Interferon and IL-27 antagonize the function of group 2 innate lymphoid cells and type 2 innate immune responses.

Group 2 innate lymphoid cells (ILC2 cells) are type 2 cytokine-producing cells of the innate immune system with important roles in helminth infection and allergic inflammation. Here we found that tissue-resident ILC2 cells proliferated in situ without migrating during inflammatory responses. Both type I and type II interferons and interleukin 27 (IL-27) suppressed ILC2 function in a manner dependent on the transcription factor STAT1. ILC2-mediated lung inflammation was enhanced in the absence of the interferon-γ (IFN-γ) receptor on ILC2 cells in vivo. IFN-γ effectively suppressed the function of tissue-resident ILC2 cells but not that of inflammatory ILC2 cells, and IL-27 suppressed tissue-resident ILC2 cells but not tissue-resident TH2 cells during lung inflammation induced by Alternaria alternata. Our results demonstrate that suppression mediated by interferon and IL-27 is a negative feedback mechanism for ILC2 function in vivo.

Comprehensive silencing of target-sharing microRNAs is a mechanism for SIRT1 overexpression in cancer.

Overexpression of SIRT1 is frequently observed in various types of cancers, suggesting its potential role in malignancies. However, the molecular basis of how SIRT1 is elevated in cancer is less understood. Here we show that cancer-related SIRT1 overexpression is due to evasion of Sirt1 mRNA from repression by a group of Sirt1-targeting microRNAs (miRNAs) that might be robustly silenced in cancer. Our comprehensive library-based screening and subsequent miRNA gene profiling revealed a housekeeping gene-like broad expression pattern and strong CpG island-association of the Sirt1-targeting miRNA genes. This suggests aberrant CpG DNA methylation as the mechanistic background for malignant SIRT1 elevation. Our work also provides an example where epigenetic mechanisms cause the group-wide regulation of miRNAs sharing a common key target.

Perturbation of copper homeostasis is instrumental in early developmental arrest of intraerythrocytic Plasmodium falciparum.

BACKGROUND: Malaria continues to be a devastating disease. The elucidation of factors inducing asexual growth versus arrest of Plasmodium falciparum can provide information about the development of the parasite, and may help in the search for novel malaria medication. Based on information from genome-wide transcriptome profiling of different developmental stages of P. falciparum, we investigated the critical importance of copper homeostasis in the developmental succession of P. falciparum with regard to three aspects of copper function. These were:1) inhibition of copper-binding proteins, 2) copper-ion chelation, and 3) down-regulated expression of genes encoding copper-binding proteins associated with a specific growth-promoting factor. RESULTS: Inhibition of copper-binding proteins with tetrathiomolybdate (TTM) caused cessation of growth of the parasite. TTM arrested the parasite irreversibly during the trophozoite to schizont stage progression. Target molecules for TTM may be present in P. falciparum. The involvement of copper ions in developmental arrest was also investigated by copper-ion chelating methods, which indicated a critical function of reduced copper ions (Cu1+) in the parasite during the early developmental stage. Copper ions, not only in the parasite but also in host cells, were targets of the chelators. Chelation of Cu1+caused blockage of trophozoite progression from the ring stage. Profound growth arrest was detected in parasites cultured in a chemically defined medium containing hexadecanoic acid alone as a growth-promoting factor. This developmental arrest was associated with down-regulated expression of genes encoding copper-binding proteins. Cis-9-octadecenoic acid completely prevented the down-regulation of gene expression and developmental arrest that were observed with the use of hexadecanoic acid. CONCLUSIONS: The critical importance of copper homeostasis in early developmental stages of P. falciparum was confirmed. Perturbation of copper homeostasis induced profound and early developmental arrest of P. falciparum. These findings should help to elucidate the mechanisms behind the development of P. falciparum, and may be applied in the development of effective antimalarial strategies.

Molecular factors that are associated with early developmental arrest of intraerythrocytic Plasmodium falciparum.

