When facing various extrinsic or intrinsic stimuli, BBB is damaged which is an early event in pathogenesis of a variety of neurological diseases in old patients including acute and chronic cerebral ischemia, Alzheimer’s disease and etc. Treatments that could maintain the integrity of BBB may prevent neurological diseases following various stimuli. Old people often face a common stress of sepsis, during which lipopolysaccharide (LPS) is released into circulation and the integrity of BBB is damaged. Of note, there is a significant decrease of melatonin level in old people and animal. Melatonin has been shown to preserves BBB integrity and permeability via a variety of pathways: inhibition of matrix metalloproteinase-9 (MMP-9), inhibition of NADPH oxidase-2, and impact on silent information regulator 1 (SIRT1) and nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome. More important, a recent study showed that melatonin supplementation alleviates LPS-induced BBB damage in old mice through activating AMP-activated protein kinase (AMPK) and inhibiting gp91 The blood brain barrier (BBB) is a regulated interface between the peripheral circulation and the central nervous system , composed of occludin, claudin and zo-1, are key components of the BBB  could produce neuroinflammation Shi, , promotiper se but also on the incidence or severity of age-related diseases /NF-κB signaling pathway in neonatal rats (Hu et al., in vitro cell model (Zhao et al., in vivo mice model (Zhou X. et al., phox up-regulation in brain capillary endothelial cells (Figure AMPK activation has been shown to play important role in maintaining the integrity of BBB (Liu et al., s Figure . AMPK acMMP-9 has been shown to play important role in BBB damage (Jin et al., SIRT1 was reported to be beneficial in sepsis. Using EX527, a SIRT1 inhibitor, the authors figured out that melatonin alleviated BBB damage in mice which subjected to cecal ligation and puncture via SIRT1 to inhibit inflammation, apoptosis and oxidative stress (Zhao et al., Aging and sepsis triggered NLRP3 inflammasome activation (Volt et al., Although acute toxicity of melatonin is extremely low in both animal and human studies, melatonin may still cause minor adverse effects, such as headache, insomnia and nightmares (Malhotra et al., In conclusion, decreased melatonin levels may account for the BBB damage in old people who often face the common stress of sepsis and neuroinflammation. Melation supplementation treatment significantly inhibits such events. Therefore, continuous daily melatonin supplementation may help prevent sepsis and neuroinflammation-related neurological diseases through maintaining the integrity of BBB in old people. Since melatonin has low toxicity profile and high efficacy in many pathophysiological states, it should be more commonly tested/used in the medical and veterinary arenas. Further studies are needed to verify the important significance of daily melatonin supplementation in old people.W-CL, XW, XZ, XC and XJ wrote the manuscript and XC, XJ obtained the funding. XW drew the figures. All authors have approved the final version of this review article.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.


Recently, a genome-wide study about gene copy number gains and corresponding expression levels has been analyzed in a cohort of 190 non-small cell lung cancer (NSCLC) patients . The auSince decades, much effort has been invested to better understand the relationship between gene expression and prognosis of tumors (Hellwig et al., 2016; Stock e


During 2014–2016, we conducted mosquito-based Zika virus surveillance in Rio de Janeiro, Brazil. Results suggest that Zika virus was probably introduced into the area during May–November 2013 via multiple in-country sources. Furthermore, our results strengthen the hypothesis that Zika virus in the Americas originated in Brazil during October 2012–May 2013. Aedes aegypti mosquito is considered the main vector of Zika virus in urban and suburban areas throughout the world, including Brazil, where the mosquito has been confirmed, together with Ae. albopictus mosquitoes, as a vector for the virus , which we pooled (n = 178) and subjected to Zika virus detection using real-time RT-PCR  clustered within the same strongly supported lineage, which included strains detected in Rio de Janeiro and other parts of Brazil in late 2015 and in 2016  Figure. Ae. aegypti mosquitoes that were collected during a mosquitoborne virus surveillance program in Rio de Janeiro. Information regarding Zika virus infection rates is lacking for female and male mosquitoes trapped in the field. However, experiments performed in the laboratory demonstrated transovarial transmission of Zika virus among Ae. aegypti mosquitoes and revealed a minimal filial infection rate of 1:290 (In this study, we detected Zika virus RNA in 2 pools of engorged Ae. aegypti mosquitoes trapped in Rio de Janeiro before the first case of autochthonous Zika virus disease was diagnosed in the city (In conclusion, we showed the presence of Zika virus in engorged Bayesian maximum clade credibility tree representing the time-scale phylogeny of the Zika virus outbreaks in the Americas.


A characteristic of post-surgery patients, particularly the more elderly, can be a persistent self-propagating cerebral inflammatory syndrome referred to as post-operative cognitive dysfunction (POCD). Changes can be analogous to those seen in Alzheimer's disease , a synthetic sedative with analgesic and anxiolytic properties, is widely used in surgery. It is a selective αPhysical trauma, including that caused by surgery, induces an innate immune response that includes release of pro-inflammatory cytokines such as TNF and interleukins (Arvin et al., Since systemic TNF has long been known to cross the blood-brain barrier (Gutierrez et al., ++ homeostasis, and thus the ionic signaling cascades on which normal function of these cells depends (Park et al., Dexmedetomidine has an extensive history of improving neurological function, for example when given preemptively in animal models tibial fracture (Zhu et al., 2-adrenoceptor agonist appears implicated, in that yohimbine, an α2-adrenoceptor antagonist, enhanced TNF levels when the two were compared in a lipopolysaccharide-induced liver damage model (Chen et al., Various pathways of TNF inhibition by dexmedetomidine have been explored. Its action as a αUsing the same mouse tibial fracture model as did others with dexmedetomidine six years later (Zhu et al., Given the pleiotropic nature of TNF, reducing its excess production with dexmedetomidine may also cast light on the mechanisms of other useful outcomes of therapy with this agent that are presently little understood. For instance dexmedetomidine is an acknowledged analgesic, particularly in surgical settings (Vaughns et al., Escherichia coli protecting them completely from harm (Tracey et al., The background information required to rationalize the contrasting outcomes reported in the two trials (Su et al., Thus it seems logical that, in the context of post-surgical delirium (Su et al., A useful step in understanding its mechanism further would be to experimentally compare preemptive use of dexmedetomidine and one of the specific anti-TNF biologicals reported to minimize POCD delirium, pain and anxiety, and to induce morphine tolerance. Because of their molecular size, these biologicals would require administering intracerebroventricularly or perispinally (Tobinick, IC proposed the scope of the review. Both authors were involved in planning and editing the manuscript, blending their complementary expertises. Both authors read, altered and approved the final 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.


Serial crystallography was developed for the use at free-electron lasers but the approach has recently also been adapted to synchrotron sources. Here we discuss how the synergy between the two X-ray sources will facilitate a wide application of the technique in microcrystallography, room-temperature structure determination and time-resolved studies. For time-resolved studies, the small crystal size allows for rapid diffusive saturation in mix-and-inject analysis of biochemical reactions, and full optical saturation of the sample by a pump laser in studies of light-driven proteins. The ability to outrun most radiation damage avoids the need for sample cooling and its artifacts, allowing studies of molecular machines at work in their correct room-temperature thermal bath or a controlled chemical environment.Recently we have seen rapid progress in the serial crystallography (SC) method at X-ray free-electron lasers (XFELs). Injection of thousands of protein microcrystals into the ∼10Despite these achievements, the XFEL community remains relatively small, due to the limited availability of XFEL beamtime which, at present, allows only two simultaneous data collections worldwide (in Japan and USA). In addition, serial femtosecond crystallography (SFX) at XFELs is experimentally demanding and complex, currently requiring sizable interdisciplinary teams of collaborating scientists and engineers. The transfer of the SC approach to next-generation synchrotrons upgraded for higher flux density and with beamlines using sophisticated focusing optics, submicron beam diameters and fast low-noise photon-counting detectors offers a way out of this dilemma. In applications such as room-temperature data collection or phasing from radiation-sensitive microcrystals, serial millisecond crystallography (SMX) at synchrotrons has developed into a viable alternative. In the near future, it may even be possible to extend the method to time-resolved studies. Already it frees beamtime at XFELs to exploit their unique capabilities, such as outrunning damage in structure analysis from nanocrystals or ultrafast time-resolved crystallography.et al., 2008et al., 2011et al.  shows many papers on SC, with some of the most cited examples devoted to the interplay between synchrotrons and XFELs.The experimental origins of fast SC can be traced to an early proposal to fire bioparticles in a continuous single-file stream across a beam for diffraction analysis using a Rayleigh jet (Spence & Doak, 2004 al. 1998. Rossmanet al., 2013The term ‘serial’ in synchrotron studies has recently become popular in a wider sense, and may also refer to taking a series of diffraction patterns along a few larger crystals, or the scanning of fixed target devices. Such serial approaches share the idea of distributing the radiation dose, to get the maximum signal from all of the available crystal volume. Short millisecond exposures may outrun slower secondary radiation damage from diffusing radicals or relaxation of individual damaged molecules (Warkentin et al., 2015et al., 2016cis–trans reaction. Yet both proteins are available in large amounts and can be grown into big crystals suitable for time-resolved Laue diffraction experiments at a synchrotron. A recent study of the light-driven proton pump bacteriorhodopsin, for which only a few micrometre thick crystals were available, resulted in a series of 13 snapshots at logarithmically spaced time points after activation (Nango et al., 2016An important application where injector-based SC has proven advantageous is fast time-resolved pump–probe crystallography. Myoglobin (Barends et al., 2014et al., 2015et al., 2015et al., 2016The work was only feasible using sample-efficient high-viscosity injectors (Weierstall


