The Role of the Transcription Factor SIM2 in Prostate Cancer

Background: Recent reports have suggested a possible involvement of Single-minded homolog 2 (SIM2) in human solid cancers, including prostate cancer. However, the exact role of SIM2 in cancer in general, and in prostate cancer in particular, remains largely unknown. This study was designed to elucidate the role of SIM2 in prostate cancer using a shRNA-based approach in the PC3 prostate cancer cell line. Methods: Lentiviral shRNAs were used to inhibit SIM2 gene and protein levels in PC3 cells. Quantitative RT-PCR and branched DNA were performed to evaluate transcript expression. SIM2 protein expression level was measured by western blot. Profiling of gene expression spanning the whole genome, as well as polar metabolomics of several major metabolic pathways was performed to identify major pathway dysregulations. Results: SIM2 gene and protein products were significantly downregulated by lenti-shRNA in PC3 cell line. This low expression of SIM2 affected gene expression profile, revealing significant changes in major signaling pathways, networks and functions. In addition, major metabolic pathways were affected. Conclusion: Taken together, our results suggest an involvement of SIM2 in key traits of prostate tumor cell biology an

MARCO variants are associated with phagocytosis, pulmonary tuberculosis susceptibility and Beijing lineage

Macrophage receptor with collagenous structure (MARCO) has an important role in the phagocytosis of Mycobacterium tuberculosis (M. tuberculosis). We hypothesized that MARCO polymorphisms are associated with phagocytosis, tuberculosis (TB) disease susceptibility and presentation, and infecting lineage. We used a human cellular model to examine how MARCO genotype mediates the immune response; a case-control study to investigate tuberculosis host genetic susceptibility; and a host-pathogen genetic analysis to study host-pathogen interactions. Two MARCO heterozygous (AG) genotypes (single-nucleotide polymorphisms rs2278589 and rs6751745) were associated with impaired phagocytosis of M. tuberculosis trehalose 6,6'-dimycolate-cord factor and β-glucan-coated beads in macrophages. The heterozygous genotypes of rs2278589 and rs6751745 were also associated with increased risk of pulmonary TB (PTB; rs2278589, P=0.001, odds ratio (OR)=1.6; rs6751745, P=0.009, OR=1.4), and with severe chest X-ray abnormalities (P=0.007, OR=1.6). These two genotypes were also associated with the Beijing lineage (rs2278589, P=0.001, OR=1.7; rs6751745, P=0.01, OR=1.5). Together, these results suggest that MARCO polymorphisms may regulate phagocytosis of M. tuberculosis and susceptibility and severity of PTB. They also suggest MARCO genotype and Beijing strains may interact to increase the risk of PT

MARCO, TLR2, and CD14 Are Required for Macrophage Cytokine Responses to Mycobacterial Trehalose Dimycolate and Mycobacterium tuberculosis

Virtually all of the elements of Mycobacterium tuberculosis (Mtb) pathogenesis, including pro-inflammatory cytokine production, granuloma formation, cachexia, and mortality, can be induced by its predominant cell wall glycolipid, trehalose 6,6′-dimycolate (TDM/cord factor). TDM mediates these potent inflammatory responses via interactions with macrophages both in vitro and in vivo in a myeloid differentiation factor 88 (MyD88)-dependent manner via phosphorylation of the mitogen activated protein kinases (MAPKs), implying involvement of toll-like receptors (TLRs). However, specific TLRs or binding receptors for TDM have yet to be identified. Herein, we demonstrate that the macrophage receptor with collagenous structure (MARCO), a class A scavenger receptor, is utilized preferentially to “tether” TDM to the macrophage and to activate the TLR2 signaling pathway. TDM-induced signaling, as measured by a nuclear factor-kappa B (NF-κB)-luciferase reporter assay, required MARCO in addition to TLR2 and CD14. MARCO was used preferentially over the highly homologous scavenger receptor class A (SRA), which required TLR2 and TLR4, as well as their respective accessory molecules, in order for a slight increase in NF-κB signaling to occur. Consistent with these observations, macrophages from MARCO−/− or MARCO−/−SRA−/− mice are defective in activation of extracellular signal-related kinase 1/2 (ERK1/2) and subsequent pro-inflammatory cytokine production in response to TDM. These results show that MARCO-expressing macrophages secrete pro-inflammatory cytokines in response to TDM by cooperation between MARCO and TLR2/CD14, whereas other macrophage subtypes (e.g. bone marrow–derived) may rely somewhat less effectively on SRA, TLR2/CD14, and TLR4/MD2. Macrophages from MARCO−/− mice also produce markedly lower levels of pro-inflammatory cytokines in response to infection with virulent Mtb. These observations identify the scavenger receptors as essential binding receptors for TDM, explain the differential response to TDM of various macrophage populations, which differ in their expression of the scavenger receptors, and identify MARCO as a novel component required for TLR signaling

