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author:("Fikrig, enol")
1.  SERUM ANTIBODIES TO BORRELIA BURGDORFERI, ANAPLASMA PHAGOCYTOPHILUM, AND BABESIA MICROTI IN RECAPTURED WHITE-FOOTED MICE 
Journal of wildlife diseases  2013;49(2):294-302.
A mark-release-recapture study was conducted during 2007 through 2010 in six, tick-infested sites in Connecticut, United States to measure changes in antibody titers for Borrelia burgdorferi sensu stricto, Anaplasma phagocytophilum, and Babesia microti in Peromyscus leucopus (white-footed mice). There was an overall recapture rate of 40%, but only four tagged mice were caught in ≥2 yr. Sera from 561 mice were analyzed for total antibodies to B. burgdorferi and A. phagocytophilum by using whole-cell or recombinant (VlsE or protein 44) antigens in a solid-phase enzyme-linked immunosorbent assay or to whole-cell B. microti by indirect fluorescent antibody staining. Antibody prevalences were highly variable for B. burgdorferi (from 56% to 98%), A. phagocytophilum (from 11% to 85%), and B. microti (from 11% to 84%) depending on the site and time of sampling. Of 463 mice with antibodies, 206 (45%) had antibodies to all three pathogens. Changes in antibody status for some mice from negative to positive (117 seroconversions) or from positive to negative (55 reversions) were observed. Seroconversions were observed in 10.1% of 417 mice for B. burgdorferi, 18.0% of 306 mice for A. phagocytophilum, and 6.6% of 304 mice for B. microti; reversion rates were 5.3, 5.9, and 4.9%, respectively. Antibodies to all pathogens persisted in some mice over several weeks while, in others, there were marked declines in titration end points to negative status. The latter may indicate elimination of a certain pathogen, such as A. phagocytophilum, or that mouse immune systems ceased to produce antibodies despite an existing patent infection.
doi:10.7589/2012-06-172
PMCID: PMC3712766  PMID: 23568904
Anaplasma phagocytophilum; antibodies; Babesia microti; Borrelia burgdorferi; Peromyscus leucopus
2.  ELF4 is critical for induction of type I interferon and the host antiviral response 
Nature immunology  2013;14(12):1237-1246.
Induction of type I interferon is a central event of innate immunity, essential for host defense. Here we report that the transcription factor ELF4 is induced by type I interferon and upregulates interferon expression in a feed-forward loop. ELF4 deficiency leads to reduced interferon production, resulting in enhanced susceptibility to West Nile virus encephalitis in mice. After viral infection, ELF4 is recruited by STING, interacts with and is activated by the MAVS-TBK1 complex, and translocates into the nucleus to bind interferon promoters. Cooperative binding with ELF4 increases the binding affinity of interferon regulatory factors IRF3 and IRF7, which is mediated by EICE elements. Thus, in addition to identifying a regulator of innate immune signaling, we uncovered a role for EICE elements in interferon transactivation.
doi:10.1038/ni.2756
PMCID: PMC3939855  PMID: 24185615
4.  Age-associated Decrease in Toll-like Receptor Function in Primary Human Dendritic Cells Predicts Influenza Vaccine Response1 
Journal of immunology (Baltimore, Md. : 1950)  2010;184(5):10.4049/jimmunol.0901022.
We evaluated Toll-like receptor (TLR) function in primary human dendritic cells from 104 young (age 21–30) and older (≥ 65 years) individuals. We used multicolor flow cytometry and intracellular cytokine staining of myeloid (mDC) and plasmacytoid (pDC) DCs and found substantial decreases in older, compared to young individuals in TNF-α, IL-6 and/or IL-12 (p40) production in mDCs and in TNF-α and IFN-α production in pDCs in response to TLR1/2, TLR2/6, TLR3, TLR5, and TLR8 engagement in mDCs and TLR7 and TLR9 in pDCs. These differences were highly significant after adjustment for heterogeneity between young and older groups (e.g. gender, race, body mass index [BMI], number of comorbid medical conditions) using mixed effect statistical modeling. Studies of surface and intracellular expression of TLR proteins, and of TLR gene expression in purified mDCs and pDCs revealed potential contributions for both transcriptional and post-transcriptional mechanisms in these age-associated effects. Moreover, intracellular cytokine production in the absence of TLR ligand stimulation was elevated in cells from older, compared to young individuals, suggesting a dysregulation of cytokine production that may limit further activation by TLR engagement. Our results provide evidence for immunosenescence in dendritic cells; notably, defects in cytokine production were strongly associated with poor antibody response to influenza immunization, a functional consequence of impaired TLR function in the aging innate immune response.
