Publications

Malaria parasites within red blood cells digest host hemoglobin into a hydrophobic heme polymer, known as hemozoin (HZ), which is subsequently released into the blood stream and then captured by and concentrated in the reticulo-endothelial system. Accumulating evidence suggests that HZ is immunologically active, but the molecular mechanism(s) through which HZ modulates the innate immune system has not been elucidated. This work demonstrates that HZ purified from Plasmodium falciparum is a novel non-DNA ligand for Toll-like receptor (TLR)9. HZ activated innate immune responses in vivo and in vitro, resulting in the production of cytokines, chemokines, and up-regulation of costimulatory molecules. Such responses were severely impaired in TLR9-/- and myeloid differentiation factor 88 (MyD88)-/-, but not in TLR2, TLR4, TLR7, or Toll/interleukin 1 receptor domain-containing adaptor-inducing interferon beta-/- mice. Synthetic HZ, which is free of the other contaminants, also activated innate immune responses in vivo in a TLR9-dependent manner. Chloroquine (CQ), an antimalarial drug, abrogated HZ-induced cytokine production. These data suggest that TLR9-mediated, MyD88-dependent, and CQ-sensitive innate immune activation by HZ may play an important role in malaria parasite-host interactions.

Lactoferrin (LF) has been implicated in innate immunity. Here we reveal the signal transduction pathway responsible for human LF (hLF)‐triggered nuclear factor‐κB (NF‐κB) activation. Endotoxin‐depleted hLF induces NF‐κB activation at physiologically relevant concentrations in the human monocytic leukemia cell line, THP‐1, and in mouse embryonic fibroblasts (MEFs). In MEFs, in which both tumor necrosis factor receptor‐associated factor 2 (TRAF2) and TRAF5 are deficient, hLF causes NF‐κB activation at a level comparable to that seen in wild‐type MEFs, whereas TRAF6‐deficient MEFs show significantly impaired NF‐κB activation in response to hLF. TRAF6 is known to be indispensable in leading to NF‐κB activation in myeloid differentiating factor 88 (MyD88)‐dependent signaling pathways, while the role of TRAF6 in the MyD88‐independent signaling pathway has not been clarified extensively. When we examined the hLF‐dependent NF‐κB activation in MyD88‐deficient MEFs, delayed, but remarkable, NF‐κB activation occurred as a result of the treatment of cells with hLF, indicating that both MyD88‐dependent and MyD88‐independent pathways are involved. Indeed, hLF fails to activate NF‐κB in MEFs lacking Toll‐like receptor 4 (TLR4), a unique TLR group member that triggers both MyD88‐depependent and MyD88‐independent signalings. Importantly, the carbohydrate chains from hLF are shown to be responsible for TLR4 activation. Furthermore, we show that lipopolysaccharide‐induced cytokine and chemokine production is attenuated by intact hLF but not by the carbohydrate chains from hLF. Thus, we present a novel model concerning the biological function of hLF: hLF induces moderate activation of TLR4‐mediated innate immunity through its carbohydrate chains; however, hLF suppresses endotoxemia by interfering with lipopolysaccharide‐dependent TLR4 activation, probably through its polypeptide moiety.

Group B streptococci, a major cause of sepsis, induce inflammatory cytokines in strict dependence on bacterial ssRNA and the host molecules MyD88 and UNC-93B. In this study, we show that NO plays an important role in Group B streptococci-induced transcriptional activation of cytokine genes. Phagocytosis induced NO in a MyD88-dependent fashion. In turn, NO propagated the acidification of phagosomes and the processing of phagosomal bacterial nucleic acids and was required for potent transcriptional activation of cytokine genes by streptococci. This NO-dependent amplification loop has important mechanistic implications for the anti-streptococcal macrophage response and sepsis pathogenesis.