Reactive oxygen and nitrogen species are generated during normal cellular metabolic activities but can also play an etiopathogenetic role in a variety of conditions, including cardiovascular diseases, neurodegenerative diseases, and cancer.Although the role of oxidative stress in human physiology and pathology has been intensely studied for several decades, it is still far from being understood. Certainly, the roles of free radicals and antioxidants have been significantly redefined. Some “negative” actions of free radicals in human biology and pathology are now known to be “positive,” and the hypothesis that “classical antioxidants”  could be always beneficial for the human health was not confirmed by several epidemiological and clinical studies. The possible reasons of the failures of the current antioxidant therapies, including methodological pitfalls in the drug development and delivery and the lack of good biological markers to select the patients, have been reviewed elsewhere , 2.We currently believe that instead of “antioxidants,” it is more appropriate to develop “modulators of oxidative stress” because, depending on the condition, it could be more beneficial to reduce or increase the oxidative stress.β-Synthase: Increased CO Reactivity as a Novel Molecular Mechanism of Pathogenicity?” by J. B. Vicente et al., “Protective Mechanisms of the Mitochondrial-Derived Peptide Humanin in Oxidative and Endoplasmic Reticulum Stress in RPE Cells” by L. Minasyan et al., “Effect of Emodin on Preventing Postoperative Intra-Abdominal Adhesion Formation” by G. Wei et al., and “Opuntia spp.: Characterization and Benefits in Chronic Diseases” by M. del Socorro Santos Díaz et al.), some cardiovascular effects of modulators of oxidative stress , some neurological effects of modulators of oxidative stress , and some effects of modulators of oxidative stress related to cell growth and cancer development  Administration Delays Hepatic Cell Proliferation by Altering Oxidative State in the Regenerating Rat Liver” by A. Butanda-Ochoa et al., “Preclinical Antileukemia Activity Of Tramesan: A Newly Identified Bioactive Fungal Metabolite” by M. R. Ricciardi et al., and “Markers of Oxidative Stress and Inflammation in Ascites and Plasma in Patients with Platinum-Sensitive, Platinum-Resistant, and Platinum-Refractory Epithelial Ovarian Cancer” by J. C. Cantón-Romero et al.).In this special issue, several aspects of the modulation of oxidative stress were examined including the role of polyphenols  and other natural substances  Opuntia spp., stressing that all Opuntia components  used as nutritional and pharmaceutical agents exhibit beneficial properties mainly resulting from their high content in antioxidants, while several other phytochemical components , which have been characterized, also contribute to the medicinal properties of Opuntia spp. that might have a great economic potential because these plants grow in arid and desert regions.In particular, this special issue contains two interesting contributions coming from INSERM Toulouse. The first one is an original research paper prepared in collaboration with CNRS and Palau Sabatier University in which C. Camare et al. investigated whether 4-hydroxynonenal (4-HNE), an aldehydic lipid oxidation product abundantly present in oxidized LDL, contributes to its proangiogenic properties. Using the immunofluorescence analysis of human atherosclerotic lesions, they found colocalization of HNE adducts with CD31 (marker of the endothelium), indicating a close relationship between 4-HNE and neovascularization. Moreover, they revealed that physiological concentrations of 4-HNE also enhance the formation of tubes by human microvascular endothelial cells (HMEC-1), through mechanisms involving reactive oxygen species (ROS) and activation of the neutral type 2 sphingomyelinase and sphingosine kinase-1 (nSMase2/SK-1) pathway. Eventually, they found that carbonyl scavengers hydralazine and bisvanillyl-hydralazone inhibited such proangiogenic effects of 4-HNE. In their second paper  prepared jointly with partners from Mexico and from INRA in Toulouse, they described in detail features of the well-known plants of In the original research paper prepared by partners from Riga and from Zagreb, we found novel information about growth-modulating effects of the dihydropyridine derivatives (DHPs) with potential antioxidant capacities. Thus, I. Bruvere et al. revealed that cell-type-specific differences in the growth-modifying effects of the DHPs in vitro can be attributed only to the novel types of the DHPs, which differentiate these substances from their well-known predecessor diludine. Since the growth-modifying effects of the novel DHPs indicate possible differential effects on cancer and on nonmalignant cells, which might be also different from their antioxidant effects, these substances deserve particular attention and further studies.The original research paper prepared by three teams from Karlsruhe, Germany, reveals the possible beneficial effects of pentaerythritol tetranitrate (PETN), which were not described before. Namely, pulmonary arterial hypertension was induced by the ranging dosage of the i.v. applied monocrotaline, which induced endothelial dysfunction, pulmonary vascular wall thickening, and fibrosis, as well as protein tyrosine nitration, thus causing an increase in pulmonary arterial pressure, followed by the increase in heart/body and lung/body weight ratios. However, in this study, S. Steven et al. found also that PETN therapy could act beneficially upon these pathophysiological processes, most likely by upregulation of heme oxygenase-1 (HO-1).


There is a natural decline in cognitive function as we age, particularly in processing speed and working memory. A range of modifiable factors can increase the risk of accelerated cognitive decline including hypertension, chronic inflammation, atherosclerosis, diabetes, atrial fibrillation, stroke, and impaired central nervous system glucose regulation. Given the lack of adequate interventions for cognitive decline and dementia, it is essential that treatments with the potential to reduce the risk of cognitive impairment are thoroughly explored.Nutraceutical and lifestyle medicines, including vitamins, herbs, supplements, physical activity, and diet, have been shown to possess anti-inflammatory, antihyperglycaemic, and antihypertensive properties, suggesting their utility in targeting the pathophysiology associated with risk for cognitive decline. The purpose of this special issue was to explore the role of such complementary medicines in the modification of risk factors for cognitive decline.Clinical trials involving nutraceutical and herbal medicine interventions for people with mild cognitive impairment and dementia are reviewed in G. Z. Steiner et al. The manuscript titled “A Systematic Review of Intervention Studies Examining Nutritional and Herbal Therapies for Mild Cognitive Impairment and Dementia Using Neuroimaging Methods: Study Characteristics and Intervention Efficacy” focuses on papers that feature neuroimaging outcome measures.D. Chang et al. provide a detailed review of the literature on herbal medicine for vascular dementia in their paper titled “Herbal Medicine for the Treatment of Vascular Dementia: An Overview of Scientific Evidence.”Vascular disease, such as cerebrovascular disease and atherosclerosis are risk factors for cognitive impairment. X. Zhou et al. review the evidence on this link and promising Traditional Chinese Medicines in their paper titled “Vascular Contributions to Cognitive Impairment and Treatments with Traditional Chinese Medicine.”The fourth review featured in this special issue summarises the evidence on the efficacy of physical activity in reducing depression. D. C. Mathersul and S. Rosenbaum's paper is titled “The Roles of Exercise and Yoga in Ameliorating Depression as a Risk Factor for Cognitive Decline.”H. Macpherson et al. outline the findings of a clinical trial on older adults in their paper titled “The Effects of Four-Week Multivitamin Supplementation on Mood in Healthy Older Women: A Randomized Controlled Trial.”β1–42.”The first preclinical paper in this special issue explores the effectiveness of Huannao Yicong extract, a Traditional Chinese Medicine, in reducing tau hyperphosphorylation in Alzheimer's disease model rats. Y. Cao et al.'s paper is titled “Traditional Chinese Medicine Huannao Yicong Decoction Extract Decreases Tau Hyperphosphorylation in the Brain of Alzheimer's Disease Model Rats Induced by Aβ and LRP1 in Hippocampus.”X. Wang et al. report their findings from an electroacupuncture treatment on Alzheimer's disease model mice in their paper titled “Improvement of Electroacupuncture on APP/PS1 Transgenic Mice in Spatial Learning and Memory probably due to Expression of AThe second electroacupuncture paper in this special issue explores the effects of this treatment on cerebral hypoperfusion model rats. C.-X. Zheng et al.'s paper is titled “Electroacupuncture Ameliorates Learning and Memory and Improves Synaptic Plasticity via Activation of the PKA/CREB Signaling Pathway in Cerebral Hypoperfusion.”In the paper titled “Preservation of Cognitive Function by Lepidium meyenii (Maca) Is Associated with Improvement of Mitochondrial Activity and Upregulation of Autophagy-Related Proteins in Middle-Aged Mouse Cortex” by S.-S. Guo et al., maca  was found to improve cognition and behavior, and mitochondrial dysfunction in middle-aged mice.β-induced neurotoxicity on SH-SY5Y cells are modulated by a the Traditional Chinese Medicine, Yi-Zhi-Fang-Dai formula. The paper is titled “Yi-Zhi-Fang-Dai Formula Protects against Aβ1–42 Oligomer Induced Cell Damage via Increasing Hsp70 and Grp78 Expression in SH-SY5Y Cells.”In an in vitro study by L. Liu et al., the effects of AThe final paper in this special issue by M. A. Akhtar et a. titled “Medicinal Plants of the Australian Aboriginal Dharawal People Exhibiting Anti-Inflammatory Activity” explores the medicinal effects of a range of Eucalyptus plants used in traditional Australian Aboriginal medicine by the Dharawal people.Authors contributed with a range of papers including original research and review articles spanning in vitro, in vivo, and human studies that improve the understanding of the pathology involved in cognitive impairment in older age and the development of evidence-based complementary treatment strategies for cognitive decline. This collection of works provides a snapshot (that is by no means exhaustive) of current research and some of the emerging trends in this field. This special issue features 11 papers including 4 reviews and 7 original research articles. The complementary therapies explored include nutritional supplements (2 papers), herbal and traditional medicines (6 papers), physical activity (1 paper), and electroacupuncture (2 papers). A brief description of these 11 works is detailed below.


In 1920, German neuropathologist Alfons Maria Jakob  describeEtymologia series is dedicated to the memory of Richard T. Johnson, MD (1931–2015), the leading prion disease authority in the United States for many years and great friend of CDC’s infectious disease programs, so many of which involve central nervous system disorders.This issue of Emerging Infectious Diseases’ long-running


Neuroinflammation is an important component of many neuroinflammatory and neurodegenerative diseases. Microglia and astrocytes play a key role in neuroinflammation. Activated glial cells under neuroinflammatory condition produce proinflammatory cytokines, chemokines, nitric oxide (NO), and other neurotoxic mediators. Thus, it has been conventionally thought that the inhibition of such glial activation might be an effective neuroprotective strategy under inflammatory or degenerative disease conditions in the central nervous system (CNS). However, recent studies indicated that activated glia could adopt either neuroprotective or neurotoxic phenotype depending on instigating stimuli present in the given microenvironment . LipopolIn a recent study by Song et al. , a phenoIn summary, based on a high-throughput screen of an approximately 3,500-member in-house library, Song et al. identified a novel small-molecule compound that has an anti-neuroinflammatory effect via the phenotypic switch towards the M2-like anti-inflammatory state in glia. The GPM is a specific PPAR-γ agonist that can be used to fine-tune glial phenotypes and functions. The GPM-mediated anti-inflammatory state of brain glia may lead to beneficial neuroprotective effects and has therapeutic potential in neuroinflammatory and neurodegenerative diseases. Recent studies suggested that functional phenotypes of macrophages and glia are closely associated with their glucose metabolism. A more recent study by Van den Bossche et al. [


