Methylation matters: binding of Ets-1 to the demethylated Foxp3 gene contributes to the stabilization of Foxp3 expression in regulatory T cells

The forkhead-box protein P3 (Foxp3) is a key transcription factor for the development and suppressive activity of regulatory T cells (Tregs), a T cell subset critically involved in the maintenance of self-tolerance and prevention of over-shooting immune responses. However, the transcriptional regulation of Foxp3 expression remains incompletely understood. We have previously shown that epigenetic modifications in the CpG-rich Treg-specific demethylated region (TSDR) in the Foxp3 locus are associated with stable Foxp3 expression. We now demonstrate that the methylation state of the CpG motifs within the TSDR controls its transcriptional activity rather than a Treg-specific transcription factor network. By systematically mutating every CpG motif within the TSDR, we could identify four CpG motifs, which are critically determining the transcriptional activity of the TSDR and which serve as binding sites for essential transcription factors, such as CREB/ATF and NF-κB, which have previously been shown to bind to this element. The transcription factor Ets-1 was here identified as an additional molecular player that specifically binds to the TSDR in a demethylation-dependent manner in vitro. Disruption of the Ets-1 binding sites within the TSDR drastically reduced its transcriptional enhancer activity. In addition, we found Ets-1 bound to the demethylated TSDR in ex vivo isolated Tregs, but not to the methylated TSDR in conventional CD4+ T cells. We therefore propose that Ets-1 is part of a larger protein complex, which binds to the TSDR only in its demethylated state, thereby restricting stable Foxp3 expression to the Treg lineage

α-Adducin Gly460Trp Gene Mutation and Essential Hypertension in a Chinese Population: A Meta-Analysis including 10960 Subjects

BACKGROUND: The α-adducin Gly460Trp (G460W) gene polymorphism may be associated with susceptibility to essential hypertension (EH), but this relationship remains controversial. In an attempt to resolve this issue, we conducted a meta-analysis. METHODS: Twenty-three separated studies involving 5939 EH patients and 5021 controls were retrieved and analyzed. Four ethnicities were included: Han, Kazakh, Mongolian, and She. Eighteen studies with 5087 EH patients and 4183 controls were included in the Han subgroup. Three studies with 636 EH patients and 462 controls were included in the Kazakh subgroup. The Mongolian subgroup was represented by only one study with 100 EH patients and 50 controls; similarly, only one study with 116 EH patients and 326 controls was available for the She subgroup. The pooled and ethnic group odds ratios (ORs) along with the corresponding 95% confidence intervals (95% CI) were assessed using a random effects model. RESULTS: There was a significant association between the α-adducin G460W gene polymorphism and EH in the pooled Chinese population under both an allelic genetic model (OR: 1.12, 95% CI: 1.04-1.20, P = 0.002) and a recessive genetic model (OR: 1.40, 95% CI: 1.16-1.70, P = 0.0005). In contrast, no significant association between the α-adducin G460W gene polymorphism and EH was observed in the dominant genetic model (OR: 0.88, 95% CI: 0.72-1.09, P = 0.24). In stratified analysis by ethnicity, significantly increased risk was detected in the Han subgroup under an allelic genetic model (OR: 1.13, 95% CI: 1.04-1.23, P = 0.003) and a recessive genetic model (OR: 1.43, 95% CI: 1.17-1.75, P = 0.0006). CONCLUSIONS: In a Chinese population of mixed ethnicity, the α-adducin G460W gene polymorphism was linked to EH susceptibility, most strongly in Han Chinese

Lack of Association of Two Common Polymorphisms rs2910164 and rs11614913 with Susceptibility to Hepatocellular Carcinoma: A Meta-Analysis

