on study population characteristics were compared between patients who had received HRV-biofeedback in addition to rehabilitation and those who had undergone rehabilitative treatment only using Student’s t–test or Mann-Whitney U test, according to distribution. The same tests were used to compare study population characteristics and post-biofeedback outcomes between patients who reported abstinence after 1 year and those who had relapse. These analyses were first undertaken in those patients that have participated in the follow-up assessment excluding those that have not returned the survey questionnaire. The same tests were then applied to undertake an intention-to-treat analysis in the entire study population, considering those who have not participated in the one-year follow-up to have relapsed. Comparisons between patients with and without achievement of on-year abstinence were performed separately for the biofeedback and the control group. Alpha level for statistical significance was set to 0.05. Bivariate logistic regression analyses were undertaken to identify possible predictor variables among demographic and outcome variables with respect to one-year abstinence in patients who completed follow-up. Those variables that emerged as predictors of abstinence were included in multivariate models with adjustment for demographic characteristics. Unstandardized B-coefficients (ß) and p-values were computed.Statistical analyses were performed using SPSS ® 21 . Dichotomous data were compared using ӼAmong 48 study participants who have been contacted by mail, 27 responded and completed the questionnaire . Demographic characteristics of patients who completed follow-up are shown in Table When including only patients who completed follow-up in the analyses, abstinence rates tended to be higher in those who had undergone HRV-biofeedback in addition to rehabilitation compared to those who received rehabilitative treatment alone. Similarly, the intention-to-treat analysis, where patients who have not completed follow-up were considered to have relapsed, also showed a trend toward higher abstinence rates among patients who had undergone HRV-biofeedback than those who underwent rehabilitation alone Fig. .Fig. 1AbIn order to identify factors important to abstinence we compared demographic characteristics as well as outcome data on psychometric tests and autonomic cardiac function at baseline and immediately after completion of the RCT between patients who showed abstinence 1 year after completion of the trial and those who had relapse. As shown in Table None of the assessed demographic and outcome variables were associated with abstinence 1 year after the HRV-biofeedback intervention. Table  The sameIn this follow-up study 1 year after a randomized controlled trial of HRV-biofeedback in patients with alcohol addiction, we observed a tendency toward higher rates of long-term abstinence in the interventional study arm when compared with patients of the control arm that have not undergone HRV-biofeedback. Viewed in conjunction with our previous observation of improvement in cardiac autonomic and neurovascular function and reduction of craving and anxiety after HRV-biofeedback in the same study population, these findings warrant follow-up research to confirm this trend in a larger study population and assess the neurophysiological mechanisms whereby HRV-biofeedback might alter long-term abstinence . AlthougA recent randomized clinical trial had confirmed that HRV-biofeedback as adjuvant therapy leads to reduced craving in patients with addiction to alcohol and drugs . SimilarMoreover, the underlying mechanism whereby improvement of the autonomic cardiac function leads to alleviation of craving, and possibly changes in abstinence, needs to be elucidated. The observed effect of HRV-biofeedback on autonomic cardiac function and craving might be explained by the physiological mechanism of action of this technique. HRV-biofeedback is a behavioral intervention that targets enhancement of the beat-to-beat fluctuations of the heart rate (HRV) due paced breathing. Physiologically, the heart rate is determined by the intrinsic sinoatrial node discharge rate as well its autonomic alteration mediated by cardiac sympathetic and parasympathetic activity . The preOur study is limited by a low return rate of the mailed questionnaires. Therefore, the observed differences in abstinence rates between HRV-biofeedback treated and control patients might not have reached statistical significance due to a type two error. In the intent-to-treat analysis, statistical power might have been compromised by the conservative approach of imputing missing data, where all patients who have not completed follow-up were considered to have relapsed. This limitation might be overcome by increasing the sample size in future studies. Alternatively, it might be necessary to increase the frequency and duration of the HRV-biofeedback treatment sessions to translate the observed trend into a significant long-term difference.We did not assess the amount of past-year drinking, therefore we cannot comment on possible associations between treatment with HRV-biofeedback and the severity of relapse. Furthermore, patients weren’t asked to continue paced breathing after the intervention and therefore we haven’t assessed the rate of those still practicing the breathing technique after discharge. This might further explain why in our study observed trends of increase in abstinence have not reached statistical significance.Taken together, our study forms a basis for a long-term investigation of HRV-biofeedback in patients with alcohol addiction which should include a larger sample size and a dose-response protocol to identify the optimal regimen and achieve a sustained improvement in craving and autonomic function. This, in turn, might translate into improved long-term abstinence.Our data suggest that HRV-biofeedback might contribute to long-term abstinence when applied in addition to rehabilitation care. Since the trends observed in this follow-up study did not reach statistical significance, further research is warranted to confirm this hypothesis in a larger study population.


Rodent models of maternal immune activation (MIA) are increasingly used as experimental tools to study neuronal and behavioral dysfunctions in relation to infection-mediated neurodevelopmental disorders such as schizophrenia and autism. One of the most widely used MIA models is based on gestational administration of poly(I:C) (= polyriboinosinic-polyribocytdilic acid), a synthetic analog of double-stranded RNA that induces a cytokine-associated viral-like acute phase response. The effects of poly(I:C)-induced MIA on phenotypic changes in the offspring are known to be influenced by various factors, including the precise prenatal timing, genetic background, and immune stimulus intensity. Thus far, however, it has been ignored whether differences in laboratory housing systems can similarly affect the outcomes of MIA models. Here, we examined this possibility by comparing poly(I:C)-based MIA in two housing systems that are commonly used in preclinical rodent research, namely the individually ventilated cage (IVC) and open cage (OC) systems.Pregnant C57BL6/N mice were kept in IVC or OC and treated with a low  or high  dose of poly(I:C), or with corresponding vehicle solution . MIA or control treatment was induced on gestation day (GD) 9 or 12, and the resulting offspring were raised and maintained in IVC or OC until adulthood for behavioral testing. An additional cohort of dams were used to assess the influence of the different caging systems on poly(I:C)-induced cytokine responses in the maternal plasma, placenta, and fetal brains 1 hr and 6 hrs post-treatment.Maternal administration of poly(I:C) on GD9 caused a dose-dependent increase in spontaneous abortion in IVC but not in OC system, whereas MIA in IVC systems during a later gestational time-point (GD12) did not do so. Maternal and fetal pro-inflammatory cytokine responses to poly(I:C) were markedly higher in animals kept in IVC as compared to OC systems. The efficacy of MIA to induce long-term behavioral deficits was influenced by the different housing conditions, the dosing of poly(I:C), and the precise prenatal timing.The present study identified the type of cage system as a novel factor that can confound the outcomes of MIA. Our findings thus urge the need to consider and report the kind of cages used in rodent MIA models. Providing this information seems pivotal to yield robust and reproducible results in these models.


