Antibodies targeting the checkpoint molecules programmed cell death 1 (PD-1) and its ligand PD-L1 are emerging cancer therapeutics. We systematically investigated PD-1 and PD-L1 expression patterns in the poor-prognosis tumor entity high-grade serous ovarian carcinoma.in silico validation of mRNA data was performed in The Cancer Genome Atlas (TCGA) dataset.PD-1 and PD-L1 protein expression was determined by immunohistochemistry on tissue microarrays from 215 primary cancers both in cancer cells and in tumor-infiltrating lymphocytes (TILs). mRNA expression was measured by quantitative reverse transcription PCR. An p < 0.035 each), and most being significant for OS. Most factors also had prognostic value that was independent from age, stage, and residual tumor. Moreover, high PD-1+ TILs as well as PD-L1+ TILs densities added prognostic value to CD3+TILs . The significant positive prognostic impact of PD-1 and PD-L1 mRNA expression could be reproduced in the TCGA gene expression datasets .PD-1 and PD-L1 expression in cancer cells, CD3+, PD-1+, and PD-L1+ TILs densities as well as PD-1 and PD-L1 mRNA levels were positive prognostic factors for progression-free (PFS) and overall survival (OS), with all factors being significant for PFS (Despite their reported immune-modulatory function, high PD-1 and PD-L1 levels are indicators of a favorable prognosis in ovarian cancer. Our data indicate that PD-1 and PD-L1 molecules are biologically relevant regulators of the immune response in high-grade serous ovarian carcinoma, which is an argument for the evaluation of immune checkpoint inhibiting drugs in this tumor entity. High-grade serous carcinoma (HGSC) is the major histological subtype (approximately 70%) of ovarian carcinomas. It is a poor-prognosis tumor , which is due to late diagnosis (75% in FIGO III/IV) as well as the development of resistance to standard platinum-based chemotherapy. However, high numbers of tumor-infiltrating lymphocytes (TILs) have repeatedly been shown to provide a significant survival advantage in ovarian carcinoma. Particularly the presence of T cells CD3+) and various T cell subpopulations  are indicators of a better prognosis, , 3 stron+ and varImmune checkpoint inhibitors constitute a novel class of cancer therapeutics that do not target the cancer cell itself but rather ligands and receptors on T cells that attenuate the anti-tumoral immune reaction and increase immune tolerance . Emevia AKT or STAT3, a mechanism termed intrinsic immune resistance, or by IFN-gamma produced by activated T cell or NK cells (adaptive resistance). [PD-L1 is induced on monocytes and epithelial cells, upon IFN-gamma stimulation , whereasstance). , 5 The mPD-1 inhibitors  have been approved in therapy-refractory malignant melanoma, and together with PD-L1 inhibitors  are investigated in clinical trials on various recurrent or metastatic malignancies to date. Early clinical trials in ovarian carcinoma are ongoing. , 10 ReliTo estimate if a tumor entity might constitute a candidate neoplasm for evaluation of a specific cancer therapeutic, the expression pattern of the drug target as well as its clinical relevance are of interest. We therefore systematically evaluated the expression of PD-1 and PD-L1 in a cohort of 215 primary ovarian high-grade serous carcinomas by determining protein expression in cancer cells and TILs as well as mRNA expression.n = 201 and n = 202 cases, respectively. If positive, both markers showed a membrane-accentuated expression, which was also often accompanied by a cytoplasmic expression . A significant number of cases did not show any membranous expression on cancer cells , and for further statistical analyses we decided to split the study group into cases with no PD-1/PD-L1 expression (IRS = 0) and cases with any expression. Membranous PD-1/PD-L1 expression were not correlated to each other . As no prognostic effect of cytoplasmic PD-1 or PD-L1 expression in cancer cells could be detected (not shown), only data on membranous expression of these markers will be given subsequently.Data on PD-1 and PD-L1 expression in cancer cells were available for n Figure . Membrann = 200 for each marker). All TILs markers showed a positively skewed distribution, which means that the median was lower than the mean: Intraepithelial CD3+ TILs numbers (per 5 HPF) ranged from 0 to 543, however most cases had rather low numbers of CD3+ TILs  for each marker. Statistical analyses are shown for these values subsequently. CD3+, CD4+, CD8+, PD-1+, and PD-L1+ TILs/mm2 highly correlated with each other . PD-1 expression in cancer cells showed a borderline positive correlation with CD3+ TILs/mm2 , but not with PD-1+, PD-L1+, CD4+, or CD8+ TILs, and PD-L1 expression in cancer was not significantly associated with TILs at all (p > 0.05 each).As stromal TILs had a weaker (CD3) or non-significant impact on prognosis  as compared to intratumoral TILs (data not shown), data for the latter only are are given subsequently. We separately counted CD3+ as well as PD-1+ and PD-L1+ TILs in identical tumor areas ). We furthermore investigated the potential dependence of CD4+ and CD8+ TILs density from combined cancer cell and TILs expression of PD-1 and PD-L1. As shown in PD-1 and PD-L1 cancer cell and TILs expressions were further investigated as to their relationship to the CD4+ and CD8+ T cell population when we grouped cancer cell expression in four groups. We did not observe a dependence of CD4+ or CD8+ TILs density and expression of PD-1 or PD-L1 in cancer cells (p < 0.0001), as well as with CD3+, PD-1+, and PD-L1+ TILs . There was a trend towards a positive correlation between PD-1 mRNA and PD-1 expression in cancer cells , however not between PD-1 mRNA and PD-L1 expression in cancer cells or between PD-L1 mRNA and PD-1/PD-L1 expression in cancer cells (p > 0.1 each).Informative mRNA expression data were available for 200 cases for PD-1 and 204 cases for PD-L1. Both markers showed a rather low expression with PD-1 40-deltaCT values ranging from 24.31 to 32.98 (median 29.49) and PD-L1 data ranging from 25.39 to 33.33 40-deltaCT , while for PD-1 expression only a trend was seen for OS  and OS (p = 0.051) were longest, in cases with only one positive marker were intermediate, and in double negative cases were worst, however as only two tumors were PD-1-/PD-L1- the data for the latter group must be interpreted with caution .Both PD-1 and PD-L1 expressions in cancer cells were significantly linked to a better PFS , p = 0.002. This significant positive impact on prognosis was also seen for PD-L1+ TILs density for both PFS (HR = 0.75 (95% CI 0.58-0.97), p = 0.026) and OS (HR = 0.72 (95% CI 0.55-0.96), p = 0.025), as well as for PD-1+ TILs density for PFS (HR = 0.78 (95% CI 0.62-0.99), p = 0.044), with a lack of significance for OS .First we investigated the prognostic impact of TILs using a cutoff-free approach. Due to the fact that TILs distribution was positively skewed, we logarithmized the data and performed Cox regression analysis using continuous data. A high density of CD3+ intratumoral TILs was a significant positive prognostic marker for PFS and OS on the continuous scale: PFS: HR = 0.65 (95% CI 0.49-0.85), 2, respectively. Using this optimal cutoff, Kaplan-Meier analysis showed that CD3+ TILs had a significantly favorable impact on PFS and OS , with borderline significance in multivariate analysis .As all TIL subpopulations in our study were highly correlated with each other, we wondered if the prognostic effect of PD-1+ and PD-L1+ TILs might only be a bystander effect of the impact of T cell infiltration by itself. We therefore investigated the prognostic effect of combined CD3+ and PD-1+/PD-L1+ TILs density. Surprisingly, tumors that had a high PD-1+ TILs or PD-L1 TILs density in addition to high CD3+ TILs had a better prognosis (both PFS and OS) than tumors with low PD-1+ or PD-L1+ TILs counts despite of a high CD3 infiltration Figure . Thus, Pp = 0.012), but PD-L1 mRNA was not (p = 0.609). Significant cutoff points - similarly to TILs density - were found over a rather large range: for PD-1 mRNA 39 out of 128 cutoff points (39.5%), and for PD-L1 mRNA 49 out of 128 cutoff points (38.3%) showed significant positive prognostic impact  as well as in the subgroup with residual tumor after surgery , and furthermore in cancers without residual tumor after surgery . PD-1 expression (syn. PDCD1 in TCGA) was also a positive prognostic factor for the total cohort, however its prognostic value was of reduced robustness as only few cutoffs were significant: Agilent: 14 out of 460 cutoffs significant (3.0%), optimal cutoff p = 0.02, Figure p = 0.013, not shown), however missed significance in RNA seq data . In the subgroup of tumors with residual tumor after surgery a trend for better OS was seen for PD-1 expression . A partially significant effect was also seen in the subgroup without residual tumor after surgery .Gene expression datasets form The Cancer Genome Atlas project on primary high-grade serous ovarian carcinomas  were anavs > 60 years) and stage (II vs III/IV). Similar results were obtained for OS (not shown).We additionally investigated the prognostic value of PD-1 and PD-L1 expression only for patients that had documented platinum-based chemotherapy. PD-1 and PD-L1 expression, both in cancer cells and TILs as well as mRNA expression were significant prognostic factors for PFS in this well-defined subgroup, too, and significance was retained for all but PD-L1+ TILs and PD-L1/CD3 TILs combination in multivariate analysis . Data foIn this study, we systematically investigated PD-1 and PD-L1 expression in primary high-grade serous ovarian carcinoma. High expression in cancer cells and mRNA expression were favorable prognostic factors, as was the density of PD-1+ and PD-L1+ TILs, moreover, this favorable prognostic effect of TILs was independent from the T cell infiltrate in general. We validated our findings on the mRNA level in the independent cohort of high-grade serous carcinomas from TCGA.n = 12 PD-1+ TILs/mm2 (tumor and stroma not separated), our study: n = 12 PD-1+ TILs/mm2 , and it is remarkable that this small subpopulation of T cells obviously has a biological significance as measured by patient prognosis. This rather surprising finding prompted us to investigate whether the positive prognostic effect of PD-1+ and PD-L1 TILs might be a bystander effect of the intratumoral T cell infiltration in general. The absolute counts of PD-1 and PD-L1 TILs in high-grade serous carcinoma were strongly associated with CD3+ TILs, but only a small fraction of T cells actually expressed PD-1 and PD-L1 and their amount seemed to increase proportionally to the intratumoral immune reaction - as reflected by the density of the lymphocyte infiltrate - potentially in terms of a feedback activation. We have made similar observations in breast cancer, where we found that mRNA levels of a plethora of immune-related genes, PD-1 and PD-L1 included, were positively correlated with each other as well as with the density of TILs, irrespective of their immune-promoting or immune-inhibitory function, and were all strong indicators of a higher response rate to neoadjuvant chemotherapy. [Our findings are in line with a recent report on a positive prognostic effect of PD-1+ TILs in a cohort of 195 high-grade serous carcinomas.  This stutherapy.  However,therapy. , 17, 18 therapy.  as well therapy.  play an vs γδ cells, an issue, which would be worth further investigations.An impact of PD-L1 on γδ T cells has been reported. One study investigated the expression and function of PD-1 in human γδ T cells that recognize phosphoantigens.  