Graphene has been studied for various applications due to its excellent properties. Graphene film fabrication from solutions of graphene oxide (GO) have attracted considerable attention because these procedures are suitable for mass production. GO, however, is an insulator, and therefore a reduction process is required to make the GO film conductive. These reduction procedures require chemical reducing agents or high temperature annealing. Herein, we report a novel direct and simple reduction procedure of GO by silicon, which is the most widely used material in the electronics industry. In this study, we also used silicon nanosheets (SiNSs) as reducing agents for GO. The reducing effect of silicon was confirmed by various characterization methods. Furthermore, the silicon wafer was also used as a reducing template to create a reduced GO (rGO) film on a silicon substrate. By this process, a pure rGO film can be formed without the impurities that normally come from chemical reducing agents. This is an easy and environmentally friendly method to prepare large scale graphene films on Si substrates. Graphene, a monolayer carbon sheet, has superior electrical and mechanical characteristics such as extremely high electron mobility and Young’s Modulus125689At this point, many researchers have studied graphene film using a variety of fabrication methods. There are two primary methods of making graphene films presently: chemical vapor deposition (CVD) and GO solution-based methods. To make a graphene film using CVD, complicated transfer steps are required for moving graphene from the catalyst substrate to the target substrate because CVD graphene can be grown on restricted substrates such as copper and nickel metal catalystset al. reported spray-coating a GO film with chemical reduction by pre-mixed hydrazineet al. suggested spin-coated a GO film followed by thermal reductionet al. presented a Meyer rod-coating process with a pre-mixed palladium chloride reducing agentet al. proposed a microliter scale solution method by meniscus-dragging deposition (MDD) with HI acid vapor reduction21For GO, many researchers have studied techniques for formation of uniform graphene films from GO solutions such as spin coating, spray coating and using chemical reactions because solution-based methods are suitable for industrial mass production with low cost. GO, however, has a naturally low conductivity, so GO has to be reduced to obtain a conductive film223242526To reduce GO, thermal and chemical reduction have been widely usedHere, we report a new technique to produce large scale graphene films on silicon substrates. In this simple method, reduction of GO and formation of graphene films occur simultaneously on a silicon wafer without any extra reducing agent or high temperature annealing. Moreover, in a comparison with the previous reducing agents, silicon is widely used materials at various applications, so this study method could be easily applied into graphene-silicon composite applications without other additive.Silicon wafers are a commonly used substrate, and graphene is also widely used on silicon substrates. Further, bare silicon can be oxidized easily, so silicon, which acts as a reducing agent in this study, can remove the oxygen groups from GO; as a result, silicon becomes silicon oxide and GO becomes rGO3940Silicon nanosheets (SiNSs) prepared by a CVD methodThe reduction effect of silicon to GO was confirmed by X-ray photoelectron spectroscopy (XPS) in −1) and a D peak (1573 cm−1). The G peak corresponds to sp2 hybridized carbon-carbon bonds, related to first-order scattering of the E2g phonon in graphene and the D peak correlates to lattice distortions, related to a breathing mode of k-point photons of A1g symmetry465152854D/IG ratio increaseD/IG ratio of GO was 0.80 and that of srGO was 1.11, which is in good agreement with the literature. The ID/IG ratio distribution is shown in Atomic structure change between GO and srGO was determined by X-ray diffraction. Graphite, GO, and srGO powder were measured. Graphite had a sharp peak at 26°, and the peak of GO, after chemical exfoliation from graphite, was shifted to 10°. These peak changes mean the interlayer distance between the carbon layer was increased, i.e., GO was successfully exfoliatedD/IG ratio of the etched wafer was 1.13 and that of the non-etched wafer was 0.79. 2D peak was observed at ~2700 cm−1, which is overtone of D peak, and a D + G peak was also observed at ~2973 cm−1, which is a combination of the D peak and G peak, as shown in Figure S6Figure S62D/ID+G was also increased after reduction process due to graphitic electronic conjugation recovery as shown in table S1Based on this evidence, a silicon wafer was also used as an efficient template to produce graphene films. The GO film, which was formed on a bare silicon wafer, can be reduced by the substrate alone. The silicon wafer, which has a native oxide layer, was etched this using 1:6 BOE and was rinsed with DI water. After the oxide etching process, the GO solution was spray coated onto the etched wafer surface immediately and was heated. As a result, the as-formed GO film on the silicon substrate was reduced to a srGO film with no further treatment or reducing agent. Film formation and reduction was done simultaneously. The resulting graphene film was investigated using Raman spectroscopy. D/IG ratio changes versus heating temperature were measured from room temperature (rt) to 140 °C. The ID/IG ratio increased with an increase in heating temperature up to 100 °C. After 100 °C, the increase in ID/IG ratio was saturated, therefore, the optimized heating temperature was 100 °C.ID/IG ratio mapping image of srGO.A graphene film on a 4-inch wafer was formed by our spray coating method. To make a uniform large area GO film, some kind of coating method such as spray coating or spin coating is required to prevent the coffee-ring effect, which occurs at the boundary of the film61Sheet resistance was measured to analyze electrical properties of our graphene film. The sheet resistance of the srGO film  was considerably lower than that of the GO film . The srGO sheet resistance is similar to other literature values even though no chemical reducing agent was used. NaOH was also used as an etchant for the silicon wafer to confirm that the reduction effect was not a result of the BOE. All other procedures were kept the same as previously described. Although the etchant was changed, there was no significant change in the Raman spectrum, as shown in Though the reduction mechanism of GO has been actively studied, but it is not sufficient2964656568692 peak (103.65 eV) was found after GO reduction process. As per our hypothesis, we are assuming that the Si-H dangling bonds could be generated on the surface of silicon, which are playing these key roles to reduced oxygen functional groups of GO as shown in 7374757778figure S8808182Based on the current experiment and characterization results, it is confirmed that GO is reduced by silicon. As the results of our experiments, silicon can absorb oxygen group from GO. That oxygen group could be formed as silicon dioxide. According to XPS data silicon dioxide was formed on silicon wafer after GO reduction as shown in Si 2p spectrums 72. The SThis study provides a demonstration of an innovative reduction method for GO. Silicon, a common material widely used throughout industry, can be used to reduce GO. CVD grown 2-dimenional SiNSs can combine with GO sheets due to similar morphology, and this advantage can enhance the reduction effect of silicon. Silicon wafers were also used as reducing templates. By this method, the reduction of GO and film formation occurred simultaneously without any additional chemicals. The graphene film, which is reduced by silicon, has no impurities such as hydrazine or hydrogen iodide, and a high quality graphene film can therefore be achieved. The reproducibility of our method was confirmed statistically. A large scale graphene film can be made by this simple method. The graphene film on a silicon wafer can be transferred to a transparent substrate by a common graphene transfer method. This suggested method is simple, easy, and eco-friendly for graphene film formation and can be used to potentially further commercialization of graphene.Silicon wafers were obtained from DASOM RMS (Korea). BOE was purchased from SAMJUN Chemicals (Korea). Graphite was obtained from Bay Carbon (USA). All other chemicals for synthesis GO were obtained from Sigma Aldrich, Korea. All chemicals were used without further purification.The whole experiment was divided into two categories. The first experiment was conducted as solution process with silicon nanosheets (SiNSs) and GO. After reduction process based on solution, GO was dried as powder form. The second experiment was performed as film process with silicon wafer and GO solutions. The result of this experiment was reduced GO film on silicon wafer.The graphene oxide solution was prepared by a modified Hummer’s method.4 as the Si precursor and H2 as the carrier gas in high gas flux environment. The substrates were placed at the center of a quartz tube reactor. The reaction temperature was maintained at 1050 °C for 30 min under a H2 (99.9999%) and an Ar (99.9999%) atmosphere. Silicon tetra-chloride  was introduced into the reactor using a bubbling system. After 30 min, the reactor was cooled to room temperature under an Ar atmosphere.SiNSs were synthesized on Si substrates through chemical vapor deposition using SiClSiNS wafers were etched using a 1:6 buffered oxide etch (BOE) for 0.5 seconds to remove the native oxide layer and were dispersed in DI water. A diluted GO solution 25 ml (0.1 mg/ml) and 5 ml of a SiNSs solution (0.1 mg/mg) were mixed well and heated up to 80 °C for 2 hours with mild stirring. Then, the mixed solution was put into a 2 M NaOH solution at 40 °C for 1 day for to fully etch the SiNSs. The solution was washed with DI water by centrifuging 5 times followed by drying.2. After drying, the wafer was placed on a spray coating device and heated up to 80 °C. During heating, the GO solution (1 mg/ml) was sprinkled on the wafer surface. The heating was continued up to 2 hours after coating.The silicon wafer was put into 1:6 BOE at room temperature for 240 or 320 seconds. Then, the wafer was rinsed with DI water 3 times quickly and dried with flowing N2 pressure of 29 psi. During the spraying process, the substrate is heated to 100 °C to obtain uniformly deposited graphene oxide layers.Graphene oxide layers were obtained by facile spray casting onto the functionalized substrate using a double-action airbrush  at a distance of 30 cm with a NChanges in chemical bonds after reduction were investigated by X-ray photoelectron spectroscopy . Adjustments of atomic and molecular structure were probed with X-ray diffraction . The reduction of GO by silicon was analyzed using a Raman Spectrometer . The absorbance differences between GO and silicon reduced graphene oxide (srGO) and the transmittance of srGO films were measured using a UV/VIS spectrophotometer . Electrical properties of srGO films were examined by using 4 probe measurement . Surface roughness and morphology of the srGO film were investigated using an atomic force microscope  and a field-emission scanning electron microscope .How to cite this article: Chan Lee, S. et al. Efficient Direct Reduction of Graphene Oxide by Silicon Substrate. Sci. Rep. 5, 12306; doi: 10.1038/srep12306 (2015).