Coxiella burnetii, Brucella abortus, Salmonella enterica serovar Typhimurium, Legionella pneumophila, Chlamydia trachomatis, and Orientia tsutsugamushi manipulate the endocytic and secretory pathways. Understanding how bacterial effector proteins manipulate host processes not only gives us keen insight into bacterial pathogenesis, but also enhances our understanding of how eukaryotic membrane trafficking is regulated.Intracellular bacteria have developed numerous strategies to hijack host vesicular trafficking pathways to form their unique replicative niches. To promote intracellular replication, the bacteria must interact with host organelles and modulate host signaling pathways to acquire nutrients and membrane for the growing parasitophorous vacuole all while suppressing activation of the immune response. To facilitate host cell subversion, bacterial pathogens use specialized secretion systems to deliver bacterial virulence factors, termed effectors, into the host cell that mimic, agonize, and/or antagonize the function of host proteins. In this review we will discuss how bacterial effector proteins from  Obligate and facultative intracellular bacteria have developed numerous methods to hijack host membranes to promote uptake, survival, and intracellular replication. Following uptake, intracellular pathogens must engage host organelles and subvert host defense mechanisms to establish their unique intracellular niches. To facilitate interactions with the host, many pathogenic bacteria deliver bacterial virulence proteins, termed effectors, into the host cell using specialized secretion systems. These proteins traffic to distinct subcellular locations within the host cell  Rabex-5  are synthesized in the rough endoplasmic reticulum (ER), delivered to the ER-Golgi intermediate compartment (ERGIC), modified as they move through the Golgi, and are ultimately packaged into transport vesicles for delivery to their final destination  associates with early and late endosomal compartments , encoded on two Salmonella pathogenicity islands  that matures by trafficking through the endocytic pathway and interacts with the secretory pathway  in the SCV, altering the membrane surface charge  that project from the SCV  2 and 3 play an integral role in maintaining SCV positioning and depletion of SCAMP2 or 3 results in dispersion of the SCV within the host cell  GTPases  or de-PCylation, respectively . Interestingly, ubiquitination of these proteins does not require host E1 or E2 enzymes, representing a method of ubiquitination that is unique to trans-Golgi network and is recruited to target membranes by GTP-bound Rab7 P and potentially modulates the activity of the retromer complex. Mutation of ridL results in decreased L. pneumophila replication in macrophages, LAMP1 accumulation on the LCV, as well as recruitment of retrograde cargo receptors (Vps10 and CIMPR) and SNX1 and 2 , ectopic pregnancy, and infertility ; IncA (CT119), InaC (CT813), and IPAM (CT223)  where it undergoes extensive interactions with the Golgi (Grieshaber et al., C. trachomatis causes relocalization of SNXs from endosomes to the inclusion membrane and induces inclusion tubulation (Aeberhard et al., L. pneumophila, knockdown of retromer components enhances C. trachomatis replication, suggesting the retromer complex controls infection (Mirrashidi et al., trans-Golgi network (Mirrashidi et al., In a study to map the host-Inc interactome, an interaction between the inclusion membrane protein IncE and SNX5/6, a component of the retromer complex, was identified (Mirrashidi et al., C. trachomatis hijacks non-vesicular ER-TGN transport to acquire ceramide, a sphingomyelin precursor. Host lipids including sphingomyelin, cholesterol, phosphatidylcholine, and phosphatidylinositol are incorporated into the bacterial cell (Hackstadt et al., In addition to acquiring nutrients by exploiting host vesicular trafficking pathways, C. trachomatis T3SS substrates localize to the inclusion membrane, an additional subset of T3SS proteins are predicted to be secreted into the host cell cytosol (Subtil et al., While a large number of Orientia tsutsugamushi is an obligate intracellular bacterium that is the causative agent of scrub typhus, a potentially fatal disease that is endemic to the Asia-Pacific region. O. tsutsugamushi is transmitted to humans via the bite of an infected trombiculid mite (Valbuena and Walker, O. tsutsugamushi is internalized by clathrin-dependent endocytosis and associates with early and late endosomes as evident by co-localization with EEA1 and LAMP2, respectively (Chu et al., O. tsutsugamushi is released into the cytoplasm where it moves along microtubules to the MTOC (Kim et al., O. tsutsugamushi escapes the phagosome is unknown, however it encodes a hemolysin, tlyC and a phospholipase D (Ge and Rikihisa, C. burnetii (Pan et al., L. pneumophila (Pan et al., Anaplasma phagocytophilum (Caturegli et al., O. tsutsugamushi Ikeda possesses 47 Ank open reading frames (ORFs; Nakayama et al., Orientia infection, the ER is distended and the Golgi is perturbed. While the exact mechanism of how this occurs and the benefit to Orientia is unknown, it is possible that at least part of this is dependent on Ank9.The ankyrin repeat domain is a 33-residue eukaryotic motif involved in mediating protein-protein interactions for numerous host cell processes including transcription, cell cycle regulation, signal transduction, and cytoskeletal rearrangements (Voth et al., Obligate and facultative intracellular bacteria have developed sophisticated strategies to modulate host endocytic and secretory trafficking to promote formation of their unique replicative niches. The adaption of genetic tools for manipulation of obligate and facultative intracellular pathogens has substantially enhanced our understanding of host-pathogen interactions and effector function. While great strides have been made toward understanding how effector proteins manipulate host processes to redirect membrane and nutrients to the parasitophorous vacuoles, the function of most effector proteins still remains ill-defined and genetic manipulation of some of these organism presents specific challenges. Large-scale screens to identify putative binding partners of ectopically produced type III secreted effectors (Mirrashidi et al., All authors listed have made a substantial, direct, and intellectual contribution to the work, and approved it for publication.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.


X-ray free-electron lasers (XFELs) provide ultra-bright femtosecond X-ray pulses that are intense enough to enable data collection of small weakly scattering objects and short enough to outrun most radiation damage effects. These beam features allow not only to study highly radiation-sensitive systems such as metalloproteins and tiny crystals but also to capture fleeting reaction intermediates in time-resolved studies. They therefore expand the structural biologist’s toolbox by a great deal . Second, there is the serial femtosecond crystallography (SFX) data collection scheme that only once probes randomly oriented microcrystals intersecting the XFEL beam and gives a still image. Thus, it had long been doubted that the resulting integrated Bragg intensities are accurate enough for de novo phasing to be successful. A number of XFEL-specific phasing approaches have been suggested  using in vivo grown nanocrystals diffracting to better than 2.5 Å resolution  becomes meaningful and fewer images (and thus less sample and beam time) are needed to obtain accurate Bragg intensities.Knowing whether one can solve a structure by SAD phasing and when to stop collecting more data because the anomalous signal in SAD phasing is high enough is still an active area of research [see, for example, Terwilliger  al. 2016 and refe al. 2017 in this de novo phasing required 60000 indexed patterns for fully automatic model building (Barends et al., 2014et al., 2016et al. (2015et al., 2017In fact, data processing programs have improved significantly in recent years. This is reflected in the finding that fewer and fewer diffraction patterns of a given dataset are needed for phasing. For example, the very first demonstration of  al. 2015 could no


Clinical observations that hyperlipidemia and diabetes are both reversed in mice and in patients taking Gleevec support the drugs' primary metabolic targets by biguanides and statins. This is evident by structural data demonstrating that Gleevec shows pyridine- and phenyl-guanidine homology with Phenformin and identical phenylcarbamoyl structural and ligand binding homology with Lipitor. The misunderstood mechanism of action of Gleevec is emblematic of the pervasive flawed reasoning that genomic analysis will lead to targeted, personalized diagnosis and therapy. The alternative perspective for Gleevec's mode of action may turn oncotargets towards metabolic channel reaction architectures in leukemia and melanoma, as well as in other cancers.Phenformin's recently demonstrated efficacy in melanoma and Gleevec's demonstrated anti-proliferative action in chronic myeloid leukemia may lie within these drugs' significant pharmacokinetics, pharmacodynamics and structural homologies, which are reviewed herein. Gleevec's success in turning a fatal leukemia into a manageable chronic disease has been trumpeted in medical, economic, political and social circles because it is considered the first successful targeted therapy. Investments have been immense in omics analyses and while in some cases they greatly helped the management of patients, in others targeted therapies failed to achieve clinically stable recurrence-free disease course or to substantially extend survival. Nevertheless protein kinase controlling approaches have persisted despite early warnings that the targeted genomics narrative is overblown. Experimental and clinical observations with Phenformin suggest an alternative explanation for Gleevec's mode of action. Using  Gleevec's  demonstrated anti-proliferative action in chronic myeloid leukemia ushered in the era of targeted therapies with claims that the drug blocks the constitutively active BCR-ABL tyrosine kinase thereby inhibiting phosphorylation of multiple downstream proteins of the mitogenic signaling pathways . Partly et al. [in vitro studies have thrown into similar question whether the single-target kinase signal blocking mechanism is indeed the primary mechanism for STI-571's mode of action [13C-guided precise flux measurements, in a comparative multiple cell line study. Targeted 13C-glucose tracer fate association studies demonstrated the drugs' downstream impact on submolecular fatty acid processing and deuterium depleting metabolic events that occurred independent of Gleevec's molecular target. Clinical observations that hyperlipidemia and diabetes are both reversed in mice and in patients taking Phenformin and Gleevec support the drugs' primary metabolic targets [In their excellent paper Petrachi et al.  reiteratf action  using 13 targets –10.et al.'s work in melanoma [The structural homology between Phenformin and Gleevec certainly links Petrachi melanoma  to Gleevmelanoma , the guamelanoma  and Gleemelanoma , which hmelanoma , 14 detemelanoma . Strikinmelanoma  so that et al.'s work [via six hydrogen bond interactions [via distinct amino acid residues that are also known to bind Lipitor, Metformin and Phenformin in their own protein targets. These low affinity and high capacity drug binding architectures are evidently shared by Lipitor, Phenformin and Gleevec, which include histidine, leucine, iso-leucine, as well as serine, as specific amino acid ligands in two dimensional visual structures [13C-glucose guided metabolomics studies [It is worth knowing that all analogues designed to treat Gleevec resistance share guanide structural similarities and target the same ATP pocket for binding, all of which result in remissions in chronic myeloid leukemia. Petrachi .'s work  may exteractions . Such hyractions . Figure ructures . Gleevec studies , 12. The studies  to Metfo studies , a struc studies –24. Thes studies , 23. MorIn summary, the excellent paper by Petrachi et al. [et al. [i et al.  that demi et al.  as well i et al. –29. The  [et al.  opens di [et al. , 14, bas [et al. .