The resuThe localization of TILs seems to be of major relevance as to their prognostic impact. Our data showing a consistent positive impact of intratumoral  TILs on survival is in line with several previous reports. , 24, 25 Currently, the expression of PD-L1 on tumor cells is regarded as an immune-escape mechanism of the tumor, as it attracts PD-1 expressing immune-inhibitory TILs. However, this mechanism is expected to rather result into a negative impact of tumorcellular PD-L1 expression on survival, and this is reported, e.g. for breast cancer, NSCLC, renal cell carcinoma , osteosToday it can only be speculated why components of the PD-1 pathway are in some instances (such as in ovarian carcinoma) linked to a favorable prognosis. A positive impact of PD-L1 expression of tumor cells as seen in our study might be explained by a compensatory up-regulation of this marker in a microenvironment that threatens the tumor by an active immune response. An association between PD-L1 on tumor cells and a high TILs density would be an argument for this hypothesis, and has as well been described by Hamanishi et al. in their study on various ovarian cancer histotypes  as well vs any expression), although we are aware of the fact that the cutoff point for response prediction might be different. A recent phase I study on pembrolizumab in NSCLC for the first time validated a previously defined cutoff point of 50% for response prediction, however responses were also seen among patients with tumors with expression below the cutoff indicating that tumoral PD-L1 expression might not constitute the definite predictive marker. [n = 1). [The fact that PD-1 and PD-L1 are expressed in two tumoral compartments (cancer cells and TILs) and that this expression has prognostic impact strongly suggest that the PD-1/PD-L1 axis has a biological relevance in high-grade serous ovarian carcinoma. This entity therefore appears as a candidate malignancy in which PD-1/PD-L1 targeting drugs should be tested. Although these agents have shown remarkable effects in subsets of patients with in poor-prognosis and therapy-resistant cancers such as NSCLC  and rena marker.  For the  marker.  as well [n = 1).  Numbers Our study has several strengths and weaknesses: We only investigated primary tumors, but the current use of immune checkpoint inhibitors is focused on metastatic or recurrent tumors to date. However, the data of Powels et al. indicate that the immunological microenvironment of a tumor might be temporally stable in terms of its predictive value.  A comparTo summarize, we report a significant prognostic impact of PD-1 and PD-L1 expression in primary high-grade serous ovarian carcinoma. PD-1+ and PD-L1+ TILs increase proportionally to the general lymphocytic infiltrate, however carry independent prognostic information. PD-1 and PD-L1 expression in cancer cells as well as mRNA expression are favorable prognostic markers. Our data indicate that PD-1 and PD-L1 molecules are biologically relevant regulators of immune response in high-grade serous ovarian carcinoma, and that the evaluation of immune checkpoint-inhibiting drugs might be of value in this poor-prognosis cancer type, for which only limited options for targeted therapy are available to date.n = 165) patients had received surgery in the Department of Gynecology of the Charité and had been included into the TOC project . Scientific use of TOC and non-TOC cases has been approved by the ethics committee of the Charité. Data on residual tumor mass after surgery (applies for stage II-IV) were available for 152 patients (76.4%). The majority of patients had been treated with standard adjuvant platinum-based chemotherapy . Data on overall survival (OS) were available for all patients. Median OS was 37.9 months, 113 patients died during follow-up (52.6%). Data on progression-free survival (PFS) were available for 185 patients .Formalin-fixed and paraffin-embedded (FFPE) surgical specimens from 215 patients with primary ovarian high-grade serous carcinoma were used (Table +), as a chromogen. CD3, PD-1, and PD-L1 were stained on consecutive TMA sections enabling the evaluation of corresponding tumor areas. Specificity of the PD-1 and PD-L1 antibodies had been validated before by evaluation of the staining patterns in normal lymphoid tissue and lymphoproliferative diseases, for which a PD-1 or PD-L1 expression had been described . Stained slides were scanned and evaluated on screen by an experienced pathologist (CAK); doubtful cases were discussed with a senior hematopathologist (KJ) until consensus was achieved.Staining was performed on tissue microarrays (TMAs) with two cores for each case, according to standard procedures. In brief, a mouse monoclonal antibody against PD-1  was used in a dilution of 1:50. For PD-L1 detection, a rabbit monoclonal antibody was used in a dilution of 1:300 (clone EPR1161(2), Abcam plc., Cambridge, UK). PD-1 and PD-L1 staining was performed using the BondMax™ device . Antigen retrieval and visualisation of bound antibodies were performed employing the manufacturer's protocols and reagents . CD3 staining was performed using a rabbit polyclonal antibody in a dilution of 1:100 , CD4 was stained with a mouse monoclonal antibody in a 1:20 dilution  and CD8 was detected with a mouse monoclonal antibody , using the BenchMark XT device  and 3,3′-diaminoenzidine peroxide substrate . Intensity and rate of stained cells were determined by two pathologists (CAK in support of KJ) on screen for both membranous and cytoplasmic staining and were combined to a semi-quantitative immuno-reactivity score (IRS).  The scor2) could be assessed. For PD-1+ and PD-L1+ TILs the same tumor areas as for CD3+ TILs were evaluated accordingly. A screen-shot of CD3 evaluation in a representative case is shown in CD3+ TILs were evaluated first: 5 tumor areas in a 400x magnification (high power fields (HPF)) were screen-shotted with the use of VM Slide Explorer 2.2 (VMscope) considering preferentially areas with higher intratumoral TILs density. Microphotographs were subsequently evaluated using the ROI Manager software (CognitionMaster) . In eachp < 0.0001), PD-L1+ , as well as for PD-1+ TILs density . Membranous PD-L1 expression in tumor cells  also correlated in TMA and large sections . In the group of cases that were scored as positive on large sections  all had been classified as positive on TMAs, too. Two of the three cases that were scored as negative on large sections  had been scored positive on TMA, one with an IRS of 2, the second with an IRS of 3, which both are in the low-expression range. Using the novel PD-1 antibody in large sections, there was no PD-1 staining in cancer cells, therefore a comparison between TMA and large sections results as to cancer cell expression was not feasible for PD-1.10 cases with high and low TILs density, respectively, were stained on paired TMAs and large sections. Large sections were stained and evaluated in exactly the same way as TMA spots had been treated before. An exception was PD-1, for which another antibody was used  because the antibody we used before was not commercially available any more. There was a strong correlation between paired TMA and large section data for CD3+  in combination with a liquid handling robot . Two 5-μm thick sections were cut from each paraffin block and transferred to a 1.5-ml tube. All tumor samples included in the study contained at least 30% tumor tissue as evaluated by H&E staining (median tumor content 60%). To achieve this tumor content, manual microdissection of the tumor area was performed if necessary. Expression of PD-1, PD-L1 as well as the normalization gene RPL37A was assessed in triplicate using a ViiA™ 7 Real-Time PCR device . Sequences of primers and probes have been published earlier. deltaCT 2) were logarithmized using the formula: log10(TILs density + 1). For the determination of cutoff points, the Cutoff Finder online tool was applied (molpath.charite.de/cutoff) [Statistical analyses were performed with IBM SPSS Statistics 22 , and GraphPad Prism v.5 . Associations were tested by Spearman, Chi square, Kruskal-Wallis or Mann-Whitney test, as indicated. Survival analysis was performed using the Kaplan-Meier method or Cox regression. All p-values were calculated two-tailed and p values < 0.05 were considered as significant. For Cox regression using continuous TILs data, TILs density  .https://tcga-data.nci.nih.gov/tcga) and investigated for the prognostic impact of PD-1 and PD-L1 gene expression using the software package R. Inclusion criteria for the analysis were: serous histology, and a G2 to G4 grading.Gene expression datasets  from TCGA  were dow


People make a city, making each city as unique as the combination of its inhabitants. However, some cities are similar and some cities are inimitable. We examine the social structure of 10 different cities using Twitter data. Each city is decomposed to its communities. We show that in many cases one city can be thought of as an amalgamation of communities from another city. For example, we find the social network of Manchester is very similar to the social network of a virtual city of the same size, where the virtual city is composed of communities from the Bristol network. However, we cannot create Bristol from Manchester since Bristol contains communities with a social structure that are not present in Manchester. Some cities, such as Leeds, are outliers. That is, Leeds contains a particularly wide range of communities, meaning we cannot build a similar city from communities outside of Leeds. Comparing communities from different cities, and building virtual cities that are comparable to real cities, is a novel approach to understand social networks. This has implications when using social media to inform or advise residents of a city. Yet, despite a century of effort, our understanding of how cities evolve is still woefully inadequate . BeneathA city has many networks, for example the road, the gas, the electric and the main drain networks. However when we think of the character of a city, these physical networks are unlikely to be our immediate thought. Instead we characterize a city by the people within the city, and the social network that they form—the essence of any society. Despite previous fears that cities impair communities , the risOne can think of each city as a unique amalgamation of social communities that differentiate it from other cities. But how unique is each city? Would an amalgamation of social communities from one city resemble another city? This paper resamples the communities from one city to create virtual cities of any size. By comparing these virtual cities to real cities of a corresponding size, we establish that many cities do not conflict with the proposition that they are indeed made up of the same social building blocks. Alternatively, we may identify which cities are inimitable.Insight into the social structure of cities may be used to ensure success of media campaigns, such as those which aim to bring social cohesion or to inform, or other types of behavioural intervention. If a social campaign was successful in one city, it is also likely to be successful in cities comprising similar social building blocks. Moreover, it may prove economical to test a campaign in a smaller city before transposing the learnings gained to a larger city.Generally physical networks evolve alongside the development of a city. Cities develop from the bottom up, and grow to facilitate a division of labour that generates scale economies . This giSize is the major determinant of most characteristics of a typical city; history, geography and design have secondary roles ,9. UsingCommunity structure within communities has been studied for a long time (e.g. ), yet thet al.  of a graph defines how easily a graph can be separated into communities, where each node belongs to only one community. A modularity close to 1 indicates clear community structure, where as modularity close to 0 indicates a complete graph, which cannot be divided into communities. The authors are unaware of any work which relates the modularity of the communities of a network to the modularity of the network as a whole.The modularity R≥1 separate networks, say {Gs)( into communities. By considering the Gs)( is the ns×ns adjacency matrix for Gs) and we may write this asConsider a set of notation , with a G has exactly G, Q, must satisfyGs) within G, thereby connecting up some communities across the Gs). Thus, equation . When α>0 we have growth which follows a power law, and when α<0 the system becomes saturated.We might have deduced the form of equation  directlyNP hard problem, there may be circumstances where it is simpler to calculate the modularity of numerous smaller networks than one large network. We now use Erdos–Rényi graphs to test