Neophocaena asiaeorientalis asiaeorientalis) is critical for its conservation. The diel biosonar behavior of the porpoise in the port areas of the Yangtze River was examined along with simultaneous observations of fish density and boat traffic. Biosonar pulses from the porpoises were detected for 1233 min (5.77%) over a 21,380 min duration of effective observations. In total, 190 (5.63%) buzzes (an indication of prey capture attempts) were recorded among the 3372 identified click trains. Of the 168 echolocation encounters (bouts of click trains less than eight min apart), 150 (89.3%) involved single animals, indicating that solitary porpoises were frequently present and feeding in the port areas. Significant diel patterns were evident involving the biosonar behavior of the porpoises (including click trains and buzzes), fish density and boat traffic. The frequencies of the click trains and buzzes were significantly lower during the day than in the evening and at night, which suggests that porpoises in this region are primarily engaged in crepuscular and nocturnal foraging. The lack of a significant diel pattern in the echolocation encounters indicates the importance of the port in porpoise conservation. A forced feeding schedule may be associated with the lack of a significant correlation between porpoise acoustics and boat traffic. Overall, prey availability appears to be the primary factor that attracts porpoises. Additionally, porpoises tend to migrate or remain downstream in the morning and migrate or remain upstream in the evening, most likely to follow their prey. The findings of this study can be used to improve the conservation of the Yangtze finless porpoise.Information on the habitat use of the critically endangered Yangtze finless porpoise ( Neophocaena asiaeorientalis asiaeorientalis) is listed as Critically Endangered on the International Union for the Conservation of Nature Red List of Threatened Species Lipotes vexillifer), which was likely driven to extinction by human activity A genetically and morphologically distinct cetacean species, the Yangtze finless porpoise  rely primarily on their sophisticated acoustic sensing biosonar for communication, navigation and foraging Passive acoustic monitoring (PAM) is a rapidly evolving discipline that facilitates the investigation of the behavior of many aquatic mammals over a range of spatial and temporal scales  Port see  from 11∶th to the 23rd of June instead of being continuously monitored. The cruise boat was anchored along the wharf boat after each day’s expedition to a random site  was used to log data while anchored at the port. The transducers of this echosounder can transmit 50 kHz as a narrow beam. However, to minimize the possible interference of the transmitted sound with the A-tags, the fish data were randomly sampled as echograms by photographing the monitoring screen of the Fishfinder at different diel phases (see below) from the 19dom site , which rth to the 23rd of June. The boat’s safety lights made photography possible during the night. The camera was fixed at an advantageous location on the second deck of the cruise vessel  shooting at 1 min intervals from the 15e vessel  and orieThe research was carried out under a research permit issued to the Institute of Hydrobiology of the Chinese Academy of Sciences by the Ministry of Agriculture of China. The approval number from the Fishery Bureau of the General Office of the Ministry of Agriculture was 2012–114.Two types of A-tags (including towed and fixed types) were used in this study. A brief description of these autonomous acoustic recorders is provided here, and a detailed description is provided by Akamatsu et al. (2005) http://aa.usno.navy.mil). The categories of night1 and night2 were combined to make up the rest of the diel phases of the night and reserved for further analysis.The 24 h of the day were divided into five diel phases  following the method proposed by Todd et al. (2009) During the off-line signal processing of the A-tag data, two-step procedures were applied using Igor Pro 5.01 software  to filter out the non-porpoise clicks and extract the porpoise sonar. A custom-made, multi-parameter filter program was used during the pilot step to classify the recorded pulses as originating from porpoises, boat traffic or other sources . In contrast, a positive or a negative value was recorded if the source was upstream or downstream (with the A-tag set as the reference point), respectively. The gradual changes in the TD of the sonar detected within each echolocation encounter indicated the underwater movement of the porpoise (acoustic trajectory). A trajectory lasting more than 20 s with at least three click trains was categorized into one of the following classes: upstream stay , fish density in the port area and boat traffic  for each day were first assigned to different diel phases , and the data from each diel phase were further grouped into 10 min bins to account for the difference in sampling effort across diel phases. Echolocation encounter and trajectory were also assigned to different diel phases. During counting, echolocation encounters and trajectories that crossed boundaries between diel phases were assigned to the dominant phase that contained the greater portion. For the analysis of diel pattern, echolocation encounters and trajectories that spanned multiple days were segmented, and each segment was assigned an appropriate diel phase.p<0.05). The Kruskal–Wallis nonparametric test th to the 23rd of June) were screened out and matched in the form of 10 min bins. As for the correlation between porpoise acoustics and fish density, the relevant data from the 19th to the 23rd of June were grouped and calculated for the 5 diel phase pattern. The Mann–Whitney U-test p<0.05.The Kolmogorov–Smirnov test and Levene’s test for equality of variances were used to analyze the distribution of the data and the homogeneity of the variance, respectively. Nonparametric statistical analyses Porpoise sonar signals (both clicks and buzzes) were monitored every day  for a toOf the 18 multi-trajectory echolocation encounters, six involved two unambiguous multi-trajectories, resulting in a combined total of 174 measured trajectories . One hunp<0.01). Specifically, the duration of the milling and up trajectory was significantly longer than the straight down trajectory , and the upstream stay was significantly shorter than the milling and up   .p<0.01; 4 phases: Kruskal-Wallis χ2 = 19.62, df = 3, p<0.01). In particular, the frequency of click trains was significantly lower during the day (mean ± SE: 1.44±0.19 per 10 min) than during the evening (mean ± SE: 2.49±0.70 per 10 min), night2 (mean ± SE: 2.03±0.36 per 10 min) and night (mean ± SE: 1.69±0.21 per 10 min)  . Buzz frequency was significantly lower during the day (mean ± SE: 0.06±0.02 per 10 min) than in the evening (mean ± SE: 0.23±0.10 per 10 min), night2 (mean ± SE: 0.17±0.06 per 10 min) and night (mean ± SE: 0.12±0.03 per 10 min)  (Significant differences in the frequency of porpoise click trains (per 10 min) were observed among the diel patterns  . The fre p<0.05) .p = 0.33; 4 phases: Kruskal-Wallis χ2 = 5.39, df = 3, p = 0.15)    or the d 0.0.53) .The stay trajectory types peaked in the morning and evening. The upstream stay, with a frequency of 0.24 counts per h , accountp = 0.02). Specifically, fish densities in the morning (mean ± SE: 0.85±0.16 fish per m3) and during the day (mean ± SE: 1.79±0.33) were significantly lower than fish densities in the evening (mean ± SE: 4.56±1.30), at night2 (mean ± SE: 4.27±0.18) and at night (mean ± SE: 4.58±0.55)   .p = 0.04; 4 phases: Kruskal-Wallis χ2 = 10.28, df = 3, p = 0.02) (p<0.05)  (p>0.05)  (p<0.05); no significant differences in frequency were observed among these other diel phases or between the day and the evening    and 9A. (p<0.05) . The fre p<0.01)  and 9B. (p>0.05) . Signifi p<0.01)  and 9C.  p>0.05) .th to the 22nd plus night1 and the morning of the 23rd of June). Significant correlations were observed between fish density and the frequency of porpoise click trains  and between fish density and the frequency of porpoise buzzes  (Twenty-two pairs of matched porpoise acoustic and fish density data were derived (including the 5 diel phases from the 19 n = 22) .p = 0.684, n = 682, Spearman’s rho = −0.045, p = 0.241, n = 682 and Spearman’s rho = 0.012, p = 0.755, n = 682, respectively) . There was also no significant association between the number of porpoise buzzes and the passage of boats upriver , downriver  or both  .A combined total of 682 simultaneously matched pairs of dolphin acoustic and boat traffic data (10 min bins) were obtained. The number of click trains was not significantly correlated with the passage of boats upriver, downriver or both  . Furthern = 682) . No signPassive acoustic monitoring showed that Yangtze finless porpoises occupy port areas  more rouDistinct diel patterns in porpoise acoustics were observed, with significantly more click trains and buzzes detected in the evening and at night than during the day. These findings suggest that porpoises are primarily engaged in crepuscular and nocturnal foraging in the port area. Considering that no significant diel patterns were observed in the frequency and duration of porpoise echolocation encounters, the diel pattern of porpoise acoustic behavior appears to be due to porpoises producing more click trains and buzzes at night Phocoenoides dalli) are thought to forage mainly at night and/or at dawn, as greater amounts of stomach content were observed in animals captured in the morning Stenella attenuata) have been shown to be primarily nocturnal and crepuscular feeders Phocoena phocoena) Ziphius cavirostris), Blainville’s beaked whales (Mesoplodon densirostris) Platanista gangetica) Grampus griseus) Lagenorhynchus albirostris) have been shown to forage only at night when monitored with bio-logging tags and passive acoustic methods Mounting evidence indicates that some marine mammals tend to feed at night. For example, Dall’s porpoises  S. attenuata) Stenella longirostris) Lagenorhynchus obscures) Stenella coeruleoalba) P. phocoena) G. griseus) Cephalorhynchus heavisidii) P. gangetica) Cephalorhynchus hectori) Diel behavioral patterns have been correlated with spatio-temporal variations in food availability in common dolphins  peak in the evening, suggesting that this is the preferred time for porpoise feeding. This finding is consistent with the plateau in fish density ,7 and 9.The number of echolocation encounters is not necessarily equal to the number of individual porpoises. Interference resulting from the movement of porpoises in and out of the detection range of the instrument and periods of silence during which the porpoises were in the vicinity of the A-tags may have resulted in duplicate acoustic detections of the same animal. Additionally, non-vocal animals, even those within range of the equipment, were not detected Porpoise sonar was frequently detected in the port areas. Recordings tended to consist of feeding buzzes, and a majority of echolocation encounters involved single animals, indicating that solitary porpoises were frequently present and feeding in the port area. The distinct diel patterns of porpoise biosonar behavior showed that the click trains and buzzes occurred much more frequently in the evening and at night than during the day, which suggests that porpoises in the region were primarily engaged in crepuscular and nocturnal foraging. Porpoise sonar was significantly correlated with fish density but not with boat traffic; it is possible that overfishing in the Yangtze River has forced the porpoises to tolerate the disturbances caused by boats in the port areas to capture prey. Overall, prey availability appears to be the primary factor attracting porpoises, and the evening, when porpoise acoustics peak, seems to represent the preferred period for porpoise feeding. The plateau in fish density and the decrease in boat traffic in the evening further support this preference. Additionally, porpoises tend to stay or migrate downstream in the morning and stay or migrate upstream in the evening; it is possible that they are following prey.No studies have examined possible seasonal , yearly or site-specific variations in patterns of porpoise activity. Longer-term and wider-ranging monitoring at more locations is needed to identify and mitigate potential threats to this species.