doi:10.4049/jimmunol.0901022
PMCID: PMC3867271  PMID: 20100933
5.  Immunity’s fourth dimension: approaching the circadian-immune connection 
Trends in immunology  2012;33(12):607-612.
The circadian system ensures the generation and maintenance of self-sustained ~24 h rhythms in physiology that are linked to internal and environmental changes. In mammals, daily variations in light intensity and other cues are integrated by a hypothalamic master clock that conveys circadian information to peripheral molecular clocks that orchestrate physiology. Multiple immune parameters also vary throughout the day and disruption of circadian homeostasis is associated with immune-related disease. Here we discuss the molecular links between the circadian and immune systems and examine their outputs and disease implications. Understanding the mechanisms that underlie circadian-immune crosstalk may prove valuable for devising novel prophylactic and therapeutic interventions.
doi:10.1016/j.it.2012.08.007
PMCID: PMC3712756  PMID: 23000010
circadian; immunity; daily rhythms
6.  Rip2 contributes to host defense against Anaplasma phagocytophilum infection 
The gram-negative obligate intracellular bacterium Anaplasma phagocytophilum is the causative agent of human granulocytic anaplasmosis (HGA), an emerging tick-borne infectious disease occuring worldwide. HGA is generally self-limiting, however, the underlying mechanisms, particularly the innate immune pathways that mediate the immune clearance of A. phagocytophilum, are less understood. We herein report an unexpected role for Receptor interacting protein-2 (Rip2), the adaptor protein for the Nod-Like Receptors (NLRs), Nod1/Nod2, in the host immune response against A. phagocytophilum infection. Although A. phagocytophilum genome is reported to lack the genes encoding the known ligands of Nod1 and Nod2, its infection up-regulated the transcription of Rip2 in human primary neutrophils. Our results revealed that Rip2 deficient mice had significantly higher bacterial load than wild type controls throughout the infection period. In addition, the Rip2 deficient mice took strikingly longer duration to clear A. phagocytophilum infection. Detailed analysis identified that interferon gamma (IFNγ) and interleukin -18 (IL-18) but not interleukin -12 (IL-12), macrophage inflammatory protein-2 (MIP-2), and KC response were diminished in A. phagocytophilum-challenged Rip2 deficient mice. Together, these results revealed that Rip2 play important roles in the immune control of A. phagocytophilum, and may contribute to our understanding of the host response to Rickettsiales.
doi:10.1111/j.1574-695X.2012.01001.x
PMCID: PMC3530031  PMID: 22747758
Rip2; Anaplasma phagocytophilum; NLR; IFNγ
7.  Ixodes scapularis JAK-STAT Pathway Regulates Tick Antimicrobial Peptides, Thereby Controlling the Agent of Human Granulocytic Anaplasmosis 
The Journal of Infectious Diseases  2012;206(8):1233-1241.