the lower bound (equation (Equation  is a gooequation ).3.Gs), for some probability p. Then G is just the union of these relatively highly connected ‘ghettos’ (the Gs). If we wish we could also add a very small number of edges so as to connect up a few vertices within the separate ghettos. Such a ghetto-model gives rise to a joint distribution for the triple . Suppose we generate R such graphs and calculate the modularity Q of the union G, using a chosen standard method, in this case one based on the successive bisection of communities [Let us assume that each munities ,16, as iR we may observe how Q increases. In fact we may generate an ensemble of such increasing virtual cities G, made up from R independent ghetto graphs, Gs). We may contrast equation . The envelope is at its widest at small and intermediate values where R<20 and is not so large as to provide a reliable sampling of the distribution assumed for the ghettos. For larger R, we see convergence of Q towards R copies of the averaged value for the s-dependent terms ).Then as we increase equation , by plugequation  are veryA, we may thus generate other virtual cities (by sampling from A’s communities with repetition) that are made of the same stuff, the same underlying distribution of component communities as City A, yet are of any size we chose. In particular, we may match the size of any other observed city, City B. So we shall be able to ask whether City B is similar or not to one made of the same underlying components as City A, while allowing for their differences in scale.After introducing some suitable dataset representing the digital social networks within distinct British cities in the next section, we shall apply the theory developed in this section. By using the community sub-structure observed from any particular city, City 4.We defined a reciprocated (Twitter) mentions network, over 10 UK cities and a four week period, as follows.We identified all those Twitter accounts that were active during 1–28 October 2014 (four full weeks), having biographical information indicating that they are associated with one of 10 cities: London, Manchester, Leeds, Sheffield, Nottingham, Birmingham, Bristol, Cardiff, Glasgow and Edinburgh. The data were collected, using the DataSift Twitter Firehose access, by selecting individuals who had one of the 10 cities stated in their user description field. (We did not look for geo-coded Tweets because the coverage would be too sparse for it to be useful.) We trapped all Tweets during that period . This resulted in 14 946 171 original Tweets made by 321 255 individual accounts.The giant components from using 14 days of data are not sufficiently different from the corresponding giant components using 28 days of data, and a diminishing returns effect becomes apparent. Nonetheless, when analysing a social network a longer time period is advantageous. However, for our purposes of demonstrating our method, 28 days is sufficient.Next we identified a subset consisting 7 241 148 of the Tweets that contained a mention of another account (@name). These Tweets were sent by 260 439 accounts.Finally, we selected the subset of accounts that mentioned any other of the accounts for which there was also a reciprocal mention. That is, those accounts, say Xname, that sent at least one Tweet containing a mention of another account, say Yname (a mention @Yname), where there is at least one reciprocal Tweet made by Yname containing a mention of Xname (a mention @Xname). This resulted in the selection of 59 789 accounts, with each having a reciprocated mention of at least one other account. Representing those accounts as vertices, and the reciprocated mentions as undirected, unweighted edges, we thus obtained a 10 city-based Twitter reciprocated mentions network defined for October 2014. It has average degree of 2.36.The employment of reciprocated mentions  shows that the accounts not only are aware of each other but have actively engaged with each other during the prescribed interval of time (28 days). In that sense the undirected network contains validated (active), two-way connections. The full dataset for this network is available in the electronic supplementary material, where the first column relates to the city , and the other two columns indicate an edge between the two listed nodes.Each of the accounts is associated with at least one city. In many cases, they may be associated with more than one city. For example, they might state ‘I am from Manchester, working in London’ implying they may be genuinely connected into more than one city-based social network. In fact out of the 59 789 accounts we have, 29 814, 16 935, 10 680, 6236, 3949, 5890, 6043, 5644, 4830 and 4061 accounts were associated with London, Manchester, Leeds, Sheffield, Birmingham, Nottingham, Cardiff, Bristol, Glasgow and Edinburgh, respectively. So on average each account claims to be associated with 1.57 cities.We may restrict the mentions network to each separate city as follows. First, we examine the subnetwork containing those accounts associated with each city. We only considered accounts that are active within a particular city, so if an account has reciprocated mentions with another account within that city, it is included within the corresponding network. This results in 10 city-based reciprocated mentions networks. Each of the city-based mentions networks contains a single giant connected component and a very large number of smaller connected components (with less than a handful of vertices).In this paper, we shall focus on these giant components as being representative of the nature of the digital social network for each city, as represented by the corresponding reciprocated Twitter mentions network. Details of the giant component for each city are described in Q. These values are relatively high compared with the modularities of random graphs conditioned solely on the corresponding degree distributions. In line with intuition, London is exceptional. The lower bound for modularity (equation (Each city may be partitioned into separate communities by the optimization of modularity. In equation ) is consQ) [There is a number of heuristic methodologies that have been designed to produce the optimal partition  –17. In aR=74. We show both the modularity and the Shannon partition entropy  as a fraction of their optimal values obtained at 74 communities. The behaviour is very similar to that observed in Onnela et al. [Q was taken while introducing a degree of freedom to control the relative scale of discrimination . The modularity is almost optimal when k=20 or so, with the remaining work, in splitting off smaller communities, representing some fine-tuning. Since modularity is not very sensitive, assuming equality within equations . If we resample each one of them exactly once and then take the union we arrive at a city very similar to City A itself, that is just missing the relatively small number of edges connecting across sub-communities . Now let us resample from X, independently with repetition for as many times, R, as we wish. For each value of R, we may obtain an ensemble of virtual cities (an ensemble where each such city contains exactly R sub-communities), that are made up of the same types and distribution of communities as we observed in City A. As with bootstrapping, we use the observed distribution, X, to represent our best sampled estimate for the underlying (and unobserved) distribution A are composed.The idea employed here is very similar to that of bootstrapping . We haveA has very few identifiable communities (a low resolution partition) forming up X, then it will be a poor representation of the underlying possibilities in X increases, we may see much more diverse behaviour inherited from Of course, if City R sub-communities from X and take their union to form-up a new city. The sth sub-community that we sample has associated observables . Let X. ThenSuppose that we independently resample exactly R=|X| then we are constructing virtual cities of a similar size to the conditioning City A, and we may create an ensemble of such virtual cities that will include the conditioning city. Now consider another city, say City B, that is observed to have nB vertices. Set R draws from X, randomly with replacement, we may construct an ensemble of virtual cities that are all approximately the same size as City B, yet that are made up of components from the same distribution, X, as City A. Thus, the deviation or otherwise of measures for B, such as Q, from the distribution for the same measure obtained from this virtual ensemble can indicate whether or not it is reasonable to assume that B may be made up of the same distribution of sub-communities as A.From , assuminA. On top of the ensemble envelope of Q-values obtained for all of the virtual cities we show the other observed cities. The size of the envelope, which is generally proportional to the number of communities R, is a guide as to the variety of communities within a city. For example, Glasgow and Edinburgh have small envelopes indicating fairly homogeneous communities. Therefore, a city with a wider envelope, such as Manchester, contains social structures not seen in Glasgow and Edinburgh.In Consider Bristol as the conditioning city. It is clear that while Manchester, Nottingham, Edinburgh and Birmingham are within the equivalent Bristol-like envelope, Leeds, especially, is not. We would contend that the social structure observed within Leeds is not consistent with that observed within Bristol; even when we allow for changes of scale, there must be some specific elements that are present or absent on one or another. Thus, the performance of any interventions or other activities designed to work within such social networks cannot be extrapolated from Bristol to Leeds. However, since Leeds contains a wider variety of social structures, a Bristol-like city could be created from Leeds communities. The transitive properties from one city to another, excluding London, are summed up in 6.NP hard problem. Therefore, representing the modularity of a graph in terms of the modularity of the communities is computationally advantageous.We have presented a way to estimate the approximate maximum modularity of a large network based on the maximum modularities of the communities within the network (independent of the method used to define communities).The lower bound estimate presented here is useful because modularity lacks sensitivity near the optimal resolution. Maximizing modularity is an We presented the null model where we built a large city from ghettos represented as a combination of Erdos–Rényi graphs. We showed that the modularity converges after the city had been divided into approximately 20 communities, irrespective of the total number of communities.We represented cities as combinations of communities from different cities, the ‘conditioning city’. This showed many cities, such as Bristol, Nottingham, Edinburgh, Manchester and Birmingham, comprise communities with similar structure—despite their size and location varying considerably. Conversely, it also identified cities like Leeds, which are not made up of similar communities to other cities we sampled. Leeds, despite being smaller than Manchester, contains social structures not observed in Manchester. At the other end of the variance spectrum, Edinburgh and Glasgow have very little variance between their communities. Therefore, recreating cities with greater community variance is challenging from these limited pools of Scottish communities. This type of analysis can aid decisions about social media activities and interventions. For example, a campaign that is successful in Bristol should also be successful in Manchester (but not Leeds). Since Bristol is considerably smaller, it may prove useful to test the campaign in Bristol first.The method is not limited to Twitter data. One could apply the same method to any form of network that could define a city, such as Facebook or e-mail. Furthermore, we are not restricted to cities. It would be interesting to compare universities to each other, or countries, or the social structure in different governments. The method is only limited by our imaginations and the data available.