Control of cell-cell coordination and communication is regulated by several factors, including paracrine and autocrine release of biomolecules, and direct exchange of soluble factors between cells through gap junction channels. Additionally, hemichannels also participate in cell-cell coordination through the release of signaling molecules, such as ATP and glutamate. A family of transmembrane proteins named connexins forms both gap junction channels and hemichannels. Because of their importance in cell and tissue coordination, connexins are controlled both by post-translational and post-transcriptional modifications. In recent years, non-coding RNAs have garnered research interest due to their ability to exert post-transcriptional regulation of gene expression. One of the most recent, well-documented control mechanisms of protein synthesis is found through the action of small, single-stranded RNA, called micro RNAs (miRNAs or miRs). Put simply, miRNAs are negative regulators of the expression of a myriad proteins involved in many physiological and pathological processes. This mini review will briefly summarize what is currently known about the action of miRNAs over Cxs expression/function in different organs under some relevant physiological and pathological conditions. Cell-cell communication and signaling is regulated by exchange of soluble factors between cells through gap junction channels (GJC) , the level of miR-1 is downregulated with a concomitant upregulation of Cx43 within the endoneurium of the sciatic nerve  the presence of Cx43 GJCs. However, after fusion, Cx43 is downregulated by both miR-206 and miR-1 in myocytes in vitro (Anderson et al., In the cell line C2C12, which is a mouse myoblast cell line, it has been shown that miR-206 promotes muscle differentiation (Kim et al., As in skeletal muscle cells, miR-1 also controls Cx43 levels in smooth muscle cells. Thus, in overactive bladder, it was shown that MYOCD downregulates Cx43 expression by controlling miR-1 levels, showing that reduction of Cx43 could be a key factor in this pathology (Imamura et al., in vivo—was observed (Inose et al., Cx43 is the main Cx expressed in osteocytes, and its presence is fundamental for their differentiation (Civitelli, GJs play a key function in propagating action potentials, and the heart is no exception to this principle. Both Cx40 and Cx43 localize along the axis of atrioventricular conduction, including atrioventricular node, atrioventricular bundle and Purkinje fibers suggesting an important role in conducting the impulse (Gourdie et al., As mentioned, miR-1 is involved in downregulation of Cx43 in skeletal muscle development (Anderson et al., miR-1 is not the sole master switch, controlling Cx43 levels in the heart. On one hand, it has been shown that miR-19 a/b decrease Cx43 levels, and that this change is associated with cardiac arrhythmia observed in a mouse constitutively overexpressing the miR-17-92 cluster in smooth muscle and cardiomyocytes (Danielson et al., in vivo and in vitro, and inhibits tumor growth in vivo (Li et al., Significant changes in gene expression patterns that promote rapid cell division are the unifying hallmark of tumorigenesis. Each different type of cancer has a distinctive signature of “driver” mutations, which are recurrent across patients and affect genes that encode key components of the cell cycle machinery (Vogelstein et al., In nasopharyngeal carcinoma associated with the Epstein-Barr virus, downregulation of miR-218 has been consistently observed (Alajez et al., In breast cancer cell line MDA-MB-231, transfection of hsa-miR-206 decrease Cx43 levels, which was correlated with a decrease of proliferation rate and cell migration (Fu et al., An interesting potential avenue of research is to better understand whether changes in expression levels of Cxs in different types of cancer are partially or fully mediated by miRNAs. Therefore, biologically accurate models are required in order to dissect the mechanism that underlies promotion of invasiveness and worsens the clinical course of different forms of cancer.This review revisits insurmountable evidence of the relevant role of Cxs in health and disease. In addition to this, we have discussed the most recent findings in microRNA-mediated regulation of Cxs for several muscle and skeletal disorders as well as rhythm-associated and structural heart defects, and several types of cancer. Table JFC and MAR wrote 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.


Finally, we demonstrate that vector-mediated expression of alpha-synuclein (SNCA), a transcript decreased in selectively vulnerable motor neurons in all four screens, can extend life span, increase weight and decrease neuromuscular junction pathology in a mouse model of SMA. In summary, we have combined multiple data sets to identify transcripts, which are strong candidates for being phenotypic modifiers, and demonstrated SNCA is a modifier of pathology in motor neuron disease.The term “motor neuron disease” encompasses a spectrum of disorders in which motor neurons are the primary pathological target. However, in both patients and animal models of these diseases, not all motor neurons are equally vulnerable, in that while some motor neurons are lost very early in disease, others remain comparatively intact, even at late stages. This creates a valuable system to investigate the factors that regulate motor neuron vulnerability. In this study, we aim to use this experimental paradigm to identify potential transcriptional modifiers. We have compared the transcriptome of motor neurons from healthy wild-type mice, which are differentially vulnerable in the childhood motor neuron disease Spinal Muscular Atrophy (SMA), and have identified 910 transcriptional changes. We have compared this data set with published microarray data sets on other differentially vulnerable motor neurons. These neurons were differentially vulnerable in the adult onset motor neuron disease Amyotrophic Lateral Sclerosis (ALS), but the screen was performed on the equivalent population of neurons from neurologically normal human, rat and mouse. This cross species comparison has generated a refined list of differentially expressed genes, including CELF5, Col5a2, PGEMN1, SNCA, Stmn1 and HOXa5, alongside a further enrichment for synaptic and axonal transcripts. As an  The term “motor neuron disease” refers to a group of disorders, causing progressive paralysis of affected patients due to the degeneration of motor neurons cells which control voluntary movements. Importantly, not all motor neurons appear to be affected in the same way, with those that control the face being affected less that those that control the abdomen. The reason why some motor neurons are more vulnerable is unknown; however, understanding this may provide new targets for therapeutics to slow motor neuron degeneration either as stand-alone therapeutics or in combination with SMN-inducing compounds. In this study, we analysed gene expression in different groups of motor neurons and compared this to previously published expression data to identify commonalities. One of the common transcripts was alpha-synuclein (SNCA), which was consistently expressed at lower levels in vulnerable motor neurons. Importantly, when SNCA levels were increased in a mouse model of motor neuron disease, the disease phenotype was significantly reduced, including an extension in survival and reduction in motor neuron pathology. Collectively, these results demonstrate that this approach can identify disease modifiers that can reduce disease severity in models of motor neuron disease and potentially identify new therapeutic targets. SMN1 gene and visualised with Cy3-conjugated secondary antibodies . Muscles were then whole-mounted in Dako Fluorescent mounting media. Confocal microscopy was performed using a Nikon A1RThe percentage of fully occupied endplates was determined by classifying each endplate in a given field of view either fully occupied  completely overlies endplate (BTX)), partially occupied ), or vacant (no pre-synaptic label overlies endplate). At least 4 fields of view were analysed per muscle totalling >100 endplates per muscle.All data was assembled and analysed using Microsoft Excel and GraphPad Prism.S1 TableTable shows the ensembl ID , the official gene symbol (gene), the chromosomal location (locus), the average normalised read count from an N of 2 samples for resistant (BS-WT) or vulnerable (SC_WT) samples, the log2 fold change, and the relevant statistics .(XLSX)Click here for additional data file.S2 TableTable includes the mouse official gene symbol and the log2 fold change identified in Brockington et al., Kaplan et al., Murray et al., and Hedlund et al.(XLSX)Click here for additional data file.S3 TableTable includes the mouse official gene symbol and the log2 fold change identified in Brockington et al., Kaplan et al., Murray et al., and Hedlund et al.(XLSX)Click here for additional data file.


I am concerned by an article “Detecting Key Genes Regulated by miRNAs in Dysfunctional Crosstalk Pathway of Myasthenia Gravis” by Cao et al., 2015 [In this study, the authors used data from public databases to compare microarray data from the thymus and miRNome data from peripheral blood mononuclear cells (PBMCs). They analyzed the regulation of mRNAs from the thymus by miRNAs from peripheral blood mononuclear cells: two different “tissues.” Of course, they ended up with results but one can really wonder about the meaning of these results.Moreover, in their manuscript, Cao et al. compiled thymic microarray data altogether while data are from different categories of patients (seropositive (for anti-AChR antibodies) and seronegative patients)  [


