International donors have increasingly shifted AIDS funding directly to community-based organizations (CBOs) with the assumption that responding to the epidemic is best achieved at the community level. The World Bank, ICF Macro, and the National Council for Population and Development in Kenya, conducted a study to evaluate the community response in Kenya. The study used a quasi-experimental design comparing seven study communities and seven comparison communities in Nyanza Province and Western Province. We examined the impact of CBO activity on individual and community-level outcomes, including HIV knowledge, awareness and perceptions, sexual risk behavior, and social transformation . The study consisted of two components: a household survey conducted in all 14 communities, and qualitative data collected in a subset of communities. Individuals in communities with higher CBO engagement were significantly more likely to have reported consistent condom use. Higher CBO engagement was associated with some measures of social capital, including participation in local and national elections, and participation in electoral campaigns. CBOs provide added value in addressing the HIV and AIDS epidemic in very targeted and specific ways that are closely tied to the services they provide ; thus, increasing CBO engagement can be an effective measure in scaling up prevention efforts in those areas. At the pandemic . CBOs arpandemic . Over thpandemic .The increased role of CBOs fighting AIDS was supported by the assumption that controlling and responding to the epidemic is best achieved through the active involvement of communities. Theoretical literature on CBOs has argued that they are well positioned to reach the previously missed elements of the society and tackle disease and poverty at their roots . In addiEmpirical literature has shown that community-based interventions in Kenya have the potential to decrease the stigma associated with HIV/AIDS , improveTo address this evidence gap, the World Bank commissioned a rigorous evaluation of the community response to HIV and AIDS in Kenya from January through September 2010. The evaluation addressed three primary research questions: Do community members in communities with stronger CBO engagement demonstrate (1) a significant difference in their access to and use of HIV and AIDS services; (2) a significant difference in knowledge, attitudes, perceptions, and behavior related to HIV and AIDS; and (3) a significant difference with respect to social transformation indicators compared to communities demonstrating weaker engagement?Recent surveys estimate the prevalence of HIV in the adult population in Kenya to be between 6.3% and 7.4% . The geoCBOs have been at the forefront of the fight against AIDS in Kenya. Their involvement was identified as a key component of the national response  and six districts Kisumu, Nyando, Kisii, Nyamira (Nyanza Province) and Butere-Mumias and Vihiga (Western Province) were included in the evaluation. A power analysis concluded that there would be 80% power to detect a medium effect size difference in 14 communities with a total of 192 households per community. Nyanza and Western provinces were selected for inclusion based on high HIV prevalence. The six districts have similar demographic characteristics, but varying levels of CBO activity. Selection of the communities occurred through several steps. First, the two provinces, Nyanza and Western, were selected for inclusion in the study based on high HIV prevalence. Within each province, the six districts that were selected were relatively close to one another and had relatively similar demographic characteristics, but were known to have varying levels of CBO activity. Within each district, a list of communities was delineated from the Kenya National Bureau of Statistics database. From this list, a random sample of 34 communities was selected. Communities in the random sample were assessed using specified criteria  in the community and individual awareness of CBO activity in the community.We used a database of CBOs maintained by the National AIDS Control Council (NACC) and information provided by local experts to capture the number of CBOs in each community. Given the potential that the NACC's CBO information was incomplete, and recognizing that the strength of the CBO activity may be determined by factors other than the number of CBOs, we used data collected from the household survey to provide additional information about the CBO reach, and to verify our initial study-comparison assignment. In the statistical analyses described below, the level of CBO engagement from the survey data was measured by the proportion of respondents who were aware of HIV/AIDS-related CBOs in their community. Communities with higher than the median proportion (46.9%) of respondents aware of services provided by CBOs were considered study communities, and communities with lower proportions of respondents aware of services provided by CBOs were considered comparison communities. Community assignment is not considered an absolute “high” versus “low,” but is rather a relative comparison of “higher” versus “lower” community engagement. A total of five communities were reclassified from the original assignment based on these post hoc criteria. Communities 9, 10, and 13 were reassigned from study to comparison, and communities 11 and 12 were reassigned from comparison to study. The final list of community assignment and their characteristics are presented in A sample of 2715 households was randomly selected using official data for household populations from the list of National Sample Surveys and Evaluation Programme clusters, created and maintained by the Kenya National Bureau of Statistics (KNBS). In each household, up to three individual adult respondents were interviewed, resulting in a total of 4378 individuals. The household respondents were chosen based on selecting the head of household (if home at the time the interviewer went to the door), and the spouse of the household head (if existing and home). If the head of the household was not home, the interviewer talked with adults in the household and asked for nominations of up to three individuals. The head of household was asked to select other adult(s) to participate to reach up to three respondents. The Household Survey was administered to one individual per household, which included items assessing household-level constructs . An individual questionnaire was administered to each individual respondent, which included individual-level constructs . The survey was fielded from May to June 2010.Service awareness and utilization was included in the individual survey and was measured using Yes-No questions whether the respondent was aware of (awareness questions) and used (utilization questions) services in the following categories: (1) prevention, (2) treatment, (3) care and support, (4) impact alleviation, and (5) community mobilization. The responses were coded as 1 if the respondent responded “Yes” and 0 otherwise.Knowledge indicators reflected the UNGASS reporting system, and focused on the knowledge of HIV transmission modes, medications, and prevention. The individual survey included the following questions : People reduce HIV chances by having one uninfected partner; People reduce chances of getting HIV by using a condom; People CANNOT get the HIV virus from a mosquito or insect bite; People cannot get HIV by sharing utensils with a person who has AIDS; It is possible for a healthy-looking person to have HIV. Correct responses were coded as 1 and incorrect (including “Don't Know”) as 0. In addition, respondents were asked the following Yes-No questions: Are there any special drugs that a doctor or a nurse can give to a woman infected with the HIV virus to reduce the risk of transmission to the baby? Have you ever heard of VCT? Would you accept VCT if a volunteer came to the house? Do you know of drugs to control HIV? The responses were coded as 1 if the respondent responded “Yes” and 0 otherwise. We analyzed each knowledge item separately because individual items measure different aspects of HIV- related knowledge . This mode of analysis provides a more nuanced assessment of the association between the community engagement and knowledge than using a single knowledge scale.Behavior was measured using two indicators. – consistent condom use with all partners in the past 12 months and ever having had an HIV test . Individuals were asked the number of sexual partners in the past 12 months, and for each partner, were asked, “Was a condom used every time you had sexual intercourse with that partner in the past 12 months?,” with response options Yes-No. Those who responded “Yes” to each partner were considered consistent condom users (coded “1”) and those who responded “No” to the question for any partner were not considered consistent condom users and coded “0”. Individuals were asked if they ever had an HIV test (response options Yes-No).Social transformation was assessed using measures from the social science and public health literature focusing on social capital and gender norms. Social capital indicators were taken from the Adapted Social Capital Assessment Tool (A-SCAT) ((A-SCAT) . Social Cognitive social capital was measured with a 20-item scale with items indicating perceived strength and cohesion of the community such as, “My neighbors would help me in an emergency; if someone in the community lost their job, there are community members who would help them with money or other resources.” Response categories included “Strongly Disagree, Disagree, Agree, and Strongly Agree.” A mean and summative scale was created and analyzed as separate outcomes. A scale of attitudes toward one's own children was measured using six items (Yes-No response): “Do you help your children to understand that they have their rights; Do you help your children to understand that their rights are legitimate and nobody can violate them; Do you help your children to react against the violation of their rights; Do you discuss with your children when others’ rights are violated; Do you encourage your children to react against violation of others’ rights; and Do you allow your children to participate in decision-making in the family?” “Yes” responses were coded “1,” and “No” responses were coded “0.” An average score across all items was calculated.gender norms were based on surveys developed by the Kenya Diffusion and Ideational Change Project and the Malawi Diffusion and Ideational Change Project , using Mplus . MultileBivariate differences in demographic factors between study and comparison communities resulting from post hoc reassignment of communities were found for HIV prevalence, rural versus urban characters, employment, age, and marital status . These dIn addition to those factors, we also controlled for education as this variable is typically associated with HIV knowledge and behavior. For binomial outcomes, logistic regression was performed, and results are reported as odds ratios with confidence intervals. For continuous outcomes, linear regression was performed, and results are reported as coefficients with standard errors and p-values.In-depth semi-structured interviews with 28 CBOs and 58 key informants (KIs) were conducted in a sub-sample of eight communities. The interviews provided more detailed information on CBO services and activities, and data on social transformation and contextual factors that facilitate and constrain the effectiveness of HIV activities and programs. Participating CBOs were identified based on the NACC database of CBOs and supplemented by information provided by the NACC staff and local community experts. KIs included (1) the director of the Constituency AIDS Control Committee  in each community, (2) a local government leader, (3) the head nurse from the largest local clinic, (4) a senior clergy person from the largest religious congregation, and (5) the principal of the largest school.The interviews with CBO staff focused on the activities of CBOs, their targets/audiences, geographic reach, and staff and volunteers involved. Questions included the following: ‘When was the organization established? What did the response to the HIV/AIDS epidemic look like before your organization was established? What activities did your organization focus on initially? How has the scope of activities changed over time? What goals have you achieved in your community? What are some of the factors that have helped you achieving your goals? How do you think your work has affected the way women/orphans/people with HIV are treated? What is your experience with the local government response to HIV/AIDS? How does your organization interact with government? How does your organization interact with other local organizations?’ The interviews with KIs provided information about social capital and transformation in the community, and the perceived role of CBOs in affecting those factors. Questions included “Over the past five years what changes have you observed in how people trust each other/voting behavior/participation in voluntary organizations/collective action/the ability of women to make economic decisions/girls’ access to education/violence against women/how people with HIV are treated? What factors do you think caused any changes?”Trained interviewers conducted the interviews in the local language, and took detailed notes, and in some cases, recorded, and transcribed the interviews. Notes and transcription were translated into English. Interviews were reviewed and coded by a team of two individuals. Initial codes were developed to match the interview topics, and additional codes were added during the review and coding process as needed.Significant associations found between the key independent variables – higher (coded “1”) or lower (coded “0”) CBO engagement – and measures of HIV knowledge, attitudes, perceptions, behavior, service utilization, and social transformation are in bold in the tables.In addition, respondents in the study group had four times higher odds of reporting consistent condom use with all sex partners in the last 12 months , and were more likely to have ever had an HIV test .We found no statistically significant association between the strength of CBO engagement and service utilization.Strength of CBO engagement was not associated with cognitive social capital or any indicators of gender norms.The average length of time a CBO was present in the community ranged between 5.5 and 8.8 years. The average number of volunteers in the past year ranged from 4 to 26. The average number of CBO clients reported in a community varied widely, and ranged from 67 to 853.CBOs were engaged in similar activities across study and comparison communities. In all eight communities, CBOs engaged in prevention activities, support for PLWHA, activities targeting OVC, and information and education activities aimed at increasing the awareness and knowledge of HIV and AIDS. In seven communities, CBOs also engaged in care; in five communities, in advocacy efforts; and in three communities, in impact mitigation. CBOs engaged in provision of medical services in only one community. In six communities, CBO leaders mentioned behavior change communication as part of their organization's mission, and in three communities, CBOs distributed condoms. The number of the clients reached as a percentage of the overall population was the most pronounced difference between the study and the comparison communities .The interviews provided historical depth to the quantitative survey and insight into how social capital, gender relations, and stigma and discrimination have changed over time. Furthermore, the interviews with KIs were designed to help determine the extent to which any observed differences in outcomes between study and comparison communities could be attributed to the activities of local CBOs.Voting, an indicator of social capital in terms of civic engagement, was reported by KIs to have increased in all communities – they noted that turnouts in national and local elections have increased over the past five years. Most attributed the increase to rising education levels and higher awareness of political and voting rights among community members. They did not, however, identify what caused the shifts in education and awareness, and none explicitly mentioned CBO activity as the cause of those changes. While KIs did not identify CBOs as a driving force behind increased voting behavior, survey data showed that this indicator was significantly higher in higher engagement communities.In all communities, most of the KIs noted increases in participation in voluntary organizations within the past five years. Self-help groups and groups engaging in income-generating activities, such as setting up fishing ponds or communal poultry-raising, were mentioned as the most common forms of organizations emerging in the research communities. In one community, KIs noted that people formed neighborhood associations assisting families in organizing and financing funerals. The fact that individuals from both study and comparison communities noted increased participation in organizations supports the lack of differences found in this measure of social capital by the household survey.In three communities, increases in participation were attributed to normative changes, although no agents driving the normative change were identified. In one community, CBOs were explicitly mentioned as the source of change. KIs noted that the CBOs in the community have “sensitized people on the importance of coming together.” In the same community, two other KIs attributed the recent changes to increase in government assistance given to voluntary associations. In one community, increases in the number of voluntary organizations and in the number of community members belonging to such organizations were attributed to the local leadership's support of such initiatives.In three communities, KIs reported an increase in economic opportunities available to women. In two communities, the KIs said that women were gaining more influence over how to spend the household money. In one community, some KIs attributed this to activities of self-help groups engaging in income-generating activities.In addition, respondents noted structural factors such as women being more aware of their rights, general increases in the level of education among women in Kenya, and migration .Across all communities, KIs noted increases in the number of girls enrolled in elementary schools. They attributed the main drivers of the increases in female enrollment in education and also perceived declines in violence against women to shifts in national policies, rather than CBO activity. KIs credited the CBOs with helping to offset some of the expenses related to education, such as paying for school uniforms or providing financial support for orphans to prevent them from dropping out. With respect to women's economic agency and their influence over household finances, KIs in two communities mentioned income-generating activities carried out by CBOs as increasing women's access to money, but not necessarily in increasing their influence on how to spend it.When asked explicitly about the impact of the CBOs on gender relations in their communities, KIs credited the CBOs with helping to offset some of the expenses related toeducation, suchaspaying for school uniforms or providing financial support for orphans to prevent them from dropping out. With respect to violence against women, KIs in all communities mentioned that CBOs educated women about their rights. In one community, one KI noted that a CBO also provided legal representation for victims of violence. The interviews, however, suggest that those activities hadalimited impactand wereofsecondary importance compared to the changes in national policies.The qualitative data regarding gender roles support the findings from the survey, in that there were no statistically significant differences between the study and comparison communities in measures of gender norms in the survey. This is consistent with perceptions of the KIs that changes gender norms have occurred across the country as a result of national policies, rather than CBO activity.The objective of this evaluation to assess, using a scientifically rigorous methodology, whether CBOs providing HIV and AIDS prevention and treatment activities add value to the government response to the epidemic and have a measurable impact on individual and community-level outcomes. While the study did not assess changes in HIV incidence or prevalence, it was able to identify how higher CBO engagement and activity affects some aspects of knowledge and behavior that partially drive the epidemic.People in communities where a high proportion of individuals were aware of CBO activity showed greater knowledge of some HIV transmission and prevention methods, were more likely to use condoms consistently, and ever have been tested for HIV, demonstrated greater likelihood to engage in community mobilization activities, including voting and participating in electoral campaigns. While CBOs provided similar services across the study and comparison communities, what differentiated communities was the number of community members aware of, and, possibly, reached by these activities. Thus, in communities where CBOs are engaging in prevention efforts, and higher numbers of community members are aware of their work, the levels of knowledge and awareness may be higher, and people may be more likely to engage in lower risk sexual behaviors.The qualitative data provide insight into the larger national effort in Kenya to address the social drivers of the HIV/AIDS epidemic. For example, national policies that have increased access to education seem to have made a significant impact on increasing the rights of women in all communities in the evaluation, regardless of existing CBO activity. It is notable that some key informants viewed CBOs as increasing awareness of policies and laws through outreach and awareness campaigns. Thus, the potential for CBOs to augment national efforts can be viewed as a means of disseminating information from the national to the local level.The study findings are subject to several limitations. First, the lack of data on CBO activities made a priori classification of the community response difficult. To address this limitation, measures of CBO engagement embedded in the household survey served to verify accuracy of a priori group assignments. The community-matching process and controlling, through multivariate analysis, for potentially confounding factors that differed across communities, reduced the likelihood of factors other than CBO engagement driving the associations detected in the analyses.The evaluation was conducted at one point in time, limiting the ability to make causal conclusions about the impact of CBO activity on the outcomes of interest. For example, the fact that individuals in some communities are more aware of CBO services may be the driving force behind, rather than the outcome of, a high level of CBO activity in that community. Similarly, greater awareness and political activism may make it easier for CBOs to establish themselves and be successful in a specific community. While the cross-sectional design does not make it possible to reach causal conclusions, the evaluation was designed with these limitations in mind. Additional evidence from qualitative interviews helped support the conclusions obtained from the quantitative analysis. Our analysis included testing several outcome variables, which may increase the risk of type 1 error. However, it also increases the level of detail and nuance of the overall analysis.We did not conduct sensitivity analysis to assess how the results presented above change if we change the cut-off we used to assign communities to study and comparison communities. The same cut-off for assignment was used in all three components of the study: the household survey, the interviews with the KIs, and the interviews with the CBO staff. This allowed us to show that the CBOs in both groups engaged in similar types of activities  and, from the KI interviews, that the contextual factors across both groups of the communities were roughly the same and the only difference between the study and comparison group was the intensity of exposure to CBO activity. Changing the cut-off point we used in the analysis of the household survey data would require reanalysis of the data from the interviews with CBO staff, to assess whether there were no systematic differences in the types of activities in which the CBOs engaged in the reassigned groups, and reanalysis of the interviews with the KIs, to assess whether there were no systematic differences in contextual factors across the newly assigned study and comparison groups. We consider the multi-method design to be the key strength of our study that allows for triangulation of information across the quantitative and qualitative components. However, it makes sensitivity analysis of a single study component exceptionally challenging. Triangulation from multiple data sources enabled us to distinguish instances in which better HIV/AIDS-related outcomes could be attributed to CBO activities from outcomes that were most likely affected by other factors, including policy changes at the national level. This evaluation suggests that CBOs provide added value in addressing the HIV/AIDS epidemic in targeted and specific ways that are closely tied to the services they provide. Increasing CBO engagement can be an effective measure in scaling up prevention efforts, especially those aimed at improving knowledge and awareness of AIDS in Kenya.