Large size graphene (LSG) and multiwall carbon nanotubes (MWCNTs) on LSG were synthesized on a copper surface via chemical vapor deposition (CVD) at low temperature and normal pressure. The LSG were formed through an easy chemical cyclic reaction in which liquid benzene was heated to a temperature below its boiling point to create benzene vapors as graphene precursor material. The reaction mechanism was observed, and the time-dependent analysis of the reaction revealed that mounds of the carbon nanotubes had grown as a result of the island that was found on the LSG sheet. The implications of the mechanism that we have introduced were investigated by coating a titanium sheet on the MWCNTs/LSG and LSG on the semiconductor electronic device. The photonic response was observed to be markedly high, which can be attributed to the positive synergetic effect between the Ti and LSG sheet of our prepared composites. Graphene consists of a single layer of carbon atoms that can be obtained from highly-oriented pyrolytic graphite through mechanical exfoliation, and it has been shown to have outstanding electronic, optical, and mechanical properties. Although various techniques have been developed to produce graphene on a solid material substrate1234678911et al. studied the growth of epitaxial graphene (EG) on insulating silicon carbide (SiC) surfaces by developing a promising route to produce large-scale high-quality graphene. The amorphous carbide precursor has been transformed to graphene nanosheets through chlorination at low temperature and ambient pressure, and some exceptional outcomes have been obtained. These results suggest the importance of the size effect when synthesizing CVD grown graphene on metal substrates16171819CVD is a relatively simple technique that can be used to obtain graphene with the desired features. CVD involves the decomposition of carbon precursor materials, that is, hydrocarbons or polymeric components, by adding heat and some metallic catalysts, such as Ni, Pt, Cu, etc., that have been found to improve the growth of graphene. The optimum conditions to produce large size graphene through CVD have yet to be determined, and therefore, various approaches have been proposed14In this study, we investigate a novel CVD growth process for LSG/MWCNTs on copper foil at low temperature of 500 °C. This procedure consists of two steps. First, we observe a large size micrometer graphene sheet on Cu foil at the appropriate temperature, and in the second step, we develop the growth mechanism for LSG and MWCNTs/LSG at very low temperature. We thus suggest a novel growth mechanism for LSG and MWCNTs/LSG through chemical vapor deposition on a copper foil at low temperature. The novel mass production of LSG and MWCNTs/LSG on Cu foil may be very useful in the fabrication of ultrafast charge transfer for next generation optical and electronic devices. Finally, we suggest a potential application by coating Ti on the MWCNTs/LSG on Cu foil and coating LSG on electronic devices.FESEM images of the as-prepared CVD-grown LSG and MWCNTs/LSG assembly are shown in When the reaction time increases to 2 and half minutes by keeping the temperature at 500 °C, a small dot appears on the surface of the LSG, and these dots are referred to as carbon nanotube seeds, as shown in The production of large area graphene and the number of layers of the G1 sample were investigated through the TEM image shown in The production of MWCNTs on the LSG sheet was verified through a transmission electron microscopy (TEM) analysis of the G4 sample. −1 from the excitation line are associated with the Cu present in the sample. Cu arises as a result of scratching the graphene composite from the Cu foil.Raman spectroscopy is a powerful technique that can be used to study the structure and electronic properties of carbon materials, including graphene/graphite and carbon nanotubes. −1 that corresponds to the 2D peak in graphene. This 2D peak was further split into four Raman peaks, and the dispersion of the peak yielded information on the number of layers and the stacking order in the graphene system. 2D peaks in bilayer graphene have been reported to be split into four components, which further increases in the layer and bulk area of graphene and leads to lower intensity peaks22The results of our study indicate that by increasing the reaction time, the Raman band intensities of the G band and D band shift towards a lower frequency. This allows the MWCNTs to start growing on the LSG sheet. The other reason for the variation in intensity is the large number of MWCNTs that roll together, encouraging phonon confinement and thus reducing the intensity24−1 while the D band frequency was observed at 1352 cm−1. For the MWCNTs, the large number of CNTs with different diameters affects the characteristic G band, and therefore theses are found close to the G band frequency of graphite−1 were attributed to the Cu that was used as a substrate3132The laser excitation also has an effect on the Raman band shift and the variation in intensity. In the MWCNTs, the bands could be attributed to the Raman resonance condition, and the incident laser on the carbon nanotubes can yield an optical resonance profile for a given mode at a vibrating frequency of the respective modes2627283435We then used AFM topography images to determine the thickness and the height of the LSG and MWCNTs/LSG composites. The LSG and MWCNTs/LSG composites are shown in The surface growth of vertically-aligned CNTs on the substrate material at a micrometer scale with nonmetric effect is very important for next generation devices. The AFM images revealed the surface construction of MWCNTs leading towards nanohill formation of carbon nanotubes, which is unique, and this formation is a complex phenomenon that may be the result of the different competing factors, such as temperature, reducing atmosphere, oxide reduction during the CVD process, and benzene decomposition. This elusive, complex phenomenon will require focused investigation in the future to explore the outstanding physical and chemical properties of the MWCNTs/LSG composites.The The scheme to form the LSG and MWCNTs is depicted in The samples were coated by titanium through the same process that was explained above by using TNB as a precursor material. During the CVD process, we observed that the Ti attached as a sheet similar to graphene that covered the maximum surface of the graphene samples. −1. The peak at around 1100 cm−1 can be attributed to the Ti lattice td symmetryThe Raman spectrum of the Ti coated G4 sample is shown in The photo-catalytic properties of the LSG/MWCNTs and Ti-MWCNTs/LSG that were obtained can be examined by using Rh.B degradation under visible light. The scratch sample from the copper foil was used as powder for the photocatalytic experiment. The Rh.B photodegradation rate was assessed under visible light . The photo activities of the LSG, MWCNTs/LSG and Ti-MWCNTs/LSG were determined according to the degradation rate of Rh.B as a function of time under visible irradiation. In order to attain adsorption/desorption equilibrium, the solution was kept in the dark for 2 h. After adsorption/desorption equilibrium had been achieved, the solution was irradiated with visible light for 30 minutes. For the analysis, the sample was periodically withdrawn, and the solids were removed via centrifugation.The dye concentration of the sample was then analyzed using a UV-visible spectrometer. Under visible light, the energy state of the electrons changed from a valance band to a conduction band, and the excited electrons reacted with the adsorbed oxygen, resulting in the formation of free radicals. These free radicals reacted with the dye, resulting in the mineralization of the dye into simple molecules. During the experiments, a decrease in the color intensity of the solution was gradually observed over time due to the adsorption and degradation of Rh.B. The Rh.B concentration in the solution was determined to be a function of the irradiation time from the change in absorbance at a wavelength of 554 nm. The results of the photo degradation  are shown in the app). The Kapp values provide the degradation rate of the Rh.B molecules by the photocatalyst materials under the influence of visible light. The values of the Kapp rate constant are listed in −3 min−1 under visible light, which was much higher than the G1 and G4 sample in our nanocomposites. These results further confirm that the Ti-MWCNTs/LSG nanocomposites are a much more effective catalyst material than the other nanocomposites in the CVD-grown LSG/MWCNTs nanocomposites.Kinetic studies were performed on the basis of the degradation rate of the organic dye, and the reactions between the dye molecules and the catalyst materials can be expressed according to the Langmuir- Hinshelwood model, as shown in et al. synthesized CNTs-Fe/Ni-TiO2 composites, and an improvement was observed in the catalytic effect due to the role of the CNTs in the compositeset al. studied the properties of CNTs-TiO2/Al2O3 membranes in terms of their photocatalytic function and provided a significant description of the use of CNTs-based materials for purification systemsThe photocatalytic properties of the CNT composites with other semiconductor materials have been extensively studied over the last two decades, and the effect of the CNTs has been made quite clear from these investigations. For example, Lei To further demonstrate the photo-stability and cyclic performance of the Ti-MWCNTs/LSG composite photo-catalysts, cyclic experiments were carried out with Rh.B as an organic dye. A semiconductor index device was considered for coating with our proposed CVD method. The actual image of the device is shown in 4344452. This high level activity may be a result of the positive synergy and high charge mobility of the MWCNTs between the Ti and graphene sheet. Thus, we believe that our low-temperature, normal-pressure CVD process provides a simple and reproducible way to produce high quality Ti-LSG/MWCNTs for the large-scale integration of next generation graphene-based devices.In summary, we have investigated a new, short route to produce large area graphene. This method can be used to produce large area graphene for industry at bulk by further controlling the experimental parameters to determine the optimum values. We demonstrate the production of large area graphene with a very short reaction time in an argon environment by using chemical vapor deposition. An increase in the reaction time to 5 minutes provides interesting results due to the production of ~350 nm long, vertically-aligned MWCNTs that are homogenously distributed through the copper foil, as shown in the AFM images. The FESEM images show that that the island found on the graphene can further give rise to vertical CNTs. The valleys that are observed between the graphene domains may be a result of the use and handling of a very thin copper foil. The UV-visible spectrum clearly shows an enhanced catalytic effect for the Ti-MWCNTs/LSG nanocomposite under visible light, suggesting an optimal loading effect of graphene to TiO16H36O4Ti) was used as a titanium precursor and was purchased from Samchun Pure Chemical Co. Ltd., Korea. The Cu foil (99.9%) was annealed and uncoated and was used as the substrate material to grow LSG/MWCNTs. Argon and Nitrogen gas were purchased from Samchun Pure Chemical Co. Ltd., Korea. A specially-designed split Si tube furnace was divided into two parts: an inner tube and an outer tube. The dimensions of the outer tube were 30 cm length and 4 cm with two nozzles, one for the inlet of benzene vapors and the other for the inlet of Ar and N2 gas. The inner furnace, which is also called the heating zone, had a 10-cm length and a 5-cm diameter and was used to grow the MWCNTs/LSG on the Cu foil. Benzene was first heated below its boiling temperature to produce vapors, and these vapors were then simultaneously transferred to the tube with Ar gas, which also acts as a carrier for the benzene vapors and prevents the Cu foil from reacting with water molecules. The vapor flow and the Ar gas ratio were controlled by using control valve, and the furnace temperature increased to 500 °C in the first step, and Ar gas was released into the Si furnace. After reaching a temperature of around 500 °C, a Cu foil was inserted in the inner tube, followed by a controlled amount of Ar gas and benzene vapors. The growth of the MWCNTs/LSG was verified at different reaction times  at 500 °C, and the samples that were obtained were labeled as G1, G2, G3 and G4, respectively.Benzene was used as a carbon precursor material and was purchased from Dae-Jung Chemical and Metals Co. Ltd Korea. Titanium (IV) n-butoxide -5200 JOEL, Japan) was used because it offers expanded imaging capabilities and is customizable in terms of the performance requirements for the analysis. High-resolution transmission electron microscopy (HRTEM) JEOL, JEM-2012, Japan was used to determine the state and the sheet morphology of the prepared graphene. A transmission electron microscope (TEM) at an acceleration voltage of 200 kV was used to investigate the number and the stacking state of the graphene layers on the various samples. Atomic force microscopy (AFM) was then carried out using Park NX 20, Korea.How to cite this article: Ullah, K. and Oh, W.-C. Fabrication of large size graphene and Ti- MWCNTs/ large size graphene composites: their photocatalytic properties and potential application. Sci. Rep.5, 14242; doi: 10.1038/srep14242 (2015).