CD4+ T cells recover primarily via thymopoiesis resulting in complete renewal of the T cell receptor (TCR) repertoire which requires years or never normalize completely. The increase in naïve T cells inducing immune tolerance, renewal of especially the regulatory TCR repertoire, and a less pro-inflammatory functional profile of the CD4+ T cells seem essential for successful immune reconstitution inducing long-term remission. There is currently a knowledge gap regarding the immune response in tissue sites post-aHSCT, as well as disease-specific factors that may determine remission or relapse. Future studies on lymphocyte dynamics and function may pave the way for optimized conditioning regimens with a more individualized approach.Autologous hematopoietic stem cell transplantation (aHSCT) for autoimmune diseases has been applied for two decades as a treatment for refractory patients with progressive disease. The rationale behind aHSCT is that high-dose immunosuppression eliminates autoreactive T and B cells, thereby resetting the immune system. Post-aHSCT the cytotoxic CD8 Autoimmune diseases in general are characterized by a loss of immune tolerance. This results in generation and activation of autoreactive T and B cells leading to inflammation and consequently tissue damage. Autologous hematopoietic stem cell transplantation (aHSCT) for autoimmune diseases aims to eliminate autoreactive T and B cells and regenerate an immune system which is self-tolerant . The fir+ selection, to purge the collected stem cells of T cells can be performed, although it is unclear whether it has clinical benefits. The next step is the immunoablative conditioning phase. In autoimmune disease high-dose CYC and occasionally, in vivo purging is performed by systemic administration of antibodies such as anti-thymocyte globulin (ATG) or rituximab. Finally, the hematopoietic stem cells are reinfused, which accelerates hematopoietic reconstitution  and recombinant G-CSF. This enables apheresis of stem cells. CD34titution . ExactlyThe innate immune system recovers within weeks post-aHSCT, in contrast to the reconstitution of the adaptive immune system which can take years .+ and CD8+ T cell counts pre-aHSCT compared with non-responders  of the inhibitory molecule programmed cell death-1 protein (PD-1) on CD8+ T cells correlated with a good clinical response post-aHSCT  cell reconstitution is unknown. While reliable data regarding incomplete immunological reconstitution post-HSCT is lacking, there are indications that this is not associated with long-term morbidity. A for instance, a study assessed outcomes in rheumatoid arthritis patients that received Alemtuzumab, an anti-CD52 antibody targeting primarily T and B cells. Follow-up for 20–25 years shows an incomplete reconstitution of T and B cells, especially central memory CD4ard care . These d+ stem cells. The influence of this selection on immune reconstitution is reported in several studies, and although broadly used in treatments, there is conflicting evidence of its benefits  aHSCT is performed relatively early in disease. Another possible confounding factor is graft manipulation by selection of CD34benefits –38. Fina+ T cells are essential for successful immune reconstitution and the induction of long-term remission. Future studies on lymphocyte dynamics and function may pave the way for optimized conditioning regimens with a more individualized approach.In the past 20 years, aHSCT has been applied for severe refractory autoimmune diseases but a comprehensive understanding of the immune reconstitution and the link with clinical outcome is still missing. Data suggest that the increase in naïve T cells, renewal of especially the regulatory TCR repertoire, and a less pro-inflammatory functional profile of the CD4Important outstanding questions regarding the immune reconstitution following transplantation include: (1) What is the most effective conditioning regime for each autoimmune disease? This possibly depends on which immune cells are important in the pathogenesis, and the tissue(s) where the disease manifests, thus differ per autoimmune disease. (2) Is a personalized approach to the conditioning regime, depending on the immune status pre-aHSCT, needed to improve the clinical outcome? The recent observation that in a hematologic malignancy setting ATG dosing should be based on lymphocyte count may also apply to autoimmune settings. Extensive monitoring of immune depletion/reconstitution in combination with pharmacokinetic/pharmacodynamic modeling can contribute to the development of pre- and post-transplantation precision treatment. (3) What is the local tissue effect of the conditioning regimen? It is not clear to which extent for example ATG penetrates all tissues to deplete tissue T cells, and it is unclear if and to what extent residual tissue T cells contribute to relapses of disease. Together, future studies may shed light on the fine balance between effectively destroying and renewing the immune system, but with limited toxicity.LL, JS, FR-B, JL, and FW have discussed, written, and edited the manuscript.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.


To compare the medium-term clinical effectiveness of total hip arthroplasty andresurfacing arthroplasty.Single centre, two-arm, parallel group, assessor blinded, randomised controlledtrial with 1:1 treatment allocation.A large teaching hospital in England.122 patients older than 18 years with severe arthritis of the hip joint, suitablefor resurfacing arthroplasty of the hip. Patients were excluded if they wereconsidered to be unable to adhere to trial procedures or completequestionnaires.Total hip arthroplasty ; hipresurfacing arthroplasty . Both procedures replaced the articular surfaceof the acetabulum.The outcome measures were hip function assessed using the Oxford Hip Score (OHS)and health-related quality of life assessed using the EuroQol (EQ-5D). Patientswere followed up annually for a minimum of 5 years. Outcome data were modelledusing the generalised estimating equation methodology to explore temporalvariations during follow-up.60 patients were randomly assigned to hip resurfacing arthroplasty and 62 to totalhip arthroplasty. 95 (78%) of the 122 original study participants provided data at5 years. There was a small decrease in both hip functions and quality of life inboth groups of patients each year during the 5-year follow-up period. However,there was no evidence of a significant difference between treatments group in theOHS (P=0.333) or the EQ-5D (P=0.501).We previously reported no difference in outcome in the first year after surgery.The current medium-term results also show no evidence of a difference in hipfunction or health-related quality of life in the 5 years following a total hiparthroplasty versus resurfacing arthroplasty.ISRCTN33354155. UKCRN 4093. The main strength of this trial is that it is pragmatic, with a large number ofsurgeons using a variety of different hip arthroplasty implants and their ownpreferred surgical technique.Other strengths include the use of validated patient-reported outcome tools, arelatively large number of participants for this type of trial and the high levelsof complete follow-up data.The key limitation of this trial was that the patients themselves could not beblind to their type of hip arthroplasty.For older patients with severe arthritis of the hip, several designs of total hiparthroplasty (THR) have shown excellent long-term results in terms of both function andvalue for money.4Resurfacing implants are more expensive than traditional  THR designsand there are potential complications associated with RSA compared with THR—mostimportantly the risk of fracture of the neck of the femur.In 2012, we reported the 1 year results of a randomised controlled trial tocompare the clinical effectiveness THR and RSA in patients with severe arthritis of thehip.This was a single-centre, two-arm, parallel group, assessor-blind randomised controlledtrial with 1:1 treatment allocation conducted in the UK. Full details of the protocolhave been described previously.In this pragmatic trial, participants were eligible if they were over 18 years ofage, medically fit for an operation and suitable for a RSA—patients suitablefor RSA are also suitable for THR. Patients were only excluded from the study ifthere was evidence that the patient would be unable to adhere to trial procedures orcomplete questionnaires. To maintain independence between observed outcomes, if arecruited patient required a contralateral hip arthroplasty during the trial periodthe second hip was not included in the study.The Warwick Arthroplasty Trial (WAT) opened in May 2007 and 126 patients wererecruited between August 2007 and February 2010 from