Cilia are microtubule-based organelles protruding from almost all mammalian cells which, when dysfunctional, result in genetic disorders called “ciliopathies”. High-throughput studies have revealed that cilia are composed of thousands of proteins. However, despite many efforts, much remains to be determined regarding the biological functions of this increasingly important complex organelle.We have derived an online tool, from a systematic network-based approach to dissect the cilia/centrosome complex interactome (CCCI). The tool integrates all current available data into a model which provides an “interaction” perspective on ciliary function. We generated a network of interactions between human proteins organized into functionally relevant “communities”, which can be defined as groups of genes that are both highly inter-connected and strongly co-expressed. We then combined sequence and co-expression data in order to identify the transcription factors responsible for regulating genes within their respective communities. Our analyses have discovered communities significantly specialized for delegating specific biological functions such as mRNA processing, protein translation, folding and degradation processes that had never been associated with ciliary proteins until now.CCCI will allow us to clarify the roles of previously unknown ciliary functions, elucidate the molecular mechanisms underlying ciliary-associated phenotypes, and apply our knowledge of the functional roles of relatively uncharacterized molecular entities to disease phenotypes and new clinical applications.The online version of this article (doi:10.1186/1471-2164-15-658) contains supplementary material, which is available to authorized users. Cilia are specialized evolutionarily conserved organelles protruding from the cell surface of most mammalian cells. The cilium consists of a basal body located under the cell surface, from which the organelle is initially assembled, a transition zone that is important for docking of proteins, and the axoneme. In non-mitotic quiescent cells the mother centriole, a centrosomal component, migrates and docks at the apical cell surface, forming the basal body as a result. The ciliary axoneme, which is nucleated and organized by the basal body, is a cytoskeletal structure, formed by a cylinder of nine doublets of microtubules and in some cases, a central pair of microtubules contained within an extension of the plasma membrane .Cilia can be broadly categorized into two subgroups. Motile cilia, protrude from the surface of cells such as tracheal cells, and promote liquid mobility along the surface. On the other hand, primary cilia, appear as individual non-motile sensory organelles, which, for example, can be found in epithelial cells of kidney tubules and neurons. The ciliary membrane contains ion channels, receptors and other signaling proteins that control axoneme bending for motility and/or sense chemical or mechanical stimuli to transduce internal signals . Primarysitus inversus, obesity, ciliary dyskinesia, mental retardation, CNS malformations and cysts in the kidney, liver and pancreas. Some examples of ciliopathies are Bardet-Biedl syndromes (BBS), Oral-facial-digital type 1 (OFD1) syndrome, polycystic kidney diseases (PKD), Joubert syndrome and related disorders, nephronophthisis and Meckel Grouber syndrome [Mutations in 103 proteins that traffic to the basal body and axoneme of cilia have been causally related to human diseases called “ciliopathies”, disorders which present overlapping phenotypes such as retinal degeneration, skeletal defects, syndrome .C. elegans, C. intestinalis, C. reinhardtii, D. melanogaster, P. tetraurelia, M. musculus and H. sapiens have revealed that cilia are composed of thousands of proteins potentially involved in ciliary function and biogenesis [in silico [http://centrosome.cnb.csic.es) the Ciliome Database [http://www.sfu.ca/~leroux/ciliome_home.htm), Ciliaproteome [http://v3.ciliaproteome.org/cgi-bin/index.php), and Cildb [http://cildb.cgm.cnrs-gif.fr/). To date, Cildb is the most comprehensive compilation as it incorporates the largest collection of experiments, including some of those from other databases, in a unified and consistent framework [Model organisms have been extensively used to investigate ciliary functions. High-throughput studies conducted in ogenesis –12. Althn silico , and thon silico  . By analysing the network we detected functionally relevant “communities”, consisting of groups of genes that are both highly inter-connected and strongly co-expressed. Intriguingly, our analysis unveiled communities significantly associated with specific biological functions that had never been associated with ciliary proteins until now.In order to resolve this disparity, the list of interactors for each specific gene may now be visualized in CentrosomeDB . Unfortuhttp://cildb.cgm.cnrs-gif.fr/), a database dedicated to proteins involved with centrioles, centrosomes, basal bodies, cilia and flagella in eukaryotes which collects the results from 32 high-throughput experiments carried out in 10 different species [We collected 3,502 human ciliary genes from Cildb ( species . Since h species ) and, afhttp://string-db.org), a resource that acts as a “one-stop shop” for all information regarding known and predicted functional links between proteins. STRING quantitatively integrates data from various sources, such as genomic context, high-throughput experiments, co-expression and previous knowledge [We obtained interactions among genes from the “Search Tool for the Retrieval of Interacting Genes” (STRING) database  [http://ccci.tigem.it).Among ciliopathy genes, yndrome) , shows thttp://www.syscilia.org). SYSCILIA Gold Standard contains genes considered to be ciliary only if evidence was published for ciliary localization , function in ciliogenesis (including cilium-specific transcription) and involvement in ciliopathies [We then compared the complete gene set  with the SYSCILIA Gold Standard (SCGSv1), a list of 303 known confirmed ciliary components curated by the multinational SYSCILIA consortium (opathies . We founhttp://ccci.tigem.it) containing, on average, 9 genes each . Thirty-nine communities contained at least 5 genes. Figure The centrosome, basal body and cilium are able to regulate their complex behaviors thanks to a set of proteins that perform different but coordinated functions. We questioned whether or not functional modules could be found within the CCCI, as they may reveal how the network is organized. We searched the network for modules, which are defined as “communities”, i.e. group of genes densely interconnected with each other and connected to few genes outside the group, via a clustering algorithm . We idenhttp://ccci.tigem.it). This percentage increases to up to 74% when only communities composed by at least 5 genes are considered. Twelve communities are composed of at least 20 genes, and each of them is enriched for a specific biological process (Table We found with Gene Ontology (GO) analysis that 45 (50%) of these communities were equipped to manage specific biological functions , respectively. All of these results are consistent with previous findings and represent internal controls that support the validity of our approach .Interestingly, four communities  are enriched for processes including mRNA processing, protein synthesis, protein folding and degradation which control the protein expression levels in eukaryotes. Members of community 7 mainly encode mRNA splicing factors, RNA binding proteins, small nuclear ribonucleoproteins (snRNPs), and RNA helicase and protein translation initiation factors; genes encoding structural constituents of ribosomes, translation elongation and termination factors, are components of the community 3. Proteins that promote correct folding of the other proteins, like chaperonin CCT complex subunits and proteins that are involved in ubiquitin-dependent protein catabolic processes, such as ubiquitin, ubiquitin-conjugating enzymes and different 26S proteasome subunits are the main components of communities 11 and 1, respectively.C. elegans [de novo protein synthesis in near proximity to centrosomes via phosphorylation [de novo protein synthesis and centrosome formation, supporting the centrosome’s dependence on components of translational machinery [Functional links between primary cilia and processes that cooperate to control the protein expression levels had never been presented before. Recently, mutations in DDX59, which encodes a member of the DEAD-box-containing the RNA helicase family of proteins, have been associated with Orofaciodigital syndrome , a clini elegans . Interes elegans . These d elegans –36, and achinery .Proteins that are involved in ubiquitin-dependent protein catabolic processes, such as ubiquitin, ubiquitin-conjugating enzymes and different 26S proteasome subunits are the main components of community 1. This non-lysosomal degradation pathway plays a crucial role in biological processes. It not only degrades misfolded and damaged proteins, but also regulates cell-signaling pathways involved in proliferation, adaptation to stress, and regulation of cell size and cell death. Indeed, it has been already shown that, although the ubiquitin proteasome system localizes and can act throughout the cytosol, the catalytic subunit 20S is also localized at the mammalian centrosome  MoreoverThe “proteasome community” is enriched in ciliary components belonging to the SYSCILIA Gold Standard SCGSv1)  and diverse groups of enzymes belonging to the cytochrome P450 and glutathione-S-transferases superfamilies (community 6 and 8).Gene products involved in nucleoside triphosphate GTP/UTP/CTP biosynthetic processes and enzymes mediating the acidification of intracellular organelles, such as subunits of vacuolar ATPase (V-ATPase), are members of two of community 10 and 12, respectively. As far we know these biological processes have never been associated with cilia physiology before and further experiments are needed in order to confirm the role of cilia in these processes.Genes belonging to different communities differ in the frequency in which they interact with each other and the communities with which they interact. Therefore communities vary in the degree to which they are interconnected and associated with one another. In