Annexin A11 is one of the oldest vertebrate annexins that has a crucial role in sarcoidosis pathogenesis. The mechanism of effect in sarcoidosis granuloma cells may be due to alterations in apoptosis. Immune cells with a specific mutation at protein location 230 are resistant to apoptosis and consequently have continued effects on inflammation and progression of sarcoidosis. The mechanism of action of annexin A11 may be based upon alterations in delivering calcium to two different apoptosis pathways (caspase and P53).Annexins are Ca They are widely distributed among eukaryotes but largely absent in prokaryotes and yeasts. The first protein of this family, discovered in 1977, was called synexin. Currently known as annexin A7, it caused aggregation of chromaffin granules in the adrenal glands in the presence of free calcium . The medmeeting” . Annexinmeeting” .A11 was linked to increasing susceptibility to sarcoidosis. It was also suggested that cells with mutant annexin A11 might have altered susceptibility to apoptosis. However, the mechanism of this effect has never been discussed. We propose here a molecular mechanism for this association.There is little information about annexin A11 and many of its structural and functional characteristics are speculated based on similarity with other members of annexin family. Recently, genetic mutation in annexin annexin genes [ANX genes with mammalian ANX genes shows that three zebrafish ANX genes are homologous with human ANX1; two are homologous with human ANX2; and two are homologous with human ANX11. This information suggests that zebrafish ANX genes may have resulted from duplications after the divergence of the zebrafish and mammalian genomes [More than a 100 annexins have been identified in many different species . Twelve in genes  that are genomes .ANXA9) to 96 kb (ANXA10) and are spread throughout the genome on chromosomes 1, 2, 4, 5, 8, 9, 10, and 15 [ANXA5 and ANXA10 genes in early vertebrate evolution. The reasons for the annexin genes or their chromosomal regions to duplicate are not well understood. Their successful preservation and the extent to which they contribute to vertebrate complexity are also not well described [The 12 human annexin genes range in size from 15 kb ANXA to 96 kb6 kb ANXA and are ANXA11 gene is located on human chromosome 10q22–q23 and is composed of 15 exons and 14 introns without the 5′ flanking region [The annexins A1, A2, A4, A5, A6, A7, and A11) to selective, such as annexin A3 in neutrophils, annexin A8 in the placenta and skin, annexin A9 in the tongue, annexin A10 in the stomach and annexin A13 in the small intestine [ANXA11 has the highest gene expression in whole blood cells, particularly CD19+ cells (B-cells), CD14+ cells (monocytes) and CD33+ cells (myeloid). However, it is found in almost all tissues including lung, heart, and intestines [Annexin gene expression levels within human organs have a broad range, from universal . Some annexins, such as annexins A11 and A2, have been found in the nucleus under particular circumstances in the cell cycle , 17. It In some circumstances, annexins can be expressed at the cell surface even without a secretory signal peptide. For example, annexin A1 translocates from the cytosol to the cell surface following exposure of cells to glucocorticoids . In a stAnnexin A2 expression at surface of vascular endothelial cells has a function in the regulation of plasmin generation . It has Annexin A11 distribution changes per cell cycle. It is more prevalent in the nucleus than the cytoplasm in interphase cells and then moves to degenerating nuclear envelope. And finally during the mitotic phase, it will be concentrated within the central spindle . It has S100 proteins, which only express in vertebrates, are a well-known group of proteins that interact with the annexins. The functions of S100 proteins are diverse and include regulating actin and microtubule networks, promoting cell survival and proliferation, calcium homeostasis, and mediating muscle contraction. The complex of annexin A2-S100A10 interacts with several membrane ion channels, such as transient receptor like potential vanilloid type 5 and 6 channels (TRPV5 and TRPV6). The complex also interacts with cystic fibrosis conductance regulator protein (CFTR) and plays a role the regulation of these ion channels , 34. It 2+ to S100A6 and apoptosis-linked gene2 (ALG-2) whose protein augments apoptosis. The significance of these interactions in the pathogenesis of sarcoidosis is discussed later in this article.Annexin A11 is involved in cellular apoptotic processes. The N-terminal domain of ANXA11 contains binding sites that deliver CaANXA1 causes changes in the inflammatory response and the effects of glucocorticoids [ANXA1 null mouse line, there was altered expression of other annexins as well as cyclooxygenase-2 and cytoplasmic phospholipase A2. In addition, there was an exaggerated response to the stimuli characterized by an increase in leukocyte emigration and IL-1β generation and a partial or complete resistance to the anti-inflammatory effects of glucocorticoids [Studies on knockout mice of annexin families show a diversity of functions among these proteins. Loss of rticoids . In the rticoids .Data supports the role of annexin 2 as a regulator of cell surface plasmin generation, fibrin homeostasis, and neovascularization in laboratory mice models . HomozygANXA7 knockout mouse. The viability of the ANXA7 null mouse was compared to the heterozygous mouse [ANXA7, was viable and able to reproduce [Another study involved the formation of the us mouse . The ANXeproduce .ANXA7 null mutant mouse proposed ANXA7 function in the fusion of vesicles as a calcium channel [ANXA7 null mice were studied. When stimulated with high frequencies, the cells showed an altered cell shortening relationship. Possibly through its role in calcium regulation, this study suggested a function for annexin A7 in electromechanical coupling [Another study of  channel . Cardiomcoupling . The othANXA2 gene polymorphism was higher in the sickle cell patients compared to controls. The SNP was also present in higher frequency in sickle cell osteonecrosis patients than those without osteonecrosis [Annexins have essential roles in the pathogenesis or progression of many human diseases. Recent genetic studies discovered single nucleotide polymorphisms (SNPs) in the genomes of this group of proteins. In a study from India, annexin A2 gene SNP (rs7170178) was found to be associated with osteonecrosis in sickle cell patients . The frenecrosis .ANXA5 gene was sequenced in 243 Japanese women with recurrent pregnancy loss and 119 fertile controls [ANXA5 gene SNPs, the carrier frequency for the minor allele was significantly higher in the pregnancy loss group [In another study from Japan, the annexin A5 gene polymorphism was found to be associated with recurrent pregnancy loss. The promoter region of the controls . In a cass group . For SNPss group .Annexins are also associated with autoimmune disorders. In rheumatoid arthritis, high extracellular annexin V levels initiates the production of annexin V autoantibodies that may have a crucial role in pathogenesis of disease . SystemiDysregulation of Annexin A11 has been found in cancer, cancer treatment, and diabetes . For exaSome members of the annexin family may also be used as biomarkers and for clinical imaging. For example, ANXA1 was investigated as a potential serum biomarker for lung cancer. Lung cancer tissues exhibited higher expression of annexin A1 than normal tissues. In addition, increased serum annexin A1 was significantly associated with pathologic grade and clinical stage of lung cancer patients . QuantitSarcoidosis is a systemic immune disorder with a characteristic accumulation of epithelioid granulomas in many organs, such as the lungs, kidney, skin and eyes , 51. Chr+ and CD19+, immune cells involved in sarcoidosis, are proposed mechanisms for sarcoidosis [ANXA11 (annexin A11) gene on chromosome 10q22.3. A common nonsynonymous SNP (rs104955) was found to be strongly associated with sarcoidosis. As it is demonstrated in Fig. ANXA11 R230C SNP were more resistant to apoptosis than the wild genotype. This association was particularly prominent in subjects with the TT ANXA11 phenotype [2+ dependent. Without calcium delivery from ANXA11 to ALG-2, the apoptosis via caspase pathway would not be activated [Genetic instability and mutation in annexin A11 has been identified in single nucleotide polymorphisms in patients with sarcoidosis compared to control groups. Decreased activation of CD8coidosis . In addicoidosis . The strhenotype . The mec SNP rs10955 was fhenotype . The MAPctivated .Fig. 3AN2+ delivery from ANXA11 [The second important protein involved in apoptosis is P53 . P53 medm ANXA11 . Indeed,m ANXA11 . With a m ANXA11 . Figure In the sarcoidosis lung, granulomas develop when the immune cells attempt to wall off an unknown antigen trigger. These cells include macrophages, lymphocytes, and multinucleated giant cells. The granuloma can be effaced by apoptosis leaving only minor scar. Without apoptosis the inflammatory and granulomatous reaction continues. Patients with ANXA11 R230C cannot clear granulomas due to altered apoptosis and may have a higher risk of onset of sarcoidosis and poorer outcome Fig. .Fig. 6DeHofmann et al. were the first group of researchers to identify the association of ANXA11 with sarcoidosis . More reIn a 2013 study, the role of annexin A11 in sarcoidosis was confirmed in a US population. This association also extended to clinical manifestation of disease. The studied involved 1689 sarcoidosis cases and 1252 controls. 25 SNPs in and around ANXA11 were significantly associated with sarcoidosis risk in both African Americans and European Americans. The most significant allelic association was the minor C allele at rs1049550 among African and European Americans . This waThese recent advances of annexin A11 in the pre-clinical stage show its significant role in the pathogenesis of sarcoidosis. In the era of personalized medicine, the annexin could be a novel focus. Annexin A11 thus may be a target for clinical trials in which a patient’s genetic risk factors are used to tailor treatment .Annexins have significant effects on human health and disease. Annexin A11 affects two apoptosis related pathways (caspase and P53) and has a crucial role in sarcoidosis pathogenesis. This theory should be tested in cell culture and animal models. Recent studies show the significant role of annexin A11 in the genetic susceptibility to sarcoidosis. If the ANXA11 could increase susceptibility to sarcoiodosis via interaction with caspase and P53, a new opportunity to develop new therapeutic targets in sarcoidosis will be recognized.