Tenascins represent key constituents of the extracellular matrix (ECM) with major impact on central nervous system (CNS) development. In this regard, several studies indicate that they play a crucial role in axonal growth and guidance, synaptogenesis and boundary formation. These functions are not only important during development, but also for regeneration under several pathological conditions. Additionally, tenascin-C (Tnc) represents a key modulator of the immune system and inflammatory processes. In the present review article, we focus on the function of Tnc and tenascin-R (Tnr) in the diseased CNS, specifically after retinal and optic nerve damage and degeneration. We summarize the current view on both tenascins in diseases such as glaucoma, retinal ischemia, age-related macular degeneration (AMD) or diabetic retinopathy. In this context, we discuss their expression profile, possible functional relevance, remodeling of the interacting matrisome and tenascin receptors, especially under pathological conditions. Numerous studies demonstrate that retina and optic nerve degeneration is highly associated with remodeling of various extracellular matrix (ECM) components. Glycoproteins and proteoglycans that surround retinal cells and optic nerve fibers represent major constituents of the ECM meshwork, known as the matrisome , Tnc exhibits high expression during early development. With ongoing maturation, it is progressively downregulated, but re-expressed under pathological conditions , diabetic retinopathy, glaucoma and retinal vascular occlusion (Mizener et al., Several studies reported on a dysregulation of Tnc following cerebral, hepatic as well as myocardial ischemia (Lu et al., In the CNS, tenascins represent main structural and functional constituents of synaptic sites (Dityatev et al., AMD is defined by a deterioration of the macula and represents a major cause of vision impairment worldwide (Jager et al., The tenascin family member Tnx was identified in AMD patients in a genome-wide association study (Cipriani et al., Additionally, high levels of Tnc were observed in choroidal neovascular membranes from AMD patients (Nicolò et al., Diabetic retinopathy is also highly associated with retinal vascular dysfunction. Tnc was found in intravitreal membranes of patients with traumatic and idiopathic proliferative vitreoretinopathy as well as in diabetic retinopathy (Hagedorn et al., RGC nerve fibers exhibit a poor regeneration capacity after injury, which often leads to irreversible vision loss. Therefore, multiple studies focused on the improvement of RGC survival as well as axonal regrowth, guidance and pathfinding (Fischer and Leibinger, After optic nerve damage, Wallerian degeneration, demyelination, immune activation and glial scar formation can be observed. In this context, it has become evident that next to the intrinsic cellular repertoire, an inhibitory environment prevents regrowth of optic nerve fibers Fischer, . ECM proin vivo. Tnr was also described as a repulsive guidance molecule of newly growing as well as regenerating optic nerve fibers in the zebrafish (Becker and Becker, in vitro. In contrast to the reduced Tnr expression levels observed in the optic nerve of the salamander (Becker et al., in vivo. In addition, Tnr and axon growth-promoting molecules were found upregulated in the regenerating visual pathway of the lizard Gallotia galloti (Lang et al., Compared to mammals, the CNS of the zebrafish displays a robust axonal regeneration capacity and allows visualization of axonal regeneration and re-myelination Since Tnr is highly associated with myelinated optic nerve fibers and nodes of Ranvier, it was proposed that it might have a functional relevance in myelination processes. Recordings of action potentials from Tnr knock-out mice revealed reduced axonal conduction velocities compared to control mice. In contrast, no significant differences in the number of myelinated optic nerve fibers or in the myelin ultrastructure were observed in Tnr knock-out compared to wild-type mice (Weber et al., A potential role of Tnc in neural repair of the injured rat optic nerve was initially reported by Ajemian et al. . Here, aA huge diversity of interacting molecules can be observed for the tenascin proteins. For Tnc this includes the cell adhesion molecules contactin-1 (Rigato et al., The signaling of integrins in RGC-glia interactions is crucial for RGC survival and process extension (Vecino et al., in vitro. Furthermore, Tnc knock-out mice show an impaired de-differentiation capacity (Besser et al., As mentioned above, CSPGs are major interaction partners of Tnc. In the CNS, CSPGs are widely recognized as major inhibitory constituents of the glial scar (Silver and Silver, in vitro (Tom et al., Likewise, Tnc displays a complex interactome with other ECM glycoproteins. For instance, its interaction with fibronectin and Tnr was reported (Chiquet-Ehrismann et al., The adhesion molecule contactin-1 was identified as an important neuronal receptor for Tnr. Interaction of these two molecules was reported to mediate the repulsion and defasciculation of neurites (Pesheva et al., In the CNS, tenascin glycoproteins are important constituents of a highly regulated and dynamic matrisome. In sum, the current literature supports the notion that Tnc and Tnr are implicated in various pathological processes following retinal and optic nerve degeneration as well as various eye diseases Table . Under pJR wrote the manuscript. LR designed the figures. LR and AF revised the manuscript. All authors have approved the final article.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.


Inflammatory response, oxidative stress, and endoplasmic reticulum (ER) stress are important pathophysiological bases of the occurrence and development of diabetes mellitus (DM) and macroangiopathy complications. Selenoprotein S (SELENOS) is involved in the regulation of these mechanisms; therefore, its association with DM and macroangiopathy has gradually received attention from scholars worldwide. SELENOS has different biological functions in different tissues and organs: it exerts antioxidant protection and has anti-ER stress effects in the pancreas and blood vessels, while it promotes the occurrence and development of insulin resistance in the liver, adipose tissue, and skeletal muscle. In addition, studies have confirmed that some SELENOS gene polymorphisms can influence the inflammatory response and are closely associated with the risk for developing DM and macroangiopathy. Therefore, comprehensive understanding of the association between SELENOS and inflammation, oxidative stress, and ER stress may better elucidate and supplement the pathogenic mechanisms of DM and macroangiopathy complications. Furthermore, in-depth investigation of the association of SELENOS function in different tissues and organs with DM and macroangiopathy may facilitate the development of new strategies for the prevention and treatment of DM and macrovascular complications. Here, we summarize the consensus and controversy regarding functions of SELENOS on currently available evidence. In 2016, the World Health Organization (WHO) declared that diabetes mellitus (DM) had become the eighth most prevalent cause of disease-related mortality. Its onset is no longer mainly observed in economically developed areas; in recent decades, the number of patients has increased significantly in middle- and low-income countries. Therefore, DM has been confirmed to be a “global” public health issue [Selenium (Se), a trace element, is a key component of selenoproteins involved in a wide range of functions, including redox homeostasis, inflammatory regulation, thyroid hormone metabolism, immunity, myocardial and tumoral diseases, and reproduction –6. MoreoPsammomys obesus (P. obesus) by Walder et al. [Selenoprotein S (SELENOS), as a member of the selenoprotein family, was first discovered in the liver of r et al.  and was r et al. , 14–16. r et al. –21. The r et al. –25. Ther188UGA codon in SELENOS variant 2 CDS can be normally encoded as Sec to produce the protein SELENOS isoform 2, with 189 amino acid residues. Because the SECIS in SELENOS variant 1 is spliced, the 188UGA in the mRNA CDS cannot be encoded as Sec and is recognized as the translation termination signal. Therefore, a short protein, SELENOS isoform 1, with 187 amino acid residues, is produced [The human SELENOS gene (GenBank: NG_013322.1) is located on 15q26.3. Its primary transcript produces two types of transcription variants after selective splicing: SELENOS variant 1 (GenBank: NM_203472.2) and SELENOS variant 2 (GenBank: NM_018445.5). The main difference between these two transcripts is that the selenocysteine insertion sequence (SECIS) in the 3′untranslated region (3′UTR) of SELENOS variant 1 is spliced –30. Therproduced , suggest174Cys and 188Sec that has reductase activity [SELENOS is a single-pass transmembrane protein with an ER region (1st–25th amino acid residues), a transmembrane region (26th–51st amino acid residues), and a cytoplasmic region (52nd–189th amino acid residues). The cytoplasmic region contains the valosin-containing protein (VCP)-interacting motif  composed of the 78th–88th amino acid residues, and there is a selenosulfide bond between In addition to localizing in the ER membrane , 18, SELWhen Walder et al.  first diZhang et al.  found thFurthermore, studies in recent years on the association between SELENOS gene polymorphisms and inflammation have also become a hot research topic. Previously, Curran et al.  showed t188Sec and was maintained through the restoration of the selenosulfide bond between 174Cys and 188Sec. In other words, in the reduction process, 188Sec of SELENOS and substrates formed a mixed selenosulfide bond to cause substrate reduction. Next, through the restoration of the selenosulfide bond between 174Cys and 188Sec, SELENOS broke the mixed selenosulfide bond between SELENOS and substrates to maintain the activity of the SELENOS reductase. Liu et al. [174Cys and 188Sec depended on the function of thioredoxin (Trx), indicating that SELENOS was a Trx-dependent reductase. In addition, they also showed that SELENOS had peroxidase activity and could break down its substrate, hydrogen peroxide (H2O2), into H2O [2O2 could up-regulate SELENOS expression in HEK293 human embryonic kidney cells, while inhibition of SELENOS expression could further aggravate the LPS-induced increase of reactive oxygen species levels in HepG2 human liver cancer cells, down-regulation of GPx expression, and inhibition of cell viability. These results suggested that SELENOS may have the function of protecting tissues and cells from oxidative stress-induced damage. Previously, Gao et al. [2O2 damage and enhance cell viability. Our study group induced SELENOS overexpression in human umbilical vein endothelial cells (HUVECs) and showed that SELENOS overexpression could significantly increase HUVEC viability after H2O2 stimulation, enhance the activity of superoxide dismutase (SOD), and reduce the production of methane dicarboxylic aldehyde (MDA) [2O2, which was accompanied by the further increase of ROS and MDA levels and further reduction of the GPx activity. They also confirmed that the antioxidant protection function of SELENOS was associated with mitogen-activated protein kinase (MAPK) and c-JUN N-terminal kinase (JNK).Christensen et al.  showed tu et al.  showed tinto H2O . The abointo H2O  and Zenginto H2O . They foo et al.  showed tde (MDA) . Gan et de (MDA)  showed tde (MDA)  indicateAs an ER membrane protein, SELENOS plays an important role in the maintenance of the morphology and distribution of ER in cells . In addiIn addition to investigating the relationship between SELENOS and inflammation, Fradejas et al.  also anaMartínez et al.  comparedP. obesus. In addition, the inhibition of SELENOS expression by glucose in a concentration-dependent manner was also confirmed in in vitro cultured HepG2 liver cancer cells. Moreover, Gao et al. [Walder et al.  showed to et al.  confirmeo et al.  induced The expression of SELENOS in 3T3-L1 adipocytes was inhibited by glucose or insulin in a concentration-dependent manner . KarlssoThe contradiction of the association of adipose tissue SELENOS with glucose metabolism in the aforementioned studies was also present in related studies on the association of skeletal muscle SELENOS with glucose metabolism. Walder et al.  confirmeSAA can interact with SELENOS, which was confirmed by both a yeast-two hybrid experiment and surface plasmon resonance analysis , and the2O2 damage and enhance cell viability, suggesting that SELENOS was a protective factor in pancreatic islet and could protect pancreatic islet β cells from oxidative stress damage. The study by Christensen et al. [174Cys and 188Sec in the molecular structure of SELENOS rendered SELENOS with reductase activity; in addition, the study by Liu et al. [The progressive apoptosis of pancreatic islet β cells is an important feature of the occurrence and development of DM, and oxidative stress damage induced by high glucose is one of the reasons for pancreatic islet β cell apoptosis. The study of Gao et al.  showed tn et al.  showed tu et al.  confirmeIn addition to the localization of SELENOS in the plasma membrane, secreted SELENOS was also detected in the culture medium of HepG2 liver cells and human serum samples. However, secreted SELENOS was not detected in the culture medium of 3T3-L1 adipocytes, L6 skeletal muscle cells, Min6 pancreatic islet β cells, HEK293 human embryonic kidney cells, HUVECs, or human aortic vascular smooth muscle cells (HA/VSMCs), suggesting that serum SELENOS was mainly secreted by hepatocytes , 33. In Therefore, SELENOS in different tissues and organs has different functions. The results are summarized in Fig. Hyrenbach et al.  used the2O2; the changes after inhibition of SELENOS expression had the opposite results [Vascular endothelial cells play an important role in the maintenance of cardiovascular homeostasis, while endothelial dysfunction is the initiating step in the development of atherosclerosis –78. Our  results . These r2O2 or TM and increase VSMC apoptosis. Therefore, it was speculated that SELENOS could increase the resistance of VSMCs to oxidative stress and ER stress [VSMCs are important components of the tunica media of vascular walls. Their abnormal proliferation is involved in the formation of early atherosclerosis plagues, and the apoptosis of VSMCs in advanced atherosclerosis plaques will induce the reduction of collagen production to cause fibrous cap thinning and reduce plague stability , which iR stress .2O2. When SELENOS expression was inhibited, Cav-1 and protein kinase Cα (PKCα) expression levels increased [2O2, indicating that SELENOS increased the resistance of VSMCs to oxidative stress damage through the MAPK/JNK pathway. In addition, the antioxidant function of SELENOS in blood vessels may also be associated with the aforementioned 188Sec in its molecular structure. Furthermore, the study of Liu et al. [2O2 into H2O, which could partially explain the study results of our group [Our study group further showed that high SELENOS expression could inhibit the increase of caveolin-1 (Cav-1) induced by Hncreased , suggestncreased  confirmeu et al.  also shour group  and thosur group . These rShibata et al.  confirmeIt has been confirmed that various SELENOS SNPs are associated with the risk of the development of autoimmune inflammatory diseases. Marinou et al.  showed tSELENOS is closely associated with inflammation, oxidative stress, and ER stress. In-depth and comprehensive understanding of its association with the above reactions may not only elucidate or supplement pathogenic mechanisms of relevant diseases but also provide new evidence for clinical physicians to develop new strategies for disease prevention and treatment. However, the current understanding of this aspect is limited; there are still some scientific questions that need to be further confirmed or studied in-depth, such as whether SELENOS can inhibit the expression of other inflammatory factors and whether the regulation of ER stress by SELENOS is different in different cell types. Furthermore, whether the human SELENOS gene promoter region has negative regulatory elements that can regulate SELENOS gene expression and whether there are transcription factors that can interact with these elements should also be further explored. These studies will help to provide new targets for drug intervention and to discover more effective disease treatment methods.The functions of SELENOS in inflammatory reactions, oxidative stress, and ER stress point to its great potential in DM and macroangiopathy. It has been shown that SELENOS expressed in different tissues and organs has different effects on the occurrence and development of DM and its macroangiopathy. The high expression of SELENOS in pancreatic islets and blood vessels can exert an antioxidant protection function and can increase the defense capacity of VSMCs to ER stress, while SELENOS expressed in liver, adipose tissue, and skeletal muscle can promote the occurrence and development of DM and insulin resistance. Based on the above functional characteristics of SELENOS, the method by which tissues and organs specifically regulate its expression to fully utilize the advantages of SELENOS and weaken its adverse effects still require further in-depth studies. SELENOS gene polymorphism is associated with DM, macroangiopathy, tumors, and autoimmune inflammatory diseases; therefore, it is expected to become one of the indicators for the prediction of the risk of the above diseases and one of the theoretical bases for the adoption of primary preventive measures for the population carrying relevant SNPs. However, SELENOS SNPs that have been discovered to be associated with disease development must still be confirmed in different populations using larger sample sizes.