Ixodes scapularis transmits the agent of human granulocytic anaplasmosis, among other pathogens. The mechanisms used by the tick to control Anaplasma phagocytophilum are not known. We demonstrate that the I. scapularis Janus kinase (JAK)–signaling transducer activator of transcription (STAT) pathway plays a critical role in A. phagocytophilum infection of ticks. The A. phagocytophilum burden increases in salivary glands and hemolymph when the JAK-STAT pathway is suppressed by RNA interference. The JAK-STAT pathway exerts its anti-Anaplasma activity presumably through STAT-regulated effectors. A salivary gland gene family encoding 5.3-kDa antimicrobial peptides is highly induced upon A. phagocytophilum infection of tick salivary glands. Gene expression and electrophoretic mobility shift assays showed that the 5.3-kDa antimicrobial peptide–encoding genes are regulated by tick STAT. Silencing of these genes increased A. phagocytophilum infection of tick salivary glands and transmission to mammalian host. These data suggest that the JAK-STAT signaling pathway plays a key role in controlling A. phagocytophilum infection in ticks by regulating the expression of antimicrobial peptides.
doi:10.1093/infdis/jis484
PMCID: PMC3448968  PMID: 22859824
8.  West Nile Virus: Biology, Transmission, and Human Infection 
Clinical Microbiology Reviews  2012;25(4):635-648.
Summary: West Nile Virus was introduced into the Western Hemisphere during the late summer of 1999 and has been causing significant and sometimes severe human diseases since that time. This article briefly touches upon the biology of the virus and provides a comprehensive review regarding recent discoveries about virus transmission, virus acquisition, and human infection and disease.
doi:10.1128/CMR.00045-12
PMCID: PMC3485754  PMID: 23034323
9.  TLR9-Targeted Biodegradable Nanoparticles as Immunization Vectors Protect against West Nile Encephalitis 
Vaccines that activate humoral and cell-mediated immune responses are urgently needed for many infectious agents, including the flaviviruses dengue and West Nile (WN) virus. Vaccine development would be greatly facilitated by a new approach, in which nanoscale modules (Ag, adjuvant, and carrier) are assembled into units that are optimized for stimulating immune responses to a specific pathogen. Toward that goal, we formulated biodegradable nanoparticles loaded with Ag and surface modified with the pathogen-associated molecular pattern CpG oligodeoxynucleotides. We chose to evaluate our construct using a recombinant envelope protein Ag from the WN virus and tested the efficiency of this system in eliciting humoral and cellular responses and providing protection against the live virus. Animals immunized with this system showed robust humoral responses polarized toward Th1 immune responses compared with predominately Th2-biased responses with the adjuvant aluminum hydroxide. Immunization with CpG oligodeoxynucleotide-modified nanoparticles resulted in a greater number of circulating effector T cells and greater activity of Ag-specific lymphocytes than unmodified nanoparticles or aluminum hydroxide. Ultimately, compared with alum, this system offered superior protection in a mouse model of WN virus encephalitis.
doi:10.4049/jimmunol.1000768
PMCID: PMC3753007  PMID: 20660705
10.  MyD88 Deficiency Markedly Worsens Tissue Inflammation and Bacterial Clearance in Mice Infected with Treponema pallidum, the Agent of Syphilis 
PLoS ONE  2013;8(8):e71388.
Research on syphilis, a sexually transmitted infection caused by the non-cultivatable spirochete Treponema pallidum, has been hampered by the lack of an inbred animal model. We hypothesized that Toll-like receptor (TLR)-dependent responses are essential for clearance of T. pallidum and, consequently, compared infection in wild-type (WT) mice and animals lacking MyD88, the adaptor molecule required for signaling by most TLRs. MyD88-deficient mice had significantly higher pathogen burdens and more extensive inflammation than control animals. Whereas tissue infiltrates in WT mice consisted of mixed mononuclear and plasma cells, infiltrates in MyD88-deficient animals were predominantly neutrophilic. Although both WT and MyD88-deficient mice produced antibodies that promoted uptake of treponemes by WT macrophages, MyD88-deficient macrophages were deficient in opsonophagocytosis of treponemes. Our results demonstrate that TLR-mediated responses are major contributors to the resistance of mice to syphilitic disease and that MyD88 signaling and FcR-mediated opsonophagocytosis are linked to the macrophage-mediated clearance of treponemes.