hip arthroplasty clinics in asingle UK hip arthroplasty centre. Eligible patients gave written informed consent.They were randomised on a 1:1 basis to receive either a THR or an RSA. Treatmentallocation was determined using a computer-generated, randomised number sequence andstratified by the supervising orthopaedic surgeon to balance any potential surgeoneffects. After patients consented to participate in the trial, an independentlyadministered, secure randomisation service was alerted by telephone of a newenrolment. The randomisation officer provided the surgeon’s secretary with thepatient’s treatment allocation, thereby keeping the research associates, whoconsented patients and collected outcome data, blinded to the allocatedtreatment.Each patient had the allocated surgery according to the preferred technique andimplants of the operating surgeon. Other perioperative interventions, such asprophylactic antibiotics and thromboprophylaxis, were the same for all patients.After the operation, all patients underwent the same standardised rehabilitationplan, including range-of-movement exercises followed by muscle strengtheningexercises. Unless the operating surgeon specifically advised otherwise, all patientswere fully weight bearing immediately.In a THR, the femoral head was removed along with most of the femoral neck. Thefemoral shaft was exposed to open up the femoral canal. The femoral component wasthen inserted into the canal and the articulating femoral head was placed onto theneck of the femoral component. The choice of components and bearing surfaces was leftto the discretion of the operating surgeon, as per their usual clinical practice.In a resurfacing arthroplasty, the articular surfaces of the femoral head was removedbut the neck was left in situ. The femoral component (cap) was then impacted onto thepatient’s own femoral neck. All resurfacing arthroplasties of the hip usedmetal-on-metal bearing surfaces, but the choice of surgical approach, implant sizeand positioning was left to the discretion of the operating surgeon.In both forms of arthroplasty, the acetabulum is prepared and the acetabularcomponent inserted into the socket.The primary outcome measure for this medium-term follow-up study was hip function,assessed using the OHS,On completion of the main phase of the trial, the trial steering group recommended toremove the additional outcomes collected during main phase  to reduce theburden on the participants and to optimise retention rates.Longitudinal data  weremodelled using generalised estimating equations (GEEs) to explore thepopulation-averaged effects of time and operative treatment on function andHRQoL.16The WAT study recruited 126 participants, 95% of whom had a primary diagnosis withosteoarthritis. Of the 126 participants recruited into the WAT study, 4 never had anoperation, leaving 122 in total available for follow-up during the original trial and inthe extended (medium-term) follow-up study that we report here. The baselinecharacteristics of these 122 participants are shown in The following medium-term follow-up results are based on the treatment received, in contrast to the previously reported study results which werebased on the allocated intervention . The amount ofmissing data at each time-point is shown in OHS scores at each year postoperation (from year 1 to 5) were strongly positivelyautocorrelated; that is, a high OHS at year 1 was predictive of a high OHS insubsequent years. The OHS scores decreased over time from year 1 to year 5 . However, this decline was unlikely to be clinically relevant in themedium term, occurring at only approximately 0.70 OHS units per year. Adding model terms to account for treatment groups (RSA vs THR) did not lead to areduction in QIC, and model parameters showed that there was no evidence of asignificant difference between treatment groups (P=0.333) during follow-up, and therate of decline of OHS score did not differ between treatment groups (P=0.317). Fullresults of analyses are shown in online 10.1136/bmjopen-2017-018849.supp1Supplementary file 1There was also evidence for a significant temporal change in EQ-5D ; EQ-5D scores decreased over time from year 1 toyear 5 (regression coefficient −0.027). However, again the decline in EQ-5Dscores was relatively small at approximately 0.027 units per year.EQ-5D scores showed considerable variability during follow-up . Adding During the 5-year follow-up, one patient in the RSA group had a revision arthroplastyand three in the THR group . TwoothFisher’s exact test provides no evidence of a statistically significantdifference in revision rates between treatment groups (P=0.619).Our previously reported randomised clinical trial found no evidence of a difference inhip function between patients having THR versus RSA for severe arthritis of the hipjoint during the first year following surgery.Only a few randomised trials have been performed comparing THR with the resurfacingtechnique. The first studies focused on the technical aspects of the procedure, such asthe position of the implants or the amount of bone removed during the resurfacingprocedures.24A further randomised trial26The only longer term follow-up study of resurfacing versus conventionalbearing-surface THR showed no difference between groups with regard to OHS or qualitylife.The main strength of this trial is that it is entirely pragmatic, with a relativelylarge number of surgeons using a variety of different hip arthroplasty implants andtheir own preferred surgical technique. Although the patients were recruited from onlyone centre, the large number of surgeons involved and the variety of implants is likelyto reflect practice in the wider surgical community. Other strengths include the use ofvalidated patient-reported outcome tools, a relatively large number of participants andthe high levels of complete follow-up data.The key limitation of this trial was that the patients themselves were not blind totheir type of hip arthroplasty. Patients undergoing RSA in the UK have generally beengiven a different preoperative information sheet and surgical consent from than thosehaving a THR; this reflects the existing evidence regarding the different risk/benefitprofile of the two procedures.How do the results of this trial inform the debate about resurfacing arthroplasty of thehip? This trial failed to show any evidence that resurfacing arthroplasty providesimproved hip function or greater quality of life when compared with THR over5 years. Given the new requirements for surveillance of metal-on-metal hiparthroplasties,


Evidence-based treatment for co-occurring disorders is needed within programs that serve homeless Veterans to assist with increasing engagement in care and to prevent future housing loss. A specialized co-occurring disorders treatment engagement intervention called Maintaining Independence and Sobriety Through Systems Integration, Outreach and Networking - Veterans Edition (MISSION-Vet) was implemented within the Housing and Urban Development - Veterans Affairs Supportive Housing (HUD-VASH) Programs with and without an implementation strategy called Getting To Outcomes (GTO). While implementation was modest for the GTO group, no one adopted MISSION in the non-GTO group. This paper reports Veteran level outcome data on treatment engagement and select behavioral health outcomes for Veterans exposed to the MISSION-Vet model compared to Veterans without access to MISSION-Vet.This hybrid Type III trial compared 81 Veterans in the GTO group to a similar group of 87 Veterans with mental health and substance use disorders from the caseload of staff in the non-GTO group. Comparisons were made on treatment engagement, negative housing exits, drug and alcohol abuse, inpatient hospitalizations, emergency department visits and income level over time, using mixed-effect or Cox regression models.p = .04). Supplemental exploratory analyses between Veterans who received “higher” and “lower” intensity MISSION-Vet services in the GTO group failed to show differences in alcohol and drug use, inpatient hospitalization and emergency department use.Treatment engagement, as measured by the overall number of case manager contacts with Veterans and others , was significantly higher among Veterans in the GTO group (B = 2.30, Despite modest MISSION-Vet fidelity among staff treating Veterans in the GTO group, differences were found in treatment engagement. However, this study failed to show differences in alcohol use, drug use, mental health hospitalizations and negative housing exits over time among those Veterans receiving higher intensity MISSION-Vet services versus low intensity services. This project suggests that MISSION-Vet could be used in HUD-VASH to increase engagement among Veterans