Programmed death ligand 1 (PD-L1) is an immunosuppressive molecule expressed by many cancer types, including a large proportion of head and neck cancers (HNC), and ligation of its receptor, programmed death 1 (PD-1), induces exhaustion of effector T cells. It has been shown that interferon gamma (IFNγ) induces PD-L1 expression in many cancer types including glioblastoma, melanoma, lung and kidney cancer. Importantly, the stimuli and mechanism for PD-L1 upregulation in HNC cells are not well characterized. IFNγ signals through Janus Kinase 1/2 (JAK1/2) heterodimer complex and mediates signal transducer and activator of transcription 1 (STAT1) phosphorylation, leading to type I cytokine expression, upregulation of antigen presentation, and tumor cell recognition by cytolytic T lymphocytes (CTL). We investigated basal PD-L1 expression and the mechanism by which IFNγ signaling upregulates PD-L1 in HNC cells including dependence on JAK/STAT pathway. We observed that IFNγ signaling increased PD-L1 expression in a JAK2 but not JAK1 dependent fashion. In addition, interferon alpha (IFNα), which signals via JAK1/TYK2 did not upregulate PD-L1 expression while still upregulated HLA class I. Specific JAK2 inhibition downregulated NK cell-derived IFNγ induced PD-L1 expression and enhanced cetuximab mediated ADCC. Our data suggest a crucial role for JAK2/STAT1 in IFNγ mediated PD-L1 upregulation. JAK2 inhibition provides a promising strategy to increase tumor cell lysis through maintaining HLA class I while suppressing tumor cell expressed PD-L1 in combination with anti-EGFR cetuximab therapy.


After successful invasion, the intron and HEG are subject to degeneration and sequential loss. It remains unclear whether these mechanisms can fully address the high dynamics and mobility of group I introns. Here, we found that HEGs undergo a fast gain-and-loss turnover comparable with introns in the yeast mitochondrial 21S-rRNA gene, which is unexpected, as the intron and HEG are generally believed to move together as a unit. We further observed extensively mosaic sequences in both the introns and HEGs, and evidence of gene conversion between HEG-containing and HEG-lacking introns. Our findings suggest horizontal transfer and gene conversion can accelerate HEG/intron degeneration and loss, or rescue and propagate HEG/introns, and ultimately result in high HEG/intron turnover rate. Given that up to 25% of the yeast mitochondrial genome is composed of introns and most mitochondrial introns are group I introns, horizontal transfer and gene conversion could have served as an important mechanism in introducing mitochondrial intron diversity, promoting intron mobility and consequently shaping mitochondrial genome architecture.Group I introns are highly dynamic and mobile, featuring extensive presence-absence variation and widespread horizontal transfer. Group I introns can invade intron-lacking alleles via intron homing powered by their own encoded  Group I introns are a special kind of self-splicing ribozyme widely found in protist nuclear ribosomal (r)DNA genes, fungal and plant organellar genomes, bacteria, and viruses  and wideOne powerful mechanism for intron mobility is intron homing, powered by active intron-encoded homing endonuclease gene (HEG) . During Another mechanism of intron invasion is reverse splicing through an RNA intermediate .Saccharomyces complex. We have examined the gain and loss of the HEG only among the intron-containing strains, since no existing theories suggest a high frequency of gains and losses of the HEG within the ω intron, and the turnover rate of HEG within the intron would be expected to be very low compared with that of the mobile intron based on the intron-homing model. To our surprise, we found fast turnover rates of the HEG comparable with those of the well-known mobile intron. We further found extensive mosaic sequences in both the HEG and intron sequences, which is inconsistent with any available mechanisms for group I intron mobility, but suggests recurrent horizontal transfer and gene conversion. This mechanism is believed to play important roles in not only introducing intron sequence diversity but also introducing intron content variation and promoting intron mobility by altering the HEG function during the evolution of group I introns.The yeast mitochondrial 21S large subunit (LSU) rRNA contains a group I intron known as the ω intron . In thisTorulaspora yeast strains from four species were obtained from the National Center of Agricultural Utilization Research  and Dr. Matthew Goddard (The University of Auckland). Genomic DNA of each strain was extracted from overnight cultures following the procedure described in cox2 sequences were also obtained to infer organismal relationships. The GenBank accession numbers for the sequences used in the study are listed in Supporting Information, Table S1, and the primers used for PCR amplification and Sanger sequencing are listed in Table S2.Ten cox2) were used to reconstruct the organismal relationship of the Saccharomyces complex. The detailed relationship among the three S. cerevisiae genomes [288C  of the intron and HEG of the LSU rRNA was available in 29 strains. The relationship of these 29 strains  function of the APE  package . Here, we only present the estimations using the simplistic model by constraining the rates of gains and losses to be the same. Furthermore, we have used custom R scripts to compute the likelihood values of given turnover rates in the same way as using the ACE and to conduct likelihood ratio tests between different turnover rates.We sought to model the gain and loss of an intron or an HEG in homologous sites across organisms related by a phylogeny using a two-state continuous-time Markov process, with states 0 (absence) and 1 (presence). In the  strains  was consSaccharomyces complex . The disrupted HEG open reading frames (ORFs) in T. delbrueckii were not attributable to sequencing error, as each type of the T. delbrueckii sequences represents sequences from at least three different T. delbrueckii strains examined in this study (Table S1). The HEGs with an intact ORF were classified as putative functional HEGs, and the ones with a disrupted ORF due to either premature stop codon or indel-associated frame-shift were classified as nonfunctional HEGs. Identical intron-HEG patterns can be found between distantly related strains, whereas very closely related strains may have different intron-HEG organizations. For instance, the ω intron is absent from the S. cerevisiae strain No7, whereas both the intron and HEG are present in the other two S. cerevisiae strains 288C and YJM789. The T. delbrueckii strains contain a copy of non-functional HEG in the intron, while all other Torulaspora species only contain an intron with no HEG.The presence-absence pattern of the ω intron in the mitochondrial LSU rRNA gene and its encoded HEG was determined in 29 strains within the  complex . There aL between μ = 3.51 and μ = 1. The latter equals the rate of nucleotide substitution , and the estimated intron turnover rate 3.51 is significantly faster than the nucleotide substitution rate . This is in good agreement with the well-known high mobility of group I introns, as ribozymes often are powered by HEGs. Both the intron and its encoded-HEG target the same recognition sequence to promote the mobility of intron/HEG as a unit. If the HEG always goes together with the intron and is lost slowly through mutation accumulation and degeneration, one would expect a very slow HEG turnover rate within the intron. We compared the HEG turnover rate against the intron turnover rate. Please note the null hypothesis here is not that the HEG rate equals the intron turnover rate; instead, the null hypothesis would be that the HEG turnover rate is close to zero if no mechanism promotes the high mobility of HEG within the intron. Surprisingly, however, the turnover rate of HEGs was estimated as μ = 21.88 ± 16.21 . On the HEG tree . A likelihood ratio test between μ = 3.51 and μ = 21.88 was also conducted on the intron tree . The different results of the two likelihood ratio tests are likely attributable to the different shapes in their likelihood surfaces . Nevertheless, our results suggest that both the intron and HEG undergo rapid turnover that is faster than the nucleotide substitution rate. The observed trend that the HEG might undergo faster turnover rates than the intron is subject to further investigation in larger datasets.To gain a better understanding of the presence-absence variation of the ω intron within the LSU rRNA and the HEG within the ω intron, we sought a quantitative measure for the intron turnover rate and HEG turnover rate. Our analysis assumed equal rates of gains and losses, since the likelihood value of letting the gain and loss rates vary is not significantly better than that of constraining the gain and loss rates to be the same for either the intron or HEG data. Analyses using the ACE function in the Analyses of Phylogenetics and Evolution package , suggest ± 16.21  and is sHEG tree , the estron tree , but theNakaseomyces bacillisporus and Candida castellii differed by one nucleotide out of the 18 HEG recognition nucleotides from the rest of the strains (not shown). The highly conserved HEG recognition sequence could have served as an excellent precondition for active ω intron invasion promoted by HEG, which is believed to be highly specific on the target sites (The HEG within the ω intron has been documented to promote intron insertion by recognizing a specific 18-nucleotide exon sequence (5′-TAGGGATAACAGGGTAAT-3′) . We haveet sites .S. paradoxus CBS432 contains the intron but does not have the signature CCT nucleotides G+15 and C+37 . In this case, a similarly fast turnover rate of HEG was observed.Intron homing might well explain the high turnover rate of introns, but it cannot explain the apparently high turnover rate of HEGs. If the mobility of HEGs was primarily due to HEG degeneration and loss by mutation accumulation, the HEG would be present in most introns, as mutation accumulation is a much slower process than the turnover of introns and the HEG turnover rate would be expected to be very low or close to zero as HEG turnover was estimated only among the intron-containing taxa. We then investigated the possibility that the unexpectedly fast turnover rate of HEG could be introduced by artifacts in the analysis. First, an intron/HEG presence-absence polymorphism within a defined species . The observed high HEG mobility, therefore, could not be explained by its own recent cleavage activity.We then considered whether any previously recognized mechanisms could explain the high HEG mobility. HEGs themselves have been suggested to be mobile elements independent of a host intron . There hHEG degeneration and sequential loss caused by mutation accumulation can contribute to HEG turnover . Our anaT. pretoriensis strains, CBS2187 and CBS5080, respectively, contain