Scientific Reports6: Article number: 3482410.1038/srep34824; published online: 10062016; updated: 12092016The Acknowledgments section in this Article is incomplete:“We are grateful to Gerrit Ansmann and Christian Geier for interesting discussions and for critical comments on earlier versions of the manuscript and by the Verein zur Foerderung der Epilepsieforschung e.V. (Bonn)”.should read:“We are grateful to Gerrit Ansmann and Christian Geier for interesting discussions and for critical comments on earlier versions of the manuscript. This study was supported by the Deutsche Forschungsgemeinschaft DFG (Grant No.: LE 660/5-2) and by the Verein zur Foerderung der Epilepsieforschung e.V. (Bonn)”.


The majority of the reported adverse events seen in these studies are mild or moderate in severity and tend to affect the gastrointestinal or nervous systems. These adverse events, which are common in both adults and children, are also typical of symptoms of malaria or concomitant infections present in these patients. The wealth of safety data on artemether/lumefantrine has not identified any neurological, cardiac or haematological safety concerns. In addition, repeated administration is not associated with an increased risk of adverse drug reactions including neurological adverse events. This finding is especially relevant for children from regions with high malaria transmission rates who often receive many courses of anti-malarial medications during their lifetime. Data are also available to show that there were no clinically relevant differences in pregnancy outcomes in women exposed to artemether/lumefantrine compared with sulphadoxine-pyrimethamine during pregnancy. The six-dose regimen of artemether/lumefantrine is therefore well tolerated in a wide range of patient populations. In addition, post-marketing experience, based on the delivery of 250 million treatments as of July 2009, has not identified any new safety concerns for artemether/lumefantrine apart from hypersensitivity and allergies, known class effects of artemisinin derivatives.This article reviews the comprehensive data on the safety and tolerability from over 6,300 patients who have taken artemether/lumefantrine (Coartem Plasmodium falciparum resistance to monotherapy , Gürkov et al [et al [et al [et al [et al [et al [There have been case reports of neurological problems  occurring after administration of herbal artemisinin  or artesov et al , and Adjl [et al , as showet al 1, Gürkov l [et al , and Mayl [et al , as showl [et al  or Makanl [et al , which aLumefantrine is chemically related to halofantrine, an anti-malarial known to be associated with significant prolongation of QTc interval. Indeed, QTc prolongation is a known class effect of many anti-malarial drugs. As such, cardiac safety has been thoroughly investigated during the preclinical and clinical development of AL.+ channels have been investigated in stably transfected human embryonic kidney cells (HEK293) using a whole cell patch-clamp technique [in vitro hERG channel assay showed that lumefantrine and desbutyl-lumefantrine have higher IC50 values (approximately 200-fold) than halofantrine  have been delivered to malaria-endemic countries. Post-marketing experience has not identified any new specific safety concerns apart from hypersensitivity and skin reactions , a recognized class effect for artemisinin derivatives [®) from published Novartis-sponsored and independently-sponsored clinical trials. Data are available to support the use of a six-dose regimen of AL as a safe and well-tolerated treatment for P. falciparum malaria or malaria due to mixed infection including P. falciparum in adults, adolescents, children and infants. Indeed, the safety profile of this drug is similar in both adults and children, and many of the reported adverse events are typical of symptoms of malaria or concomitant infections that are commonly seen in these patient populations. Reported adverse events are mainly of mild or moderate severity, with very few serious adverse events reported and even fewer serious adverse events considered related to treatment. In addition, no notable neurological adverse events or cardiac safety concerns were identified in the many studies that examined the safety of AL. As patients  often take repeated courses of treatment for malaria, it is reassuring to note that data are available that show repeated administration of AL is not associated with an increased risk of adverse drug reactions, in particular with regard to neurological adverse events. In addition, a thorough review of the clinical data for AL does not identify any potential cardiac safety issues. Data are also available to show that there were no clinically relevant differences in perinatal mortality, neonatal mortality, still birth, pre-term delivery, low birth weight and spontaneous abortion in women exposed to AL compared with SP during pregnancy.This review summarizes some of the safety and tolerability data on AL  treatments have been delivered and post-marketing experience confirms the favourable safety and tolerability profile for this drug. In addition, this wealth of safety evidence has not identified any safety concerns for AL apart from rare type 1 hypersensitivity reactions, a recognized class effect of artemisinin derivatives [In conclusion, AL is a safe and well-tolerated treatment for 50: Inhibitory concentration 50 (the concentration that gives 50% inhibition); ICH: International Conference on Harmonization; IKr: Rectifying K+ current; MAS: Mefloquine plus artesunate; MedDRA: Medical Dictionary for Regulatory Activities; SAE: Serious adverse event; SP: Sulphadoxine-pyrimethamine; vs.: Versus; WHO: World Health Organization.ACT: Artemisinin-based combination therapy; AE: Adverse event; AL: Artemether/lumefantrine; AQ: Amodiaquine; AQSP: Amodiaquine plus sulphadoxine-pyrimethamine; AS: Artesunate: ASAQ: Artesunate plus amodiaquine; ASTMH: American Society of Tropical Medicine and Hygiene; CQ: Chloroquine; CQSP: Chloroquine plus sulphadoxine-pyrimethamine; DNP: Dihydroartemisinin plus napthoquine plus trimethoprim; DP: Dihydroartemisinin-piperaquine; ECG: Electrocardiogram ; hERG: Human ether-a-go-go related gene; ICThe authors would like to acknowledge that Novartis Pharma AG sponsored this supplement. However, none of the authors works for, or represents in any way, Novartis Pharma AG.All authors met International Committee of Medical Journal Editors criteria for authorship.