Genetic Variants in MARCO Are Associated with the Susceptibility to Pulmonary Tuberculosis in Chinese Han Population

BACKGROUND: Susceptibility to tuberculosis is not only determined by Mycobacterium tuberculosis infection, but also by the genetic component of the host. Macrophage receptor with a collagenous structure (MARCO) is essential components required for toll like receptor-signaling in macrophage response to Mycobacterium tuberculosis, which may contribute to tuberculosis risk. PRINCIPAL FINDINGS: To specifically investigated whether single nucleotide polymorphisms (SNPs) in MARCO gene are associated with pulmonary tuberculosis in Chinese Han population. By selecting tagging SNPs in MARCO gene, 17 tag SNPs were identified and genotyped in 923 pulmonary tuberculosis patients and 1033 healthy control subjects using a hospital based case-control association study. Single-point and haplotype analysis revealed an association in intron and exon region of MARCO gene. One SNP (rs17009726) was associated with susceptibility to pulmonary tuberculosis, where the carriers of the G allele had a 1.65 fold (95% CI = 1.32-2.05, p(corrected) = 9.27E-5) increased risk of pulmonary tuberculosis. Haplotype analysis revealed that haplotype GC containing G allele of 17009726 and haplotype TGCC (rs17795618T/A, rs1371562G/T, rs6761637T/C, rs2011839C/T) were also associated with susceptibility to pulmonary tuberculosis (p(corrected) = 0.0001 and 0.029, respectively). CONCLUSIONS: Our study suggested that genetic variants in MARCO gene were associated with pulmonary tuberculosis susceptibility in Chinese Han population, and the findings emphasize the importance of MARCO mediated immune responses in the pathogenesis of tuberculosis

Protective Contributions against Invasive Streptococcus pneumoniae Pneumonia of Antibody and Th17-Cell Responses to Nasopharyngeal Colonisation

The nasopharyngeal commensal bacteria Streptococcus pneumoniae is also a frequent cause of serious infections. Nasopharyngeal colonisation with S. pneumoniae inhibits subsequent re-colonisation by inducing Th17-cell adaptive responses, whereas vaccination prevents invasive infections by inducing antibodies to S. pneumoniae capsular polysaccharides. In contrast, protection against invasive infection after nasopharyngeal colonisation with mutant S. pneumoniae strains was associated with antibody responses to protein antigens. The role of colonisation-induced Th17-cell responses during subsequent invasive infections is unknown. Using mouse models, we show that previous colonisation with S. pneumoniae protects against subsequent lethal pneumonia mainly by preventing bacteraemia with a more modest effect on local control of infection within the lung. Previous colonisation resulted in CD4-dependent increased levels of Th17-cell cytokines during subsequent infectious challenge. However, mice depleted of CD4 cells prior to challenge remained protected against bacteraemia, whereas no protection was seen in antibody deficient mice and similar protection could be achieved through passive transfer of serum. Serum from colonised mice but not antibody deficient mice promoted phagocytosis of S. pneumoniae, and previously colonised mice were able to rapidly clear S. pneumoniae from the blood after intravenous inoculation. Thus, despite priming for a Th17-cell response during subsequent infection, the protective effects of prior colonisation in this model was not dependent on CD4 cells but on rapid clearance of bacteria from the blood by antibody-mediated phagocytosis. These data suggest that whilst nasopharyngeal colonisation induces a range of immune responses, the effective protective responses depend upon the site of subsequent infectio