Toll-like receptor 9 (TLR9) induces an inflammatory response by recognition of unmethylated CpG dinucleotides, mainly present in prokaryotic DNA. So far, TLR9-deficient mice have been shown to be more sensitive than wild-type mice to viral, but not to bacterial infections. Here, we show that mice deficient in TLR9 but not in TLR1, TLR2, TLR4 and TLR6 or IL-1R/IL-18R are more susceptible to a respiratory tract bacterial infection caused by Streptococcus pneumoniae. Intranasal challenge studies revealed that TLR9 plays a protective role in the lungs at an early stage of infection prior to the entry of circulating inflammatory cells. Alveolar as well as bone marrow-derived macrophages deficient in either TLR9 or the myeloid adaptor differentiation protein MyD88 were impaired in pneumococcal uptake and in pneumococcal killing. Our data suggest that in the airways, pneumococcal infection triggers a TLR9 and MyD88-dependent activation of phagocytic activity from resident macrophages leading to an early clearance of bacteria from the lower respiratory tract.

MyD88 is a general adaptor protein that plays an important role in the Toll/IL-1 receptor family signalings. Recently, Toll-like receptors 2 and 4 (TLR2 and TLR4) have been suggested to be the signaling receptors for lipopolysaccharide (LPS). In this study, we demonstrate that MyD88 knockout mice lack the ability to respond to LPS as measured by shock response, B cell proliferative response, and secretion of cytokines by macrophages and embryonic fibroblasts. However, activation of neither NF-κB nor the mitogen-activated protein (MAP) kinase family is abolished in MyD88 knockout mice. These findings demonstrate that signaling via MyD88 is essential for LPS response, but the inability of MyD88 knockout mice to induce LPS-dependent gene expression cannot simply be attributed to lack of the activation of MAP kinases and NF-κB.

G‐CSF specifically stimulates the proliferation and differentiation of cells that are committed to the neutrophil‐granulocyte lineage. Although Stat3 was thought to be essential for the transduction of G‐CSF–induced cell proliferation and differentiation signals, mice deficient for Stat3 in hematopoietic cells show neutrocytosis and infiltration of cells into the digestive tract. The number of progenitor cells in the neutrophil lineage is not changed, and G‐CSF–induced proliferation of progenitor cells and prolonged neutrophil survival were observed in Stat3‐deficient mice. In hematopoietic cells from Stat3‐deficient mice, trace levels of SOCS3, a negative regulator of granulopoiesis, were observed, and SOCS3 expression was not induced by G‐CSF stimulation. Stat3‐null bone marrow cells displayed a significant activation of extra‐cellular regulated kinase 1 (ERK1)/ERK2 under basal conditions, and the activation of ERK was enhanced and sustained by G‐CSF stimulation. Furthermore, the augmented proliferation of Stat3‐deficient bone marrow cells in response to G‐CSF was dramatically decreased by addition of a MEK1 inhibitor. These results indicate that Stat3 functions as a negative regulator of G‐CSF signaling by inducing SOCS3 expression and that ERK activation is the major factor responsible for inducing the proliferation of hematopoietic cells in response to G‐CSF.

No abstract is provided for this article.

No abstract is provided for this article.

VOLUME 283 (2008) PAGES 36211–36220 On page 36212, the sequences of PCR primers of IFN-α are incorrect. The correct sequences are as follows: IFN-α, 5′-ATGGCTAGGCTCTGTGCTTTCCT-3′ and 5′-AGGGCTCTCCAGACTTCTGCTCTG-3′.

Background We reported previously that serum levels of IL-33 are significantly increased in patients with allergic rhinitis (AR). However, very little is known about the role of IL-33 for the development of AR. Objective We thought to develop a novel murine model of ragweed pollen–specific AR and examined the pathologic role for ragweed-induced IL-33 in the development of AR manifestation using IL-33–deficient (il33 −/− ) mice. Methods Ragweed-immunized and ragweed-challenged mice were examined for early- and late-phase nasal responses. IL-33 protein expression in the nasal epithelial cells of the AR murine model and patients with AR were assessed by using confocal microscopy. Results After nasal challenge with ragweed pollen, ragweed-immunized wild-type mice manifested early-phase (sneezing) and late-phase (eosinophilic and basophilic accumulation) responses. In contrast, il33 −/− and FcεRI−/− mice did not have both early- and late-phase AR responses. IL-33 protein was constitutively expressed in the nucleus of nasal epithelial cells and was promptly released into nasal fluids in response to nasal exposure to ragweed pollen. In human subjects we revealed constitutive expression of IL-33 protein in the nasal epithelial cells of healthy control subjects and downregulated expression of IL-33 protein in inflamed nasal epithelial cells of patients with AR. IL-33–stimulated mast cells and basophils contributed to the early- and late-phase AR manifestation through increasing histamine release and production of chemoattractants for eosinophils/basophils, respectively. Conclusions Ragweed pollen–driven endogenous IL-33 contributed to the development of AR responses. IL-33 might present an important therapeutic target for the prevention of AR.