doi:10.1371/journal.pone.0071388
PMCID: PMC3734110  PMID: 23940747
11.  Impaired Toll-Like Receptor 3-Mediated Immune Responses from Macrophages of Patients Chronically Infected with Hepatitis C Virus 
Hepatitis C virus (HCV) is the most common chronic blood-borne infection in the United States, with the majority of patients becoming chronically infected and a subset (20%) progressing to cirrhosis and hepatocellular carcinoma. Individual variations in immune responses may help define successful resistance to infection with HCV. We have compared the immune response in primary macrophages from patients who have spontaneously cleared HCV (viral load negative [VL−], n = 37) to that of primary macrophages from HCV genotype 1 chronically infected (VL+) subjects (n = 32) and found that macrophages from VL− subjects have an elevated baseline expression of Toll-like receptor 3 (TLR3). Macrophages from HCV patients were stimulated ex vivo through the TLR3 pathway and assessed using gene expression arrays and pathway analysis. We found elevated TLR3 response genes and pathway activity from VL− subjects. Furthermore, macrophages from VL− subjects showed higher production of beta interferon (IFN-β) and related IFN response genes by quantitative PCR (Q-PCR) and increased phosphorylation of STAT-1 by immunoblotting. Analysis of polymorphisms in TLR3 revealed a significant association of intronic TLR3 polymorphism (rs13126816) with the clearance of HCV and the expression of TLR3. Of note, peripheral blood mononuclear cells (PBMCs) from the same donors showed opposite changes in gene expression, suggesting ongoing inflammatory responses in PBMCs from VL+ HCV patients. Our results suggest that an elevated innate immune response enhances HCV clearance mechanisms and may offer a potential therapeutic approach to increase viral clearance.
doi:10.1128/CVI.00530-12
PMCID: PMC3571267  PMID: 23220997
12.  Caspase-12 controls West Nile virus infection via the viral RNA receptor RIG-I 
Nature immunology  2010;11(10):912-919.
Caspase-12 has been shown to negatively modulate inflammasome signaling during bacterial infection. Its function in viral immunity, however, has not been characterized. We now report an important role for caspase-12 in controlling viral infection via the pattern-recognition receptor RIG-I. After challenge with West Nile virus (WNV), caspase-12-deficient mice had greater mortality, higher viral burden and defective type I interferon response compared with those of challenged wild-type mice. In vitro studies of primary neurons and mouse embryonic fibroblasts showed that caspase-12 positively modulated the production of type I interferon by regulating E3 ubiquitin ligase TRIM25–mediated ubiquitination of RIG-I, a critical signaling event for the type I interferon response to WNV and other important viral pathogens.
doi:10.1038/ni.1933
PMCID: PMC3712356  PMID: 20818395
13.  UBXN1 Interferes with Rig-I-like Receptor-Mediated Antiviral Immune Response by Targeting MAVS 
Cell reports  2013;3(4):10.1016/j.celrep.2013.02.027.
SUMMARY
RNA viruses are sensed by RIG-I-like receptors (RLRs), which signal through a mitochondria-associated adaptor molecule, MAVS, resulting in systemic antiviral immune responses. Although RLR signaling is essential for limiting RNA virus replication, it must be stringently controlled to prevent damage from inflammation. We demonstrate here that among all tested UBX-domain-containing protein family members, UBXN1 exhibits the strongest inhibitory effect on RNA-virus-induced type I interferon response. UBXN1 potently inhibits RLR- and MAVS-induced, but not TLR3-, TLR4-, or DNA-virus-induced innate immune responses. Depletion of UBXN1 enhances virus-induced innate immune responses, including those resulting from RNA viruses such as vesicular stomatitis, Sendai, West Nile, and dengue virus infection, repressing viral replication. Following viral infection, UBXN1 is induced, binds to MAVS, interferes with intracellular MAVS oligomerization, and disrupts the MAVS/TRAF3/TRAF6 signalosome. These findings underscore a critical role of UBXN1 in the modulation of a major antiviral signaling pathway.
doi:10.1016/j.celrep.2013.02.027
PMCID: PMC3707122  PMID: 23545497
14.  Characterization of Ixophilin, A Thrombin Inhibitor from the Gut of Ixodes scapularis 
PLoS ONE  2013;8(7):e68012.