remarkably different intron sequences. T. pretoriensis CBS2187 is very similar to the two T. delbrueckii strains and S. cerevisiae. T. pretoriensis CBS5080 is similar to K. thermotolerans and L. mirantina in different genera. Significant phylogenetic incongruence was observed between the intron tree and organismal tree  was identical with T. delbrueckii CBS5448, but differed from either K. thermotolerans or L. mirantina by at least 12 nucleotides plus one indel. In T. pretoriensis CBS2187, at least two intron regions  were different from T. delbrueckii but identical with the two S. cerevisiae strains. These findings suggest that horizontal transfer and gene conversion can take place recurrently in the intron sequences with multiple donor strains and can be at a fine-scale. We have previously demonstrated that gene conversion between foreign and native homologs can significantly confound phylogenetic analysis  in T. delbrueckii differs from the two S. cerevisiae strains by eight nucleotides, all of which could be found in either K. thermotolerans or L. mirantina. It is important to mention that along the T. delbrueckii HEG sequence there are additional regions highly similar or identical with S. cerevisiae, and some other regions that are significantly different from S. cerevisiae but for which the donor species could not be successfully identified . These findings strongly suggest that frequent gene conversion has taken place within the HEG sequence.Like the mosaic intron sequences, the HEG sequences are also highly mosaic . The H1 P < 0.001, PHI test) were found in both the intron and HEG sequences. In a contrast, no significant recombination signal was detected in the exon sequences, nor was significant phylogenetic incongruence observed between the exon tree and organismal tree . In this study, we focused primarily on the demonstration of the highly mosaic nature of the intron and HEG sequences and did not attempt to identify all the possible gene conversion breakpoints. For illustration purposes, the regions in We used the PHI (pairwise homoplasy index) package . It is very likely that the T. delbrueckii HEG resulted from gene conversion between a native HEG-lacking intron and a foreign S. cerevisiae-like HEG-containing intron. It is also noteworthy that the HEG-containing intron in S. cerevisiae itself is likely of foreign origin, as the intron sequences in the two S. cerevisiae strains are not clustered with their sister species, S. paradoxus, S. cariocanus, and S. mikatae, which formed a phylogenetic clade  do not support the scenario of HEG loss after homing of an HEG-containing intron because intron homing would have introduced the HEG and intron as a unit from the very same donor. One possible explanation is that horizontal transfer of an HEG-containing intron has taken place independently in each of the two T. pretoriensis strains; each foreign HEG-containing intron (K. thermotolerans-L. mirantina-like or T. delbrueckii-like) has been separately converted to an HEG-lacking intron by the presumably native HEG-lacking intron via gene conversion.The overall intron sequence in Gene conversion occurs within group I introns independently of the HEG-recognition sequence and HEG function, which therefore increases the chance of genetic exchange. As a consequence, group I introns can show high sequence diversity and high HEG turnover rates. Furthermore, gene conversion also takes place within the HEG sequence, which could potentially alter HEG function in two ways. (1) Gene conversion could pseudogenize a previously functional HEG open reading frame, leading to HEG degeneration, HEG loss, and ultimately intron loss. (2) Gene conversion could rescue a degenerated HEG back to an intact and functional HEG and promote further intron invasion. Together with the gene conversion between the HEG-containing and HEG-lacking introns, gene conversion can be a powerful force driving HEG and intron evolution .A growing body of evidence has shown that, after horizontal transfer of foreign homologs, gene conversion takes place between the foreign and native homologs to introduce sequence diversity, sequence content variation, and even the gain and loss of adjacent dispensable genes . Considering the wide spread of mobile introns and elements, horizontal transfer and gene conversion could have a significant impact on eukaryotic genomes. All of this could be tested using fast growing genomic data.Group I intron invasion is generally believed to introduce the intron as a whole into an intron-less allele. Up to date, few studies questioned the presumption that the whole intron is of a single origin. In this study, our maximum likelihood analysis supports that HEGs undergo faster or at least comparable turnover compared with the intron in the mitochondrial LSU rRNA gene from the I intron , rescue I intron . Our con


Sarcoidosis is a multisystemic disorder of unknown etiology which is characterized by the formation of non-caseating granulomas in involved tissues. Cardiac involvement is one of the least common manifestations and it can occur at any point of time during the course of sarcoidosis. Here we present the case of 2 patients with known sarcoidosis who develop cardiac abnormalities in the absence of known primary cardiac cause. In our report, we would like to draw attention to the importance of considering heart involvement in any case with systemic sarcoidosis especially in young age. Sarcoidosis, an enigmatic multisystemic disease, is characterized by the formation of granulomas in many tissues mainly the lungs, the lymphoreticular system, the eyes, and the skin, with cardiac involvement being a rare entity . Many reA 40-year-old Tunisian woman, with a history of diabetes mellitus in diet and Lofgren's syndrome  diagnosed in February 2013, came to our department, in February 2014, for exploration of exertional dyspnea of one month evolution and progressively worsening without fever, chest pain, or syncope. The patient denied any past history of hypertension, coronary artery, or thyroid disease. She did not smoke or consume alcohol. On physical examination, she was conscious, cooperative, and afebrile. She was hemodynamically stable (blood pressure (BP) 130/70 mmHg and had a regular pulse of 88 beats /minute.). She had good color and was slightly polypneic with no peripheral cyanosis . She presented no palpable adenopathy or signs of jugular distension. Cardiac auscultation revealed rhythmic sounds with no murmur or friction rub. There were no signs of peripheral edema or deep vein thrombosis. Pulmonary, abdominal, and neurological examination revealed no abnormalities. The admission electrocardiogram (ECG) was normal. On investigation, blood biochemistry, liver, renal function, thyroid function and, hemogram showed no abnormalities except lymphopenia. Her angiotensin converting enzyme (ACE) level was 132 units . The immunological markers were negative. A contrast-enhanced computed tomography (CT) chest scan showed almost complete regression of mediastinal lymphadenopathies without pleuroparenchymal lesion. Respiratory functional tests and bronchoscopy with transbronchial lung biopsy were normal. The transthoracic echocardiography with Doppler showed abnormal left ventricular relaxation and systolic dysfunction of the left ventricle with ventricular ejection fraction (EF) of 40%. In the light of these symptoms and echocardiographic abnormalities, for which other possible causes have been excluded, the diagnosis of sarcoidosis with heart involvement was suspected. Thus, more specialized cardiac evaluation was needed. 24 hour holter monitoring showed recurrent episodes of 2000 monomorphic ventricular extrasystoles. Cardiac magnetic resonance imaging (MRI) with gadolinium contrast showed evidence of delayed hyper enhancement in both the basal and inferolateral left ventricular regions, suspicious for fibrogranulomatous tissue of sarcoidosis . The lefA 48-year-old Tunisian woman, presented to our consultations with a history of intermittent palpitations and exertional dyspnea of 4 months duration, progressively worsening for 2 weeks before, without any history of chest pain or syncope. She had a past history of systemic sarcoidosis diagnosed in April 2009 on the basis of the following compatible clinical and radiologic findings: systemic symptoms , parotid enlargement, negative tuberculin skin test, lymphopenia, hypercalciuria, increased serum ACE level, intrathoracic lymphadenopathies which were hilar , bilaterSarcoidosis is a multisystemic disorder of unknown etiology which is characterized by the formation of non-caseating granulomas in involved tissues. To this day, it remains something of enigma, the search for an etiologic agent is ongoing . SarcoidMaking a diagnosis of cardiac sarcoidosis remains one of the main challenges in the management of the disease: in fact, if cardiac manifestations occur in patients with multisystemic sarcoidosis, the diagnosis, although circumstancial, is strongly suspected, which is in keeping with our 2 cases. However, when cardiac dysfunction is the only manifestation of sarcoidosis, the diagnosis is frequently not entertained. Clinical manifestations of cardiac sarcoidosis, ranging from asymptomatic conduction abnormalities to fatal ventricular arrhythmia or sudden death, are dependent on both the profusion and location of granulomas. Granulomas can involve the left ventricular free wall, basal ventricular septum, right ventricle, papillary muscle, right atrium, and left atrium . Main reConsidering the increased risk of sudden death, cardiac sarcoidosis is an indication for early treatment. Steroids should be started as soon as possible: many reports showed that, in early or middle stage disease, steroid therapy may be protective or therapeutic but may not be as effective in the late stages . In bothSome authors recommended ICD implantation in any patient with sarcoidosis and non-sustained ventricular tachyarrhythmia because of the high rate of recurrence of ventricular tachycardia despite medical treatment , 9. As fCardiac sarcoidosis should always be suspected in any patient who is younger than expected presenting with cardiac symptoms without evidence of primary heart disease particularly if he has already been diagnosed with sarcoidosis. To date, early diagnosis of cardiac sarcoidosis remains a challenge because clinical manifestations are non specific and the diagnostic modalities lack sensitivity and specificity in spite of recent advances. A negative endomyocardial biopsy should in no way be regarded as evidence of absence of cardiac sarcoidosis, especially if diagnostic suspicion is strong. Cardiac sarcoidosis is on occasion a medical emergency that may lead to sudden death due to the rhythm disturbances or progressive heart failure from a dilated cardiomyopathy; therefore, in addition to corticosteroid treatment, early consideration for implantation of an ICD should be a priority.