struggling with homelessness, a group often disconnected from care.NCT01430741, registered July 26, 2011.Clinicaltrials.gov, registration number:  The Department of Veterans Affairs has undergone a major initiative to end Veteran Homelessness, which included the development and implementation of many new programs and services . One sucMaintaining Independence and Sobriety through Systems Integration, Outreach, and Networking-Veterans Edition (MISSION-Vet) is an evidence-based intervention for co-occurring mental health and substance abuse developed specifically to increase treatment engagement and address the psychosocial needs of homeless Veterans –16. WhilThe VA’s Quality Enhancement Research Initiative funded a Hybrid type III implementation and effectiveness trial of MISSION-Vet within HUD-VASH . This trn = 87).A recent paper compared the implementation of MISSION-Vet between the GTO and IU groups. Sixty-eight percent of the case managers in the GTO group adopted MISSION-Vet whereas none adopted it in the IU group; implementation uptake was modest with 70% of Veterans with case managers in the GTO group receiving at least one MISSION-Vet session but none receiving a full dose of the intervention . Among tThe study was approved by the VA Central IRB and case managers consented to participate. The study received a waiver of consent to use the client-level data from the VA’s existing HOMES database to assess MISSION-Vet outcomes.The MISSION-Vet treatment intervention is listed in the Substance Abuse and Mental Health Services Registry of Evidence Based Practices . MISSIONGetting To Outcomes in services for homeless Veterans: 10 steps for achieving accountability [GTO is an implementation support approach used to strengthen the knowledge, attitudes and skills case managers and peer specialists need to carry out MISSION-Vet. GTO involves guiding practitioners to complete steps in three general areas: (1) planning – e.g., developing goals and performance targets, ensuring staff are trained in the evidence-based program; (2) implementation – e.g., monitoring progress, maintaining adherence to an evidence-based program model, and (3) evaluation – e.g., tracking patient outcomes, using data to improve program operations , 23. Eactability . (2) Tratability . The GTOWe examined the following set of outcome measures: 1) Treatment engagement ; 2) Drug and alcohol use, which was a dichotomous measure based on case managers’ clinical assessment of Veterans’ drug or alcohol use as either dependence or severe dependence; 3) Any inpatient hospitalization for medical and mental health conditions; 4) Any emergency department visit for medical and mental health conditions; and 5) negative housing exits from HUD-VASH. The first four of these outcomes  measures were assessed monthly for 12 months following study enrollment. These measures were obtained from the standardized monthly and quarterly status reports completed by case managers, which were extracted from HOMES. We obtained information on negative housing exits from the HUD-VASH Exit form, which is available in HOMES and tracks the dates and reasons for all exits from the program. We measured whether and when a Veteran experienced a negative housing exit over the period from study enrollment (this was the first date of MISSION-Vet for those served by case managers in the GTO group and the date of the first MISSION-Vet session provided to any Veteran at the appropriate study site for those in the comparison group) until the end of the study observation period. The maximum follow-up time for capturing exits was 2.4 years, with variable follow-up times due to different dates  of entry into the study and corresponding differences in the availability of follow-up data on housing outcomes from HOMES. A negative exit from HUD-VASH was defined as one that occurred due to non-compliance with case management, eviction, Veteran dissatisfaction with housing, inability to locate Veteran and incarceration.We compared changes in the first four outcomes  among Veterans served by case managers in the GTO group relative to the comparison group of Veterans, by fitting a series of mixed-effects regression models to account for clustering of observations within individuals and case managers, using linear models and SAS PROC MIXED for continuous outcomes and binary logistic models and SAS PROC GLIMMIX for dichotomous outcomes. Final mixed-effect models for all outcomes included fixed terms for group (GTO group vs. comparison), time, group x time interaction as well as control covariates for age, year of move-in to HUD-VASH, and the following measures obtained from a VA homeless program intake form: need for treatment for a medical problem; employment status ; and housing status at time of assessment . We used Kaplan-Meier survival curves to estimate the extent and timing of negative housing exits over time and Cox proportional hazards regression to assess the relationship between membership in the GTO group and risk of experiencing a negative housing exit, adjusting for a set of relevant covariates.Inspection of data collected on MISSION-Vet fidelity suggested low to moderate fidelity, with 20% of Veterans served by case managers in the GTO condition receiving only 1 or 2 MISSION-Vet Sessions, two thirds receiving 19 or fewer of the proposed 64 sessions and none receiving the full intervention. As such, we conducted a supplemental exploratory set of analyses to assess the relationship between intensity of MISSION-Vet and the outcomes of interest in which we: 1) treated the number of MISSION-Vet sessions received per month as a continuous variable, and 2) stratified Veterans in the GTO case manager group into “higher” and “lower” intensity groups, using the median number of MISSION-Vet sessions received (11 sessions) as the delimiting factor.p < .001), had been in HUD-VASH housing longer , were less likely to need treatment for a medical problem , but were no different in terms of employment  and housing status at time of assessment .Bivariate tests found that, relative to the comparison group, Veterans served by the GTO group were significantly older . However, these measures of treatment engagement declined significantly over time for both groups, and the negative time by GTO interaction term  indicates that the difference in treatment engagement between Veterans in the GTO group and those in the comparison group attenuated over time. Of note, this gradual reduction in service intensity in the GTO group is consistent with the MISSION-Vet service delivery schedule, whereas the comparison group had consistently lower intensity throughout the observation period. Neither the main effect of membership in the MISSION GTO group, nor the interaction with time were statistically significant predictors in the other outcome models, as shown in Tables As shown in Table With the categorical predictor, those in the GTO group who met the criteria for higher intensity MISSION-Vet had significantly more face to face case manager contacts than those in the lower intensity MISSION-Vet group and the magnitude of this relationship did not change over time. With respect to the continuous MISSION-Vet predictor, the number of MISSION-Vet sessions was positively associated with all three case manager contact measures, although the strength of this relationship attenuated over time for face to face and case manager contacts. There were no significant relationships between the categorical and continuous MISSION-Vet measures (or their interaction with time) and the other outcome measures, although in both cases there were consistently non-significant negative relationships between receipt of MISSION-Vet services and the probability of alcohol use, drug use, and mental health hospitalizations.p = .481).The Kaplan-Meier estimates found that 11.6% of Veterans in the GTO group and 19.2% of Veterans in the comparison group had experienced a negative housing exit at 2 years following study enrollment. However, the Cox regression model found no significant association between group membership and risk of negative housing exit . With regards to future directions, we recognize that MISSION-Vet is a complex intervention that includes the delivery of five evidence-based practices, which could have made it more difficult for HUD-VASH case managers to fully implement. Future research might systematically vary the MISSION-Vet components in a disaggregate study design to assess the relative contribution of each component. This would ultimately help to examine whether delivering only a portion of the MISSION-Vet services would demonstrate a robust effect as was the case on our prior trials with clients receiving the full dose of the MISSION-Vet services.