Innate Immune Response of Human Alveolar Macrophages during Influenza A Infection

Alveolar macrophages (AM) are one of the key cell types for initiating inflammatory and immune responses to influenza virus in the lung. However, the genome-wide changes in response to influenza infection in AM have not been defined. We performed gene profiling of human AM in response to H1N1 influenza A virus PR/8 using Affymetrix HG-U133 Plus 2.0 chips and verified the changes at both mRNA and protein levels by real-time RT-PCR and ELISA. We confirmed the response with a contemporary H3N2 influenza virus A/New York/238/2005 (NY/238). To understand the local cellular response, we also evaluated the impact of paracrine factors on virus-induced chemokine and cytokine secretion. In addition, we investigated the changes in the expression of macrophage receptors and uptake of pathogens after PR/8 infection. Although macrophages fail to release a large amount of infectious virus, we observed a robust induction of type I and type III interferons and several cytokines and chemokines following influenza infection. CXCL9, 10, and 11 were the most highly induced chemokines by influenza infection. UV-inactivation abolished virus-induced cytokine and chemokine response, with the exception of CXCL10. The contemporary influenza virus NY/238 infection of AM induced a similar response as PR/8. Inhibition of TNF and/or IL-1β activity significantly decreased the secretion of the proinflammatory chemokines CCL5 and CXCL8 by over 50%. PR/8 infection also significantly decreased mRNA levels of macrophage receptors including C-type lectin domain family 7 member A (CLEC7A), macrophage scavenger receptor 1 (MSR1), and CD36, and reduced uptake of zymosan. In conclusion, influenza infection induced an extensive proinflammatory response in human AM. Targeting local components of innate immune response might provide a strategy for controlling influenza A infection-induced proinflammatory response in vivo

Scavenger receptors and β-glucan receptors participate in the recognition of yeasts by murine macrophages

Objectives: Numerous receptors have been implicated in recognition of pathogenic fungi by macrophages, including the $\beta$-glucan receptor dectin-1. The role of scavenger receptors (SRs) in anti-fungal immunity is not well characterized. Methods: We studied uptake of unopsonized Saccharomycetes cerevisiae (zymosan) and live Candida albicans yeasts as well as zymosan-stimulated $H_2O_2$ production in J774 macrophage-like cells and peritoneal exudate macrophages (PEMs). The role of different receptors was assessed with the use of competitive ligands, transfected cells and receptor-deficient macrophages. Results: The uptake of zymosan by untreated J774 cells was mediated approximately half by SRs and half by a $\beta$-glucan receptor which was distinct from dectin-1 and not linked to stimulation of $H_2O_2$ production. Ligands of $\beta$-glucan receptors and of SRs also inhibited uptake of C. albicans by macrophages (J774 cells and PEMs). In macrophages pretreated with a CpG motif-containing oligodeoxynucleotide (CpG-ODN) the relative contribution of SRs to yeast uptake increased and that of $\beta$-glucan receptors decreased. Whereas the class A SR MARCO participated in the uptake of both zymosan and C. albicans by CpG-ODN-pretreated, but not untreated macrophages, the related receptor SR-A/CD204 was involved in the uptake of zymosan, but not of C. albicans. The reduction of zymosan-stimulated $H_2O_2$ production observed in DS-pretreated J774 cells and in class A SRs-deficient PEMs suggest that class A SRs mediate part of this process. Conclusions: Our results revealed that SRs belong to a redundant system of receptors for yeasts. Binding of yeasts to different receptors in resting versus CpG-ODN-pre-exposed macrophages may differentially affect polarization of adaptive immune responses