Ixodes scapularis, the black-legged tick, vectors several human pathogens including Borrelia burgdorferi, the agent of Lyme disease in North America. Pathogen transmission to the vertebrate host occurs when infected ticks feed on the mammalian host to obtain a blood meal. Efforts to understand how the tick confronts host hemostatic mechanisms and imbibes a fluid blood meal have largely focused on the anticoagulation strategies of tick saliva. The blood meal that enters the tick gut remains in a fluid state for several days during the process of feeding, and the role of the tick gut in maintaining the blood-meal fluid is not understood. We now demonstrate that the tick gut produces a potent inhibitor of thrombin, a key enzyme in the mammalian coagulation cascade. Chromatographic fractionation of engorged tick gut proteins identified one predominant thrombin inhibitory activity associated with an approximately 18 kDa protein, henceforth referred to as Ixophilin. The ixophilin gene was preferentially transcribed in the guts of feeding nymphs. Expression began after 24 hours of feeding, coincident with the flow of host blood into the tick gut. Immunity against Ixophilin delayed tick feeding, and decreased feeding efficiency significantly. Surprisingly, immunity against Ixophilin resulted in increased Borrelia burgdorferi transmission to the host, possibly due to delayed feeding and increased transmission opportunity. These observations illuminate the potential drawbacks of targeting individual tick proteins in a functional suite. They also underscore the need to identify the “anticoagulome” of the tick gut, and to prioritize a critical subset of anticoagulants that could be targeted to efficiently thwart tick feeding, and block pathogen transmission to the vertebrate host.
doi:10.1371/journal.pone.0068012
PMCID: PMC3706618  PMID: 23874485
15.  Anaplasma phagocytophilum Asp14 Is an Invasin That Interacts with Mammalian Host Cells via Its C Terminus To Facilitate Infection 
Infection and Immunity  2013;81(1):65-79.
Anaplasma phagocytophilum, a member of the family Anaplasmataceae, is the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis. The life cycle of A. phagocytophilum is biphasic, transitioning between the noninfectious reticulate cell (RC) and infectious dense-cored (DC) forms. We analyzed the bacterium's DC surface proteome by selective biotinylation of surface proteins, NeutrAvidin affinity purification, and mass spectrometry. Transcriptional profiling of selected outer membrane protein candidates over the course of infection revealed that aph_0248 (designated asp14 [14-kDa A. phagocytophilum surface protein]) expression was upregulated the most during A. phagocytophilum cellular invasion. asp14 transcription was induced during transmission feeding of A. phagocytophilum-infected ticks on mice and was upregulated when the bacterium engaged its receptor, P-selectin glycoprotein ligand 1. Asp14 localized to the A. phagocytophilum surface and was expressed during in vivo infection. Treating DC organisms with Asp14 antiserum or preincubating mammalian host cells with glutathione S-transferase (GST)–Asp14 significantly inhibited infection of host cells. Moreover, preincubating host cells with GST-tagged forms of both Asp14 and outer membrane protein A, another A. phagocytophilum invasin, pronouncedly reduced infection relative to treatment with either protein alone. The Asp14 domain that is sufficient for cellular adherence and invasion lies within the C-terminal 12 to 24 amino acids and is conserved among other Anaplasma and Ehrlichia species. These results identify Asp14 as an A. phagocytophilum surface protein that is critical for infection, delineate its invasion domain, and demonstrate the potential of targeting Asp14 in concert with OmpA for protecting against infection by A. phagocytophilum and other Anaplasmataceae pathogens.
doi:10.1128/IAI.00932-12
PMCID: PMC3536139  PMID: 23071137
16.  Identification of Genes Critical for Resistance to Infection by West Nile Virus Using RNA-Seq Analysis 
Viruses  2013;5(7):1664-1681.