Malaria continues to be a devastating disease. We investigated the factors that control intraerythrocytic development of the parasite Plasmodium falciparum by using a chemically defined medium (CDM) containing non-esterified fatty acid(s) (NEFA) and phospholipids with specific fatty acid moieties, to identify substances crucial for parasite development. Different NEFAs in the CDM played distinct roles by altering the development of the parasite at various stages, with effects ranging from complete growth to growth arrest at the ring stage. We used genome-wide transcriptome profiling to identify genes that were differentially expressed among the different developmental stages of the parasite, cultured in the presence of various NEFAs. We predicted 26 transcripts that were associated with the suppression of schizogony, of which 5 transcripts, including merozoite surface protein 2, a putative DEAD/DEAH box RNA helicase, serine repeat antigen 3, a putative copper channel, and palmitoyl acyltransferase, were particularly associated with blockage of trophozoite progression from the ring stage. Furthermore, the involvement of copper ions in developmental arrest was detected by copper-ion-chelating methods, implying a critical function of copper homeostasis in the early growth stage of the parasite. These results should help to elucidate the mechanisms behind the development of P. falciparum.

Natural helper cells: a new player in the innate immune response against helminth infection.

The Th2-type immune response, characterized by the production of IL-4, IL-5, and IL-13, is a critical immune response against helminths invading cutaneous or mucosal sites. Th2 cytokines are induced soon after helminth infection, even before a pathogen-specific adaptive immune response is established. Although the expulsion and clearance of helminths usually requires pathogen-specific Th2-mediated immunity, early induction of Th2 cytokines during the innate immune phase is important for host protection from helminth invasion. Recent studies have shed light on such Th2 cytokine production by formerly uncharacterized innate immune cells such as a newly identified natural helper cell. We discuss here the mechanisms of innate production of Th2 cytokines in host immune responses against helminth infection.

Innate production of T(H)2 cytokines by adipose tissue-associated c-Kit(+)Sca-1(+) lymphoid cells.

Innate immune responses are important in combating various microbes during the early phases of infection. Natural killer (NK) cells are innate lymphocytes that, unlike T and B lymphocytes, do not express antigen receptors but rapidly exhibit cytotoxic activities against virus-infected cells and produce various cytokines. Here we report a new type of innate lymphocyte present in a novel lymphoid structure associated with adipose tissues in the peritoneal cavity. These cells do not express lineage (Lin) markers but do express c-Kit, Sca-1 (also known as Ly6a), IL7R and IL33R. Similar lymphoid clusters were found in both human and mouse mesentery and we term this tissue 'FALC' (fat-associated lymphoid cluster). FALC Lin(-)c-Kit(+)Sca-1(+) cells are distinct from lymphoid progenitors and lymphoid tissue inducer cells. These cells proliferate in response to IL2 and produce large amounts of T(H)2 cytokines such as IL5, IL6 and IL13. IL5 and IL6 regulate B-cell antibody production and self-renewal of B1 cells. Indeed, FALC Lin(-)c-Kit(+)Sca-1(+) cells support the self-renewal of B1 cells and enhance IgA production. IL5 and IL13 mediate allergic inflammation and protection against helminth infection. After helminth infection and in response to IL33, FALC Lin(-)c-Kit(+)Sca-1(+) cells produce large amounts of IL13, which leads to goblet cell hyperplasia-a critical step for helminth expulsion. In mice devoid of FALC Lin(-)c-Kit(+)Sca-1(+) cells, such goblet cell hyperplasia was not induced. Thus, FALC Lin(-)c-Kit(+)Sca-1(+) cells are T(H)2-type innate lymphocytes, and we propose that these cells be called 'natural helper cells'.

Mammalian target of rapamycin and glycogen synthase kinase 3 differentially regulate lipopolysaccharide-induced interleukin-12 production in dendritic cells.