A simple one-pot process to exfoliate graphite and synthesize nanographene-dimesitylporphyrin hybrids has been developed. Despite the bulky mesityl groups, which are expected to hinder the efficient π–π stacking between the porphyrin core and graphene, the liquid-phase exfoliation of graphite is significantly favored by the presence of the porphyrins. Metallation of the porphyrin further enhances this effect. The resulting graphene/porphyrin hybrids were characterized by spectroscopy  and microscopy . The −1) to NMP-exfoliated graphene (86 cm−1). Significantly, this effect is more pronounced in FLG/DMPP (78 cm−1) and even more in FLG/ZnMPP (75 cm−1). These results indicate that not only the yield, but also the efficiency of the exfoliation process is enhanced upon addition of the porphyrins. Meanwhile, the 2D band intensity diminishes due to the electron doping of graphene by porphyrin molecules, as corroborated by the significant shifts in the frequency of the G band and the decrease of the 2D/G ratio . Graphite powder  and all reagents were purchased from Aldrich . Solvents were purchased from commercial suppliers and used as received. UV-visible spectra of three different samples of each graphene solution were measured by using a Cary 50 UV-visible spectrophotometer  in quartz cuvettes with a pathlength of 1 cm. Fluorescence experiments were carried out at least three times for each sample. Emission spectra were recorded on a NanoLog 4 HORIBA . Raman spectra were acquired with a Bruker Senterra confocal Raman microscopy instrument , at excitation laser wavelength of 532 nm. The laser power at the sample was 10 mW, and the data acquisition time was 20 s. The sample preparation involved drop casting of the solutions on a Si substrate with a 300 nm oxide layer. STEM images were obtained with an Auriga Carl ZEISS operating at 25 kV . The FLG/porphyrin hybrids were deposited by drop casting of the dispersions onto holey carbon mesh grid. −1. The solutions were sonicated  for various periods from 60 to 240 min. In order to exclude the effect of temperature and to avoid the degradation of the solvent due to an increase of the bath water during sonication, a cooling system was connected to the bath sonicator performing the experiments at 20 °C. At different times during this period, aliquots of the suspensions were centrifuged at 3000 rpm for 30 min . After that, the supernatant was carefully collected for further characterization. For FLG/porphyrin hybrids, the same procedure was followed but graphite was added to solutions of 5,15-dimesityl-10,20-diphenylporphyrin (0.01 mg·mL−1 in NMP) and Zn-5,15-dimesityl-10,20-diphenylporphyrin (0.01 mg·mL−1). In this case, the supernatant of FLG/porphyrin hybrids was directly measured as obtained.Graphene dispersions were prepared via a liquid-phase exfoliation technique by adding graphite powder to 1-methyl-2-pyrrolidone (NMP) at a concentration of 1 mg·mLThe one-pot exfoliation of graphite and formation of FLG/DMPP hybrids has been demonstrated. The presence of the two bulky mesityl groups in the porphyrins does not impede their interaction with FLG. In fact, UV-vis measurements show that the presence of the porphyrins enhances the concentration of exfoliated graphene by up to 140%. We have also observed a very significant effect of the metal core of the porphyrin on the interaction with the surface of graphene. Our results show that the decrease in the efficiency of π–π interactions between the aromatic core of porphyrins and FLG can be compensated for by adequate CH–π and van der Waals interactions. This expands the range of porphyrins that can be used to synthesize FLG/porphyrin hybrids to include those with bulky substituents in the meso positions, which in turn should facilitate their use in electronic devices.


Two hundred and twenty-three CRC patient specimens were used to determine both BDNF and TrkB mRNA levels. The expression of these proteins in their primary and metastatic tumors was investigated by immunohistochemistry. CRC cell lines and recombinant BDNF and K252a  were used for in vitro cell viability, migration, invasion, anoikis resistance and in vivo peritoneal metastasis assays. Tissue BDNF mRNA was associated with liver and peritoneal metastasis. Tissue TrkB mRNA was also associated with lymph node metastasis. The co-expression of BDNF and TrkB was associated with liver and peritoneal metastasis. Patients with higher BDNF, TrkB, and co-expression of BDNF and TrkB had a significantly poor prognosis. BDNF increased tumor cell viability, migration, invasion and inhibited anoikis in the TrkB-expressing CRC cell lines. These effects were suppressed by K252a. In mice injected with DLD1 co-expressing BDNF and TrkB, and subsequently treated with K252a, peritoneal metastatic nodules was found to be reduced, as compared with control mice. BDNF/TrkB signaling may thus be a potential target for treating peritoneal carcinomatosis arising from colorectal cancer.Tropomyosin-related receptor kinase B (TrkB) signaling, stimulated by brain-derived neurotrophic factor (BDNF) ligand, promotes tumor progression, and is related to the poor prognosis of various malignancies. We sought to examine the clinical relevance of BDNF/TrkB expression in colorectal cancer (CRC) tissues, its prognostic value for CRC patients, and its therapeutic potential  The outstanding progress in the treatment of metastatic colorectal cancer (mCRC) has been founded on the recent multi-drug combination chemotherapies, which now produce median survival times exceeding 20 months NTR) with low affinity NTR which exerts diverse functions such as the regulation of cell survival and differentiation during neuronal development Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin (NT) family. BDNF plays an important role in the development and repair of the nervous system NTR involved in proliferation, differentiation, survival and apoptosis of neuronal and non-neuronal tumors NTR preferentially acts as an interacting partner of TrkB, modulates TrkB activation by BDNF, and influences prosurvival effect by BDNF/TrkB signaling Although both TrkB and p75NTR seems to have either tumor-promoting or tumor-suppressing functions according to tumor types BDNF/TrkB signaling has been reported to be associated with tumor progression, metastasis, and response to chemotherapy in several human malignancies such as neuroblastoma Previously, studies in our laboratory have revealed: an association between TrkB levels tumor progression and patient prognosis in gastric cancer In vitro studies showed that BDNF/TrkB signaling was involved in proliferative or invasive properties With regard to the BDNF/TrkB signaling in CRC, BDNF or TrkB was overexpressed in both clinical tumor samples and associated with aggressive tumor phenotypes The aim of this study was to examine: the association between BDNF/TrkB expression and clinicopathological variables in a series of human CRC tissues; the prognostic value of BDNF/TrkB signaling in CRC patients; and its therapeutic potential in vitro and in vivo.This study was reviewed and approved by the Institutional Review Board and the Local Ethics Committee of the Mie University Graduate School of Medicine (No. 2126). Written informed consent was obtained from all the patients (adults and parents of children) enrolled onto the study.The experimental protocols of in vivo studies were reviewed and approved by the Animal Care and Use Committee at the Mie University Graduate School of Medicine.A total of 223 patients with CRC, who were treated at the Department of Gastrointestinal and Pediatric Surgery in the Mie University Graduate School of Medicine from 2000 to 2008, were included in this study. Patients with incomplete clinical data, inadequate follow-up or inadequate tissue sampling were excluded from the study. All patients had histologically confirmed adenocarcinoma of the colon or rectum. The median age of the patients was 67 years (range: 12–91 years). The median follow-up time was 30.6 months (range: 1.5–107.2). A total of 49 patients died of CRC-related causes during this period.Staging was based on clinical assessment and histopathological analysis using the International Union Against Cancer (UICC) TNM staging system. Written informed consent was obtained from all the patients enrolled onto the study, according to local ethics guidelines. This study was approved by the Institutional Review Board.Cancer tissues were frozen in liquid nitrogen immediately after surgical resection and stored at −80°C until use. The diagnosis of CRC was confirmed for all 223 patients based on clinicopathologic findings.Cancer tissues were minced and homogenized with a Mixer Mill MM 300 homogenizer . Total RNA was isolated using an RNeasy mini kit (Qiagen) according to the manufacturer's instructions. cDNA was synthesized from 5 µg total RNA with a random hexamer primer and Superscript III reverse transcriptase  according to the manufacturer's instructions.Quantitative PCR (qPCR) analysis was performed using a TaqMan Universal PCR Master Mix . The expression levels of the target gene transcripts, measured using TaqMan probes for BDNF  and TrkB , were normalized to GAPDH  and evaluated using Applied Biosystems StepOne Software (v2.1).2). The level of expression of the target gene was calculated from the standard curve, and normalized against GAPDH. Finally, the target gene mRNA level was expressed as a ratio relative to the GAPDH mRNA level. Real-time quantitative PCR assays were performed in triplicate for each sample and the mean value was used in calculating mRNA expression levels.The relative BDNF and TrkB gene expression levels were determined using a standard curve. The standard curve was generated using a 5-fold serial dilution of random-primed qPCR Human Reference cDNA . All standard curves were linear within the range used for analysis, with an acceptable corresponding correlation coefficient . The cell line authentication testing has been performed for these cell lines. These cell lines were maintained in RPMI-1640 medium supplemented with 10% fetal bovine serum, 100 IU/mL penicillin, and 100 µg/mL streptomycin at 37 °C and 5% CO2.in vitro assays.Recombinant human BDNF was purchased from PeproTech  and prepared according to the manufacturer's instructions. Recombinant human BDNF was dissolved in PBS (10 µg/ml) for the in vitro and in vivo assays.K252a was purchased from Calbiochem  and stored at −20°C before use. K252a was dissolved in PBS (10 µg/ml) for the The CRC cell lines were washed in ice-cold PBS whilst on the dish. Cold lysis buffer  was then added directly to the dish. The cells were then scraped off the dish, collected, and homogenized using a Mixer Mill MM 300 homogenizer . The supernatants were collected and frozen at −20°C until use. The protein concentration was measured using the BCA protein assay . 