Haptoglobin and Sickle Cell Polymorphisms and Risk of Active Trachoma in Gambian Children

BACKGROUND: Susceptibility and resistance to trachoma, the leading infectious cause of blindness, have been associated with a range of host genetic factors. In vitro studies of the causative organism, Chlamydia trachomatis, demonstrate that iron availability regulates its growth, suggesting that host genes involved in regulating iron status and/or availability may modulate the risk of trachoma. The objective was to investigate whether haptoglobin (Hp) haplotypes constructed from the functional polymorphism (Hp1/Hp2) plus the functional promoter SNPs -61A-C (rs5471) and -101C-G (rs5470), or sickle cell trait (HbAS, rs334) were associated with risk of active trachoma when stratified by age and sex, in rural Gambian children. METHODOLOGY AND PRINCIPAL FINDINGS: In two cross sectional surveys of children aged 6-78 months (n = 836), the prevalence of the clinical signs of active trachoma was 21.4%. Within boys, haplotype E (-101G, -61A, Hp1), containing the variant allele of the -101C-G promoter SNP, was associated with a two-fold increased risk of active trachoma (OR = 2.0 [1.17-3.44]). Within girls, an opposite association was non-significant (OR = 0.58 [0.32-1.04]; P = 0.07) and the interaction by sex was statistically significant (P = 0.001). There was no association between trachoma and HbAS. CONCLUSIONS: These data indicate that genetic variation in Hp may affect susceptibility to active trachoma differentially by sex in The Gambia

Proteomics Mapping of Cord Blood Identifies Haptoglobin “Switch-On” Pattern as Biomarker of Early-Onset Neonatal Sepsis in Preterm Newborns

Intra-amniotic infection and/or inflammation (IAI) are important causes of preterm birth and early-onset neonatal sepsis (EONS). A prompt and accurate diagnosis of EONS is critical for improved neonatal outcomes. We sought to explore the cord blood proteome and identify biomarkers and functional protein networks characterizing EONS in preterm newborns.We studied a prospective cohort of 180 premature newborns delivered May 2004-September 2009. A proteomics discovery phase employing two-dimensional differential gel electrophoresis (2D-DIGE) and mass spectrometry identified 19 differentially-expressed proteins in cord blood of newborns with culture-confirmed EONS (n = 3) versus GA-matched controls (n = 3). Ontological classifications of the proteins included transfer/carrier, immunity/defense, protease/extracellular matrix. The 1(st)-level external validation conducted in the remaining 174 samples confirmed elevated haptoglobin and haptoglobin-related protein immunoreactivity (Hp&HpRP) in newborns with EONS (presumed and culture-confirmed) independent of GA at birth and birthweight (P<0.001). Western blot concurred in determining that EONS babies had conspicuous Hp&HpRP bands in cord blood ("switch-on pattern") as opposed to non-EONS newborns who had near-absent "switch-off pattern" (P<0.001). Fetal Hp phenotype independently impacted Hp&HpRP. A bayesian latent-class analysis (LCA) was further used for unbiased classification of all 180 cases based on probability of "antenatal IAI exposure" as latent variable. This was then subjected to 2(nd)-level validation against indicators of adverse short-term neonatal outcome. The optimal LCA algorithm combined Hp&HpRP switch pattern (most input), interleukin-6 and neonatal hematological indices yielding two non-overlapping newborn clusters with low (≤20%) versus high (≥70%) probability of IAI exposure. This approach reclassified ∼30% of clinical EONS diagnoses lowering the number needed to harm and increasing the odds ratios for several adverse outcomes including intra-ventricular hemorrhage.Antenatal exposure to IAI results in precocious switch-on of Hp&HpRP expression. As EONS biomarker, cord blood Hp&HpRP has potential to improve the selection of newborns for prompt and targeted treatment at birth