The West Nile virus (WNV) is an emerging infection of biodefense concern and there are no available treatments or vaccines. Here we used a high-throughput method based on a novel gene expression analysis, RNA-Seq, to give a global picture of differential gene expression by primary human macrophages of 10 healthy donors infected in vitro with WNV. From a total of 28 million reads per sample, we identified 1,514 transcripts that were differentially expressed after infection. Both predicted and novel gene changes were detected, as were gene isoforms, and while many of the genes were expressed by all donors, some were unique. Knock-down of genes not previously known to be associated with WNV resistance identified their critical role in control of viral infection. Our study distinguishes both common gene pathways as well as novel cellular responses. Such analyses will be valuable for translational studies of susceptible and resistant individuals—and for targeting therapeutics—in multiple biological settings.
doi:10.3390/v5071664
PMCID: PMC3738954  PMID: 23881275
anti-viral gene expression; immune response; macrophage; RNA-Seq; West Nile virus
17.  Fusion loop peptide of the West Nile virus envelope protein is essential for pathogenesis and recognized by a therapeutic cross-reactive human monoclonal antibody 
West Nile virus is an emerging pathogen that can cause fatal neurological disease. A recombinant human monoclonal antibody, mAb11, has been described as a candidate for the prevention and treatment of West Nile disease. Using a yeast surface display epitope mapping assay and neutralization escape mutant, we show that mAb11 recognizes the fusion loop, at the distal end of domain II of the West Nile virus envelope protein. Antibody mAb11 cross-reacts with all four dengue viruses and provides protection against dengue (serotype 2 and 4) viruses. In contrast to the parental West Nile virus, a neutralization escape variant failed to cause lethal encephalitis (at higher infectious doses) or induce the inflammatory responses associated with blood-brain barrier permeability in mice, suggesting an important role for the fusion loop in viral pathogenesis. Our data demonstrate that an intact West Nile virus fusion loop is critical for virulence, and that human mAb11 targeting this region is efficacious against West Nile virus infection. These experiments define the molecular determinant on the envelope protein recognized by mAb11 and demonstrate the importance of this region in causing West Nile encephalitis.
doi:10.4049/jimmunol.0900093
PMCID: PMC3690769  PMID: 19535627
Viral; antibodies; cytokines; inflammation; vaccination
18.  Anaplasma phagocytophilum Outer Membrane Protein A Interacts with Sialylated Glycoproteins To Promote Infection of Mammalian Host Cells 
Infection and Immunity  2012;80(11):3748-3760.
Anaplasma phagocytophilum is the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis (HGA). A. phagocytophilum binding to sialyl Lewis x (sLex) and other sialylated glycans that decorate P selectin glycoprotein 1 (PSGL-1) and other glycoproteins is critical for infection of mammalian host cells. Here, we demonstrate the importance of A. phagocytophilum outer membrane protein A (OmpA) APH_0338 in infection of mammalian host cells. OmpA is transcriptionally induced during transmission feeding of A. phagocytophilum-infected ticks on mice and is upregulated during invasion of HL-60 cells. OmpA is presented on the pathogen's surface. Sera from HGA patients and experimentally infected mice recognize recombinant OmpA. Pretreatment of A. phagocytophilum organisms with OmpA antiserum reduces their abilities to infect HL-60 cells. The OmpA N-terminal region is predicted to contain the protein's extracellular domain. Glutathione S-transferase (GST)-tagged versions of OmpA and OmpA amino acids 19 to 74 (OmpA19-74) but not OmpA75-205 bind to, and competitively inhibit A. phagocytophilum infection of, host cells. Pretreatment of host cells with sialidase or trypsin reduces or nearly eliminates, respectively, GST-OmpA adhesion. Therefore, OmpA interacts with sialylated glycoproteins. This study identifies the first A. phagocytophilum adhesin-receptor pair and delineates the region of OmpA that is critical for infection.