Phosphoinositide 3-kinase (PI3K) negatively regulates Toll-like receptor (TLR)-mediated interleukin-12 (IL-12) expression in dendritic cells (DCs). We show here that 2 signaling pathways downstream of PI3K, mammalian target of rapamycin (mTOR) and glycogen synthase kinase 3 (GSK3), differentially regulate the expression of IL-12 in lipopolysaccharide (LPS)-stimulated DCs. Rapamycin, an inhibitor of mTOR, enhanced IL-12 production in LPS-stimulated DCs, whereas the activation of mTOR by lentivirus-mediated transduction of a constitutively active form of Rheb suppressed the production of IL-12. The inhibition of protein secretion or deletion of IL-10 cancelled the effect of rapamycin, indicating that mTOR regulates IL-12 expression through an autocrine action of IL-10. In contrast, GSK3 positively regulates IL-12 production through an IL-10-independent pathway. Rapamycin-treated DCs enhanced Th1 induction in vitro compared with untreated DCs. LiCl, an inhibitor of GSK3, suppressed a Th1 response on Leishmania major infection in vivo. These results suggest that mTOR and GSK3 pathways regulate the Th1/Th2 balance though the regulation of IL-12 expression in DCs. The signaling pathway downstream of PI3K would be a good target to modulate the Th1/Th2 balance in immune responses in vivo.

An evolutionarily conserved mechanism for microRNA-223 expression revealed by microRNA gene profiling.

Many microRNAs (miRNAs) are evolutionarily conserved and have intriguing expression patterns. Tissue and/or time-specific expressions of some miRNAs are presumably controlled by unique cis-acting regulatory elements that coevolved with the miRNA sequences. Exploiting bioinformatics, we identified several miRNAs whose primary transcripts could be regulated by conserved genomic elements proximal to their transcription start sites. Such miRNAs include microRNA-223 (miR-223), which is reportedly controlled by a unique regulatory mechanism during granulopoiesis. Here, we define a mechanism distinct from that previously proposed to regulate miR-223 expression. We find that the mir-223 gene resembles a "myeloid gene" and might be driven by the myeloid transcription factors, PU.1 and C/EBPs. This mechanism is specified by the conserved proximal cis-regulatory element and might be common among different species. Hence, it needs to be considered that two distinct mechanisms that would play critical roles in myeloid functions and differentiation are actually concerned with the regulation of miR-223.

Antibody isotype responses to paramyosin, a vaccine candidate for schistosomiasis, and their correlations with resistance and fibrosis in patients infected with Schistosoma japonicum in Leyte, The Philippines.

We examined whether antibody isotype responses to paramyosin (PM), a vaccine candidate for schistosomiasis, are associated with age-dependent resistance and pathology in liver fibrosis using human sera collected from 139 individuals infected with Schistosoma japonicum in Leyte, The Philippines. We report that IgA and IgG3 responses to PM showed a positive correlation with age and that the epitopes responsible were localized predominantly within the N-terminal half of PM. In addition, the IgG3 response to PM was associated with serum level of procollagen-III-peptide (P-III-P), an indicator of progression of liver fibrosis. These results imply that IgG3 against PM may not only provoke age-dependent resistance to S. japonicum infection but also enhance liver fibrosis. In contrast, levels of IgE to PM and to multiple PM fragments showed a negative correlation with P-III-P level. Thus, in contrast to IgG3, increases in PM-specific IgE may contribute to suppression of liver pathogenesis in schistosomiasis.

IL-7 exacerbates chronic colitis with expansion of memory IL-7Rhigh CD4+ mucosal T cells in mice.