particular, we expect communities with related genes, e.g. involved in related biological process, to be more likely to be interconnected. We explicitly investigated these types of interactions in order to better elucidate community relationships and obtain a general overview of the processes in which the CCCI is involved.We calculated the Interaction Strength (IS) between two communities, which is defined as the number of connections between genes belonging to two different communities, divided by the expected number of connections. If the IS is different from 0, then a connection exists between genes belonging to two different communities. Similarly to the gene-wise network, also the community-wise network can also be mapped out by assigning each node to a community. There is an edge connecting two communities only if the IS is different from 0. We obtained a community-wise network composed of 90 communities represented as nodes and 237 interactions. Using the same approach initially used to find communities of genes, we then grouped the communities with at least 5 genes into sets of highly interconnected communities that we call “super-communities” (SCs), i.e. communities of communities. We obtained 2 SCs  to be abundant in ciliary genes  and 7 communities (18%) in centrosomal genes   < 0.05)  complex, interact with pericentrin to regulate centrosome integrity . These rWe finally explored whether communities were enriched in specific transcription factors (TFs) targets. In order to get the most accurate annotation, we combined sequence and expression data. In particular, to keep as many meaningful gene-gene connections as possible, we considered a gene to be a target of a TF if (i) there was a conserved binding site within the gene (plus a flanking region 10Kb upstream and 5Kb downstream) , 49 and More specifically, we collected a list of TFs and relative targets with at least one binding site within the genomic region of a ciliary gene from the UCSC Genome Browser. We further refined this list by only retaining targets co-expressed with the TF located in the CCCI. Figure We used this list to explore target density for each community. We found 11 communities to be significantly enriched (FDR < 0.05 and at least 5 targets within the community), in 30 out of the 102 tested different TFs. Additional file As expected, we found that some communities contained members of Regulatory factor X (RFX) transcription factors, which are known to play a conserved role in ciliogenesis throughout evolution , 50–52 aWe collected the information from the primary literature and classified the functions and biological roles of the transcription factors that we had identified. Most (54%) of the transcription factors, such as COUP-TF1, AP2REP, GCNF, PAX4, HMX1, HNF1A, LHX3, OCT1, RFX, NFYA, YY1, NKX3, MZF1, GATA1, GATA2 and RUNX1, are involved in developmental processes, and 10 of the 11 communities  are enriched in these developmental TFs. Interestingly, 4 of these, MZF1, GATA1, GATA2 and AML1, are regulators of transcriptional events during hematopoietic development.OFD1, BBS1 and BBS4 both in vivo and in vitro [Communities 2, 4 and 5 host TFs, such as NFAT, AREB6, NF-κB and STAT1, which play a central role in inducible gene transcription during immune response. NF-κB is a TF that is crucial in promoting inflammation through its ability to induce transcription of pro-inflammatory genes . Interesin vitro . Moreovein vitro .Communities 4, 5, 11, 15 and 26 include transcription factors involved in cell cycle control such as CREB1, c-MYC-MAX complex, ELK1, E2F1, ER-alpha, FREAC7, CREBP1, RREB1 and MAZR. They represent about a quarter (30%) of the TFs identified.Communities 4 and 5 also include MYOD, a nuclear protein belonging to the myogenic factors subfamily that regulates muscle cell differentiation and muscle regeneration.http://ccci.tigem.it). The CCCI can be searched by gene of interest and the tool interactively shows information on that gene (e.g. the community to which it belongs to) as well as its interacting genes. It is possible to explore the different communities and their properties.Our resource is publicly available as an online tool  which allows the user to navigate and explore the interactome.We integrated information from different sources to provide a comprehensive view on the genes in the cilia/centrosome complex and the interactions among them. By using a network-based approach, we identified communities of genes more closely interacting and showed that most of these communities are enriched in proteins that carry out specific biological processes. Strikingly, we identified several communities related to mRNA processing, protein synthesis, protein folding and degradation. We also showed that some of the communities contain genes that had been, until now, known to be preeminently expressed in the nucleus. Finally, we identified TF targets in some communities, but the roles of these targets and how they contribute to ciliogenesis need to be elucidated. All the results are freely available through an online tool  were extracted from STRING 9.0 . We starribed in  but discThe final interactome consists of 11,608 interactions between 1,695 ciliary genes plus 1,845 singletons (ciliary genes with no interactions).We investigated the main categories of genes present in the CCCI by using GO . GO termNetwork analysis  and visualization were performed using Cytoscape ver. 2.8.2 . CommuniThe IS between communities was calculated using the statistical framework R by using the approach defined in .http://www.R-project.org).Throughout the study a p-value of 0.05 was considered to be the statistically significant value after false discovery rate correction. All statistical analyses were performed using R ver. 2.12.1 , and 7 were designed to investigate basal body/centriole/MTOC and centrosome proteome . For each gene we counted: (A) the number of experiments in the “ciliary group” in which the gene was present and (B) the number of experiments in the “centrosomal group” in which the gene was present. If (A) was larger than (B) plus one (A > B + 1), the gene localization was set to “ciliary”. Conversely, if (B) was larger than (A) plus one (B > A + 1), the gene localization was set to “centrosomal”. If the difference between (A) and (B) was either 0 or 1, the gene was annotated as “unknown”.http://www.uniprot.org) [The list of nuclear genes was obtained from UniProtKB release 2012_07 (available at rot.org) . We extrThe presence of ciliary, centrosomal and nuclear genes in the communities was calculated in R, using the “fisher.test” and “p.adjust” functions, whilst considering CCCI genes as the background.For the TF analysis we used both sequence (binding motif) and expression (co-expression) data. We obtained the sequence information from the DAVID Bioinformatics Resources 6.7 “UCSC TFBS” table, which is based on the “tfbsConsSites” track of the UCSC Genome Browser. In this table, a gene is considered to be target of a TF if it has a binding site for the TF conserved in the human/mouse/rat alignment (i) within 10Kbp-region of 5′ (upstream) end, (ii) within 3Kbp-region of 3′ (down stream) end, or (iii) with in exon/intron region. Any TF without at least one binding site within the genomic region of a ciliary gene (i.e. present in the CCCI) was removed, resulting in a list of 140 TFs.This list was further filtered by only considering genes/targets that were also significantly co-expressed with the TF according to . To thisThe presence of TF targets in the communities was then assessed using this list, and corrected p-values were calculated in R using the “fisher.test” and “p.adjust” functions, considering CCCI genes as background.Table S2 includes information about the communities of CCCI. (XLSX 284 KB)Additional file 1: Table S1: Includes information about all the genes included in the analysis;


The RNA/DNA-binding protein, TDP-43, is the key component of ubiquitinated inclusions characteristic of amyotrophic lateral sclerosis (ALS) and the majority of frontotemporal lobar degeneration (FTLD-TDP) referred to collectively as TDP-43 proteinopathies. To further elucidate mechanisms of pathological TDP-43 processing and identify TDP-43 epitopes that could be useful as potential biomarkers of TDP-43 proteinopathies, we developed a panel of novel monoclonal antibodies (MAbs) directed at regions extending across the length of TDP-43. Here, we confirm previous observations that there is no or minimal accumulation of TDP-43 N-terminal domains in neocortical inclusions in human TDP-43 proteinopathy tissues and we identify a subset of these MAbs that are specific for human versus mouse TDP-43. Notably, one of these MAbs recognized an epitope that preferentially detected pathological TDP-43 inclusions with negligible reactivity for normal nuclear TDP-43 resembling anti-phospho-TDP-43 specific antibodies that only bind pathological TDP-43. Hence, we infer that this new MAb recognizes a phosphorylation independent but disease-specific pathologic conformation in abnormal TDP-43. These data suggest that the novel MAbs reported here will be useful for patient-oriented research as well as for studies of animal and cell-based models of TDP-43 proteinopathies including ALS and FTLD-TDP. Since the landmark discovery of the RNA/DNA binding protein, TDP-43, as the main constituent of ubiquitin-positive, tau/synuclein-negative inclusions in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) referred to as FTLD-TDP , there hE. coli cells using the pCOLD vector system . TDP-43 proteins were extracted from E. coli 16 h after IPTG induction, and the vast majority of the rTDP-43 proteins were present in inclusion bodies. The bacterial pellet was sequentially extracted with PBS and 1% Triton X-100 to remove soluble bacterial proteins. For immunization, the Triton X-100 insoluble rTDP-43 was solublized with 1% Sarkosyl buffer or 4 M urea. Sarkosyl was removed by exhaustive dialysis before use. For epitope mapping, the Triton X-100 insoluble rTDP-43 was solublized directly in Laemmli sample buffer, and immunoblot (IB) analysis was done as described previously [Human recombinant TDP-43 (rTDP-43) proteins including full length TDP-43 (FL-rTDP-43), i.e. amino acids (aa) 1–414, N-terminally truncated TDP-43 (Nt-rTDP-43), aa 1–261, and C-terminally truncated TDP-43 (Ct-rTDP-43), aa 182–414 were expressed in BL21 (DE3) eviously ,14.Murine MAbs were raised against human rTDP-43 proteins using similar methods described previously -18. MiceThe TDP-43 domains harboring the epitopes recognized by these new MAbs were first estimated using FL-rTDP-43, Nt-rTDP-43 and Ct-rTDP-43 in IB studies similar to our characterization of MAbs we previously generated to tau and alpha-synuclein ,19. AfteThe 384-well format sandwich ELISA method used to evaluate the MAbs was similar to those described previously for a 96-well format, except the sample volumes used were reduced to 30 μL/well . BrieflyHuman CNS tissues from the superior temporal cortex (FTLD-TDP subtypes A-D) and spinal cord (ALS) (one patient each) were examined to evaluate the reactivity of TDP-43 MAbs in pathological aggregations and neuronal nuclei . Slides were coverslipped with CytoSeal 60 mounting medium (Thermo Scientific).IHC of mouse CNS tissues was performed in the same manner described above, and in addition, we used a mouse on mouse (MOM) immunodetection kit (Vector Laboratories) to minimize endogenous mouse IgG background. For all sections, labeling was detected using species-specific biotinylated secondary antibodies incubated for 1 h and the Vectastain ABC kit followed by the ImmPACTThe degree of non-pathological nuclear TDP-43 and pathological TDP-43 deposits in human lower motor neurons (LMN), white-matter glial cell inclusions (GCI), neuronal cytoplasmic inclusions (NCI), dystrophic neurites (DN) and neuronal nuclear inclusions (NII) were reviewed by a trained examiner (DJI) and rated on a semi-quantitative scale  for each MAb. Digital images of IHC results were obtained using an Olympus BX 51 microscope equipped with a bright-field and fluorescence light source with a DP-71 digital camera  and DP manager software (Olympus).To gain further insights into the molecular and neuropathological roles of TDP-43 in neurodegeneration, we generated new MAbs for future studies of TDP-proteinopathies. Approximately 4500 potential MAb clones from three hybridoma fusion procedures were screened by FL-rTDP-43 indirect ELISA of which 168 clones were positive on this initial screen and all were evaluated further in IB and IHC studies. Only 45 clones continued to show immunoreactivity toward TDP-43 in the secondary screens and all of these MAbs were studied further to determine the epitopes and their antibody isotype as described ,19. UsinTo determine whether or not the selected 18 MAbs detected both human and mouse TDP-43, IHC was performed using normal human and non-transgenic mouse brain sections. All 18 MAbs detected nuclear TDP-43 in sections of human hippocampus Figure a,c, but As of now, there is no reliable method to quantify TDP-43 proteins in biofluids such as plasma or cerebrospinal fluid (CSF) samples which would be useful as a biomarker assay. In order to determine if the MAbs generated in this study might be useful for developing an assay to detect TDP-43 in CSF or plasma, we evaluated them in a sandwich ELISA using previously developed Nt- or Ct-TDP-43 pAbs  as captuTDP-43 MAbs were screened for reactivity in human CNS tissues using IHC in the spinal cord of ALS and superior temporal cortex of morphological subtypes of TDP-43 (Type A-D) . Semi-quThe reactivity to neocortical TDP-43 pathology by MAbs specific for C-terminal TDP-43 epitopes was accompanied by a moderate to high reactivity for normal nuclear TDP-43 by all MAbs except MAb 138. MAb 138, which is specific for region C in the RRM2, displayed high reactivity to pathological TDP-43 deposits similar to other TDP-43 MAbs specific for Regions C-F, in the absence of significant normal nuclear TDP-43 staining  showed robust reactivity for neocortical TDP-43 pathology across FTLD-TDP subtypes and ALS spinal cord inclusions. These MAbs directed against epitopes in the mid- and C-terminus of TDP-43 detected a moderate to abundant normal nuclear TDP-43 with the exception of MAb 138 (Region C) (Figure There are several possible explanations for the unique IHC staining by MAb 138. First, there is a disease-specific pathological modification to one or more aa within the epitope recognized by MAb 138, but this seems unlikely as all these novel MAbs were generated using un-modified human rTDP-43. Second, a recent report suggests that the RRM2 region can undergo self-association when interacting with nucleic acids and this could alter the conformation of TDP-43 or the accessibility of the epitope recognized by MAb 138 . FinallyProtein misfolding and prion-like spread of neurodegenerative disease proteins has emerged in the forefront of research for drug-development in FTLD-TDP and ALS ,29. AlthThe authors declare that there are no competing interests to disclose.LKK, DJI, AKW, VML, and JQT designed the study and wrote the manuscript. DMR carried out the hybridoma fusion and subcloning as well as the ICC experiments. LKK, AKW, and YX performed the MAb characterizations and evaluations. DJI evaluated the human neuropathology. All authors read and approved the final manuscript.Figure S2. Serial Dilutions of MAb 138 show minimal nuclear reactivity and Table S1. Summary of patient demographics.Representative ELISA results for a newly developed MAb, Click here for file


To the general public, the Internet is an acceptable method of obtaining information. It also plays an important role in guiding patients and solving their problems.We investigated the clinical characteristics of an Internet website to provide guidelinesand tips for consultation.In this retrospective observational study, we analyzed the useof a free public Internet perinatal consultation website provided by the Ministry of Healthand Welfare of Korea. We evaluated 2,254 Internet perinatal consultations and assessments of prenatal and obstetrics from August 2006 to December 2009. We evaluated thepatients’ questions based on Williams’ textbook categories and their clinical characteristics.The mean age of patients seeking consultation was 33.9 ± 13.2 years, and paritywas 1.2 ± 0.5. The most commonly asked questions were about drug safety during pregnancy (20.4%), and questions regarding prenatal care (18.7%) and labor pain (15.4%)were the second and third most commonly asked questions, respectively.The Internet can provide good information to patients. Thus, guidelines regarding pregnancy-related questions and answers should be created. Obstetricians coulduse our data to identify question tendencies. Women who wish to become pregnant usuallyseek educational methods such as books and magazinesfor safe fertility and delivery. The Internetis currently a common and acceptable method bywhich patients seek answers to their questions.Based on 33 websites about Internet usage by pregnantwomen, 96.6% of women have Internet accessat home . PregnanIn the present study, we investigated the clinicalcharacteristics of questions on an Internet websiteto provide guidelines and tips for consultation.In this retrospective observational study, we analyzedthe use of a free public Internet perinatalconsultation website provided by the Ministry ofHealth and Welfare of Korea. This website providesinformation on pregnancy, delivery, childcare, and infant health programs and provides amultilingual system for immigrants as well.The website also offers perinatal consultationsand assessments of gynecologic problems free of charge. On average, there were 25,192 logins per day. The service was accessible via a Korean web domain. Fourteen obstetricians conducted the consultations using a private personal computer. Pediatric doctors also consulted with patients about pediatric problems. The consultant had a duty to reply to each patient’s questions. Women asked questions on the site, and an automatic, immediate, repetitive alarm sounded on the consultant’s telephone until a reply was updated on the website. The consultant was required to complete the reply while on duty. Duty days covered all 365 days of the year. All replies were completed within 24 hours.We evaluated 2,254 consultations from August 2006 to December 2009. A total of 122 consultations occurred during the initiation year, whereas 802 consultations were performed in 2009. Two research nurses and one obstetrician evaluated each consultation. We evaluated the questions based on Williams’ textbook categories and patients’ clinical characteristics.Approval for this study was given by the Human Ethics Committee at SCH (Soonchunhyang University) Medical Center . We mention that no personal data is published and the privacy of the users was respected.Results are presented as number and percentage values. Variables were expressed using the number and the percentage. Statistical data were generated with the SPSS version 12 for Windows .The mean age of patients seeking consultation was 33.9 ± 13.2 years, and parity was 1.2 ± 0.5. Most of the consultations were with women  , and theIn Korea, there are almost 80 known websites about pregnancy, delivery, and child care . Our datThe Internet is useful for pregnant women and plays a role in decision-making. In a Web survey of Internet use in 24 countries, most women (97%) used the Internet for pregnancy-related questions . MankataThe preconception period was also a common topic of consultation based on our data. Mankata reported that 5% of those asking questions were husbands, similar to our study 4.7%). Women in other countries were interested in and had many questions regarding pregnancy, but the topics in which they were interested were not the same. Pregnancy-associated questions are common on the Internet, which is a very cost-effective way to obtain information (.7%. WomeThe Internet can provide good information, but that information is sometimes confusing and may be inaccurate. If the website evaluated in this study had an automatic system for differentiating questions based on keywords, trimester, previous related questions, and so forth, the questions could easily be categorized and the evaluation time could be reduced. Thus, such a system should be created for pregnancy-related questions and answers. Culture and health care systems might influence the nature of the most frequent questions. We inferred that Korean women are very anxious about the health of the baby, so they were very eager to obtain information about drug safety and preconception care.