doi:10.1128/IAI.00654-12
PMCID: PMC3486060  PMID: 22907813
19.  Horizontal and Vertical Transmission of West Nile Virus Genotype NY99 by Culex salinarius and Genotypes NY99 and WN02 by Culex tarsalis 
Culex tarsalis is a superior horizontal and vertical vector of West Nile virus (WNV) compared with Culex salinarius. Culex salinarius transmitted WNV genotype NY99 (CT 2741-99 strain) horizontally to suckling mice at significantly lower rates than Cx. tarsalis on Days 8, 9, 10, and 12 post-infection, and Cx. salinarius transmitted WNV genotype NY99 to offspring at a lower vertical transmission infection rate than Cx. tarsalis. Culex tarsalis transmitted WNV genotypes NY99 and WN02 (CT S0084-08 strain) with equal efficiency. Daily percent horizontal transmission of genotype NY99 by Cx. tarsalis-infected per os and by intra-thoracic infection was not significantly different from daily transmission of genotype WN02 from Days 5–23 and Days 2–9 post-infection, respectively. Our findings do not support the previously published hypothesis that genotype NY99 was replaced in the New World by WN02 because of a shorter extrinsic incubation of WN02.
doi:10.4269/ajtmh.2012.11-0473
PMCID: PMC3247122  PMID: 22232464
20.  Age-associated elevation in TLR5 leads to increased inflammatory responses in the elderly 
Aging cell  2011;11(1):104-110.
Summary
Aging is accompanied by a progressive decline in immune function. Studies have shown age-related decreases in expression and signaling efficiency of Toll-like receptors (TLRs) in monocytes and dendritic cells and dysregulation of macrophage TLR3. Using a multivariable mixed effect model, we report a highly significant increase in TLR5 induced production of IL-8 from monocytes of older individuals (p<0.0001). Elevated IL-8 is accompanied by increased expression of TLR5, both protein and mRNA, and by increased levels of TLR5 mediated phosphorylation of MAPK p38 and ERK. We noted incomplete activation of NF-κB in response to TLR5 signaling in monocytes of elderly donors, as reflected by the absence of an associated increase in the production of TNF-α. Elevated TLR5 may provide a critical mechanism to enhance immune responsiveness in older individuals.
doi:10.1111/j.1474-9726.2011.00759.x
PMCID: PMC3257374  PMID: 22023165
Aging; Toll-like receptors; monocytes; immunosenescence; IL-8; flagellin
21.  Innate Immune Control of West Nile Virus Infection 
Cellular microbiology  2011;13(11):1648-1658.
West Nile virus (WNV), from the Flaviviridae family, is a re-emerging zoonotic pathogen of medical importance. In humans, WNV infection may cause life-threatening meningoencephalitis or long-term neurologic sequelae. WNV is transmitted by Culex spp mosquitoes and both the arthropod vector and the mammalian host are equipped with antiviral innate immune mechanisms sharing a common phylogeny. As far as the current evidence is able to demonstrate, mosquitoes primarily rely on RNA interference, Toll, Imd and JAK-STAT signaling pathways for limiting viral infection, while mammals are provided with these and other more complex antiviral mechanisms involving antiviral effectors, inflammatory mediators, and cellular responses triggered by highly specialized pathogen detection mechanisms that often resemble their invertebrate ancestry. This mini-review summarizes our current understanding of how the innate immune systems of the vector and the mammalian host react to WNV infection and shape its pathogenesis.
doi:10.1111/j.1462-5822.2011.01649.x
PMCID: PMC3196381  PMID: 21790942
22.  Postgenomic Analyses Reveal Development of Infectious Anaplasma phagocytophilum during Transmission from Ticks to Mice 
Journal of Bacteriology  2012;194(9):2238-2247.