MicroRNAs were detected in the plasma samples by specifically-primed RT-PCR followed by multivariate analyses of the miR expression dynamics. We herein established the plasma miR-classifier for detecting CTCL based on the miR-155 upregulation and miR-203/miR-205 downregulation with 100% specificity and 94% sensitivity. The 3-miR-score in the consecutive samples coincided with the clinical outcome of MF and SS patients such as the therapy response or changes in the clinical stage or tumor size. Quantitation of the selected microRNAs in plasma is a specific and straightforward approach for evaluating CTCL outcome representing, thus, a valuable tool for CTCL diagnostics and therapy response monitoring.Primary cutaneous T-cell lymphomas (CTCL) affect the skin and tend to transform and spread. CTCL involves primarily the Mycosis fungoides (MF) and more aggressive Sezary syndrome (SS). Oncogenic microRNAs (miRs) are stable epigenetic inhibitors often deregulated in the tumour and detectable as biomarkers in non-cellular fractions of peripheral blood. The tumour-specific expression of miR-155, miR-203, and miR-205 was shown to correctly diagnose CTCL. We herein asked whether these microRNAs can be used as plasma biomarkers for clinical CTCL monitoring. Patients with CTCL ( Primary cutaneous T-cell lymphomas (CTCL) affect the skin in form of lesions that tend to transform and spread into lymph nodes, organs or blood. CTCL is a heterogeneous group of lymphomas that in up to 3/4 of cases present as Mycosis fungoides (MF). Another form of CTCL with overlapping and aggressive features is represented by the Sezary syndrome (SS). Differential diagnosis involves other types of lymphomas or benign lesions (BL) such as Psoriasis vulgaris, atopic dermatitis and other chronic skin diseases all requiring repeated skin histology examinations in order to differentiate the BL from CTCL. Staging systems for MF/SS distinguish surface lesions , as well as lymph node or systemic involvement of the tumour cells . ImproviCTCL pathogenesis may be a result of chronic antigenic stimulation by viral or bacterial infection resulting into clonal expansion and transformation of T-cells also involving their epigenetic reprograming . There eWork from others, as well as from our group, demonstrated that miRs produced by a tumour  can be very reproducibly detected in non-cellular fractions of peripheral blood and, thus, can serve as tumour biomarkers . It is ap = 0.001 utilizing exact binomial test) and sensitivity was 94% .As pointed above the expression of certain miRs in tumour tissue distinguishes CTCL from BLs with a very high accuracy . It is aalignant B, right.To further understand which miRs added the most to the miR-classifier, we display the data of each miR separately in the To further study the role of the miR classifier utilizing the most influential miRs , we asked how it reflects the clinical context of CTCL. As indicated already by PCA analyses some samples tend to stay closer to the BLs and we noted that these samples were those obtained from the clinically-responding patients (CR). Therefore, we next analysed the 3-miR-score of MF patients in three consecutive samples in respect to the clinical outcome . The patWe next evaluated the following orders of benefit from therapy using the multivariate ordinal logistic regression analysis in CTCL patients. These orders included the therapy response , clinical stages (IA < IB < IIA < IIB < IIIA), and tumour size (skin: 1 < 2 < 3 < 4). Values of miRs analysed in plasma of the control (Ctrl), BLs, and CTCL also represented such order of diagnoses (Ctrl < BL < CTCL). Results summarized in the miRs are very stable epigenetic modifiers of posttranscriptional expression profiles, often involved in tumourigenesis. miR stability precludes its use as the disease biomarker. We herein evaluated the expression of candidate miR-oncomarkers in CTCL in the plasma of patients with MF and SS in several disease time-points, which allowed us to compare how dynamic changes in miR level relate to CTCL biology and clinical outcomes. miR-155, miR-203, and miR-205 were indeed differentially regulated in CTCL compared to control samples from patients with BLs or normal individuals. In addition, the predictive 3-miR-score previously generated for tumour tissues  successfIt is very important to correctly diagnose CTCL from benign lesions, as well as to establish sensitive and specific CTCL monitoring of potential relapse. Our work suggests that this can be achieved via specific microRNA detection in plasma. As shown in Taken together, the tool involving the miR detection in plasma allows both to correctly diagnose CTCL from benign lesions as well as to establish sensitive and specific monitoring of CTCL progression. Although our work represents initial findings on relatively small patient cohort we are confident that associating the miR levels with clinical outcome allows to initiate further validation studies in the CTCL patients.n = 7) of median age was 66 years (61–83) and three SS patients  as well as the controls of either five healthy volunteers  or six BL patients with Psoriasis vulgaris or atopic dermatitis  participated in the study. Samples were collected between 2012 and 2016 in collaboration between the Haematology and Dermatology clinical departments of General Hospital Prague. All samples were obtained upon written informed consent according to the Helsinki Declaration and approved by the internal ethics committee . Patients’ characteristics are shown in the Patients with CTCL  donated three plasma samples with a median interval between samples of five months (2–14 Mo.). The diagnosis of CTCL was established according to the WHO2008 criteria. MF patients  from 200 μL of plasma ) and processed with several modifications exactly as published previously [−(ΔCp) equation in duplicate samples [RNA was isolated using an miRNeasyeviously . Reverse samples . PCA wasMiR expression data were evaluated using a multivariate proportional odds regression model together with the likelihood ratio test to associate their levels to belonging to one of the four orders of benefit . The applied analysis is appropriate as the model takes into account the ordinary character of the categorical variables.The determination of plasma levels of oncogenic microRNA miR-155 and two tumour-suppressor microRNAs, miR-203 and miR-205, represents a very useful tool for both making diagnostic distinctions from benign skin lesions, as well as for monitoring of CTCL patients utilizing peripheral blood plasma samples. Furthermore, this 3-miR-score relates to CTCL clinical responses and in the near future may be used when introducing novel therapies to evaluate their efficacy and assessing the remaining tumour burden.