BACKGROUND: Single nucleotide polymorphisms (SNPs) in microRNA-coding genes may participate in the process of carcinogenesis by altering the expression of tumor-related microRNAs. It has been suggested that two common SNPs rs2910164 in miR-146a and rs11614913 in miR-196a2 are associated with susceptibility to hepatocellular carcinoma (HCC). However, published results are inconsistent and inconclusive. In the present study, we performed a meta-analysis to systematically summarize the possible association between the two SNPs and the risk for HCC. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a search of case-control studies on the associations of SNPs rs2910164 and/or rs11614913 with susceptibility to HCC in PubMed, EMBASE, ISI Web of Science, Cochrane Central Register of Controlled Trials, ScienceDirect, Wiley Online Library and Chinese National Knowledge Infrastructure databases. Data from eligible studies were extracted for meta-analysis. HCC risk associated with the two polymorphisms was estimated by pooled odds ratios (ORs) and 95% confidence intervals (95% CIs). 5 studies on rs2910164 and 4 studies on rs11614913 were included in our meta-analysis. Our results showed that neither allele frequency nor genotype distribution of the two polymorphisms was associated with risk for HCC in all genetic models. Similarly, subgroup analysis in Chinese population showed no association between the two SNPs and the susceptibility to HCC. CONCLUSIONS/SIGNIFICANCE: This meta-analysis suggests that two common SNPs rs2910164 and rs11614913 are not associated with the risk of HCC. Well-designed studies with larger sample size and more ethnic groups are required to further validate the results

TIAF1 self-aggregation in peritumor capsule formation, spontaneous activation of SMAD-responsive promoter in p53-deficient environment, and cell death

Self-aggregation of transforming growth factor β (TGF-β)1-induced antiapoptotic factor (TIAF1) is known in the nondemented human hippocampus, and the aggregating process may lead to generation of amyloid β (Aβ) for causing neurodegeneration. Here, we determined that overexpressed TIAF1 exhibits as aggregates together with Smad4 and Aβ in the cancer stroma and peritumor capsules of solid tumors. Also, TIAF1/Aβ aggregates are shown on the interface between brain neural cells and the metastatic cancer cell mass. TIAF1 is upregulated in developing tumors, but may disappear in established metastatic cancer cells. Growing neuroblastoma cells on the extracellular matrices from other cancer cell types induced production of aggregated TIAF1 and Aβ. In vitro induction of TIAF1 self-association upregulated the expression of tumor suppressors Smad4 and WW domain-containing oxidoreductase (WOX1 or WWOX), and WOX1 in turn increased the TIAF1 expression. TIAF1/Smad4 interaction further enhanced Aβ formation. TIAF1 is known to suppress SMAD-regulated promoter activation. Intriguingly, without p53, self-aggregating TIAF1 spontaneously activated the SMAD-regulated promoter. TIAF1 was essential for p53-, WOX1- and dominant-negative JNK1-induced cell death. TIAF1, p53 and WOX1 acted synergistically in suppressing anchorage-independent growth, blocking cell migration and causing apoptosis. Together, TIAF1 shows an aggregation-dependent control of tumor progression and metastasis, and regulation of cell death

A Low T Regulatory Cell Response May Contribute to Both Viral Control and Generalized Immune Activation in HIV Controllers

HIV-infected individuals maintaining undetectable viremia in the absence of therapy (HIV controllers) often maintain high HIV-specific T cell responses, which has spurred the development of vaccines eliciting HIV-specific T cell responses. However, controllers also often have abnormally high T cell activation levels, potentially contributing to T cell dysfunction, CD4+ T cell depletion, and non-AIDS morbidity. We hypothesized that a weak T regulatory cell (Treg) response might contribute to the control of viral replication in HIV controllers, but might also contribute to generalized immune activation, contributing to CD4+ T cell loss. To address these hypotheses, we measured frequencies of activated (CD38+ HLA-DR+), regulatory (CD4+CD25+CD127dim), HIV-specific, and CMV-specific T cells among HIV controllers and 3 control populations: HIV-infected individuals with treatment-mediated viral suppression (ART-suppressed), untreated HIV-infected “non-controllers” with high levels of viremia, and HIV-uninfected individuals. Despite abnormally high T cell activation levels, controllers had lower Treg frequencies than HIV-uninfected controls (P = 0.014). Supporting the propensity for an unusually low Treg response to viral infection in HIV controllers, we observed unusually high CMV-specific CD4+ T cell frequencies and a strong correlation between HIV-specific CD4+ T cell responses and generalized CD8+ T cell activation levels in HIV controllers (P≤0.001). These data support a model in which low frequencies of Tregs in HIV controllers may contribute to an effective adaptive immune response, but may also contribute to generalized immune activation, potentially contributing to CD4 depletion

