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1.
Am J Trop Med Hyg ; 2022 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-35226872

RESUMO

Monocyte dysfunction in helminth infection is one of the mechanisms proposed to explain the diminished parasite antigen-specific T cell responses seen with patent filarial infection. In fact, monocytes from filariae-infected individuals demonstrate internalized filarial antigens and, as a consequence, express inhibitory surface molecules and have diminished cytokine production. To investigate the mechanisms underlying monocyte dysfunction in filarial infections, purified human monocytes were exposed to live microfilariae (mf) of Brugia malayi, and the mRNA and protein expression of important inhibitory and/or autophagy-related molecules were assessed. Our data indicate that mf-induced autophagy in human monocytes shown by the formation of autophagic vesicles, by the upregulation in the mRNA expression of autophagy-related genes BCN1, LC3B, ATG5, ATG7 (P < 0.05), and by increase in the levels of LC3B protein. Furthermore, this mf-induced autophagy increased the levels of monocyte CD206 expression. In addition, mf significantly induced the frequency of interferon (IFN)-γ+ human monocytes and at the same time induced the mRNA expression of indoleamine 2,3-dioxygenase (IDO) through an IFN-γ-dependent mechanism; significantly enhanced tryptophan degradation (an indicator of IDO activity; P < 0.005). Interestingly, this autophagy induction by mf in monocytes was IFN-γ-dependent but IDO-independent as was reversed by anti-IFN-γ but not by an IDO inhibitor. Our data collectively suggest that mf of Brugia malayi regulate the function of monocytes by induction of IDO and IFN-γ, induce autophagy through an IFN-γ-dependent mechanism, and increase M2 phenotype through induction of autophagy; all acting in concert to drive monocyte dysfunction.

2.
Proc Natl Acad Sci U S A ; 118(34)2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34417304

RESUMO

Despite widespread yearly vaccination, influenza leads to significant morbidity and mortality across the globe. To make a more broadly protective influenza vaccine, it may be necessary to elicit antibodies that can activate effector functions in immune cells, such as antibody-dependent cellular cytotoxicity (ADCC). There is growing evidence supporting the necessity for ADCC in protection against influenza and herpes simplex virus (HSV), among other infectious diseases. An HSV-2 strain lacking the essential glycoprotein D (gD), was used to create ΔgD-2, which is a highly protective vaccine against lethal HSV-1 and HSV-2 infection in mice. It also elicits high levels of IgG2c antibodies that bind FcγRIV, a receptor that activates ADCC. To make an ADCC-eliciting influenza vaccine, we cloned the hemagglutinin (HA) gene from an H1N1 influenza A strain into the ΔgD-2 HSV vector. Vaccination with ΔgD-2::HAPR8 was protective against homologous influenza challenge and elicited an antibody response against HA that inhibits hemagglutination (HAI+), is predominantly IgG2c, strongly activates FcγRIV, and protects against influenza challenge following passive immunization of naïve mice. Prior exposure of mice to HSV-1, HSV-2, or a replication-defective HSV-2 vaccine (dl5-29) does not reduce protection against influenza by ΔgD-2::HAPR8 This vaccine also continues to elicit protection against both HSV-1 and HSV-2, including high levels of IgG2c antibodies against HSV-2. Mice lacking the interferon-α/ß receptor and mice lacking the interferon-γ receptor were also protected against influenza challenge by ΔgD-2::HAPR8 Our results suggest that ΔgD-2 can be used as a vaccine vector against other pathogens, while also eliciting protective anti-HSV immunity.


Assuntos
Citotoxicidade Celular Dependente de Anticorpos/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Herpes Simples/imunologia , Vacinas contra Influenza/administração & dosagem , Infecções por Orthomyxoviridae/prevenção & controle , Orthomyxoviridae/imunologia , Animais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Feminino , Herpes Simples/prevenção & controle , Herpesvirus Humano 1/fisiologia , Herpesvirus Humano 2/fisiologia , Vacinas contra Influenza/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae/imunologia
3.
PLoS Negl Trop Dis ; 12(4): e0006404, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29668679

RESUMO

A number of features at the host-parasite interface are reminiscent of those that are also observed at the host-tumor interface. Both cancer cells and parasites establish a tissue microenvironment that allows for immune evasion and may reflect functional alterations of various innate cells. Here, we investigated how the phenotype and function of human monocytes is altered by exposure to cancer cell lines and if these functional and phenotypic alterations parallel those induced by exposure to helminth parasites. Thus, human monocytes were exposed to three different cancer cell lines (breast, ovarian, or glioblastoma) or to live microfilariae (mf) of Brugia malayi-a causative agent of lymphatic filariasis. After 2 days of co-culture, monocytes exposed to cancer cell lines showed markedly upregulated expression of M1-associated (TNF-α, IL-1ß), M2-associated (CCL13, CD206), Mreg-associated (IL-10, TGF-ß), and angiogenesis associated (MMP9, VEGF) genes. Similar to cancer cell lines, but less dramatically, mf altered the mRNA expression of IL-1ß, CCL13, TGM2 and MMP9. When surface expression of the inhibitory ligands PDL1 and PDL2 was assessed, monocytes exposed to both cancer cell lines and to live mf significantly upregulated PDL1 and PDL2 expression. In contrast to exposure to mf, exposure to cancer cell lines increased the phagocytic ability of monocytes and reduced their ability to induce T cell proliferation and to expand Granzyme A+ CD8+ T cells. Our data suggest that despite the fact that helminth parasites and cancer cell lines are extraordinarily disparate, they share the ability to alter the phenotype of human monocytes.