20 µg of the protein lysate was mixed with an equal volume of 2× Laemmli loading buffer containing 2-mercaptoethanol, and heated at 100 °C for 5 min. Samples were electrophoretically separated on 12.5% gradient polyacrylamide gels containing 0.1% SDS, followed by semi-dry transfer to an Immun-Blot PVDF membrane . The membrane was then blocked using 5% skimmed milk in Tris-buffered saline, pH 7.5, supplemented with 0.1% Tween 20 (TBS-T).The blots were then incubated with mouse monoclonal anti-TrkB antibody  at a 1∶1000 dilution, rabbit polyclonal anti-BDNF antibody  at a 1∶100 dilution, and mouse monoclonal anti-actin (clone C4) antibody  at a 1∶400 dilution in 5% skimmed milk in TBS-T overnight at 4 °C. After washing three times in TBS-T, the blots were incubated with alkaline-phosphatase-conjugated goat anti- mouse IgG  at a 1∶200 dilution in 5% skimmed milk in TBS-T. Following treatment with an enhanced chemiluminescence detection solution, chemiluminescent signals were visualized in a CS Analyzer and AE-6962 light capture .To assess the effect of BDNF on cell viability, migration, invasion, and anoikis resistance, recombinant human BDNF and K252a, a selective pharmacological pan-Trk inhibitor, were used.Cell viability, migration, invasion, and anoikis resistance were compared among non-treated cells (control), BDNF-treated cells, K252a-treated cells, and K252a followed by BDNF-treated cells.Cytotoxicity was evaluated using a WST-8  colorimetric assay. Cancer cells (5000 cells/well) were seeded onto 96-well cell plates  in 100 µl of culture medium for 24 h. After preincubation, the cells were treated with K252a (100 nM) or serum-free medium for 2 h, and then treated with either recombinant human BDNF (100 ng/ml) or serum-free medium. 48 h later, the medium was discarded and replaced with 90 µl of fresh medium followed by the addition of 10 µl WST-8 reagent solution  and incubated for 2 h at 37 °C in an incubator.Cell viability was determined by colorimetric comparison by reading optical density (OD) values from a microplate reader  at an absorption wavelength of 450 nm. Cytotoxicity was evaluated using the Cell Counting Kit according to the manufacturer's instructions. The data were obtained from similar results of at least three independent experiments. Results are presented as mean ± standard error (SE).Confluent cancer cells were serum-deprived for 48 h. Wounds were generated using a sterile 200-µL pipette tip after preincubation with K252a (100 nM) or serum-free medium for 2 h. Recombinant human BDNF (100 ng/ml) or serum-free medium was added and the cells were incubated at 37 °C for an additional 46 h. Wound closure was assessed using an Olympus IX71 microscope  at 10× magnification. Cell migration distance was measured using Adobe Photoshop 9.0.2 software and compared with baseline measurements. Each independent experiment was performed at least three times. Results are presented as mean ±SE.Cell invasion was evaluated using Biocoat Matrigel invasion chambers and control inserts (Becton Dickinson Labware). A total of 50000 cells/well were seeded in the invasion and control chambers, and pre-incubated with 100 nM K252a or serum-free medium for 2 h. Fresh medium alone or medium containing recombinant human BDNF (100 ng/ml) was then added to the Falcon companion plates . The Matrigel invasion chambers and control inserts were incubated for 24 h at 37 °C. The incubation medium containing cells was removed from the top chamber using cotton swabs and serum-free medium. The membranes were fixed in methanol, stained with Mayer's hematoxylin, dehydrated in ethanol, and mounted on glass slides. The number of cells that invaded the underside of the membrane was then determined. Each independent experiment was performed three times. Results are presented as mean ±SE.Anoikis, detachment-induced apoptosis, is known to be induced when adherent tumor cells are forced to grow in a non-adherent fashion. Anoikis resistance was evaluated by the proportion of viable tumor cells that proliferated non-adherently in low-attachment dishes.5 cells/ml, respectively. Their cell suspensions (2 ml) were added to each well and incubated for 24 h in a humidified atmosphere (37 °C and 5% CO2).Anoikis assays were performed in six-well Costar Ultra Low Attachment Microplates . DLD1, LoVo, and SW480 cells were suspended in RPMI-1640 with BDNF (100 ng/ml), K252a (100 nM), or K252a+BDNF at a concentration of 5×103 cells/well in microtiter plates  in a final volume of 100 µL culture medium per well. To assess the viable tumor cells that proliferated non-adherently in low-attachment dishes, spectrophotometric absorbance of each well was measured using WST-8 reagent solution as described above (cell viability assay). Each independent experiment was performed six times. Results are presented as mean ±SE.After induction of anoikis, cells were seeded at 5×105 cells were washed and resuspended in 0.5 ml of 1×binding buffer, and Annexin V: fluorescein isothiocyanate/Propidium Iodide (PI) labeling performed according to the manufacturer's protocol . The analysis was performed using a FACSCalibur flow cytometer  to quantify the proportion of viable or apoptotic tumor cells under non-adherent growth condition using low-attachment dishes. Each sample contained 5×105 cells. The data were analyzed using CellQuest Pro software (BD Biosciences). Viable tumor cells that were negative for both Annexin V and PI staining (lower left quadrant) were considered as anoikis-resistant cells with non-adherent growth. Apoptotic tumor cells that were positive for Annexin V staining (lower right quadrant+upper right quadrant) were considered as anoikis-induced cells. Tumor cells treated with 2.5% formalin solution were used as a positive control for apoptosis induced cells. Flow cytometric analysis was performed to confirm the results of cell viability assay for anoikis resistance.After induction of anoikis, 5×107 cells/500 µl PBS) were injected intraperitoneally for peritoneal metastatic formation. Either K252a (500 µg/kg) or PBS (control) was injected intraperitoneally three times a week to examine the effect of K252a on the peritoneal metastatic formation. Four weeks later, the mice were sacrificed, and then the size and number of peritoneal metastatic nodules was evaluated . Results are presented as mean ±SE.Male nude mice (BALB/c) at 8 weeks of age were purchased from Japan SLC Inc. . The experimental protocols were reviewed and approved by the Animal Care and Use Committee at the Mie University Graduate School of Medicine. DLD1 cells . The results are expressed as the mean ±SE (standard error). Differences between groups were estimated using the Pearson's chi-square test, repeated-measures analysis of variance (ANOVA) analysis, or an unpaired Student's t-test. Actuarial survival curves were obtained using the Kaplan–Meier method, and comparisons were made using log-rank tests. Univariate and multivariate analyses were done using the Cox proportional hazards model to investigate the effects of the histopathological and molecular factors (BDNF and TrkB expression status) present in the primary tumor specimen on overall survival (OS). Variables associated with survival with a P-value less than 0.05 in univariate analysis were used for multivariate analysis. P-values of less than 0.05 were considered statistically significant. Two-sided P-values <0.05 were considered to be statistically significant.The BDNF mRNA expression ratio, relative to GAPDH, in CRC tissues was 0.122±0.230 (mean±SD), ranging from 0 to 1.357. For statistical analysis purposes the BDNF mRNA expression values were dichotomized as low or high. A cutoff value for the BDNF mRNA level was determined as a maximum predictive value using an OS based on receiver–operating characteristic (ROC) curve analysis. One hundred and two patients had high BDNF mRNA expression (>0.037), whereas 121 patients had low BDNF mRNA expression. As shown in According to the TaqMan Gene Expression Assay , probe of TrkB  is designed to span the exon 6/7 boundaries. Corresponding primers amplify the fragments containing exons 6–7 which encodes the extracellular portion of the TrkB receptor. Therefore, this primer and probe set detects both the full-length TrkB (TrkB.FL) and the truncated TrkB (TrkB.T1) The TrkB mRNA expression ratio, relative to GAPDH, in CRC tissues was 0.054±0.141 (mean±SD), ranging from 0 to 1.149. ROC curve analysis showed that the cutoff value for TrkB mRNA was 0.002 . One hundred and seventy-one patients had high TrkB mRNA expression, whereas 52 patients had low TrkB mRNA expression. As shown in Ninety-four patients showed both high BDNF and TrkB mRNA expression. This patient group was classified as a co-expressing both BDNF and TrkB. As shown in Immunohistochemistry showed that the BDNF protein  was expressed in the cytoplasm of human CRC cells and that the TrkB protein  was expressed in the nucleus of human CRC cells . In patiAs shown by the above results, both BDNF and TrkB expression in tumor cells seems to be involved in both primary and metastatic tumor progression in human CRC. To explore the involvement of an autocrine BDNF/TrkB signaling in CRC progression, we first examined BDNF and TrkB expression in 5 CRC cell lines including CaCO2, DLD1, HT29, LoVo, and SW480.BDNF mRNA expression was detected in all 5 cell lines by RT-PCR analysis . In contWestern blotting analysis  showed tBased on our results and evidence from previous reports, we selected 3 cell lines  which showed both BDNF and TrkB mRNA expression with either TrkB.FL or TrkB.T1 protein expression for further examination.To explore the possibility of an autocrine BDNF/TrkB signaling loop occurring in CRC, we investigated the effect of BDNF on tumor cell viability in TrkB-expressing CRC cells. As shown in These results suggest that exogenous BDNF increases tumor cell viability in TrkB-expressing CRC cells, and that TrkB receptor blockade may provide a potent means of inhibiting tumor growth.in vitro migration assays. Forty-eight hours later, BDNF significantly increased the migration of DLD1 cells, as compared with the non-treated controls. K252a decreased the migratory ability of the DLD1 cells, which was nearly identical to the non-treated controls. K252a followed by BDNF inhibited the migratory ability of DLD1 cells, as compared with BDNF treatment. The representative images of the above results were shown in To examine the effect of BDNF on tumor cell motility, DLD1 cells were used for in vitro invasion assays. Twenty-four hours later, BDNF significantly increased the invasion of SW480 cells, as compared with the non-treated controls. K252a decreased the invasive ability of SW480 cells, which was nearly identical to the non-treated controls. K252a followed by BDNF treatment inhibited the invasive ability of SW480 cells, as compared with BDNF treatment. The representative images of the above results were shown in To examine the effect of BDNF on tumor cell invasion, SW480 cells were used for in vitro experimental setting, it is induced when adherent tumor cells are forced into non-adherent growth. To examine the effect of BDNF on anoikis resistance (detachment-induced apoptosis