The humidity sensing characteristics of different sensing materials are important properties in order to monitor different products or events in a wide range of industrial sectors, research and development laboratories as well as daily life. The primary aim of this study is to compare the sensing characteristics, including impedance or resistance, capacitance, hysteresis, recovery and response times, and stability with respect to relative humidity, frequency, and temperature, of different materials. Various materials, including ceramics, semiconductors, and polymers, used for sensing relative humidity have been reviewed. Correlations of the different electrical characteristics of different doped sensor materials as the most unique feature of a material have been noted. The electrical properties of different sensor materials are found to change significantly with the morphological changes, doping concentration of different materials and film thickness of the substrate. Various applications and scopes are pointed out in the review article. We extensively reviewed almost all main kinds of relative humidity sensors and how their electrical characteristics vary with different doping concentrations, film thickness and basic sensing materials. Based on statistical tests, the zinc oxide-based sensing material is best for humidity sensor design since it shows extremely low hysteresis loss, minimum response and recovery times and excellent stability. The regulation of humidity is important for human comfort, storage of various goods, industrial process control, high-tech instruments, and plenty of advanced sectors . In induIn 1450, Nicolas Cryfts invented a hygrometer which is the first humidity measurement instrument on record. This hygrometer used wool to determine the changes of humidity in air ,3. Over Humidity is evaluated by different functions such as vapor pressure, saturation vapor pressure, dew/frost point temperature, and relative humidity . Vapor pRelative humidity  is a ratio of the vapor pressure (VP) of moist air to its saturation vapor pressure (SVP) at a given temperature, which is expressed by 3), defined as the amount of water vapor contained in a unit volume of dry air, i.e., mixing ratio mr  and the ratio with respect to saturation is defined as saturation deficit psd (mbar).Humidity can also be expressed in other ways like absolute humidity , food processing and storage, medicine, ecology, agriculture, mineral processing, fuel quality control, and aerospace. Finally the concluding remarks are provided in Section 5.2.For many sophisticated applications, there has been a keen attempt to reduce the size of the sensors using the most advanced technologies collectively called miniaturization. Miniaturized humidity sensors mainly depend on five different transduction principles: hygrometry, capacity, resistance, gravimetry and optical properties.2.1.For many decades hygrometric sensors have been using in old devices that are classified into four basic types—mechanical, dry bulb-wet bulb, chilled mirror, LiCl dew point—for the measurement of the humidity of air, which is converted into mechanical energy –10. ThesAdvantages:Low cost;No power supply is required;It provides a primary rather than a secondary measurement of dew point;High accuracy;High repeatability;Traceable to N.I.S.T. or other national standards labs;Wide dew point range;Wide temperature range;Low hysteresis;No drift.Disadvantages:More complex, may be more expensive;Narrow flow rate range;Periodic cleaning may be required;High uncertainty (±2% to 5%).Generally, hygrometric sensors have two different materials with different thermal expansions coefficients, such as a polysilicon membrane and a hygroscopic material such as polyimide. The polyimide as a hygroscopic coating expands due to the absorption of water. Due to the unequal expansion that occurs between the polyimide and polysilicon, a change in the bending structure of the membranes is observed. The change in bending is converted into an electrical signal via a piezoresistor. The hygrometric sensor has some advantages as well as some disadvantages which are listed below:2.2.R(hr) represents the resistance at relative humidity (RH) at a specific humid condition h, Rr0 is the resistance at zero humidity, and a and b are the constants.The most common resistive sensors are based on electrical resistance. The electrical resistance of a material is the opposition to the passage of an electric current through that conducting material. Since the humidity can change the electrical resistance or impedance of a material noticeably, resistive humidity sensors have been developed using this principle. Generally the all resistive sensors have four layers: substrate, interdigital electrode, humidity sensitive film and protective layer which are shown in RS) can be measured using an expression given in Rh refers to the resistance at certain humidity and R0 represents the original resistance of the sensors at air of 3% RH.The sensing response (4BF4-PEG600 thin film resistive type humidity sensor is based on cellulose acetate (CA), ammonium tetrafluoroborate (NH4BF4) and polyethylene glycol (PEG600) . L. L106]. et al., had shown that 0.45 mol % of Li-doping in a γ-Fe2O3-based humidity senor is critical, since it showed maximum linearity . M. M2O3 se3.3.2.+ addition in a LaCo0.3Fe0.7O3 thick film sensor, the resistance decreases effectively as the frequency increases at a low RH range and the difference in resistance value between two working frequencies becomes smaller at a high RH range . T. T+ addi3.3.3.i.e., in the range of 10−11–10−12 F). The capacitance value increases with increased Li doping concentration in iron oxide film over the full RH range at 1 kHz frequency and at high RH (90%), the capacitance value increased by about one order in magnitude  in the low frequency range  and in the high frequency region , the capacitance becomes almost independent of RH , the value of capacitance increases with the increasing value of RH  and in the high frequency region (10–100 kHz), the capacitance becomes almost independent of RH, is illustrated in Due to the addition of K3.3.4.+ ion may improve the response time of an iron-based humidity sensor. The response times of MgFe2O4 (in the 53%–98% RH range) and Mg0.9Sn0.1Fe2O4 (in the 0%–53% RH range) sensors, which are 5 min and 3 min, respectively, is shown in + doping in an iron-based humidity sensor, the response and recovery times in a humidity range of 33%–95% RH are 100 s and 120 s, respectively . I. I+-ion ty range .3.3.5.+-ion-doped iron oxide perovskite type nanocrystals show narrow hysteresis loops during cyclic humidity operation. The hysteresis of a 2 wt % K2CO3-doped sample between humidification and desiccation processes over the whole humidity range with 1 V, 100 Hz and at room temperature is shown in 1−xKxCo0.3Fe0.7O3−δ perovskite humidity sensor is observed . O. O125]. 2 nanoparticles, polypyrrole and AgNO3 based humidity sensor, impedance varies significantly with RH with addition of different doping concentration. The impedance decreases with increases of doping concentration in all relative humidity range. The main advantages of the conducting polypyrrole polymers (PPy) are relatively good environmental stability and most importantly its surface charges can easily be modified by different dopants by easy synthesis method . P. P128]. s method . It has y sensor . The basng, 2007 . It has ng, 2007  gives fang, 2007  sensor.5 to 7.0 × 104 Ω with an increase in RH from 11% to 95% with greater linearity and good sensitivity (0.0370), but copolymer film has poor linearity and less sensitivity (0.0305) ..170].3.9.2) has being selected for advanced humidity sensors due to its unique properties, such as high thermally and excellent chemically stability, high toughness, high strength, and good catalytic activities, which make it more attractive compared to other materials in wide range of applications. Very little work has been reported on ZrO2-based humidity sensors so far. Wang et al. [2 thick film humidity sensor, which was made of ZrO2 nanoparticles with an average grain size 20 nm, was used to deposit it as a thick film on a silicon substrate. Y3+-doped and Mg2+-doped zirconia thick film humidity sensors have also been investigated by Su et al. [Many kinds of materials like ceramics, organic polymers, semiconductors, and solid electrolytes have been studied for humidity analysis. However, recently zirconia -doped ZrO2 humidity sensors [Like the other materials, the impedance decreases as the frequency increases and the impedance became independent of the humidity with increasing frequency. It has been found that the best linearity of the impedance–RH characteristic appears at frequencies between 100 Hz and 1 kHz for ZrO sensors , however sensors , as depi3.9.3.2-based sensors change strongly with doping agent. For example, the response time of a Y3+-doped ZrO2 humidity sensor is 30 s and for a Mg2+-doped sensor it is 5 s; and the recovery time for both the Y3+ and Mg2+-doped ZrO2 sensor is 5 s [2 humidity sensor are 130 s and 60 s, respectively [2+-doped ZrO2 sensor has the smallest response and recovery time, and can be considered as the best compared to the pure ZrO2 or Y3+-doped ZrO2 sensors.The response and recovery times of ZrOr is 5 s . The resectively . Therefo3.9.4.2 sensor is about 8% RH [3+- and Mg2+-doped ZrO2 sensors, respectively [2-based ceramic sensors, for less hysteresis value of a Y3+-doped sensor can be selected as the most suitable material for humidity measuring devices.The humidity hysteresis of the pure ZrOut 8% RH , while oectively . Therefo3.10.et al. [3. The changes in electrical characteristics  with the variation of frequency, temperature and relative humidity of sodium-based sensor materials are explained in the following subsections.Oxide ceramic materials have mostly been used in humidity sensor applications. Sodium oxide-based sensors also are being tried in humidity sensing devices due to its attractive thermal, physical and chemical stability like that of other ceramic materials. However, the main drawback of normal ceramics-based humidity sensors is their insufficient sensitivity over a wide humidity range, as well as lack of reversibility and drift in base resistance with time due to chemisorption of water molecules. Therefore, to overcome this problem many researchers are trying to use sodium-based materials to promote the sensitivity. Among them Na- and K-montmorillonite  sensors et al.  develope3.10.1.et al. [Su et al.  observed3.10.2.3-based humidity sensors [3-based humidity sensor has a better linearity than the pure Na-montmorillonite one.In both pure Na-montmorillonite-  and NaTa sensors , the imp3-based sensor the best linearity is observed at 100 Hz  is an important biomarker which has a significant role in the human body. Ammonia is present in all body fluids, mainly as ammonium ion (NH4+) but also in the form of NH3. A high concentration of NH3 is toxic to the human body. During the last decades, considerable effort