All arthropod-borne flaviviruses generate a short noncoding RNA (sfRNA) from the viral 3′ untranslated region during infection due to stalling of the cellular 5′-to-3′ exonuclease XRN1. We show here that formation of sfRNA also inhibits XRN1 activity. Cells infected with Dengue or Kunjin viruses accumulate uncapped mRNAs, decay intermediates normally targeted by XRN1. XRN1 repression also resulted in the increased overall stability of cellular mRNAs in flavivirus-infected cells. Importantly, a mutant Kunjin virus that cannot form sfRNA but replicates to normal levels failed to affect host mRNA stability or XRN1 activity. Expression of sfRNA in the absence of viral infection demonstrated that sfRNA formation was directly responsible for the stabilization of cellular mRNAs. Finally, numerous cellular mRNAs were differentially expressed in an sfRNA-dependent fashion in a Kunjin virus infection. We conclude that flaviviruses incapacitate XRN1 during infection and dysregulate host mRNA stability as a result of sfRNA formation.

The cytoplasmic helicase RIG-I is an established sensor for viral 5′-triphosphorylated RNA species. Recently, RIG-I was also implicated in the detection of intracellular bacteria. However, little is known about the host cell specificity of this process and the bacterial pathogen-associated molecular pattern (PAMP) that activates RIG-I. Here we show that RNA of Salmonella enterica serovar Typhimurium activates production of beta interferon in a RIG-I-dependent fashion only in nonphagocytic cells. In phagocytic cells, RIG-I is obsolete for detection of Salmonella infection. We further demonstrate that Salmonella mRNA reaches the cytoplasm during infection and is thus accessible for RIG-I. The results from next-generation sequencing analysis of RIG-I-associated RNA suggest that coding bacterial mRNAs represent the activating PAMP. IMPORTANCE S. Typhimurium is a major food-borne pathogen. After fecal-oral transmission, it can infect epithelial cells in the gut as well as immune cells (mainly macrophages, dendritic cells, and M cells). The innate host immune system relies on a growing number of sensors that detect pathogen-associated molecular patterns (PAMPs) to launch a first broad-spectrum response to invading pathogens. Successful detection of a given pathogen depends on colocalization of host sensors and PAMPs as well as potential countermeasures of the pathogen during infection. RIG-I-like helicases were mainly associated with detection of RNA viruses. Our work shows that S. Typhimurium is detected by RIG-I during infection specifically in nonimmune cells.

Caffeine intake is associated with a reduced risk developing non-alcoholic fatty liver disease (NAFLD), but the underlying molecular mechanisms remain to be fully elucidated. We report here that caffeine markedly improved high fat diet-induced NAFLD in mice resulting in a 10-fold increase in circulating IL-6 levels, leading to STAT3 activation in the liver. Interestingly, the expression of IL-6 mRNA was not increased in the liver, but increased substantially in the muscles of caffeine-treated mice. Caffeine was found to stimulate IL-6 production in cultured myotubes but not in hepatocytes, adipocytes, or macrophages. The inhibition of p38/MAPK abrogated caffeine-induced IL-6 production in muscle cells. Caffeine failed to improve NAFLD in IL-6 and hepatocyte-specific STAT3 knockout mice, indicating that the IL-6/STAT3 pathway is vital for the hepatoprotective effects of caffeine in NAFLD. The possibility that IL-6/STAT3-mediated hepatic autophagosome induction and hepatocytic oxygen consumption are involved in the anti-NAFLD effects of caffeine cannot be excluded, based on the findings presented here. Our results reveal that caffeine ameliorates NAFLD via crosstalk between muscle IL-6 production and liver STAT3 activation.

Rare, circulating dendritic cells differentially shape the immunogenicity mechanisms for protection against H1N1 influenza.