Expansion of CD4+CD25+ and CD25- T-Bet, GATA-3, Foxp3 and RORγt Cells in Allergic Inflammation, Local Lung Distribution and Chemokine Gene Expression

Allergic asthma is associated with airway eosinophilia, which is regulated by different T-effector cells. T cells express transcription factors T-bet, GATA-3, RORγt and Foxp3, representing Th1, Th2, Th17 and Treg cells respectively. No study has directly determined the relative presence of each of these T cell subsets concomitantly in a model of allergic airway inflammation. In this study we determined the degree of expansion of these T cell subsets, in the lungs of allergen challenged mice. Cell proliferation was determined by incorporation of 5-bromo-2′-deoxyuridine (BrdU) together with 7-aminoactnomycin (7-AAD). The immunohistochemical localisation of T cells in the lung microenvironments was also quantified. Local expression of cytokines, chemokines and receptor genes was measured using real-time RT-PCR array analysis in tissue sections isolated by laser microdissection and pressure catapulting technology. Allergen exposure increased the numbers of T-bet+, GATA-3+, RORγt+ and Foxp3+ cells in CD4+CD25+ and CD4+CD25- T cells, with the greatest expansion of GATA-3+ cells. The majority of CD4+CD25+ T-bet+, GATA-3+, RORγt+ and Foxp3+ cells had incorporated BrdU and underwent proliferation during allergen exposure. Allergen exposure led to the accumulation of T-bet+, GATA-3+ and Foxp3+ cells in peribronchial and alveolar tissue, GATA-3+ and Foxp3+ cells in perivascular tissue, and RORγt+ cells in alveolar tissue. A total of 28 cytokines, chemokines and receptor genes were altered more than 3 fold upon allergen exposure, with expression of half of the genes claimed in all three microenvironments. Our study shows that allergen exposure affects all T effector cells in lung, with a dominant of Th2 cells, but with different local cell distribution, probably due to a distinguished local inflammatory milieu

Infected erythrocyte-derived extracellular vesicles alter vascular function via regulatory Ago2-miRNA complexes in malaria

Malaria remains one of the greatest public health challenges worldwide, particularly in sub-Saharan Africa. The clinical outcome of individuals infected with Plasmodium falciparum parasites depends on many factors including host systemic inflammatory responses, parasite sequestration in tissues and vascular dysfunction. Production of pro-inflammatory cytokines and chemokines promotes endothelial activation as well as recruitment and infiltration of inflammatory cells, which in turn triggers further endothelial cell activation and parasite sequestration. Inflammatory responses are triggered in part by bioactive parasite products such as hemozoin and infected red blood cell-derived extracellular vesicles (iRBC-derived EVs). Here we demonstrate that such EVs contain functional miRNA-Argonaute 2 complexes that are derived from the host RBC. Moreover, we show that EVs are efficiently internalized by endothelial cells, where the miRNA-Argonaute 2 complexes modulate target gene expression and barrier properties. Altogether, these findings provide a mechanistic link between EVs and vascular dysfunction during malaria infection

Prevalence, Distribution and Functional Significance of the −237C to T Polymorphism in the IL-12Rβ2 Promoter in Indian Tuberculosis Patients