Objective: although the preponderance of research on functional brain activity investigates mean group differences, mounting evidence suggests that variability in neural activity is beneficial for optimal central nervous system (CNS) function. Independent of mean signal estimates, recent findings have shown that neural variability diminishes with age and is positively associated with cognitive performance, underscoring its adaptive nature. The present investigation sought to employ functional near infrared spectroscopy (fNIRS) to derive two operationalizations of cerebral oxygenation, representing mean and variability [using standard deviation (SD)] in neural activity, and to specifically contrast these mean- and SD-oxyhemoglobin (HbO) estimates as predictors of cognitive function. Method: a total of 25 older adults (71 to 81 years of age) completed a test of cognitive interference (Multisource Interference Task) while undergoing fNIRS recording using a multichannel continuous-wave optical imaging system (TechEn CW6) over bilateral prefrontal cortex (PFC). Time-varying covariation models were employed to simultaneously estimate the within- and between-person effects of cerebral oxygenation on behavioral performance fluctuations. Results: mean effects were predominantly observed at the between-person level and suggest that greater concentrations of HbO are associated with slower and less accurate performance. Greater HbO variability at the between-person level was associated with slower performance, but was associated with faster performance at the within-person level. Conclusions: these findings are in keeping with assertions that mean and variability confer complementary (as opposed to redundant) sources of information regarding the effective functioning of a neural system and suggest that fNIRS is a viable methodology for capturing meaningful variance in the hemodynamic response that is characteristic of adaptive CNS function. In stark contrast to the number of studies examining cognitive behavioral and physiological variability, there is a paucity of research on variability in neural activity, despite the longstanding knowledge that neural variability is not merely noise, but rather a central feature of a stable and well-functioning neural system.–,,Empirical support for the functional significance of neural variability includes examples from lifespan developmental phenomena as well as linkages to behavioral performance. McDonnell and Ward–,,,,–,,,,,,Neural variability also appears to be sensitive to developmental phenomena at both ends of the lifespan and to recovery following injury. EEG data from infants1.1Given that the majority of the extant neural variability literature is based on average between-person effects , it is of particular interest to investigate whether associations between behavior and neural function are also observed within persons . It has long been known that analyses executed at separate levels of nested-data hierarchies  do not necessarily yield equivalent results. Referred to as the ecological fallacy,1.2In the present study, we examined mean and variability operationalizations of the cortical hemodynamic response, at both the within- and between-person levels, for older adults performing an executive functioning task. Several key research aims were explored. First, functional estimates for mean and variability during an executive function task were derived. Relative to fMRI, the faster fNIRS sampling frequency arguably yields a more precise and reliable estimate of neural variability. Although several operationalizations of neural variability have been employed including a block normalized SD approach (fMRI literature) as well as multivariate multiscale entropy (MSE: EEG literature), these various approaches and the signals they index may not be equivalent.Time-varying covariation models were then employed to estimate the within-person  and between-person (at the sample average level) associations for these two oxyhemoglobin (HbO) indicators on behavioral cognitive performance [Multisource Interference Task (MSIT) response latency and accuracy] across all performance blocks within the experiment. For variables such as functional activation that are studied over any longitudinal interval, an estimate for any given cross-sectional sample  comprises both between- and within-person sources of variance.,,Finally, a related research objective pertains to understanding whether the complementary operationalizations (mean and standard deviation) of cortical hemoglobin concentrations were differentially modulated as a function of cognitive load (control versus interference conditions) on the MSIT task. Neural variability remains poorly understood for higher-order cognitive measures, such as executive function, and the modulation of neural variability has not been explored within persons. Consistent with some interpretations offered in the functional literature for the impact of load modulation on mean neural activations,22.1This study was approved by the University of Victoria Human Research Ethics Board and was conducted in accordance with institutional guidelines. Data were collected from a narrow-age cohort 71- to 81-years of age , one of which is a different numerical value than the others. Using a serial response input device  to ensure nate see . A rest 2.3fNIRS data were recorded using a continuous-wave TechEn CW6 system , with a sampling frequency of 50 Hz (corresponding to ex (PFC)  containi,The optical array was positioned relative to several 10 to 20 landmarks . 3-D coordinates of scalp reference and optode locations were obtained using a Polhemus Fastrak digitizer system , to perform probabilistic spatial registration.The methods undertaken as part of this study were well situated to investigate the within- and between-person associations between HbO variability and behavioral performance, as well as the modulation of neural variability by cognitive load. Although fNIRS is limited in its spatial resolution to cortical regions, the greatest age-related group differences in BOLD variability (between young and old) have emerged in relation to cortical regions.2.4,Preprocessing of the fNIRS data was performed using Homer 2 software.2.5Mean- and SD-based operationalizations of HbO were derived for the purpose of this study. Outliers were identified as values ,,Mean estimates of HbO were estimated by aggregating across all samples contained within a given experimental condition. We employed a block design to index HbO within each condition. Specifically, we derived single estimates of HbO within a given block. This equated to eight segments for the MSIT (four control and four interference). Signal variance was estimated with block independent estimates, based on in-house software following the approach used by Garrett and colleagues.33.1MSIT scores were first screened for blocks with accuracy performance 3.2a priori hypotheses regarding expected directional effects, we employed one-tailed  6.08 software was used to fit linear mixed models to examine the time-varying covariation (or “coupling”) between each operationalization of hemoglobin  and behavioral performance , with separate models for each fNIRS channel . In order to avoid a type II error and incorrectly rejecting a true finding, we did not correct for multiple comparisons.HbO mean: The within- and between-person associations of HbO mean on variation in behavioral performance were tested separately for each behavioral metric and fNIRS channel. ,HbO SD: The within- and between-person associations of HbO SD on variation in behavioral performance were tested separately for each behavioral metric and fNIRS channel.4,–,,,,,–This investigation sought to examine complementary operationalizations of the cortical hemodynamic response for older adults completing a measure of executive functioning. Neuroimaging data have historically been examined through central tendency computations, in order to derive what have been perceived as more stable estimates based upon block averaging of neural activity. Recent work has revisited previous assertions that neural variability is not merely noiseUsing fNIRS, two operationalizations of HbO were derived, based on central tendency (mean) and variability (SD). Behavioral associations with each operationalization of HbO differed across task difficulty (control or interference), behavioral metric (accuracy or response latency), cortical area, and level of analysis (between- versus within-person). In general, greater amounts of mean HbO  were associated with less accurate and slower response in both MSIT conditions, in lateral regions of PFC (including DLPFC). Greater amounts of mean HbO were associated with faster performance in the easier control condition, but with slower performance in the more challenging interference condition. One interpretation of this finding is that recruiting additional neural tissue may have been beneficial as the task remained relatively easy, but that as it became more challenging, additional recruitment of neural tissue was no longer able to compensate for the increased task demands.–,–On balance, these results are in keeping with fMRI findings that mean and variability in the HbO signal confer complementary sources of information.4.1The observed effects that have been reported in this study are limited due to sample size constraints and duration of task , resulting in decreased power. To ascertain fully the extent to which each operationalization of HbO is driven by within- or between-person effects, a greater sample size is required; ideally, one in which the incidence of a potential cognitive impairment is apparent . Similarly, additional experimental blocks would allow for greater exploration of the within-person associations between HbO and behavior. Event-related designs will also allow for more precise yoking of neural variability with behavioral performance, with an increased number of blocks increasing statistical power for the coupling analyses. The use of short-separation channels would have allowed for a more precise estimate of high-frequency physiological artifacts ,5,Variability operationalizations of neuroimaging data are emerging as complementary metrics to conventionally employed mean-based computations. Our results demonstrate that variability in HbO recorded using fNIRS is sensitive to behavioral performance and further substantiate claims that increased neural variability is adaptive. Notably, these associations were apparent at the within-person level, suggesting that they were not driven by between-person confounds such as age differences or individual differences in practice or learning. These patterns are consistent with suppositions that increased neural variability connotes moment-to-moment fluctuations indicative of a nimble, responsive, and dynamic system.


