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The multiprotein Mediator coactivator complex functions in large part by controlling the formation and function of the promoter-bound preinitiation complex (PIC), which consists of RNA polymerase II and general transcription factors. However, precisely how Mediator impacts the PIC, especially post-recruitment, has remained unclear. Here, we have studied Mediator effects on basal transcription in an in vitro transcription system reconstituted from purified components. Our results reveal a close functional interplay between Mediator and TFIIH in the early stages of PIC development. We find that under conditions when TFIIH is not normally required for transcription, Mediator actually represses transcription. TFIIH, whose recruitment to the PIC is known to be facilitated by the Mediator, then acts to relieve Mediator-induced repression to generate an active form of the PIC. Gel mobility shift analyses of PICs and characterization of TFIIH preparations carrying mutant XPB translocase subunit further indicate that this relief of repression is achieved through expending energy via ATP hydrolysis, suggesting that it is coupled to TFIIH's established promoter melting activity. Our interpretation of these results is that Mediator functions as an assembly factor that facilitates PIC maturation through its various stages. Whereas the overall effect of the Mediator is to stimulate basal transcription, its initial engagement with the PIC generates a transcriptionally inert PIC intermediate, which necessitates energy expenditure to complete the process. (C) 2016 Elsevier Ltd. All rights reserved.

There is no consensus regarding the best practice for detecting murine pinworm infections. Initially, we evaluated 7 fecal concentration methods by using feces containing Aspiculuris tetraptera (AT) eggs (n = 20 samples per method). Sodium nitrate flotation, sodium nitrate centrifugation, Sheather sugar centrifugation, and zinc sulfate centrifugation detected eggs in 100% of samples; zinc sulfate flotation and water sedimentation detected eggs in 90%. All had better detection rates than Sheather sugar flotation (50%). To determine optimal detection methods, Swiss Webster mice were exposed to Syphacia obvelata (SO; n = 60) or AT (n = 60). We compared the following methods at days 0, 30, and 90, beginning 21 or 28 d after SO and AT exposure, respectively: fecal concentration (AT only), anal tape test (SO only), direct examination of intestinal contents (cecum and colon), Swiss roll histology (cecum and colon), and PCR analysis (pooled fur swab and feces). Detection rates for SO-exposed mice were: PCR analysis, 45%; Swiss roll histology, 30%; intestinal content exam, 27%; and tape test, 27%. The SO detection rate for PCR analysis was significantly greater than that for the tape test. Detection rates for AT-exposed mice were: intestinal content exam, 53%; PCR analysis, 33%; fecal flotation, 22%; and Swiss roll histology, 17%. The AT detection rate of PCR analysis combined with intestinal content examination was greater than for PCR analysis only and the AT detection rate of intestinal content examination was greater than for Swiss roll histology. Combining PCR analysis with intestinal content examination detected 100% of infected animals. No single test detected all positive animals. We recommend combining PCR analysis with intestinal content examination for optimal pinworm detection.

Background/Aims: The role of cytomegalovirus (CMV) reactivation during exacerbations of ulcerative colitis (UC) is yet a matter of debate, and assessment of CMV infection in UC patients remains an ongoing challenge. We aimed to identify associated parameters and compare detection methods for CMV infection during UC exacerbation. Methods: Clinical, pathological and virological parameters were retrospectively analyzed in all patients hospitalized in our institution for UC exacerbation between January 2009 and April 2015, who underwent full evaluation for CMV infection in colonic tissue by histopathology, immunohistochemistry (IHC) and CMV-PCR. Results: Of 28 patients who underwent full examination for tissue CMV-infection, 13 (46.4%) were found to be positive for CMV. Tissue CMV-PCR was more sensitive for the detection of CMV infection than histopathology and IHC. CMV-positive patients had a statistically higher frequency of recent steroid treatment and fever, with higher mean partial Mayo scores and lower mean albumin levels. There were no significant differences between CMV-positive and CMV-negative patients in terms of age, severity of colitis and disease duration. In a multivariable model, only recent steroid treatment and fever were independently associated with colonic CMV infection. Conclusions: This study provides a clinical model to detect the presence of CMV infection in patients hospitalized with UC exacerbation, which could direct proper investigation and facilitate timely empirical therapy (C) 2017 S. Karger AG, Basel

How type I and II interferons prevent periodic reemergence of latent pathogens in tissues of diverse cell types remains unknown. Using homogeneous neuron cultures latently infected with herpes simplex virus 1, we show that extrinsic type I or II interferon acts directly on neurons to induce unique gene expression signatures and inhibit the reactivation-specific burst of viral genome-wide transcription called phase I. Surprisingly, interferons suppressed reactivation only during a limited period early in phase I preceding productive virus growth. Sensitivity to type II interferon was selectively lost if viral ICP0, which normally accumulates later in phase I, was expressed before reactivation. Thus, interferons suppress reactivation by preventing initial expression of latent genomes but are ineffective once phase I viral proteins accumulate, limiting interferon action. This demonstrates that inducible reactivation from latency is only transiently sensitive to interferon. Moreover, it illustrates how latent pathogens escape host immune control to periodically replicate by rapidly deploying an interferon-resistant state.