The author conducted a review of studies that compared the efficacy, tolerability and indication for the use of clozapine in current perspectives for the treatment of resistant schizophrenia/ partial responders. In the early 1960s, German psychiatrists working with G. Stille at Wander Pharmaceuticals in Bern, Switzerland, worked to refute the concept that EPS and antipsychotic efficacy were linked.3 ClinicaHowever, enthusiasm for the drug was maintained by a small cadre of clinical investigators including G. Honigfeld at Sandoz, who observed that clozapine, was remarkably effective in treatment-resistant patients. This led to a landmark double-blind study of clozapine in a well-defined group of treatment-resistant patients whose blood cell counts were closely monitored during treatment and ulti1The mechanisms of action which account for the effectiveness of clozapine as a pharmacotherapy for the treatment of neuroleptic non-responders and neuroleptic intolerant schizophrenic subjects remain elusive. It is characterized by generally lower affinities for D2 receptors and relatively greater affinities for serotonin (5-hydroxytryptamine) 5-HT2A receptors in particular, but also for noradrenergic receptors (α1 and α2), muscarinic acetylcholine receptors, histamine and other dopamine (DA) subtype receptors.et al,[11C] SCH23390 and [11C]raclopride to investigate D1 and D2 receptor occupancy in vivo in 25 schizophrenic patients receiving atypical antipsychotic treatment with clozapine, olanzapine, quetiapine or risperidone. They concluded that among the atypical antipsychotics, clozapine appeared to have a simultaneous and equivalent occupancy of dopamine D1 and D2 receptors. Whether its effect on D1 receptors represents agonism or antagonism is not yet clear. This issue is still unresolved in the preclinical arena. The distinctive effect on D1/D2 receptors may be responsible for clozapine's unique effectiveness in patients with schizophrenia refractory to other typical and atypical antipsychotics.Tauscher et al, used PosMeta analysis of controlled trials involving patients who had treatment resistant schizophrenia–12 and aet al.,[In an open trial by Agarwal et al., drug-resMoncrieff comparedSeveral authors have questioned whether clozapine should be indicated as a first-line treatment for early psychosis.–17 The ret al., reported the interesting observation that clozapine, in comparison with fluphenazine, was similarly efficacious for new-onset patients with schizophrenia or schizoaffective disorder.[2 receptors in their brains. Typical antipsychotics may cause the upregulation because they dissociate slowly from the D2 receptor[Woerner disorder. It is frdisorder. that pat receptor and the  receptor may allo receptor Because  receptoret al.,[Pragmatic trials, supposed to provide more complete information for physician in clinical practice, support et al., in a phaet al., studied et al., Patientset al,[In another pragmatic trial Lewis et al, studied a second generation antipsychotic, Tuunainen et al,[In a Meta analysis comparing clozapine with en et al, found a et al., found that the magnitude of improvement in mean BPRS and CGI scores from baseline to end of the study was significantly greater in the clozapine group than in the risperidone group. Statistically significant differences in favor of clozapine were also seen for most of the secondary efficacy measures . The adverse event profile was similar for both treatment groups, with a lower risk of extrapyramidal symptoms in the clozapine group.In a Double-Blind Comparative Study, includinClozapine has shown consistent clinical benefit in schizophrenic patient with persistent aggressive and violent behavior.30 Whetheet al.,[McGurk et al., investiget al.,[Bilder et al., examinedet al.,[Purdon et al., examinedIt is estimated that approximately 30% of patients treated with clozapine do not respond adequately, remaining with persistent psychotic symptoms despite taking adequate treatment for sufficient period. Such patients are called “partial responders to clozapine,” “clozapine resistant,” or even “super refractory”. Despite 353745In a study conducted at psychiatry center, S.M.S. medical college, Jaipur, by Dr. R.K. Solanki and Dr. Paramjeet Singh, 100 diagnosed cases of schizophrenia on clozapine maintenance (not showing adequate response) were augmented with risperidone (2-4 mg/d). Patients showed significant improvement after 4 week on CGI, BPRS and PANSS scales.et al.,[Lieberman et al., reviewedIn international suicide prevention trial includinet al.,[Turetz et al., treated et al.,[et al.[Green et al., examined.,[et al.It is als.,[et al.Clozapine is least likely to worsen the neurological disorder so it isA number of authors have investigated the factors associated with response to clozapine but findings are contradictory. Reviewing the evidence Chung and Remington suggesteP<0.001), in 12 (60%) it lasted longer (P=0.0368) and in 17 (85%) it occurred more quickly on rechallenge (P<0.001).they concluded that after risks and benefits have been considered, rechallenge may well be justified in some patients.Apart from common side effects like sedation, dizziness, syncope, tachycardia, hypotension, ECG changes, nausea and vomiting, fatigue, weight gain, constipation, anticholinergic effects and subjective muscle weakness, several rare side effects has been described during clozapine treatment. One most important is neutropenia which may lead to fatal agranulocytosis, occurs in about 1% patient. Its mechanism is unclear but may be the result of direct toxicity or an immune response. Annan LJIn a review Wetterling suggested that Average weight gain with clozapine is 1.72 kg /month, occurring most frequently during first 6 to 12 month of treatment. There haet al.,[2. Thirty patients (36.6%) were diagnosed with diabetes during the 5-year follow-up. Weight gain, use of Valproate and total daily dose of clozapine were not significant risk factors for developing diabetes mellitus. Patients experienced significant weight gain that continued until approximately month 46 from initiation of clozapine. There was a nonsignificant increase in total serum cholesterol and a significant increase in serum triglycerides level.In a 5 year naturalistic study Henderson et al., examinedet al.,[Howes et al.,examined et al., Similarlet al.,et al.,[Akathisia occurs even occasionally with clozapine. Very raret al., reviewedet al.,et al.,[Cohen et al., conducteet al.,69 Myocaret al., There iset al.,Clozapine is known to increase seizure frequency. In a review seizure 74Clozapine specifically has two important risks of intestinal dysfunction: potentially severe ileus and sialDe novo emergence or exacerbation of obsessive-compulsive (OC) symptoms during treatment with clozapine has been described in the literature. Lykouras et al.,[s et al., revieweds et al.,Toxic delirium can occur in about 3% of patient.ClozapineData regarding the potential teratogenacity of clozapine are not unanimous. There are some reports of congenital malformations and other complications during pregnancy. But there is report of succeThe ten years of clinical experience of clozapine use in the dose range 100-400 mg/day as monotherapy or augmentation has been significantly safe and acceptable at large by patient subgroups. The side effect profile remained comparable to other SGA's, as evident from the unpublished data of the ongoing study at psychiatry center, SMS medical college, jaipur.The result of the different studies indicated that clozapine exhibit superiority over typical antipsychotics in terms of both efficacy  and safety. However the magnitude of its effect is not consistent. Efficacy data for other SGA's in the treatment of refractory schizophrenics were inconclusive.So there is growing need to consider different treatment strategies. Whether monotherapy or adjuvant therapy for non responsive or partially responsive schizophrenics.


Sir,et al for their interest in my letter published in 2003 on traditional behavioural practices and exchange of saliva among sub-Saharan African populations. The comments by M Coluzzi et al on my letter highlight a potentially important behavioural practice associated with human herpesvirus 8 (HHV-8) transmission; the use of saliva to soothe blood-sucking arthropod bites. As mentioned in my letter, I found some evidence of this practice in reviewing ethnographic material from the Human Relations Area Files (HRAF). However, neither the extent nor frequency of saliva-associated behavioural practices, including those highlighted by Coluzzi et al, has been investigated among sub-Saharan African populations. The HRAF files present only descriptive, ethnographic data. Scientifically oriented epidemio-logical studies need to be developed to better characterise the frequency and distribution of saliva-associated behavioural practices and evaluate the potential association between these practices and risk of infection with HHV-8.I thank M Coluzzi et al discuss ecological data from a previous study The ‘promoter arthropod’ hypothesis raised by


An outbreak of acute hemorrhagic conjunctivitis occurred in Delhi, India, during August and September 1996. The etiologic agent was confirmed as enterovirus type 70 by a modified centrifugation-enhanced culture method followed by immunofluorescence and neutralization tests. After nearly a decade, this virus is reemerging as a cause of acute hemorrhagic conjunctivitis in India.


Newer techniques of Laser Trabeculoplasty have revived the procedure and gained widespread acceptance by the ophthalmic community. This review was undertaken to address the evolution of different laser trabeculoplaty techniques, proposed mechanisms of action as well as review current studies of the therapeutic effects of these interventions. Laser trabeculoplasty (LT), first described in 1974 by Worthen and Whickham,Several other lasers including krypton (647.1 or 568.2 nm) diode (810 nm), DLT and continuous wave, frequency-doubled Nd: YAG 532 nm), the work by Latina and colleagues (1998)2 nm, the4Most glaucoma specialists regard LT as an adjunct to maximum tolerated medical therapy. Alternatively, it can be used as an initial glaucoma therapy. LT provides successful results in POAG, pseudo exfoliation and pigmentary glaucoma and in eyes with combined mechanism glaucoma. It does not appear to be effective in glaucoma secondary to uveitis, congenital glaucoma, juvenile open angle glaucoma, iridocorneal endothelial syndrome, iridocorneal mesodennal dysgenesis, steroid induced glaucoma, and glaucoma secondary to elevated episcleral venous pressure.SLT results in selective absorption of energy by trabecular pigmented cells sparing adjacent cells and tissues from thermal damage. It delivers a 400 um diameter treatment spot in three nanoseconds, by using short bursts at low power settings . SLT delivers energy fluence levels to the TM cells that are several thousand times lower than with ALT. One consequence of this energy reduction is that SLT seems to produce less thermal tissue damage to TM than ALT.5MLTMLT utilizes an 810-nm diode laser emitting a train of repetitive short pulses to allow the surgeon to control and spatially confine the laser-induced thermal elevation to produce the intended sub lethal photo thermal effects to elicit a thermal stress response in the TM cells. It is typically performed to deliver 60 to 65 (or 120-130) confluent 300-um diameter invisible laser applications covering the whole height of the TM over 180- (or 360-) angle. Due to the combination of (a) low absorption of the 810-nm laser wavelength by the TM and (b) low irradiance of 2W over the relatively large 300 um spot, MLT seems to interact and thermally affect all superficial and deep TM-pigmented cells without producing visible photo thermal changes, tissue blanching, or bubble formation. The treatment is invisible to the surgeon and uneventful for the patient with no pain and no dazzling laser flashes (810 nm is invisible).Many prospective and retrospective studies compare SLT with ALT indicating that a similar short- and long-term efficacy of ALT and SLT can be expected in patients with OAG with a similar and a mild risk profile.The GLT results demonstrate that ALT could be considered a primary therapy for OAG. Several other studies have also demonstrated that ALT is effective at lowering IOP.18Studies investigating the use of SLT as initial therapy using a 180-treatment report a mean reduction of 7.7 plus 3.5 mmHg (30%)19et al.et al.et al.McIlraith et al.et al.et al.Other variables such as age, sex, type of OAG and degree of trabecular meshwork pigmentation are not associated with improved outcomes in the group of participants receiving SLT.et al.et al.et al.et al.Repeat ALT effectiveness was assessed in a few studies. Feldman et al.et alet al.8Since SLT does not induce trabecular meshwork scarring, Latina et al.et al.Repeat SLT was assessed by Shah The findings are encouraging and suggest that 360-degree SLT may be repeated after an initially successful 360-degree SLT fails and with IOP reduction that suggests only slightly diminishing returns. Findings also suggest that these results may be achieved as early as six months after the first treatment.In a randomized pilot study conducted in the University of Missouri Kansas city,Along with medications, LT effectively and safely lowers elevated IOP. SLT and MLT can be performed with minimum iatrogenic damage to the TM- lowest intraoperative and postoperative complications and side-effects. This will eventually gain widespread acceptance by the ophthalmic community. Newer techniques may assume an increasingly important role of LT in our management of glaucoma.