Cytokine/cytokine receptor gene polymorphisms related to structure/expression could impact immune response. Hence, the −237 polymorphic site in the 5′ promoter region of the IL-12Rβ2 (SNP ID: rs11810249) gene associated with the AP-4 transcription motif GAGCTG, was examined. Amplicons encompassing the polymorphism were generated from 46 pulmonary tuberculosis patients, 35 family contacts and 28 miscellaneous volunteers and sequenced. The C allele predominated among patients, (93.4%, 43/46), and in all volunteers and contacts screened, but the T allele was exclusively limited to patients, (6.5%, 3/46). The functional impact of this polymorphism on transcriptional activity was assessed by Luciferase-reporter and electrophoretic mobility shift assays (EMSA). Luciferase-reporter assays showed a significant reduction in transcriptional efficiency with T compared to C allele. The reduction in transcriptional efficiency with the T allele construct (pGIL-12Rb2-T), in U-87MG, THP-1 and Jurkat cell lines, were 53, 37.6, and 49.8% respectively, compared to the C allele construct (pGIL-12Rb2-C). Similarly, densitometric analysis of the EMSA assay showed reduced binding of the AP-4 transcription factor, to T compared to the C nucleotide probe. Reduced mRNA expression in all patients (3/3) harboring the T allele was seen, whereas individuals with the C allele exhibited high mRNA expression (17/25; 68%, p = 0.05). These observations were in agreement with the in vitro assessment of the promoter activity by Luciferase-reporter and EMSA assays. The reduced expression of IL-12Rβ2 transcripts in 8 patients despite having the C allele was attributed to the predominant over expression of the suppressors (IL-4 and GATA-3) and reduced expression of enhancers (IFN-α) of IL-12Rβ2 transcripts. The 17 high IL-12Rβ2 mRNA expressers had significantly elevated IFN-α mRNA levels compared to low expressers and volunteers. Notwithstanding the presence of high levels of IL-12Rβ2 mRNA in these patients elevated IFN-α expression could modulate their immune responses to Mycobacterium tuberculosis

Differentially expressed microRNAs in experimental cerebral malaria and their involvement in endocytosis, adherens junctions, FoxO and TGF-β signalling pathways

Cerebral malaria (CM) is the most severe manifestation of infection with Plasmodium, however its pathogenesis is still not completely understood. microRNA (miRNA) have been an area of focus in infectious disease research, due to their ability to affect normal biological processes, and have been shown to play roles in various viral, bacterial and parasitic infections, including malaria. The expression of miRNA was studied following infection of CBA mice with either Plasmodium berghei ANKA (causing CM), or Plasmodium yoelii (causing severe but non-cerebral malaria (NCM)). Using microarray analysis, miRNA expression was compared in the brains of non-infected (NI), NCM and CM mice. Six miRNA were significantly dysregulated between NCM and CM mice, and four of these, miR-19a-3p, miR-19b-3p, miR-142-3p and miR-223-3p, were further validated by qPCR assays. These miRNA are significantly involved in several pathways relevant to CM, including the TGF-β and endocytosis pathways. Dysregulation of these miRNA during CM specifically compared with NCM suggests that these miRNA, through their regulation of downstream targets, may be vitally involved in the neurological syndrome. Our data implies that, at least in the mouse model, miRNA may play a regulatory role in CM pathogenesis.This work was funded by the National Health and Medical Research Council (#1099920 for GEG). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S

The Characteristics of Blood Glucose and WBC Counts in Peripheral Blood of Cases of Hand Foot and Mouth Disease in China: A Systematic Review

Background: Outbreaks of Hand Foot and Mouth Disease (HFMD) have occurred in many parts of the world especially in China. We aimed to summarize the characteristics of the levels of blood glucose and white blood cell (WBC) counts in cases of HFMD in Mainland China and Taiwan, using meta-analysis based on systematic review of published articles. Methods: We systematically reviewed published studies, from the MEDLINE and WANFANG Data, about the levels of blood glucose and WBC counts in cases of HFMD until 15 th June 2011, and quantitatively summarized the characteristics of them using meta-analysis. Results: In total, 37 studies were included in this review. In Mainland China and Taiwan, generally, the average level of blood glucose, the prevalence of hyperglycemia, WBC counts and the prevalence of leukocytosis increased with the severity of the illness. There was no significant difference in the prevalence of leukocytosis between ANS (autonomic nervous system dysregulation)/PE (pulmonary edema) group and CNS (central nervous system) group, and in the average level of blood glucose between healthy controls and mild cases of HFMD. WBC counts in cases infected by EV71 were less than those in cases infected by CA16. Conclusions: our analyses indicated that blood glucose and WBC counts increased with the severity of HFMD disease, which would help doctors to manage patients efficiently