Bald heads may soon not be a sign that identifies a cancer patient receiving treatment. Indeed, therapies for cancer patients are improving dramatically leading to increased survival rates, and most are associated with a different toxicity profile. Recently, antibody-based therapy has transformed the therapeutic landscape and biology of non-small cell lung cancer (NSCLC) and other solid tumors. This may also reshuffle the playing cards for an intensive care unit (ICU) admission policy due to improved outcomes. In November 2016, the results of the KEYNOTE-024 trial showed for the first time the superiority of immunotherapy over chemotherapy as first-line treatment for NSCLC [Immunotherapy with immune checkpoint inhibitors has ushered in a new chapter in the treatment of solid tumors. In place of directly targeting cancer cells, these drugs stimulate immune cells to enhance the host immune response against cancer. Two classes of immune checkpoint inhibitors with blocking antibodies have demonstrated a high level of antitumor activity in a variety of malignancies: (i) anti-cytotoxic T-lymphocyte antigen-4 antibodies (not yet approved in NSCLC); and (ii) inhibitors of PD-1 —all established second-line therapeutic options for patients with metastatic NSCLC .Antibody-based therapy in NSCLC is a therapeutic breakthrough—the first mAb was approved only a decade ago . It is d


Salmonella enterica serovar Typhimurium LT2.Chitinases are glycosyl hydrolases that catalyze the hydrolysis of the β-1,4 linkages in complex carbohydrates and those that contain GlcNAc. These enzymes are considered emerging virulence factors during infection because the host glycan changes. This is the release of four single chitinase deletion mutants in  N-glycosylated proteins in the host membrane, which enable host association as well as glycan digestion, to gain access to the cell membrane to initiate invasion  that belong to the GH18 and GH19 families 16. GH enzinvasion . Glycansinvasion  are founinvasion . This prListeria monocytogenes led to a reduction in bacterial counts in the liver and spleen of infected mice (Escherichia coli (AIEC) LF82 deletion of the chiA gene significantly reduced the adhesion to IECs compared to that of the wild type (N-glycosylated chitin-binding protein (CHI3L1) on the host cell to mediate close interaction between the host membrane and bacterial cell, which is regulated in animal models of colitis and in human inflammatory bowel diseases (IBDs) (SL0018 (chiA) gene in the Salmonella SL1344 strain was strongly induced during the infection of murine macrophage cells  . Microarge cells , 16. Thehttp://www.100kgenomes.org) is a large-scale sequencing consortium that offers the use of new next-generation sequencing methods to provide cutting-edge methods for pathogen detection and control in the food supply. This project is focused on sequencing genomes of bacteria from the environment, plants, animals, and humans worldwide, providing new insights into the genetic diversity of pathogens and the microbiome. Four chitinase deletions  were constructed in the Weimer laboratory   . CultureDavis) 1825. Genomhttp://www.ncbi.nlm.nih.gov/sra). NCBI GenBank accession numbers for the genome assemblies are listed in All sequences are publicly available and can be found at the 100K Pathogen Genome Project BioProject (NCBI PRJNA186441) in the Sequence Read Archive (


The analysis of viral vector genomic integration sites is an important component in assessing the safety and efficiency of patient treatment using gene therapy. Alongside this clinical application, integration site identification is a key step in the genetic mapping of viral elements in mutagenesis screens that aim to elucidate gene function.We have developed a UNIX-based vector integration site analysis pipeline (Ub-ISAP) that utilises a UNIX-based workflow for automated integration site identification and annotation of both single and paired-end sequencing reads. Reads that contain viral sequences of interest are selected and aligned to the host genome, and unique integration sites are then classified as transcription start site-proximal, intragenic or intergenic.https://sourceforge.net/projects/ub-isap/.Ub-ISAP provides a reliable and efficient pipeline to generate large datasets for assessing the safety and efficiency of integrating vectors in clinical settings, with broader applications in cancer research. Ub-ISAP is available as an open source software package at  LMO2 gene, leading to deviant expression of the LMO2 protein, a proto-oncogene implicated in the causation of T cell leukaemias [The goal of many gene therapy strategies is to stably integrate new DNA sequences into the genome of therapeutically relevant target cell populations, and their progeny. Engineered viral vectors are capable of performing this function, either through their underlying biological properties, as in the case of retroviral vectors, or through combinatorial approaches whereby elements driving integration are incorporated into vector design and delivery, such as the piggyBac system for adeno-associated viral (AAV) vectors . One impukaemias . Similarukaemias .The mapping of ISs to their genomic location is an important step in understanding the potential genotoxicity associated with the use of GT vectors in the clinic, in understanding the mechanisms of insertional mutagenesis (IM), and in enabling development of improved vectors in preclinical studies . IS idenhttp://seqmap.compbio.iupui.edu/) no longer provide mapping to the most recent human genome assembly. For this reason, we developed a stand-alone and user-friendly UNIX-based IS analysis pipeline, Ub-ISAP [Although web-based tools are available for IS analysis using NGS data, problems arise when access becomes difficult for example, Quickmap tool,  or when  Ub-ISAP . In addiUb-ISAP was designed to analyse junction fragments generated from a range of custom library preparation methods including LM-PCR  or Mu trFor our work, fragment libraries generated using Mu transposase methodology, and using primers designed to amplify from the 3’TR of the integrated cassette into downstream gDNA were size-selected to yield fragments ranging from 100 to 400 bp in length. Re-amplification of the library fragments then facilitated the incorporation of sequencing platform-specific adaptors and an additional six-nucleotide sequence barcode to enable sample recognition for multiplexing. Junction fragment DNA libraries were sequenced using the Thermofisher Ion-torrent proton Personal Genome Machine (PGM). As described above, the sequencing reads comprised both the genomic sequence required for IS mapping and viral sequence for identification of candidate junction fragments.Ub-ISAP accepts multiple raw NGS reads in a single FASTQ file (with .fastq file extension) for single reads or two FASTQ files (with .fastq file extensions) for paired-end reads, which can be selected on being prompted by the command line or as command line parameters. The only other inputs required from the user are the 5′ TR primer and the 3′ distal primer sequences used in library preparation. The command line parameters for processing of both single and paired end reads are as follows:./Ub-ISAP.sh <1(Single) or 2 (Paired) > <Directory> <Indexedfile> <bedfile> <fastqfile1> <Forwardprimer> <ReversePrimer> <fastqfile2> <Forwardprimer> <ReversePrimer>The initial step, using the Python program cutadapt  takes thTo identify the ISs within the host genome, trimmed reads are mapped to the reference genome using Bowtie2 with pre-defined filters. These pre-defined filters could be altered with basic understanding of Unix scripting. For our datasets, Ub-ISAP aligned the selected reads against the Genome Reference Consortium Human build 37 (GRCh37/hg19), which is set as the default genome file. Selection of an alternative genome (for example an updated human genome or another species) is possible at this point by selection of the “other” option in the software and provision of the appropriate filenames for the indexed genome and its associated bedfile. The bed file must be in the order of chromosome number, starting position, end position, gene accession number, gene symbol and strand (+/−).Single end reads are aligned to the reference genome allowing zero mismatches and reads that align to more than one location are discarded.Reads from step 1 are aligned again, this time allowing one mismatch. Any reads aligning at more than one genomic location are again discarded.Reads from step 2 are aligned again, but allowing two mismatches. Any reads aligning at more than one genomic location are again discarded.The process for end-to-end alignment proceeds as follows:The