We have previously demonstrated that mucosal CD4(+) T cells expressing high levels of IL-7 receptor (IL-7R(high)) are pathogenic cells responsible for chronic colitis. Here we investigate whether IL-7 is directly involved in the expansion of IL-7R(high) memory CD4(+) mucosal T cells and the exacerbation of colitis. We first showed that CD4(+) lamina propria lymphocytes (LPLs) from wild-type, T cell receptor-alpha-deficient (TCR-alpha(-/-)), and recombinase-activating gene (RAG)-2(-/-)-transferred mice with or without colitis showed phenotypes of memory cells, but only CD4(+) LPLs from colitic mice showed IL-7R(high). In vitro stimulation by IL-7, but not by IL-15 and thymic stromal lymphopoietin, enhanced significant proliferative responses and survival of colitic CD4(+), but not normal CD4(+) LPLs. Importantly, in vivo administration of IL-7 mice accelerated the expansion of IL-7R(high) memory CD4(+) LPLs and thereby exacerbated chronic colitis in RAG-2(-/-) mice transferred with CD4(+) LPLs from colitic TCR-alpha(-/-) mice. Conversely, the administration of anti-IL-7R monoclonal antibody significantly inhibited the development of TCR-alpha(-/-) colitis with decreased expansion of CD4(+) LPLs. Collectively, the present data indicate that IL-7 is essential for the expansion of pathogenic memory CD4(+) T cells under pathological conditions. Therefore, therapeutic approaches targeting the IL-7R pathway may be feasible in the treatment of human inflammatory bowel disease.

Haemostatic abnormalities in hepatosplenic schistosomiasis mansoni.

Hepatosplenic schistosomiasis is a complex immuno-regulatory disease and is major health problem in endemic countries. Acute bleeding is one of its most serious complications and often life-threatening. Clinical studies have demonstrated that the patients with hepatosplenic schistosomiasis are prone to develop complex haemostatic abnormalities that may be linked to the potential risk of bleeding from ruptured esophageal varices in these patients. The deficit in haemostatic parameters is more pronounced with the advancement of the disease and is maximal in the patients with experience of haematomesis. Evidences of enhanced generation of thrombin and plasmin indicate the presence of low-grade DIC in advanced hepatosplenic schistosomiasis, which is considered as a principal cause of haemostatic abnormalities in this endemic disease. Demonstration of procoagulant expression in peripheral blood monocytes of the patients and in the livers, spleens and intestines of S. mansoni-infected mice suggest their possible implication in the causation of DIC in S. mansoni infections. Moreover, because in vitro analysis indicates a participation of immune mechanisms in the localized procoagulant expression, it seems likely that the immune responses to schistosomes play a major role in the pathogenic mechanisms of haemostatic abnormalities in hepatosplenic schistosomiasis.

Mucosal T cells expressing high levels of IL-7 receptor are potential targets for treatment of chronic colitis.

The IL-7/IL-7R-dependent signaling pathway plays a crucial role in regulating the immune response in intestinal mucosa. Here we demonstrate the pivotal role of this pathway in the development and treatment of chronic colitis. T cells expressing high levels of IL-7R were substantially infiltrated in the chronic inflamed mucosa of TCR alpha-chain knockout mice and IL-7 transgenic mice. Transfer of mucosal T cells expressing high levels of IL-7R, but not T cells expressing low levels of IL-7R, from these mice into recombinase-activating gene-2(-/-) mice induced chronic colitis. Selective elimination of T cells expressing high levels of IL-7R by administrating small amounts of toxin-conjugated anti-IL-7R Ab completely ameliorated established, ongoing colitis. These findings provide evidence that therapeutic approaches targeting mucosal T cells expressing high levels of IL-7R are effective in the treatment of chronic intestinal inflammation and may be feasible for use in the therapy of human inflammatory bowel disease.

PI3K-mediated negative feedback regulation of IL-12 production in DCs.

Although interleukin 12 (IL-12) production by dendritic cells (DCs) confers protection against harmful invasions by regulating both innate and adaptive immunity, its dysregulation may have detrimental effects on the host. We show here that phosphoinositide 3-kinase (PI3K) negatively regulates IL-12 synthesis by DCs. We found that numerous stimuli that induced IL-12 production concomitantly elicited PI3K activation in DCs, but both PI3K(-/-) and PI3K inhibitor#150;treated DCs showed increased IL-12 production. Accordingly, an enhanced T helper type 1 (T(H)1) response was observed upon Leishmania major infection in PI3K(-/-) mice. Our findings indicate that a negative feedback mechanism exists that regulates IL-12 production during DC activation and may help prevent the excessive T(H)1 polarization that causes undesirable immune responses.