Obligate intracellular bacteria of the Rickettsiales order have evolved to colonize both arthropod and mammalian hosts, but few details are known about the bacterial adaptations that occur during transmission from blood-feeding arthropods to mammals. Here we apply proteomics and transcriptome sequencing to Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis, in Ixodes scapularis tick salivary glands, to detect proteins or genes expressed by the pathogen during transmission feeding by the tick. We detected expression of 139 genes, representing 11% of the open reading frames (ORFs) in the A. phagocytophilum genome. The predominant categories of proteins were ribosomal proteins, cell surface proteins, chaperones, and uncharacterized proteins. There was no evidence of DNA replication enzymes, suggesting that most of the A. phagocytophilum cells were no longer dividing. Instead, protein expression reflected conversion to the extracellular, infectious “dense-core” (DC) form. High expression of a DC-specific marker, APH_1235, further suggested this developmental transition in ticks. We showed that blocking APH_1235 with antibodies reduced A. phagocytophilum infection levels in mammalian cell culture. This work represents a starting point for clarifying essential proteins expressed by A. phagocytophilum during transmission from ticks to mammals and demonstrates that the abundantly expressed, DC-associated APH_1235 protein is important during in vivo infection by A. phagocytophilum.
doi:10.1128/JB.06791-11
PMCID: PMC3347074  PMID: 22389475
23.  prM-antibody renders immature West Nile virus infectious in vivo 
The Journal of General Virology  2011;92(Pt 10):2281-2285.
West Nile virus (WNV) is a member of the family Flaviviridae and is a neurotropic pathogen responsible for severe human disease. Flavivirus-infected cells release virus particles that contain variable numbers of precursor membrane (prM) protein molecules at the viral surface. Consequently, antibodies are produced against the prM protein. These antibodies have been shown to activate the infectious potential of fully immature flavivirus particles in vitro. Here, we provide in vivo proof that prM antibodies render immature WNV infectious. Infection with antibody-opsonized immature WNV particles caused disease and death of mice, and infectious WNV was found in the brains and sera.
doi:10.1099/vir.0.031427-0
PMCID: PMC3347797  PMID: 21697345
24.  Circadian expression of clock genes in mouse macrophages, dendritic cells, and B cells 
Brain, behavior, and immunity  2011;26(3):407-413.
In mammals, circadian and daily rhythms influence nearly all aspects of physiology, ranging from behavior to gene expression. Functional molecular clocks have been described in the murine spleen and splenic NK cells. The aim of our study was to investigate the existence of molecular clock mechanisms in other immune cells. Therefore, we measured the circadian changes in gene expression of clock genes (Per1, Per2, Bmal1, and Clock) and clock-controlled transcription factors (Rev-erbα and Dbp) in splenic enriched macrophages, dendritic cells, and B cells in both mice entrained to a light-dark cycle and under constant environmental conditions. Our study reveals the existence of functional molecular clock mechanisms in splenic macrophages, dendritic cells, and B cells.
doi:10.1016/j.bbi.2011.10.001
PMCID: PMC3336152  PMID: 22019350
Mouse splenic macrophages; dendritic cells; B cells possess functional circadian molecular clocks
25.  IL-22 Signaling Contributes to West Nile Encephalitis Pathogenesis 
PLoS ONE  2012;7(8):e44153.
The Th17 cytokine, IL-22, regulates host immune responses to extracellular pathogens. Whether IL-22 plays a role in viral infection, however, is poorly understood. We report here that Il22−/− mice were more resistant to lethal West Nile virus (WNV) encephalitis, but had similar viral loads in the periphery compared to wild type (WT) mice. Viral loads, leukocyte infiltrates, proinflammatory cytokines and apoptotic cells in the central nervous system (CNS) of Il22−/− mice were also strikingly reduced. Further examination showed that Cxcr2, a chemokine receptor that plays a non-redundant role in mediating neutrophil migration, was significantly reduced in Il22−/− compared to WT leukocytes. Expression of Cxcr2 ligands, cxcl1 and cxcl5, was lower in Il22−/− brains than wild type mice. Correspondingly, neutrophil migration from the blood into the brain was attenuated following lethal WNV infection of Il22−/− mice. Our results suggest that IL-22 signaling exacerbates lethal WNV encephalitis likely by promoting WNV neuroinvasion.
doi:10.1371/journal.pone.0044153
PMCID: PMC3429482  PMID: 22952908

Results 1-25 (121)