To the Editor: We thank Su and Chee (The range of dengue-related ophthalmic complication is still being investigated, and we agree with Su and Chee that other ophthalmic manifestations may occur in patients with dengue fever. In a retrospective observational case series involving 22 eyes of 13 patients with visual impairment from dengue infection, carried out in our hospital, Chan et al. (


Environmental Health Perspectives, La Pensee et al. (2009) postulated that bisphenol A (BPA), at nanomolecular doses, confers chemo resistance in estrogen receptor (ER)-α–positive and –negative breast cancer cells. Certainly, drug resistance is well-known to be an important complication in a variety of cancer chemotherapy options. Several molecular mechanisms have been suggested to explain the onset of drug resistance. Determining the exact mechanism in a particular case is challenging both at the clinical and preclinical research levels because the genome-wide and proteomic approaches to mechanistic studies are still at a developing stage , suggesting that direct or rapid response to E2 is widespread at the mRNA levels in these genes. 2 and environmental endocrine-disrupting chemicals in the uterus of women. Involvement of these gene transcripts, which are present in breast, uterine, and ovarian tissues, in the environment–endocrine inter action suggests the possibility of a utero-ovarian feedback control of breast cancer chemo sensitivity effected by npcRNAs.Recent reports from other laboratories have tended to support a role of npcRNAs in BPA-mediated mechanisms involved in breast cancer and a possible physiochemical interaction of BPA with estrogen and non-estrogen-mediated chemosensitivity-inducing pathway elements. For example, These observations suggest that there may be a utero-ovarian feedback control mediated by ERs on the uterus that cross-talk with vasoneural pathways; the feedback control may mediate estrogen involved in chemoresponsive pathways of breast cancer. Furthermore, these pathways may be regulated by noncoding npcRNAs whose functions may be physiochemically modified by environmental toxicants such as BPA and other related chemials.in Vitro and Laboratory Animal Studies for Assessing Risks to Human Health” in Chapel Hill, North Carolina, on 28–30 November 2006 (Since the forum titled “Bisphenol A: An Expert Panel Examination of the Relevance of Ecological, ber 2006 , there h


Time-resolved structural studies of proteins have undergone several significant developments during the last decade. Recent developments using time-resolved X-ray methods, such as time-resolved Laue diffraction, low-temperature intermediate trapping, time-resolved wide-angle X-ray scattering and time-resolved X-ray absorption spectroscopy, are reviewed. Proteins undergo conformational changes during their biological function. As such, a high-resolution structure of a protein’s resting conformation provides a starting point for elucidating its reaction mechanism, but provides no direct information concerning the protein’s conformational dynamics. Several X-ray methods have been developed to elucidate those conformational changes that occur during a protein’s reaction, including time-resolved Laue diffraction and intermediate trapping studies on three-dimensional protein crystals, and time-resolved wide-angle X-ray scattering and X-ray absorption studies on proteins in the solution phase. This review emphasizes the scope and limitations of these complementary experimental approaches when seeking to understand protein conformational dynamics. These methods are illustrated using a limited set of examples including myoglobin and haemoglobin in complex with carbon monoxide, the simple light-driven proton pump bacteriorhodopsin, and the superoxide scavenger superoxide reductase. In conclusion, likely future developments of these methods at synchrotron X-ray sources and the potential impact of emerging X-ray free-electron laser facilities are speculated upon. Currently approximately 20 new structures are deposited daily within the Protein Data Bank, adding to an accumulated total of more than 60 000 structural entries. This mass of structural knowledge has provided detailed biological insight into protein structure and function relationships and uncountable details into the specific biochemistry of cellular reactions. Despite this impressive history of success, understanding the details of protein conformational changes presents a major challenge for structural biology. This is because X-ray diffraction, by far the most successful technique for elucidating new protein structures at high resolution, intrinsically relies upon the presence of well ordered arrangements of identical copies of the same protein. Flexible regions of a protein are frequently not visible within a crystal structure and proteins known to display large levels of flexibility, such as membrane protein transporters and receptors : the first is that the maximum available X-ray flux from the synchrotron insertion device may be utilized; the second is that the broad spectrum of the X-ray beam enables a large number of full X-ray diffraction reflections to be collected  without the need to rotate the crystal, which is essential when one wishes to record snapshots on sub-millisecond timescales. Nevertheless, this approach encompasses a large number of technical challenges which have been widely discussed  at a given time delay following reaction triggering within three-dimensional crystals. An alternative approach to real-time room-temperature studies is to control the kinetics of the reaction by using temperature or the chemical environment, and thereby find conditions for which a large population of a desired intermediate becomes trapped within three-dimensional crystals. Once conditions have been identified under which the population of a specific intermediate builds up, standard monochromatic X-ray diffraction data can be collected. While conceptually less elegant than time-resolved Laue diffraction, intermediate trapping methods have proven very popular in the search to understand the structural pathways of protein-mediated reactions F            obs − F            calc electron density map) is visible close to the iron atom, indicating a reactive hydrogen peroxide species within the active site, but the resolution of the electron density map does not allow an atomic model to be built without additional chemical restraints. To address this issue, off-resonance Raman spectra were recorded from both crystals and solutions of superoxide reductase treated with hydrogen peroxide   show very good agreement between the low-temperature structure -induced conformational changes in bacteriorhodopsin [see Hirai & Subramaniam 2009 and Ande al. 2009 for over4.a).Although time-resolved Laue diffraction and intermediate trapping studies have provided several detailed insights into protein-mediated reactions, these methods necessarily probe protein conformational dynamics within the crystalline state. Structural probes applicable to liquid phases would explicitly side-step this fundamental limitation of time-resolved crystallographic studies of proteins. Time-resolved wide-angle X-ray scattering is an emerging technique that addresses this shortcoming, whereby X-ray scattering data are recorded as a function of time from an ensemble of samples in a liquid environment. Thus the breakage and formation of chemical bonds within small molecules, or the rearrangement of secondary structural elements within proteins, can be visualized since the internal distances between the atoms of the sample change with time. On the other hand, all structural information accessible using wide-angle X-ray scattering is averaged over all orientations of a randomly ordered ensemble of molecules within the sample. As such, the level of structural detail that can be envisioned, even in principle, is significantly less than what can be gleaned using X-ray diffraction. The experimental set-up at the ESRF is shown in Fig. 4et al., 2001et al., 2004et al., 2005 et al., 2005et al., 2006et al., 2006et al., 2004et al., 2005et al., 2005et al., 2009et al., 2006et al., 2008et al., 2008Time-resolved wide-angle X-ray scattering was first successfully applied to probe the structural dynamics of a number of photosensitive small molecules in solution et al., 1998Time-resolved wide-angle X-ray scattering has also recently been extended to probe the conformational dynamics of proteins (Cammarata b)et al. (2008b            b            c)Difference time-resolved wide-angle X-ray scattering data recorded from solubilized haemoglobin are reproduced in Fig. 4t, following photoactivation. These data were fitted to three basis spectra and their characteristic time constants determined. Structural refinement against the intermediate and late state basis spectra  records the modulation of an X-ray absorption spectrum in the energy region from 50 eV to approximately 1000 eV above the absorption edge, whereas X-ray absorption near-edge structure (XANES) focuses upon the smaller energy region up to approximately 50 eV above the edge et al., 2005Myoglobin in complex with carbon monoxide again provided a convenient system for proof-of-principle X-ray spectroscopy studies of protein conformational dynamics about a metal centre. It is more than two decades since the first time-resolved X-ray absorption near-edge spectra of the photodissociation of carbon monoxide from the haem iron of myoglobin were described  and Japan (http://www-xfel.spring8.or.jp/) and the first American XFEL facility has recently reported lasing at 8 keV (http://lcls.slac.stanford.edu/). These fourth-generation X-ray sources promise extremely intense hard X-ray bursts of approximately 100 fs in duration, and will thereby create new opportunities for imaging of biological molecules from extremely small samples  are currently under construction in Europe (et al., 2005et al., 2008et al., 2007Within the realm of time-resolved pump–probe structural studies, the most obvious benefit of these emerging sources will be their remarkable capability to probe the structural dynamics of light-driven reactions with a temporal resolution of 100 fs. Several landmark measurements have already been reported on this timescale at linear-accelerator-based X-ray sources, including key technical innovations for synchronizing the femtosecond laser and X-ray pulses (Cavalieri et al., 2003et al., 2003et al., 2004et al., 2005et al., 2005et al., 2004et al., 2000et al., 2000While taking a very optimistic viewpoint of these emerging opportunities, it seems likely that several technical challenges will be encountered along this path. For example, when using pump–probe methods to study protein reaction dynamics in the crystalline phase, it may well prove advantageous to use the full width of the background undulator spectrum of the XFEL to perform ‘pink’ time-resolved Laue-diffraction (Bourgeois et al., 2008et al., 2009et al., 2005et al., 2006et al., 2006Time-resolved wide-angle X-ray scattering (Cammarata In closing, the last decade has witnessed several significant technical improvements to existing methods, as well as the development of novel approaches, for elucidating the structural details of protein catalyzed biochemical reactions as a function of time. Even a conservative extrapolation of these recent developments can foresee intermediate trapping and time-resolved wide-angle X-ray scattering studies becoming increasingly widely applied approaches within structural biology. We also foresee that that domain of applications of time-resolved Laue diffraction and X-ray absorption spectroscopy will grow, albeit more slowly. Finally, the imminent application of ultra-fast extremely intense XFEL-generated X-ray pulses to probe the reaction pathways of light-sensitive proteins will offer unique opportunities, delivering many surprises as it opens new structural windows upon fundamental photochemical events on the timescale at which they occur.


Indirect immunofluorescence with serum is used in the diagnosis of pemphigus. We report a case in whom blister fluid was used as the specimen for indirect immunofluorecscence. Immunofluorescence is one of the diagnostic tests done in cases of pemphigus. In 1941, Coons et al. developed the technique of immunofluorescence. In 1964, Beutner and Jordon used the indirect immunofluorescence (IIF) technique to demonstrate antibodies in the sera of pemphigus patients.[Pemphigus vulgaris is an autoimmune blistering disease characterized histologically by suprabasal blister and immunopathologically by A 30-year-old female a known case of pemphigus was biopsied for histopathology and direct immunofluorescence (DIF). Blister fluid along with serum was taken for indirect immunofluorescence (IIF). DIF showed strongly positive IgG deposits. Serial dilutions of blister fluid using phosphate buffered saline (PBS) were incubated with cryostat sections of normal skin (substrate). Subsequently sections were incubated with FITC-labeled anti-human IgG for 30 min at 37°C. Slides were washed with PBS and mounted in buffered glycerol. Fluorescent microscopy revealed moderately strong ICS deposits of IgG  and focal weak ICS deposits of C3  Figures . IIF witet al.[et al.[et al.[Immunofluorescence is nowadays commonly used in the diagnosis of vesicobullous disorders. Direct immunofluorescence requires perilesional skin, whereas serum is used for indirect immunofluorescence. Blisters are formed as a result of a breakdown of tissue integrity and fluid accumulation in a specific compartment of the skin. In pemphigus, simple binding of the antibody to the ectodomain of the antigen (desmoglein 3) triggers blister formation, perhaps by impairing the function of the molecule. Blister l.[et al. comparedl.[et al. had desc