Very few cases of scar sarcoidosis affecting the eyes and bone together have been reported in the last few years.We report a case of a 49-year-old Spanish man with recurrent bilateral granulomatous uveitis and a fistulous nodular lesion in the left pre-tibial region (scar granuloma) on the site of an 8-year-old scar. He presented with bilateral hilar adenopathies and elevation of inflammatory markers and angiotensin-converting enzyme. A histologically confirmed sarcoid of the tibia with a radiologic appearance unusual for long tubular bone involvement was observed. He also had bilateral ophthalmologic involvement.Sarcoidosis is a disease of unknown cause histologically characterized by non-caseating granulomas that can involve any organ or tissue. Osseous sarcoidosis is a relatively rare presentation. However, on the basis of cases reported in the literature, sarcoid lesions on bones are generally asymptomatic. Biologic agents are considered an alternative therapy for sarcoidosis resistant to conventional treatment. Scar sarcoidosis of the tibia is considered a rare disease. It is usually described in old scars 10 years) [0 years .In the last few years, very few cases of scar sarcoidosis affecting the eyes or bone have been described in the literature. The appropriate treatment consists of steroids with gradual dose increases until the condition is controlled, followed by immunosuppressive agents such as methotrexate and azathioprine for control of inflammation . SeveralA 49-year-old Spanish man was referred to our hospital by an ophthalmologist in June 2010 with a symptom of red eyes. A uveitis protocol was implemented. A chest X-ray was ordered. The X-ray showed mediastinal adenopathy, for which the patient was referred to the respiratory medicine department. The results of his respiratory function tests were normal, so a high-resolution computed tomographic (CT) scan of the lungs was ordered. As he had no interstitial disease, he was diagnosed with stage I pulmonary sarcoidosis, and no systemic treatment was prescribed. A sacroiliac X-ray and a magnetic resonance imaging scan showed sclerotic changes that appeared chronic. No bone edema was observed. His human leukocyte antigen B27 test was negative, and his angiotensin-converting enzyme (ACE) level was elevated. His physical examination showed that he was hemodynamically stable and afebrile and that his overall health status was good. His musculoskeletal examination revealed a raised fistulous erythematous lesion with serous bloody secretion. He had no erythema or joint effusion. In an examination of his skin, no erythematous bullous lesions were found on his lower extremities.Toxoplasma, Treponema pallidum, Borrelia, Mycoplasma and Chlamydia). To rule out the possibility of tuberculosis, a Mantoux test (negative) and serial urine cultures with auramine staining (negative) were ordered. His high-resolution CT scan was compatible with stage I sarcoidosis .The patient’s blood examination results showed elevated acute-phase reactants  but no leukocytosis or left shift. His kidney and liver function, electrolytes and clotting were normal. His ACE level was somewhat elevated, and his urine sediment was normal. The results of his serologic tests were negative  that do not generally require treatment. However, soft tissues can be affected, with formation of fistulas on the skin that require curettage and corticosteroid therapy or chloroquine, which generally work very well with cutaneous sarcoidosis. The incidence of these lesions is 1–13%, and bone lesions due to sarcoid granulomas are diagnosed by biopsy .Approximately 38% of all cases of erythema nodosum are related to sarcoidosis in Löfgren syndrome, and they occur with skin lesions such as plaques, nodules and scars –19. WhenIn 50−70% of sarcoidosis cases related to other sites, the eyes are also involved and the eye condition often becomes chronic and recurring –22.ACE values are elevated in 88% of patients with active sarcoidosis, but using these levels to diagnose the disease can be misleading because approximately 10% of results are false positives and about 40% are false negatives. However, measuring the ACE level is useful to monitor disease progression .The cornerstone treatment is based on corticosteroids with gradual dose decreases until the disease is controlled. If the patient’s sarcoidosis is still active, immunosuppressive agents followed by second-line treatments such as methotrexate or azathioprine, as well as biologic agents such as infliximab, can be used for several clinical manifestations, although the evidence for these is based on small studies, so more extensive studies are needed to establish use of these treatments , 14, 23.Four other scar sarcoidosis clinical cases have been reported to date. Scar sarcoidosis was found in all of them. This is an uncommon but specific manifestation of sarcoidosis. One of the patients developed scar sarcoidosis on his right finger . AnotherTo the best of our knowledge, this is the first case of tibia sarcoidosis scar reported in the literature. Sarcoidosis generally responds well to corticosteroid therapy, but it is not self-limiting and often also requires the use of an immunosuppressant to avoid corticosteroids and to achieve remission without recurrence. When the eyes are involved, it is often related to other sarcoidosis symptoms. If the disease is not controlled, these symptoms will not be controlled either. Sarcoidosis is usually treated with methotrexate, cyclosporine, azathioprine or cyclophosphamide. New biologic therapies have also been used, with infliximab producing the best response, although the number of studies on the subject is limited , 27, 28.Written informed consent was obtained from the patient for publication of this case report and any accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal.


Since the tunable ranges for λ and μ in graphene are experimentally reachable, we thus open the possibilities for its device application. The symmetry in spatial distribution is broken by the vertical and the transversal spin-spin correlations due to the effect of spin-orbit coupling, leading to the spatial anisotropy of spin exchange, which distinguish our findings from the case in normal Fermi liquid.Two magnetic impurities on the edge of a zigzag graphene nanoribbon strongly interact with each other via indirect coupling, which can be mediated by conducting carriers. By means of Quantum Monte Carlo (QMC) simulations, we find that the spin-orbit coupling  Fk as well as decay behavior Rγ−Due to the unusual electronic quasi-particles and the flexible carrier concentration46k ↔ −k is forbidden without a spin flip. Due to this scenario, there is spin-current vortex around the single local spin on the helical edge132Se3 Recently, there is a surge of interest in effect of spin-orbit coupling (SOC) on electronic properties, which opens a class of novel time-reversal invariant topological insulators (TI)In this paper, we consider two Anderson impurities on the edge of a zigzag graphene ribbon, and investigated the effect of Kane-Mele SOC and chemical potential on the local spin exchange, which can be achieved by heavy metal doping or electric field. Through Quantum Monte Carlo simulation at finite temperature, we carry out a comprehensive study on the spin and charge coupling strengthes of this impurity pair. For the spin-spin correlation, which measures the RKKY interaction, we found that, the in-plane components of correlations favor ferromagnetism but the out-plane correlation can be tuned from ferromagnetism to antiferromagnetism by the SOC, while such transition is much dependent on the on-site Coulomb interaction of impurity orbital. From recent first-principal and experimental results of enhanced SOC in graphene, we conclude that spin-spin exchange among impurities in graphene is experimentally tunable by SOC. In contrast, the charge fluctuation between two impurities is always suppressed. Meanwhile, in-plane components of correlation will decay with an oscillation behavior when the distance between two impurities is increased, but the out-plane spin and charge correlations will decay smoothly, which is totally different from the Fermi liquid behavior, and leading to the spatial anisotropy of spin exchange.To simulate the model, we construct a model with width of 20 Å zigzag-edge graphene ribbon and along the horizontal direction we adopt periodical condition. As indicated in Kane-Mele investigationHK-M is the Kane-Mele model in a zigzag edge graphene ribbonHK-M = tH + Hso, with tH being the usual nearest-neighbor hopping of tight-binding model in grapheneHso is the SOC term and zσ is the z Pauli matrix. Hso thus has opposite signs for opposite electron spins. The parameters ijν = −jiν = ±1 depend on the orientation of the two nearest neighbor bonds as the electron hops from site i to j: ijν = +1 if the electron makes a left turn to the second bond. It is negative if it makes a right turn. H1 is the part for two impuritiesH2 describes the hybridization between impurities and conducting electronsm is the index of two impurities, in H1 and H2, mε is the energy of the impurity orbital and mU is Coulomb repulsion inhibiting the simultaneous occupancy of the orbital by two electrons and mV is the hybridization strength. In this work, we consider the two identical impurities, so the above parameters in Hamiltonian are m-independent.The QMC simulation is based on two-Anderson-impurity model, the total Hamiltonian reads asτ = 0.1 to suppress the systemic error. The QMC naturally returns the imaginary-time Green's function α = x, y, z directions versus distance asαS (R) directly reflect the RKKY interaction which is mediated by conducting electrons on the helical edge, while charge-charge correlation C(R) indicates the indirect Coulomb repulsion.We use the single-impurity QMC base on discrete-time algorithmμ = 0 and zS highly depend on SOC, which can be reduced in a linearly way and have a wide variation range. When SOC is increased, the value of vertical correlation zS can be changed from positive to negative, it means that a transition from a ferromagnetic (FM) to anti-ferromagnetic (AFM) correlation is achieved when the SOC reaches a critical value. It is interesting to note that, the critical SOC strength for such a transition increases as Coulomb interaction is increased, from the QMC simulations. For example, the critical SOC value is about 100 meV when U is 1 eV, but it is increased to 300 meV when U is is 5 eV. The underlying mechanism can be attributed to the competition of spin and charge fluctuations between two impurities. When they are close to each other, the changing in carriers SOC can directly result in the spin-flip process which reflects in zS, but the increasing Coulomb interaction U can block spin flip, so it is seen that with a larger U in zS but in xSy) at SOC strength λ = 0. But when λ ≠ 0, the SOC effect on both spin correlation is different, leading to non-degenerated zS and xSy)( (μ is in the range of −0.2 t to −0.8 t). But when μ is further reduced, both zS and xSy) and transversal one xSy)((R) have different spatial distribution. xSy)((R) has both the behavior of decaying and oscillating, but zS(R) is always decaying with the increased distance. Similar to zS(R), the charge-charge correlation is also decaying with distance (f1(R), f2(R) and f3(R) are decaying function of distance has the formula as 1/γR.The distance between two impurities is expected to influence