has been put into detecting gaseous exhaled breath ammonia (eNH3) as a surrogate for blood or urine detection. Schmidt et al. [Advanced telehomecare (THC) systems use humidity sensors to monitor serious changes in chronic conditions as well as other health risks including floods, fires and gas leaks. A team led by Haick at the Israel Institute of Technology (Technion) made a humidity sensor breathing tests to detect lung cancer which contains multiple gold nanoparticle-based chemiresistors that can detect chemicals through changes in electrical resistance. The approach for breath-volatile organic compound (VOC) collection and preconcentration by applying needle traps was developed and optimized by Filipiak et al. . Here hut et al.  developeBreath monitoring is essential during certain imaging and surgical procedures where the patient needs to be sedated or anesthetized . Humiditet al. [et al. [To study the progression of a diagnosed illness or to evaluate the health of a person it is very important to monitor the breath. It is not at all wise to use electronic breathing sensors when patients are in a magnetic resonance imaging (MRI) system, or during any oncological treatment, which may create hazards for the patient . A simplet al.  and an a [et al. .et al. [The most familiar and common form of communication is voice communication. Due to some hereditary reasons or acquired impediments or due to other reasons like an accident, there are many people who suffer from speech/hearing difficulties. A humidity sensor-based language recognition system that focuses on the moisture included in devoiced breaths as a method for communication support in persons with speaking difficulties was developed by Morisawa et al. .4.2.Humidity controlled incubators are one of the essential pieces of equipment in tissue engineering applications. Normally 50%–60% RH is maintained for growing animal cells in the incubator.4.3.The shelf life of fruits and vegetables may change with humidity. Excessive water loss from fruits during storage or transportation often reduces its shelf-life. Depending on the type of fruits and the storage methods, the environment can be infected with different bacteria . Therefo4.4.et al. [Analysis of the root distribution of plant and its water absorbing ability is an important subject of considerable interest in ecology and agriculture. It gives a better understanding of the behaviour of different crops under sub-optimal environments, so that the improvement of the quality of modelling of root water uptake in different hydrological environment is possible . The effet al.  and it w4.5.The most widely used areas of humidity and moisture sensors are in structural health monitoring (SHM) –197 appl4.6.To reduce erosion in mountain areas humidity sensors is used by different regulated methods such as check dams, often made of concrete or wood-logs, which decrease the water speed during storm events, allowing sediment to settle .4.7.The study of moisture content in soil has also been a subject of considerable interest in mineral processing plants. Manual gravimetric drying moisture determination methods are still currently employed by most mineral processing plants, but they fails to provide timely and accurate information . An effi4.8.et al. [in situ wireless sensing system for a proton exchange membrane fuel cell (PEMFC) application.In many ways, biomass is a new source of power. Biomass is the energy which is contained inside plants and animals. The variation of moisture contents affects its energy-producing ability. The moisture content in biomass fuel is therefore an important parameter, which often fluctuates . By usinet al. ,189 deve4.9.in situ measurements in space, most spectacularly of the near-surface atmospheric water content on Mars [Recently, humidity sensors have been used to perform  on Mars –203. The4.10.et al. [The Conseil Européen pour la Recherche Nucléaire (European Council for Nuclear Research or CERN)  is the wet al. . This wo4.11.Humidity controlled dark rooms are used for many electronics materials or electronic devices during manufacturing. Power circuit boards (PCBs) are manufactured in controlled humidity dark rooms. Nowadays for low power consumptions in most of the industrial application, radio frequency identification (RFID) tags  based fl5.i.e., >3% for hysteresis, >19 s for response and recovery times), very good  and excellent . Since stability is the change in resistance or capacitance curves with respect to humidity over a long time, according to observation, it is directly graded as good, very good and excellent according to our flatness for a particular humidity sensor material.Humidity sensors for measuring relative humidity based on different types of materials, including carbon, vanadium, iron, silicon, polymer, tin, titanium, zinc, zirconia, and sodium have been reviewed extensively. The electrical properties of humidity sensors such as resistance and capacitance, the effects of temperature, frequency, and relative humidity, sensitivity, response time, hysteresis and stability have been compared in detail for various materials. For different sensor materials, the electrical properties are changed significantly with the doping concentration of different materials, film thickness of the substrate and the resulting morphological changes. The basic principle of six different types of humidity sensors and their typical advantages as well as disadvantages are discussed in the miniaturization section. Based on our overall review, the general electrical responses such as resistance, capacitance, hysteresis, response and recovery times, stability and their best characteristic features for different materials with respect to relative humidity, frequency and temperature are listed in Although response and hysteresis loss are a little less in polymeric sensors compared to zinc oxide-based humidity sensors, the recovery time is significantly larger in polymeric sensors than that of zinc oxide-based sensor materials. Based on our review, it has been found that the zinc oxide-based sensor material is the best for humidity sensor design due to its extremely low hysteresis loss, minimum response and recovery times, and excellent stability. Humidity sensors have a tremendous range of applications in different important fields such as biomedicine, SHM, food processing and storage, medicine, ecology, agriculture, mineral processing, fuel quality control, aerospace, the electronics industry and high-energy physics applications. The humidity conditions, including temperature and % RH, are also very important for different manufacturing or processing sectors as well as their specific areas. Therefore, the development of more integrated humidity sensors for multi-field applications would be highly fascinating topic for the new generation of humidity sensing devices. The review has provided examples that represent the diversity of applications and the demand for RH or moisture sensors in industry today. Therefore, it can be expected that different advanced humidity sensors made of hybrid or doped materials using the latest nanotechnology will be able to monitor a wide range of application sectors.


Confocal Raman spectroscopy has emerged as a major, versatile workhorse for the non-invasive characterization of graphene. Although it is successfully used to determine the number of layers, the quality of edges, and the effects of strain, doping and disorder, the nature of the experimentally observed broadening of the most prominent Raman 2D line has remained unclear. Here we show that the observed 2D line width contains valuable information on strain variations in graphene on length scales far below the laser spot size, that is, on the nanometre-scale. This finding is highly relevant as it has been shown recently that such nanometre-scaled strain variations limit the carrier mobility in high-quality graphene devices. Consequently, the 2D line width is a good and easily accessible quantity for classifying the crystalline quality, nanometre-scale flatness as well as local electronic properties of graphene, all important for future scientific and industrial applications. Raman spectroscopy has become an invaluable tool for graphene characterisation, yet the nature of the broadening of the Raman 2D line remains unclear. Here, Stampfer et al. show that the Raman 2D line width is a measure of nanometre-scale strain variations in graphene on insulating substrates. Graphene combines several highly interesting material properties in a unique way, promising unprecedented material functionality. This makes graphene increasingly attractive for fundamental research as well as industrial applications2456810731415In this article, we show that the experimentally observed Raman 2D line width is a measure of nanometre-scale strain variations in graphene on insulating substrates, that is, it contains valuable information on local  flatness, lattice deformations and crystal quality of graphene. Our findings solve the long-standing question of the nature of the observed broadening of the Raman 2D line and also link this quantity to the electronic transport properties of graphene, making it a valuable quantity for classifying the quality of graphene. To prove that the experimentally observed 2D line width depends on sub-spot-size strain variations and lattice deformations, we employ the following strategy.B-field dependent. In agreement with existing theory19202123B-field of ∼8 T, the G line becomes almost independent of electronic properties such as charge carrier doping, screening, or electronic broadening.We start by showing that by combining Raman spectroscopy with magnetic fields, electronic broadening contributions for the Raman G line width can be strongly suppressed. Since in perpendicular magnetic fields the electronic states in graphene condense into Landau levels (LLs), the interaction between electronic excitations and lattice vibrations becomes We observe that, under these conditions, the G line width nevertheless exhibits strong variations across graphene flakes. In particular, we show that the G line width is significantly increased in regions where the graphene flake features bubbles and folds, that is, in correspondence with increased structural deformations.B-fieldFinally, we show that at 8 T, there is a (nearly) linear dependence between the G line width and the 2D line width, implying that there is a common source of line broadening. According to the previous points, the broadening must be related to structural lattice deformations. This finding is further supported by a detailed analysis of the relation between the area of the 2D peak and its line width. By analysing the relation between the G and 2D line width, we find that nanometre-scale strain variations constitute a dominant contribution to the observed line broadenings. Importantly, the 2D line has been shown to only very weakly depend on the 2 and hBN as illustrated in 2 usually feature lower carrier mobilities of around 103–104 cm2/(Vs), indicating a detrimental