reads that have aligned only at one location following three alignment rounds are considered for calling as Unique IS and subsequent annotation. This strategy is implemented to reduce false-positive mapping of reads containing one or two base position errors generated during the sequencing reaction . For paihttp://bedtools.readthedocs.io/en/latest/).Recent NGS-based IS analyses have demonstrated that different types of viral vectors have distinct integration patterns . For exaUb-ISAP functions on LINUX command line and has been developed and tested on the UBUNTU operating system version 14.0. The required installed programs for running Ub-ISAP are cutadapt, Bowtie2, Bedtools and Perl.Ub-ISAP provides researchers with a consistent methodology for the extraction of vector ISs from NGS data generated from gDNA (regardless of species origin) without the need to develop custom scripts. In addition to evaluating the computational performance of Ub-ISAP using datasets derived from complex junction fragment libraries (prepared as described above), we also tested Ub-ISAP against a web-based tool for IS analysis (VISA) , and alsThe computation performance of Ub-ISAP was investigated using NGS data sets . Removal of reads less than 30 bp reduced the number of TR-selected and trimmed reads to 200,526 (29.7% of input for data set 1) and 162,394 (27.8% of input for dataset 2). Alignment to hg19/GRCh37 (Feb 2009) proceeded in a stepwise manner, as described above, resulting in 101,622 reads , 78,236 reads  for datasets 1 and 2 respectively. Since these datasets comprised single end reads, further alignment rounds (as described above) yielded 67,201 reads (9.94% of input reads) and 51,210 reads (8.76% of input reads) respectively.Reasons for exclusion of reads throughout these filtering steps include the lack of a valid match on the reference genome, the presence of trimmed reads that contain only vector sequence, as previously described, or the presence of reads that could not be unambiguously mapped to the reference genome .Following alignment, reads were analysed to identify 1981 and 1789 unique ISs from data sets 1 and 2 respectively, giving an average IS read coverage of 33.9 (dataset 1) and 28.6 (dataset 2) (range 1 – 1789). Variability in read depth of individual ISs can be ascribed to either the repeated recovery of a specific IS due to PCR amplification bias during library preparation and/or clonal expansion of vector-transduced cell populations after integration . It is lhttps://visa.pharmacy.wsu.edu/bioinformatics/) using single end reads in dataset 1 and 2 (VISA is unable to process paired end reads). Use of VISA required uploading of sequence files in the FASTA format, and input of vector and TR sequences. By contrast, Ub-ISAP was able to process FASTQ files directly from the sequencer platform, with input of 5′ TR and 3′ primer sequences. In Ub-ISAP, trimming of 3′ primer sequences from TR-selected and trimmed reads allows for better alignment to the reference genome. VISA does not allow for input of these sequences for trimming. We are unable to ascertain whether VISA can align such reads, as VISA output contains final ISs identity but not intermediate alignment files for review.In order to compare IS data generated from Ub-ISAP with data generated via alternative methods, we compared both the workflow and outcome of raw sequence read processing via Ub-ISAP with that obtained using the web based Vector Integration Site Analysis  by VISA  generateThe greatest advantage of Ub-ISAP over VISA in our hands was the significant reduction in processing time in addition to the stringency of alignment calling, with a similar IS yield. Other advantages relating to the running of Ub-ISAP on the UNIX operating system on a local computer included the ability to customise Ub-ISAP for alternative IS selection criteria, and the possibility of further custom tool development. Another advantage of Ub-ISAP over VISA is the capacity to process paired end reads, which may provide greater certainty of mapping ISs within repetitive regions of the genome, and an overall greater confidence in the reliability of output ISs datasets, since only concordant reads from each end of a given junction fragment proceed to the alignment process.Ub-ISAP was further validated as an efficient IS analysis tool by re-analysis of two existing, independent sets of raw sequence reads, analysed using alternative custom workflows, for which published ISs data were available , 24. FirP < 0.0001) than the L conditions. Also in agreement with these published data, was the presence of significantly fewer ISs outside gene coding regions (intergenic) under the P conditions compared with the L conditions . In contrast to the published analysis for intergenic location, in which there was no statistically significant difference between the P and L culture conditions, the number of ISs identified by Ub-ISAP were, however, significantly different in favour of a greater number within gene coding regions (intragenic) under the P culture conditions compared with the alternative L conditions . This result further supports the hypothesis suggested by Hallwirth et al. that the P conditions for transduction may impart a higher risk of insertional mutagenesis than do the alternative L conditions. The small increase in number of ISs recovered and annotated using Ub-ISAP (2.11% and 11.99% increase in P and L respectively) was sufficient to resolve a statistically significant difference between the conditions for this category of IS location.Annotation of the ISs extracted for the P and L datasets via Ub-ISAP into TSS-proximal, intragenic and intergenic location, followed by pair-wise statistical comparisons of IS distribution between the three categories upheld the statistically significant differences demonstrated by Hallwirth and colleagues  in relatSince the above-mentioned comparison is limited to the total number of annotated ISs recovered after analysis rather than their identity, we also sought to compare the identity of ISs recovered using the two different pipelines by extracting data for the top 20 IS-containing genes for both the P and L datasets derived from each different method. This exercise demonstrated that Ub-ISAP extracted an identical list for the top 18/20 genes targeted for integration /piggyBac combination vector system that drives integration of an otherwise predominantly non-integrating AAV vector cassette into the liver cell genome in mice through a transposon mediated mechanism . The libAs data for extraction of the top 20 targeted genes for the published data were unavailable, we instead compared the top 20 targeted genes for integration between the PBR-I and PBR_II ISs generated by Ub-ISAP, predicting concordance between these lists, given that both ends of the same integrated vector cassette would have been represented within the pooled DNA junction fragment library that underwent sequencing. This exercise verified that the list of top 20 genes containing ISs was identical for PBR-I and PBR_II generated paired-end reads Fig.  and in fThe parameters discussed above to filter the raw and the aligned reads to identify unique ISs using Ub-ISAP are pre-defined and recommended, but these could be changed by users, with basic knowledge of Unix scripting. The usage of dataset 1 and 2 from the Thermofisher PGM platform, datasets PBR_I and PBR_II from Illumina Mi-Seq and datasets P and L from Illumina Genome Analyser IIx (GAIIx), and the flexibility to change the parameters, proves the capability of Ub-ISAP to analyse datasets from varying NGS platforms. Ub-ISAP provides a concise result, with chromosomal positioning and closest gene accession number and symbol from the processed reads in tabular format as.txt file, and containing the unique ISs characterised as TSS-proximal, intragenic or intergenic. Being a UNIX-based tool, Ub-ISAP is designed to be installed on a local computer allowing shorter processing times. Another additional advantage for Ub-ISAP is that the reference genome database and version used for alignment (human or other) can be user-defined and can be easily updated. Ub-ISAP is therefore of utility across all species and into the future as additional genomic databases become available.Integration site (IS) analysis is an important step for assessing the safety and efficiency of gene therapies that use integrating viral vectors. Ub-ISAP is a time and memory efficient UNIX-based pipeline that allows researchers to analyse NGS datasets for ISs in a consistent manner. It can be readily updated with the current reference genome. Results are returned in a simple format to allow easy analysis of integration profiles of viral vectors.