the couplings of spin-spin and charge-charge due to the fact that the RKKY interation is sensitive to the distance. We preformed the QMC calculations to address the problem when varying the distance. In distance . These cxSy)((R)/C(R) and zS(R)/C(R). We find that zS(R)/C(R) is a trivial constant, but xSy)((R)/C(R) is an oscillating function of distance, as shown in To more clearly display these correlations, we calculate the ratios U′ in graphene and hybridization. In realistic graphene zigzag nanoribbon, the Coulomb interaction plays a signification role and stabilizes a spin-polarized ground stateRγ− cos 2FRk, here Fk is the Fermi momentum. The decaying factor γ is the function of Hubbard U′: when U′ = 0, γ = 1, and when U′ ≠ 0, 0 < γ < 1 31U′ = 0. The methods used to enhance the graphene's SOC in π bands still dominate near the Fermi energy and Kane-Mele Hamiltonian is also valid and the effect of hybridization can be absorbed in the enhanced SOC through second order approximationFinally, we discuss the model Hamiltonian of In conclusion, we investigate two Anderson impurities on a zigzag graphene nanoribbon by QMC simulation in present work. When the system is under a topological phase transition driven by Kane-Mele SOC, we find that, the spin rotation symmetry in RKKY interaction between two impurities can be partly broken because of the SOC. As the two impurities are close to each other and the Fermi energy is near the Dirac point, the in-plane RKKY interactions favor ferromagnetism but the out-plane component can be tuned from ferromagnetism to anti-ferromagnetism by the SOC. The distance effect on RKKY interaction is also systermatically investigated, the in-plane interaction has both behavior of decaying and oscillation, but the out-plane interaction and charge-charge correlation only has decaying as the distance between them is increased. This behavior is totally different from the conventional Fermi liquid. Our theoretical findings here provide some useful suggestions of the spin-polarized STM measurement to RKKY interactionF.H. conceived the idea and do the QMC calculation, L.K. made the first-principle discussion and collected the data in the Table 1, and F.H. and L.K. wrote the manuscript together. F.H., L.K. and T.F. discussed the results and reviewed the manuscript.


Idiopathic pulmonary fibrosis (IPF) is a devastating condition with a variable course. Not uncommonly, IPF patients are hospitalized for respiratory-related causes, including disease worsening. This study aimed to characterize the prevalence, and economic and health care burden of IPF.Retrospective insurance claims data collected yearly between January 1, 2009 and December 31, 2011, were used to determine prevalence and calculate all-cause and respiratory-related resource utilization and costs. Patients had at least one inpatient claim or two outpatient claims for IPF (ICD-9-CM code 516.3). Results for health care burden are reported for the 2011 cohort (similar findings in 2009–2010). Costs are reported in 2011 US dollars ($).Patients with IPF had a mean age of 69.8–71.3 years. Overall prevalence for IPF was 28.8, 28.1 and 19.8 per 100,000 insured persons in 2009, 2010 and 2011. In each year, prevalence increased with age. In 2011, 37.7 % of patients were hospitalized at least once for any cause; 19.5 % for respiratory-related reasons. Also in 2011, the mean number of all-cause outpatient visits and respiratory-related office visits was 18.5 and 5.7 per patient, respectively. All-cause health care costs in 2011 were $59,379 per patient; 36.6 % of costs  were respiratory related.The prevalence of IPF in this claims database increased with age, with a notable increase in those over 65 years. IPF is associated with a large economic and health care burden. Additional research is needed to determine how such burden might be reduced. Idiopathic pulmonary fibrosis (IPF) is a chronic disease in which normal lung parenchyma is replaced with fibrotic tissue, leading to dyspnea, cough and impaired lung function . IPF preAlthough data are limited, compared with patients without IPF, those with IPF require greater health care resource utilization, have a greater number of comorbidities , 8 and gIPF incidence rate and prevalence estimates vary across studies because of differences in design and data sources , 12. TheData for this study were obtained from a large Health Insurance Portability and Accountability Act (HIPAA) of 1996-compliant insurance claims database. The database used was the Humedica (from Optum) database and access was via a license agreement (not openly available). The database provides information on medical and pharmacy claims for approximately 30 million lives in the USA. In this study, extracted data covered four geographical regions in the USA . In compliance with HIPAA 1996, all data were void of identifying information and therefore the study is exempt from institutional review board approval.This study included three cohorts, each formed by capturing data spanning a 1 year period , and other patients may have been diagnosed in the last month of a study period (e.g. December) and were therefore included in the cohort for that entire year. In 2012, the ICD-9 code for IPF changed from 516.3 to 516.31. Due to the possibility of coding errors in 2012 resulting from this change, the study was limited to 2009–2011. For each year, insurance claims were extracted from the database for all patients. Respiratory-related medical claims were classified as those associated with ICD-9-CM codes 460.xx–519.xx.Total health care costs, non-pharmacy costs (including inpatient and outpatient services costs) and pharmacy/medication costs were analyzed for all-cause and respiratory-related health care utilization. The number of inpatient hospitalizations, emergency department (ED) visits and physician office visits was also assessed. Respiratory-related medication costs include all outpatient pharmacy claims of respiratory therapy, oral antibiotics and all possible IPF medication. In these analyses, medication costs were assessed and include any of the eight groups of prescription medications used to possibly treat IPF . No FDA-approved treatments were available for IPF at the time of this study. The usual physician specialty for IPF care was defined as the physician specialty with the largest plurality of IPF office visits (office visits with primary diagnosis of IPF) with evaluation and management services. For some patients, there was no office visit meeting that definition (e.g. with a primary IPF diagnosis) and they could not be classified. In other patients, there were visits but the physician specialty was not listed, and they could not be classified either.Prevalence estimates for each year were calculated based on the number of patients with an ICD-9-CM code for IPF (516.3) and no other type of ILD after their last IPF claim as the numerator. The number of members continuously enrolled in a given calendar year, regardless of diagnosis, was used as the denominator.Descriptive statistics were generated for baseline data. Means and standard deviations are reported for continuous variables, and counts and percentages for categorical variables. Economic and health care burden are reported for the 2011 cohort (findings were similar in 2009–2010). Estimated costs are reported in 2011 US dollars ($). The Charlson Comorbidity Index, which includes a total of 22 conditions and can predict 10-year mortality risks , was useDemographics are reported for all 3 years and for prevalence estimates, but only 2011 data were used for the utilization analyses. All figures and tables show data for all 3 years.P = 0.917). The study population had a high (mean of six) number of chronic conditions and comorbidities contributing to this score , 28.1  and 19.8  per 100,000 insured persons in 2009, 2010 and 2011, respectively. Similar values were observed between genders  consistent with an attempt to confirm a diagnosis of IPF. Use of oxygen at home was observed in 52.6 % of patients; 44.6 % were treated with oral corticosteroids and 1.7 % had a lung transplant. A total of 12.1 % of patients participated in pulmonary rehabilitation.P = 0.690) in the mean number of all-cause outpatient clinic visits, which in 2011 was 18.5 per patient. Similarly, there was no significant difference over time in the mean number of clinic visits specifically due to respiratory-related conditions . A total of 37.7 % of patients were hospitalized at least once due to any cause, and 7.7 % were hospitalized (for any cause) at least three times. Overall, 30.5 % of patients visited the ED on at least one occasion for any cause. For respiratory-related conditions, 19.5 % of patients (n = 221) were hospitalized and 15.8 % (n = 179) visited the ED at least once .There was no significant difference over the three periods —the denominator—is likely to have contributed to the lower prevalence of IPF in 2011, since the numbers of IPF patients (the numerator) were similar over the 3-year period. In fact, the higher number of insured people in 2011 was composed of a large number of individuals from the youngest age group (unlikely to have IPF). The number of those aged between 12 and 44 years differed from 1.7 and 1.9 million in 2009 and 2010, to 2.5 million in 2011. Data from the Centers of Disease Control and Progression support our observation, and show that the proportion of uninsured adults aged 19–25 years fell from 35.6 % in the third quarter of 2010 to 27.0 % in the fourth quarter of 2012 . In addiThe prevalence of IPF is usually reported as higher among males –21; howeFew previous studies have reported on the economic burden of IPF. Most recently, Wu et al.  found thDatabase analyses come with inherent limitations, and use of secondary data, without direct involvement of the patient or physician, may decrease confidence in the conclusions drawn , 27. InsFurthermore, approximately 50 % of IPF patients also had a diagnosis of COPD, including emphysema; whether these were incorrect diagnoses in patients who previously smoked (but did not have COPD) or correctly coded comorbid conditions cannot be determined from the available data. To help reduce the effect of coding inconsistencies, data on all-cause and respiratory-related conditions were considered. This included individuals with at least 1 inpatient claim or two outpatient claims with IPF as one of the diagnosis codes, in hopes of capturing all patients with IPF, including those who may have been misdiagnosed. To limit miscoding, patients who had a non-IPF ILD claim after the last IPF claim in that calendar year were excluded from the analysis. Similar criteria have been used in previous studies , 8, 18.Because the ICD-9 code for IPF changed from 516.3 to 516.31 in 2012, the current analysis was limited to 2009–2011 to further limit possible coding errors. Other limitations include the cross-sectional study design and the fact that patients were not required to have IPF for the entire calendar year; thus, given that some patients may only have started to incur costs later in the year, the total economic burden of disease may be underestimated. A cross-sectional view of patients with IPF offers a true-to-life picture of what might be expected in any given year in which some patients are diagnosed early and others later in the year.IPF presents a large burden on health care utilization resources. Additional research is needed to determine whether and how this burden might be reduced.