influence of SiO2 on the electronic properties of graphene. In this regard, our structure gives us the invaluable capability of probing a single graphene sheet exposed to two different substrates  sheet is partly encapsulated in hexagonal boron nitride (hBN) and partly sandwiched between SiO2 areas can be clearly distinguished in the map (see highlighted regions I and II). While analysing the Raman spectra of every point on the map, it is evident that the G lines recorded in the hBN-encapsulated area are broader than in the SiO2 supported area  is used to suppress Landau damping of the G mode, leaving ΓG unaffected from electronic contributions. The coloured data points stem from Raman maps (B=0 T) of chemical vapour deposition (CVD)-grown graphene flakes that were transferred onto SiO2 by a wet chemistry-based transfer. These graphene sheets contain doping values of nel>3 × 1012 cm−2, which corresponds to Fermi energies EF>200 meV  or to the presence of defects (increasing the electron-defect scattering)38402D and the integrated area of the 2D peak (area2D) we can exclude such a hypothesis. In 2D versus the region-normalized area2D for both B=8 T and B=0 T, highlighting the very weak B-field dependence of Γ2D. More importantly, we observe that the area of the 2D peak does not depend on Γ2D, contrary to what is expected in presence of a variation of the electronic broadening γ  delivered through a single-mode optical fibre, where the spot size is limited by diffraction. Using a long working distance focusing lens with a numerical aperture of 0.80, we obtained a spot size of ∼400–500 nm. For the low-temperature Raman measurements, we employ a commercially available confocal Raman setup that allows us to perform spatially resolved experiments at a temperature of 4.2 K and magnetic fields of up to 9 T. We use an excitation laser wavelength of 532 nm with a spot diameter on the sample of ∼500 nm. For detection, we use a single-mode optical fibre and a charge-coupled spectrometer with a grating of 1,200 lines mmet al.−1 wide Lorentzianset al.2D as a function of area2D. For a laser energy of 2.33 eV, the electron–phonon contribution for the electronic broadening is 81.9 meV, which leads to Γ2D=12.1 cm−1. However, when we choose γ to be twice this value, to account for additional electron–electron interaction2D equal to 17.9 cm−1. These values can be understood as a theoretical expectation of the 2D line width for a perfect graphene lattice disregarding any broadening from averaging effects over different strain values within the laser spot.For the computation of the double-resonant Raman cross-section we employ an approach based on Fermi's golden rule generalized to the fourth perturbative order as described in detail in Reference Venezuela How to cite this article: Neumann, C. et al. Raman spectroscopy as probe of nanometre-scale strain variations in graphene. Nat. Commun. 6:8429 doi: 10.1038/ncomms9429 (2015).Supplementary Figures 1-3


The molecular machinery responsible for cytosolic accumulation of misfolded TDP-43 in amyotrophic lateral sclerosis (ALS) remains elusive. Here we identified a cullin-2 (CUL2) RING complex as a novel ubiquitin ligase for fragmented forms of TDP-43. The von Hippel Lindau protein (VHL), a substrate binding component of the complex, preferentially recognized misfolded TDP-43 at Glu246 in RNA-recognition motif 2. Recombinant full-length TDP-43 was structurally fragile and readily cleaved, suggesting that misfolded TDP-43 is cleared by VHL/CUL2 in a step-wise manner via fragmentation. Surprisingly, excess VHL stabilized and led to inclusion formation of TDP-43, as well as mutant SOD1, at the juxtanuclear protein quality control center. Moreover, TDP-43 knockdown elevated VHL expression in cultured cells, implying an aberrant interaction between VHL and mislocalized TDP-43 in ALS. Finally, cytoplasmic inclusions especially in oligodendrocytes in ALS spinal cords were immunoreactive to both phosphorylated TDP-43 and VHL. Thus, our results suggest that an imbalance in VHL and CUL2 may underlie oligodendrocyte dysfunction in ALS, and highlight CUL2 E3 ligase emerges as a novel therapeutic potential for ALS. Amyotrophic lateral sclerosis (ALS) is a life-threatening neurodegenerative disorder that is characterized by progressive muscle atrophy and weakness. The majority of ALS cases are caused by abnormal conformation of ALS-linked proteins and by defective RNA handling, both of which induce various downstream cascades, including excitotoxicity, endoplasmic reticulum stress, mitochondrial dysfunction, glial dysfunction, proteasome impairment, unexpected secretion, and neuroinflammation12467The exact mechanism of RNA mishandling in TDP-43 proteinopathy is unclear, but a loss of nuclear TDP-43 due to cytosolic mislocalization is considered a likely mechanism. Indeed, aggregate-prone TDP-43 proteins are readily mislocalized in the cytosol and sequester nuclear TDP-4391045VHL is an anti-oncogenic gene that was originally identified as a cause of von Hippel Lindau disease, a dominantly inherited tumorigenic disorder in multiple organs. The VHL protein is a substrate binding protein in the CUL2 E3 complex and has been implicated in diverse pathways, including ubiquitination of hypoxia inducing factors (HIF)151819Here, we found that the von Hippel Lindau (VHL)/cullin-2 (CUL2) E3 complex recognized and ubiquitinated misfolded TDP-43 and promoted clearance of fragmented forms of TDP-43. in vitro ubiquitination and a reversible covalent immunoprecipitation assayBecause of the potential failure to seize labile protein complexes in specific conditions such as ubiquitination, we integrated Based on the finding that VHL and CUL2 colocalize with cytosolic inclusions of TDP-43, we investigated whether VHL/CUL2 recognizes misfolded forms of TDP-43 more than normally folded TDP-43. We previously reported that the conformation of TDP-43 is tightly regulated by two cysteine residues in RRM1, the substitution of which (C173S and/or C175S) leads to formation of aggregate-prone TDP-43 that shares diverse features of pathogenic inclusions in ALSWe next attempted to identify the site in TDP-43 molecules that is recognized by VHL. First, a pull-down study using domain deletion mutants revealed that RRM2, but not RRM1 or the C-terminus, was required for the interaction with VHL . Interesin vivo ubiquitination assay showed no clear trend toward increased ubiquitination of FL TDP-43 by CUL2  rather than in motor neurons by immunohistochemistry. Why TDP-43 inclusions in motor neurons and oligodendrocytes showed different colocalization patterns for VHL remains elusive, the abundant expression and the more active roles of VHL/CUL2 in oligodendrocytes for their maturation from OPCs could underlie the interaction between VHL and misfolded TDP-43 than in motor neurons1819In conclusion, we identified VHL/CUL2 proteins as a novel E3 ligase for misfolded forms of TDP-43. The substrate-binding protein, VHL, plays bidirectional roles by promoting ubiquitin-mediated degradation of fragmented TDP-43 at proteasomes with CUL2, and by stabilizing TDP-43 at JUNQ when upregulated. The deregulation of this pathway would cause oligodendrocyte failure in ALS. Further investigation of regulation of the VHL/CUL2 balance may suggest therapeutic strategies for ALS.in vitro ubiquitination2, 2 mM DTT, 0.5 μg ubiquitin aldehyde , 5 μg ubiquitin, 30 μg S100 lysate 0.5 mM ATP, 10 mM creatine phosphate, and 0.5 μg of creatine phosphokinase in a total volume of 20 μl) was incubated for 30 min at 37 °C. The disulfide-dependent cross-linker, Dithiobis[succinimidyl propionate] (DSP), was then added to the mixture at a concentration of 40 mM and was allowed to cross-link for another 30 min. Disulfide formation was terminated with excess l-cysteine . TDP-43-FLAG proteins were pulled down with anti-FLAG affinity beads , and disulfide-mediated binding proteins were released by mild reducing conditions (50 mM DTT at 37 °C for 30 min). The eluates were separated on an acrylamide gel and visualized with a fluorescent dye . Positive bands, which disappeared in the absence of ATP or TDP-43-FLAG, were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), as previously describedTDP-43-interacting proteins during the ubiquitin-proteasome reaction (UPR) were investigated with a combination of the reversible covalent-linked immunoprecipitation (ReCLIP) assayRabbit polyclonal anti-DYKDDDD antibody and mouse anti-GFP antibody were purchased from Cell Signaling Technology  and Nacalai Tesque , respectively. Rabbit polyclonal anti-Cullin2 (CUL2) and anti-TDP-43 antibodies were purchased from Abcam  and Proteintech , respectively. Rat monoclonal anti-HA antibody was obtained from Roche . Mouse monoclonal antibodies against actin (C4), elongin B (D-5), and elongin C (56) were obtained from Santa Cruz Biotechnology . Mouse monoclonal anti-FLAG antibody (M2) was obtained from Sigma. Rabbit anti-ubiquitin, Lys48-specific antibody was obtained from EMD Millipore . Mouse anti-Hsc70/Hsp70 antibody was purchased from StressGen . Mouse anti-HIF1α antibody was obtained from Abcam. Goat anti-mouse IgG Alexa fluor488 and fluor405 were purchased from Invitrogen , and goat anti-rabbit IgG CF568 was from BIOTIM . Mouse monoclonal anti-human VHL was generated as previously describedMammalian expression plasmids for TDP-43 tagged with FLAG (pcDNA3-TDP-43-FLAG) or EGFP (pEGFP-N3-TDP-43) were constructed using a conventional PCR technique as described previouslyThe mutant C-terminal fragment 35 kDa  tagged with FLAG (pcDNA3-CTF35-FLAG) and CUL2 tagged with Myc (pCMV-Myc-CUL2) were constructed using conventional PCR and subcloned into pcDNA3-FLAG using the BamHI/XhoI sites or pCMV-Myc using the BamHI/KpnI sites. Human VHL tagged with HA (pHM6-HA-VHL) was constructed using conventional PCR and subcloned into pHM6-HA using the HindIII/EcoRI sites. The primer pairs are described in 2 and 100% humidity. HeLa cells and HEK293A cells (Invitrogen) were maintained in Dulbecco’s modified Eagle’s medium  containing 10% fetal bovine serum and penicillin/streptomycin . LipofectamineTM 2000 (Invitrogen) and FuGene HD Transfection Reagent  were used for transfection of the plasmid according to the manufacturer’s protocols. For the RNA knockdown experiment, short interference RNA pairs were transduced into cells using Lipofectamine2000 and Magnetofection system . Incubation time for the overexpression and siRNA knockdown transfection before the harvest, cells were 48 h and 96 h, respectively, unless mentioned.All cultured cells were maintained at 37 °C in 5% COAt 48 h after the transfection, cultured cells were lysed in co-immunoprecipitation buffer  or RIPA buffer , containing protease inhibitor cocktail (Roche) and PR-619 , a de-ubiquitination inhibitor for ubiquitination assay. In several experiments, RIPA-insoluble fractions were solubilized in 8 M urea. Ten percent of each cell lysate was analyzed as the total cell lysate, and the remaining 90% was incubated with anti-FLAG M2 affinity gel (Sigma) or beads coated with anti-HA