Background: Assisted Reproductive Technology (ART) has undergone considerable changes over the last decade, with consequences on ART outcomes in different regions of the world being unknown. Methods: We conducted a systematic review of published national and regional ART registry data to assess how changes in clinical practice between 2004 and 2013 have impacted outcomes in Australia and New Zealand, Canada, Continental Europe, the United Kingdom (U.K.), Japan, Latin America, and the United States (U.S.). The data reflect 7,079,145 total ART cycles utilizing both fresh and previously cryopreserved embryos from autologous oocytes that resulted in 1,454,724 live births. This review focused on the following measures: ART cycle volume, use of cryopreserved embryos, single embryo transfer (SET), live birth rates in fresh and frozen-thawed cycles, and perinatal outcomes in recent years. Results: SETs and utilization of frozen-thawed embryos increased worldwide over the study period. In 2012 SET utilization in all ART cycles was highest in Japan and Australia/New Zealand (82.6% and 76.3% respectively) and lowest in Latin America (16.0%). While gradual improvements in live birth rates were observed in most regions, some demonstrated declines. By 2012-2013, fresh cycle live birth rates were highest in the U.S. (29%) and lowest in Japan (5%). In Japan, the observed decline in fresh cycle live birth rate coincided with transition to minimal stimulation protocols, transfer of frozen-thawed rather than fresh embryos, and implementation of an SET policy. Similarly, implementation of an SET policy in parts of Canada was followed by a decline in fresh cycle live birth rate. Increasing live birth rates in frozen-thawed embryo cycles, seen all over the world, partially compensated for declines in fresh ART cycles. During 2012-2013 Australia/New Zealand and Japan reported the lowest multiple delivery rates of 5.6 and 4% respectively while the US had the highest of 27%. In recent years, preterm delivery rates in all regions ranged between 9.0 to 16.6% for singletons, 53.9 to 67.3% for twins, and 91.4 to 100% for triplets and higher order multiples. Inconsistencies in the way perinatal outcome data are presented by various registries, made comparison between regions difficult. Conclusions: ART practices are characterized by outcome differences between regions. International consensus on the definition of ART success, which accounts for perinatal outcomes, may help to standardize worldwide ART practice and improve outcomes.

In this study, we describe four patients from two unrelated families of different ethnicities with a primary immunodeficiency, predominantly manifesting as susceptibility to Epstein-Barr virus (EBV)-related diseases. Three patients presented with EBV-associated Hodgkin's lymphoma and hypogammaglobulinemia; one also had severe varicella infection. The fourth had viral encephalitis during infancy. Homozygous frameshift or in-frame deletions in CD70 in these patients abolished either CD70 surface expression or binding to its cognate receptor CD27. Blood lymphocyte numbers were normal, but the proportions of memory B cells and EBV-specific effector memory CD8(+) T cells were reduced. Furthermore, although T cell proliferation was normal, in vitro-generated EBV-specific cytotoxic T cell activity was reduced because of CD70 deficiency. This reflected impaired activation by, rather than effects during killing of, EBV-transformed B cells. Notably, expression of 2B4 and NKG2D, receptors implicated in controlling EBV infection, on memory CD8(+) T cells from CD70-deficient individuals was reduced, consistent with their impaired killing of EBV-infected cells. Thus, autosomal recessive CD70 deficiency is a novel cause of combined immunodeficiency and EBV-associated diseases, reminiscent of inherited CD27 deficiency. Overall, human CD70-CD27 interactions therefore play a nonredundant role in T and B cell-mediated immunity, especially for protection against EBV and humoral immunity.

Futility disputes are more likely to be resolved and relational breaches repaired by engaging in a process that fosters communication between clinicians, patients, and families. This essay calls for mediative fluency. The preemptive use of a futility definition can stifle conversation when it is needed most, exacerbating the very power imbalances and associated health disparities that often precipitate futility disputes. When clinicians, patients, and families engage in dialogue, clinicians can appreciate what motivates requests for what is thought to be futile care, and patients and families can better understand the limits of available therapies. This sharing of knowledge, values, and attitudes cannot be achieved through the unilateral invocation of a futility definition. Furthermore, futility definitions are prone to interpretative judgment by clinicians and can be informed by the norms and attitudes attendant to a practitioner's medical specialty. They also need to be interpreted in the context of emerging trends in medical therapeutics and in relation to the clinical details of each case. In the aggregate, these challenges make the application of a futility definition futile.

Sex hormones act throughout the entire brain of both males and females via both genomic and nongenomic receptors. Sex hormones can act through many cellular and molecular processes that alter structure and function of neural systems and influence behavior as well as providing neuroprotection. Within neurons, sex hormone receptors are found in nuclei and are also located near membranes, where they are associated with presynaptic terminals, mitochondria, spine apparatus, and postsynaptic densities. Sex hormone receptors also are found in glial cells. Hormonal regulation of a variety of signaling pathways as well as direct and indirect effects on gene expression induce spine synapses, up-or downregulate and alter the distribution of neurotransmitter receptors, and regulate neuropeptide expression and cholinergic and GABAergic activity as well as calcium sequestration and oxidative stress. Many neural and behavioral functions are affected, including mood, cognitive function, blood pressure regulation, motor coordination, pain, and opioid sensitivity. Subtle sex differences exist for many of these functions that are developmentally programmed by hormones and by not yet precisely defined genetic factors, including the mitochondrial genome. These sex differences and responses to sex hormones in brain regions, which influence functions not previously regarded as subject to such differences, indicate that we are entering a new era of our ability to understand and appreciate the diversity of gender-related behaviors and brain functions. (C) 2016 Wiley Periodicals, Inc.