Archaea, specifically the Thermoproteales (phylum Crenarchaeota). Environmental sequencing (16S rRNA gene and metagenome) from geothermal habitats in Yellowstone National Park (YNP) has expanded the available datasets for investigating 16S rRNA gene introns. The objectives of this study were to characterize and curate archaeal 16S rRNA gene introns from high-temperature habitats, evaluate the conservation and distribution of archaeal 16S rRNA introns in geothermal systems, and determine which “universal” archaeal 16S rRNA gene primers are impacted by the presence of intron sequences.Intron sequences are common in 16S rRNA genes of specific thermophilic lineages of Pyrobaculum, Thermoproteus, and Caldivirga 16S rRNA genes contained the most abundant and diverse intron sequences. Phylogenetic analysis of introns revealed that sequences within the same locus are distributed biogeographically. The most diverse set of introns were observed in a high-temperature, circumneutral (pH 6) sulfur sediment environment, which also contained the greatest diversity of different Thermoproteales phylotypes.Several new introns were identified and their insertion loci were constrained to thirteen locations across the 16S rRNA gene. Many of these introns encode homing endonucleases, although some introns were short or partial sequences. The widespread presence of introns in the Thermoproteales indicates a high probability of misalignments using different “universal” 16S rRNA primers employed in environmental microbial community analysis.This article was reviewed by Dr. Eugene Koonin and Dr. W. Ford Doolittle.The online version of this article (doi:10.1186/s13062-015-0065-6) contains supplementary material, which is available to authorized users. Archaea contain 16S rRNA gene introns, which are mobile genetic elements that do not appear to impact the host’s growth or metabolism. 16S rRNA gene introns have been identified in two genera of the order Desulfurococcales (Aeropyrum and Staphylothermus) ).Pyrobaculum spp. contained the majority of known introns and these are found at diverse loci across the 16S rRNA gene . Other genera in the Thermoproteales also contain intron sequences at many of the same loci: Thermoproteus , Caldivirga , and Vulcanisaeta (locus 1391). Desulfurococcales-like introns were identified in six loci .Intron sequences were confined to 13 loci across the 16S rRNA gene Fig. . PyrobacP. oguniense 16S rRNA homing endonuclease (Pog.S1213) promoted the homing of its own intron and guaranteed the co-conversion of both the downstream hairpin-forming intron (Pog.S1205) and the intervening eight nucleotides by cleaving at the intron− locus 1205 [Most 9/13) intron regions contained two or more predicted homing endonuclease genes, which code for either one or two of the canonical LAGLI-DADG motifs . The nucleotide sequences comprising the DNA insertion sites and the BHB motif vary among loci and it is unclear how sequence specificity of the homing endoribonuclease or the splicing ligase contributes to intron propagation and distribution. Several intron loci were identified in both the Crenarchaeota and Aigarchaeota , which suggests that these loci have been sites of intron insertions since the divergence of these two phyla. Based on extant insertion loci, intron sequences radiated throughout the Thermoproteales, specifically in the Pyrobaculum and Thermoproteus spp.Comparison of intron sequences within the same loci identified highly-conserved nucleotide residues near the intron-exon junctions, which represent conserved intron cores , 25. ThePyrobaculum sp. 1860 [P. neutrophilum [P. yellowstonensis strain WP30 [Vulcanisaeta distributa IC-065 clone from Japan [Pyrobaculum and Thermoproteus). Introns found at different 16S rRNA gene loci were too different for phylogenetic comparison (<20 % nt/aa identity).Intron sequences within the same loci were successfully aligned at the nucleotide level and subsequent phylogenetic analysis revealed both intra-locus and geographic separation Fig. . Intron sp. 1860  and P. nrophilum , respectain WP30  that is Pyrobaculum contain the most abundant 16S rRNA introns in the order Thermoproteales and although they predominate in high-temperature, near-neutral to alkaline (pH > 6) geothermal systems, populations of the other Thermoproteales genera  are found with Pyrobaculum in > 65 °C, pH 5 - 7 hot springs [The phylogenetic distribution of intron sequences by geographic location is consistent with the hypothesis that once a homing endonuclease becomes fixed in a genome, selection pressure for the gene significantly decreases resulting in degeneration (mutation) and enforcing the requirement of frequent horizontal transmissions between populations to maintain intron propagation . Populat springs , 48. The springs , these e springs . Very liPrimer sequences designed for archaeal 16S rRNA genes (n = 51) were manually aligned to intron-containing 16S rRNA gene sequences to determine which primers were interrupted by, or spanned intron sequences . All transcribed introns form predictable secondary structure including the characteristic bulge-helix-bulge motif. Many intron sequences encoded homing endonucleases, although other introns were short and/or non-coding sequences. Phylogenetic analysis revealed that intron sequences grouped together by geographic location and then by host taxa. The presence of introns in highly conserved regions of 16S rRNA introns directly interferes with use of “universal” primers often used in environmental gene surveys.Intron sequences were identified by querying (blastn/blastx) the NCBI nr and the DOE-Joint Genome Institute (DOE-JGI) Integrated Microbial Genomes (with Microbiome Samples (IMG/M) databases) with previously identified 16S gene introns  before phylogenetic analysis and/or Weblogo3 analysis . PhylogeReport form: Jay and Inskeep report the distribution of Group I introns in 16S RNA genes from hyperthermophilic archaea, and in particular Thermoproteles, where these introns are most common. The analysis is carefully performed, and there is an important conclusion on the evolution of self-splicing introns, namely that they group by geographic location, i.e. spread primarily horizontally. Also, this analysis is of practical value because the authors carefully assess the effect of introns on the use of universal primers.Authors’ response: We thank the reviewer for these comments.Quality of written English: Acceptable.Report form: This is a perfectly fine and straightforward report and summary of the distribution of introns in the rRNA genes of members of the Thermoproteales. It will provide a basis for future experimental and comparative studies, and raises some interesting questions, such as the means by which introns are transferred between different lineages in a given environment and the structural adaptations of the intron RNA to high temperature, but does not attempt to answer them. It also points out how intron presence might result in failure to amplify specific regions of rRNA genes by PCR. Members of Thermoproteales might thus be underestimated in environmental sampling, and one wonders whether there are abundant major groups that go undetected because they always have such introns. Indeed, is there anything to keep introns from becoming colonized by genes essential for cellular survival, so that pieces of the rRNA gene become permanently separated, and the cells that bear them become undetectable with standard molecular methods?Authors response: We thank the reviewer for these comments, and for the very interesting question regarding possible ‘colonization’ of introns by genes coding for essential function, and subsequent separation of rRNA gene fragments . The question regarding intron colonization by other genes is difficult to answer due to the complexities of intron propagation. As we currently understand 16S rRNA gene introns in the Thermoproteales, a homing endonuclease (gene or transcript) is required for intron insertion into a functional rRNA. Consequently, any disruption to the homing endonuclease would not allow for insertion and propagation of the intron sequence. Any cell that may bear intron fragments by definition will still have a functioning rRNA somewhere in the genome and therefore would be detectable with standard methods.Quality of written English: Acceptable.