antibody  at 4 °C overnight. The immunoprecipitates were eluted into sodium dodecyl sulfate (SDS) sample buffer. Samples were reacted in one-third volume of 4× SDS buffer, denatured at 95 °C for 5 min, separated on a 5–20% or 15% polyacrylamide gel (Wako), and transferred onto a PVDF membrane (Millipore) for Western blotting. Proteins were detected using an enhanced chemiluminescence system , and the densitometric analysis of protein was performed using Image J softwareAt 48 h after the transfection, cultured cells were fixed in 4% paraformaldehyde in PBS, pH 7.2, for 20 min at room temperature and permeabilized with 0.1% Triton X-100 containing 5% normal goat serum as blocking agent. Cells were reacted with primary antibody (4 °C overnight) and subsequently with fluorescence-tagged secondary antibody  for 1 h at room temperature. Cells were counter-stained with 4′,6-diamidino-2-phenylindole (DAPI). Fluorescent images were obtained using a confocal laser microscope FV1000-D IX81  or fluorescence microscope .Information about the antibodies used is described in Protein half-life was estimated 48 h after transfection by chronological chase analysis at 5, 10, and 24 h after CHX treatment (100 μg) of HEK293A cells to inhibit protein synthesis. The cells were directly lysed in the SDS sampling buffer containing 100 mM DTT and were denatured at 70 °C for 20 min. The remaining amount of protein as estimated by densitometry was expressed as the percentage of the residual protein level compared with that at the starting point (0 h). To determine the effect of CUL2 on transgene expression, cells were transfected with siRNAs for CUL2 , incubated for a further 48 h, and subcultured before transfection with TDP-43 plasmids for the chase study.The total RNA samples were purified from the transfected cells using a commercially available kit (Invitrogen) and were converted to cDNA with reverse transcriptase . The levels of mRNAs for VHL and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in the synthesized cDNAs were analyzed with the SYBR quantitative PCR kit (Toyobo) and real-time PCR Detection Systems (BIO-RAD). GAPDH was used as an internal standard, and the relative expression levels of mRNAs were calculated by the ΔCT method according to the manufacturer’s protocol using the included software (BIO-RAD). The primer sequences are described in 6 cells/dish. At 48 h after the transfection, all lysates were analyzed for proteasome activity, which was determined at 360 nm excitation and 480 nm emission using a multi-plate reader .Chymotrypsin-like activity was measured as proteasome activity using a commercially available kit . HEK293A cells were seeded in 6-cm culture dishes at a density of 1 × 10All experiments were approved by and performed under the guidelines of the Kyoto University ethics committee. Informed consent was obtained from all individuals or their guardians before the autopsy analysis. Postmortem lumbar spinal cords from five patients with a definite diagnosis of sporadic ALS and five neurological disease controls other than ALS  were analyzed. At autopsy, spinal cord blocks from the lumbar level were paraffinized after fixation in 10% buffered formalin. Sections with 6 μm-thickness were deparaffinized and antigen retrieved by autoclaving (20 min at 120 °C), using Histofine deparaffinizing antigen retrieval buffer, pH 6 . Sections were then incubated overnight with primary antibodies in PBS containing 3% bovine serum albumin at 4 °C. The bound antibodies were visualized by the peroxidase polymer-based method using a Histofine Simple Stain MAX-PO kit (Nichirei) with diaminobenzidine as the chromogen. Double immunofluorescence was performed with the similar protocols, except fluorescence-tagged secondary antibodies were used, as mentioned above.Primary cultures of embryonic rat oligodendrocytes and their precursor cells (OPCs) were prepared as previously descriedet al. 2010). The spinal cord was homogenized in a tissue lysis buffer (20 mM HEPES–KOH (pH 7.4), 120 mM NaCl, 2 mM EDTA, 10% glycerol, 1% Triton X-100, and protease inhibitor cocktail) and then incubated for 1 h at 4 °C, and the supernatant was collected for Western blotting. The protein amount was adjusted according to Bradford assay . All study protocols were performed respecting the dignity of animal lives and were approved by the animal experimental committee of the Shiga University of Medical Science (Document# 2009-12-2).Transgenic mice harboring human G93A SOD1  were housed and bred as previously reported . The difference between the two groups was assessed by Stuent’s t test. How to cite this article: Uchida, T. et al. CUL2-mediated clearance of misfolded TDP-43 is paradoxically affected by VHL in oligodendrocytes in ALS. Sci. Rep.6, 19118; doi: 10.1038/srep19118 (2016).


Graphene devices for radio frequency (RF) applications are of great interest due to their excellent carrier mobility and saturation velocity. However, the insufficient current saturation in graphene field effect transistors (FETs) is a barrier preventing enhancements of the maximum oscillation frequency and voltage gain, both of which should be improved for RF transistors. Achieving a high output resistance is therefore a crucial step for graphene to be utilized in RF applications. In the present study, we report high output resistances and voltage gains in graphene-on-silicon (GoS) FETs. This is achieved by utilizing bare silicon as a supporting substrate without an insulating layer under the graphene. The GoSFETs exhibit a maximum output resistance of 2.5 MΩ∙μm, maximum intrinsic voltage gain of 28 dB, and maximum voltage gain of 9 dB. This method opens a new route to overcome the limitations of conventional graphene-on-insulator (GoI) FETs and subsequently brings graphene electronics closer to practical usage. The enormous interest in graphene for electronic device applications originates from its outstanding charge transport properties and atomic-scale thickness1214fT) with the help of gate length scalingfT values as high as 400 GHz were reported from state-of-the-art graphene RF FETs789101112131415fT, the maximum oscillation frequency (fmax) and intrinsic voltage gain (AV0) are more important factors for RF devicesfmax values as high as 105 GHz were reportedmax and AV0 values of conventional GoIFETs mainly originate from the absence of current saturation in the output characteristics due to the absence of a band gap and the Klein tunneling effectIntensive research activities examining graphene RF FETs thus far have mainly focused on achieving higher cut-off frequencies  and thereby degraded RF performance. Thus, by suppressing the generation of the other charged carrier in the channel near the drain, the graphene device is able to obtain a higher ro.It is therefore highly desirable to obtain robust current saturation in the output characteristics from graphene FETs, which would result in a higher n0)22One of the methods that has been attempted to suppress the generation of the other charged carriers is the strengthening of the gate electrostatic control. In other words, the influence of the drain voltage is weakened with respect to the graphene channel near the drain side. Thus, previous attempts have focused on increasing the gate coupling by increasing the gate capacitance1920h-BN) has been adopted as a bottom gate dielectric to minimize n022h-BN exhibits lower charged impurities and provide higher optical phonon energy than a conventional SiO2 substrate19n0, which was proportional to the charged impurities, disorder, and thermal excitation, was found to significantly affect the drain conductance , indicating that the silicon substrate successfully acted as an insulator under the reverse bias condition. In contrast to the reverse bias condition at VDS < 0 , which results in the doping of the graphene and degradation of the graphene devices2 were reported to be around 2 ~ 5 × 1011 cm−2, while those on the silicon surface were around 0.3 ~ 2.8 × 1010 cm−2 312 than graphene/Si interfaces.The cumulative results of the maximum ro is inversely proportional to RC for the GoSFETs, while ro exists irrespective of RC for the GoIFET. The lower contact resistance in the GoSFETs compared to that of the GoIFETs is attributed to the presence of additional charge injection into the graphene from the Si bottom substrate, in addition to the metal pad. Both Pd and Si atoms are able to inter-diffuse through the defects in the graphene, such as carbon atom vacancies, thereby form a conductive Pd silicide (ro and μ can be observed in 2 (50 meV)34The suppression capabilities that prevent the generation of the other polarity carrier in the channel near the drain are directly affected by the electron potential energy in the Si, which is influenced by the metal/graphene voltage drop. Therefore, it is expected that the maximum silicide  in the Sro was proportional to the maximum AV0, and that a maximum AV0 as high as 28 dB was achieved from the GoSFETs, which is comparable to that of Si-based RF FETs. In order to verify the performance of the voltage amplifier, a common-source voltage amplifier was implemented with a P-GoSFET, as shown in V as high as 9 dB was obtained at a load resistance (RD) of 50 kΩ  on a heavily doped p-type silicon wafer (resistivity of ~0.01–0.05 Ω∙cm). For the GoSFETs, the thermal oxide (50 nm) in the active region on lightly doped n- or p-type silicon (resistivity of ~1–5 Ω∙cm) was patterned using a buffered oxide etchant, and the graphene was transferred immediately after the removal of the photoresist and the native oxide. The subsequent processes were identical for both the GoSFETs and GoIFETs. After patterning the channel graphene, the source and drain were defined using a Pd (5 nm)/Au (50 nm) lift-off process. Next, 20 nm of Al2O3 was deposited by atomic layer deposition (ALD) process on the graphene surface. Prior to the ALD process, 0.5 nm aluminum layer was evaporated on the graphene surface as a nucleation layer. The ultra-thin aluminum nucleation layer is soon oxidized during the ALD process. The 150 nm Al gate metal was formed using a thermal evaporator and the lift-off process. Finally, the device went through a thermal budget of 300 °C for 12 hrs.The graphene used in this work was synthesized using the inductive coupling plasma chemical vapor deposition method on a thin copper film, and it was transferred using the typical wet transfer method with PMMA. For the conventional GoIFETs, the graphene was transferred onto thermally grown SiORG = 1 MΩ, RD = 10, 30, 50 kΩ) and capacitor (CG = 1 μF) set on a breadboard, as well as a DC supplier and function generator, were connected. The transistors were staged in a probe station and connected to the circuit with a biaxial cable. The input signal was sinusoidal with an amplitude of 20 mV and a frequency of 1 kHz.All of the devices were measured at room temperature in ambient air with a B1500 parameter analyzer. In order to implement a common source voltage amplifier with a graphene–on–silicon transistor, the external resistances .