resistance), DLD1, LoVo, and SW480 cells were used for in vitro anoikis assays. The number of anoikis resistant cells was evaluated by counting viable tumor cells that proliferated non-adherently in low-attachment dishes (cell viability assay using a WST-8 reagent solution). As shown in Anoikis is a form of detachment-induced apoptosis. In the These results suggest that exogenous BDNF enhances anoikis resistance in TrkB-expressing CRC cells, and that a TrkB receptor blockade may inhibit the anoikis resistance of these tumor cells.in vivo peritoneal metastasis assay. in vivo peritoneal metastasis assay. The representative images of PC arising from DLD1 cells and peritoneal metastases were shown in The results presented above have shown that BDNF enhances the cell viability, migration, and anoikis resistance of TrkB-expressing CRC cells, and that the TrkB inhibitor, K252a, inhibited these effects. These results suggest that an autocrine BDNF/TrkB loop may be involved in the progression of both primary and peritoneal metastatic tumors. We next investigated whether a TrkB receptor blockade suppress the peritoneal metastasis of BDNF/TrkB-co-expressing DLD1 cells using an These results suggest that a TrkB receptor blockade may inhibit either anoikis resistance, adhesion onto the peritoneal surface, or the colonization of the peritoneal cavity.Previously, we reported an inverse correlation between TrkB and E-cadherin mRNA levels in CRC patients, and an inhibition of tumor cell proliferation, migration, and invasion by TrkB knockdown in CRC cell lines The mRNA levels of BDNF, TrkB and their co-expression in CRC tissues were associated with tumor progression and poor prognosis of CRC patients. Although we couldn't show that co-expression of BDNF and TrkB had an independent prognostic significance by multivariate analysis (data not shown), BDNF/TrkB expression status of the primary CRC tissues may provide prognostic information for CRC patients.Previous reports showed that BDNF was higher in CRC tissues than adjacent normal tissues at the mRNA We found that BDNF/TrkB signaling was significantly associated with proliferation, migration, invasion, and anoikis resistance in BDNF/TrkB co-expressing CRC cells. Recent studies also demonstrated the molecular mechanisms of BDNF/TrkB signaling in tumor cell migration, invasion, and anoikis resistance. They showed the involvement of β5 integrin in BDNF/TrkB signaling mediated cell migration In this study, BDNF was detected at the mRNA and protein level in all 5 cell lines, although cell line-dependent difference was observed at the protein expression level. In contrast, TrkB was detected at the mRNA level in 3 out of 5 cell lines. Additionally, these 3 cell lines showed higher protein expression of TrkB.T1 than that of TrkB.FL.TrkB.T1, one of the truncated forms of TrkB receptor, is generated by alternative splicing during gene transcription and lacks the intracellular tyrosine kinase domain with the extracellular BDNF binding region TrkB.T1 was also frequently detected in several human malignancies rather than TrkB.FL in vitro studies.In CRC cell lines, TrkB.T1 has been also reported to be preferentially expressed at both the mRNA and protein levels, as compared with TrkB.FL Since TrkB.T1 regulates BDNF/TrkB.FL signaling in vivo. PC arising from CRC has been reported to be found in approximately 7% of CRC patients at diagnosis, or at the time of primary tumor surgery (synchronous PC) We also demonstrated that treatment of mice with the pharmacological pan-Trk inhibitor K252a suppressed the PC formation of BDNF/TrkB-co-expressing CRC cells In our series, synchronous PC was the most powerful predictive factor of overall survival, with the highest risk ratio during the last 5 years of life (data not shown). These lines of evidence indicate that PC is regarded as a primary target of CRC treatment. Although the effects of chemotherapeutic treatment on the prognosis of metastatic CRC have been well reported, it remains unclear whether these treatments provide the same benefit to the subset of CRC patients with PC.Recently, we showed that the blockade of BDNF/TrkB signaling by K252a suppressed the PC formation arising from gastric cancer cells with co-expression of BDNF and TrkB However, further studies will be needed to reveal the therapeutic potential of BDNF/TrkB signaling in CRC, especially for PC arising from CRC. To clarify the anti-tumor efficacy of the blockade of BDNF/TrkB signaling in CRC, we would first need to confirm whether a BDNF blockade, such as an anti-BDNF antibody, and a TrkB blockade, such as a pharmacological pan-Trk inhibitor or an antagonistic anti-TrkB antibody, show anti-tumor effects on CRC in patients with various BDNF/TrkB expression statuses.In conclusion, we have demonstrated that BDNF/TrkB signaling in tumor tissues is associated with tumor progression and poor prognosis in CRC patients, and that it is also associated with proliferation, migration, invasion, and anoikis resistance in BDNF/TrkB co-expressing CRC cells. The BDNF and TrkB expression status in our resected specimens may provide prognostic information for CRC patients. Finally, BDNF/TrkB signaling may be a promising potential target for PC arising from CRC.


Since graphene has been taken as the potential host material for next-generation electric devices, coexistence of high carrier mobility and an energy gap has the determining role in its real applications. However, in conventional methods of band-gap engineering, the energy gap and carrier mobility in graphene are seemed to be the two terminals of a seesaw, which limit its rapid development in electronic devices. Here we demonstrated the realization of insulating-like state in graphene without breaking Dirac cone. Using first-principles calculations, we found that ferroelectric substrate not only well reserves the Dirac fermions, but also induces pseudo-gap states in graphene. Calculated transport results clearly revealed that electrons cannot move along the ferroelectric direction. Thus, our work established a new concept of opening an energy gap in graphene without reducing the high mobility of carriers, which is a step towards manufacturing graphene-based devices. This fascinating character of extremely high carrier mobility endows graphene great advantage for electronic applications.Two-dimensional atomic materials, which are consisted of one or several atomic layers, have attracted great research interests in recent years124578101112zp-zp bonding −7 μC/cm.For the experimentally synthesized hydroxyl-functionalized BNSL (OH-BNSL), hydroxyl groups are connected with boron atoms. However, the orientation of hydroxyl groups on BNSL is not explored. As shown in Now we turn to explore the electronic properties of graphene/OH-BNSL bilayer, in which graphene is covered on the surface of ferroelectric OH-BNSL. As shown in 6 m/s).As stated in our previous reportsAlthough the character of linear band dispersion and zero-energy gap in graphene is not disturbed by the OH-BNSL, it is cursory to deduce that OH-BNSL has little effect on the electronic property of graphene. Since OH-BNSL introduces a periodic electric potential along the ferroelectric direction (the orientation of hydroxyls) in graphene, the charge density of graphene should be modulated by this periodic electric potential. As shown in Generally, the carriers transport will be modified by the charge density wave. As we know, along the direction of chain-like charge patterns, the carriers can much easier be transported because of the strong overlapping of nearest-neighbor wave function. Besides, the calculated band dispersion has demonstrated the existence of Dirac fermions, namely, such carriers have a high mobility. While for the direction perpendicular to the chain-like charge patterns, carriers are difficult to be transferred from one chain to another one because of the weaker overlapping of nearest-neighbor wave function, namely, there is a “pseudogap” along this direction although there is no real energy gap in the electronic structures.PJ and VJ, respectively. The calculated zero-bias transmission functions of two proposed graphene/OH-BNSL bilayer junctions are plotted in VJ are significant larger than that of PJ within the energy range from −0.25 to 0.12 eV. For example, at the Fermi level, the transmission coefficient of VJ is about 1.6, and the corresponding value of PJ is very small (less than 10−4), which stands for the metallic and insulating states, respectively. These results show that the transport direction of graphene/OH-BNSL bilayer system depends on the orientation of hydroxyl groups on BNSL. The metallic channel coexists with an insulating-like one in graphene/OH-BNSL bilayer system, which well confirms our above electronic structure calculations. Moreover, the metallic state can be switched into the insulating-like state by changing the orientation of hydroxyl groups on BNSL.To verify the above predictions, we perform first-principles calculations combined with non-equilibrium Green's function method to explore the transport properties of two graphene/OH-BNSL bilayer junctions, as shown in inserts of It should be noted that the “pseudogap” explored in graphene/OH-BNSL bilayer is totally different with that of previous reports181920211819202113/cm2. Therefore, in the following studies, we will limit the electron concentration below 1.0 × 1013/cm2. As shown in 13/cm2, 0.5 × 1013/cm2, 1.0 × 1013/cm2). Importantly, in all cases, the charge density around Fermi level has the chain-like patterns  code. The projected augmented wave (PAW) method25−4 eV, and 0.01 eV/Å, respectively. The accuracy of our simulation was tested through a comparison of the presented results with previous theoretical results.Our first-principles calculations were based on spin-polarized density functional theory (DFT) using the generalized gradient approximation (GGA) known as PW91LGΓRG+], in which ΓL(R) stands for the coupling matrix between the left(right) electrode and the central scattering region, and G is the retarded Green's function of the scattering region. In our calculations, we adopted Troullier-Martins nonlocal pseudopotential and double-zeta plus polarization basis sets for all elements in the junctions. The grid integration was defined by an energy cutoff of 150 Ry, while the exchange and correlation functional was treated at the level of Ceperley-Alder local-density approximation. Test calculations with more k-points and larger cutoff energy yield the similar results.The transport calculations for graphene/OH-BNSL bilayer were carried out by using the fully self-consistent non-equilibrium Green's function method combined with first-principles calculations, implemented in ATK package27E.K. conceived the project. F.W. performed the calculations of electronic properties. J.H. studied the electric conductivity and calculated the I–V curve. F.W., J.H., Q.L., K.D. and E.K. co-wrote the paper, and all authors contributed to the discussion and preparation of the manuscript.