Selective loss of gastrointestinal mast cells and impaired immunity in PI3K-deficient mice.

Mice that lack the p85alpha regulatory subunit of phosphatidylinositol-3 kinase (PI3K) are deficient in gastrointestinal and peritoneal mast cells but have dermal mast cells. Accordingly, these mice show impaired bacterial clearance in response to acute septic peritonitis and are highly susceptible to infection by the intestinal nematode Strongyloides venezuelensis. Systemic anaphylactic shock responses, however, are intact. We found that although reconstitution of PI3Kminus sign/minus sign mice with bone marrow--derived mast cells (BMMCs) restored anti-bacterial immunity, only T helper type 2 (TH2)-conditioned BMMCs, not "standard" BMMCs, were able to restore anti-nematode immunity. This finding highlights the importance of the TH2 response in the control of nematode infection. Thus, PI3K likely plays an essential role in host immune responses by regulating both the development and induction of mast cells.

Parasitological and serological studies on amoebiasis and other intestinal parasitic infections in the rural sector around Recife, Northeast Brazil

Foram realizados exames parasitológicos em 485 habitantes de quatro vilarejos da cidade de Säo Lourenço da Mata, distante 25 Km à noroeste de Recife-PE, Brasil, no período de julho à dezembro de 1989. Aproximadamente 99,6% dos examinados mostraram-se infectados com pelo menos uma espécie de parasita intestinal. Observou-se ainda uma alta prevalência de Shistosoma mansoni (82,1%), ancilostomídeos (80,2%), Trichuris trichiura (69,9%), Ascaris lumbricoides (61,9%) e Entamoeba coli36,7%). A cultura de larvas de ancilostomídeos nas fezes - método de Harada - revelou que Necator americanus (84,4%) é a espécie mais comum nesta regiäo seguida pelo Strongyloides stercoralis. Foram ainda realizados testes sorológicos imunodifusäo em gel (GDP) e enzima imunoensaio (ELISA) - em 334 soros, para o diagnóstico de amebíase, todavia apenas 24 (7,2%) dos soros apresentaram positivdade no teste de ELISA e nenhuma positividade foi encontrada nos mesmos soros testados pelo GDP

Parasitological and serological studies on amoebiasis and other intestinal parasitic infections in Recife and its suburban area, northeast Brazil

Exames parasitológicos foram realizados em 187 pacientes do Hospital do IMIP e 464 habitantes de vários vilarejos no município do Cabo, 50 Km à sudeste de Recife, durante os meses de abril a agosto. Aproximadamente 71% dos pacientes examinados do IMIP e 92% dos examinados do Cabo apresentavam-se infectados com, no mínimo, uma espécie de parasita intestinal. Houve uma diferença mínima na taxa de prevalência de trichiura entre as duas áreas, entretanto a prevalência de Ascaris lumbricoides, família Ancylostomidae, Strongyloides stercoralis, Schistosoma mansoni e Entamoeba histolytica foi maior entre os habitantes do Cabo. Somente a Giardia lamblia apresentou uma taxa de prevalência maior nos pacientes do IMIP. O cultivo em tubo de ensaio revelou que a prevalência do Necator americanus em relaçäo a do Ancylostoma duodenale era muito maior em ambas as áreas e que a do S. stercoralis entre os pacientes do IMIP e dos habitantes do Cabo era, respectivamente, 4.5% e 9.6%. A amebíase foi verificada através de exames sorológicos, imunodifusäo em gel (GDP) e enzima imunoensaio (ELISA), usando como antígeno extrato bruto preparado a partir dos trofozoitos de E. histolytica (cepa HM-1: IMSS), realizados em 615 soros, onde nenhuma reaçäo positiva aparente foi observada através da imunodifusäo, contudo foram observados resultados positivos em 32 dos 615 casos através da enzima imunoensaio