Assuntos
Brugia Malayi/imunologia , Filariose/imunologia , Evasão da Resposta Imune , Monócitos/imunologia , Monócitos/parasitologia , Neoplasias/imunologia , Animais , Brugia Malayi/genética , Brugia Malayi/fisiologia , Linhagem Celular Tumoral , Filariose/parasitologia , Humanos , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Ativação Linfocitária , Fagocitose , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
4.
BMC Genomics ; 17: 406, 2016 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-27229469

RESUMO

BACKGROUND: Next-generation sequencing of transposon-genome junctions from a saturated bacterial mutant library (Tn-seq) is a powerful tool that permits genome-wide determination of the contribution of genes to fitness of the organism under a wide range of experimental conditions. We report development, testing, and results from a Tn-seq system for use in Streptococcus agalactiae (group B Streptococcus; GBS), an important cause of neonatal sepsis. METHODS: Our method uses a Himar1 mini-transposon that inserts at genomic TA dinucleotide sites, delivered to GBS on a temperature-sensitive plasmid that is subsequently cured from the bacterial population. In order to establish the GBS essential genome, we performed Tn-seq on DNA collected from three independent mutant libraries-with at least 135,000 mutants per library-at serial 24 h time points after outgrowth in rich media. RESULTS: After statistical analysis of transposon insertion density and distribution, we identified 13.5 % of genes as essential and 1.2 % as critical, with high levels of reproducibility. Essential and critical genes are enriched for fundamental cellular housekeeping functions, such as acyl-tRNA biosynthesis, nucleotide metabolism, and glycolysis. We further validated our system by comparing fitness assignments of homologous genes in GBS and a close bacterial relative, Streptococcus pyogenes, which demonstrated 93 % concordance. Finally, we used our fitness assignments to identify signal transduction pathway components predicted to be essential or critical in GBS. CONCLUSIONS: We believe that our baseline fitness assignments will be a valuable tool for GBS researchers and that our system has the potential to reveal key pathogenesis gene networks and potential therapeutic/preventative targets.


Assuntos
Genoma Bacteriano , Genômica , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/genética , Elementos de DNA Transponíveis , Biblioteca Gênica , Vetores Genéticos/genética , Genômica/métodos , Mutagênese Insercional , Transdução de Sinais , Streptococcus agalactiae/metabolismo
5.
J Infect Dis ; 210(2): 265-73, 2014 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-24474814

RESUMO

BACKGROUND: Maternal vaginal colonization with Streptococcus agalactiae (Group B Streptococcus [GBS]) is a precursor to chorioamnionitis, fetal infection, and neonatal sepsis, but the understanding of specific factors in the pathogenesis of ascending infection remains limited. METHODS: We used a new murine model to evaluate the contribution of the pore-forming GBS ß-hemolysin/cytolysin (ßH/C) to vaginal colonization, ascension, and fetal infection. RESULTS: Competition assays demonstrated a marked advantage to ßH/C-expressing GBS during colonization. Intrauterine fetal demise and/or preterm birth were observed in 54% of pregnant mice colonized with wild-type (WT) GBS and 0% of those colonized with the toxin-deficient cylE knockout strain, despite efficient colonization and ascension by both strains. Robust placental inflammation, disruption of maternal-fetal barriers, and fetal infection were more frequent in animals colonized with WT bacteria. Histopathologic examination revealed bacterial tropism for fetal lung and liver. CONCLUSIONS: Preterm birth and fetal demise are likely the direct result of toxin-induced damage and inflammation rather than differences in efficiency of ascension into the upper genital tract. These data demonstrate a distinct contribution of ßH/C to GBS chorioamnionitis and subsequent fetal infection in vivo and showcase a model for this most proximal step in GBS pathogenesis.


Assuntos
Morte Fetal/induzido quimicamente , Morte Fetal/etiologia , Proteínas Hemolisinas/metabolismo , Nascimento Prematuro/induzido quimicamente , Nascimento Prematuro/etiologia , Infecções Estreptocócicas/patologia , Streptococcus agalactiae/fisiologia , Animais , Modelos Animais de Doenças , Feminino , Histocitoquímica , Humanos , Fígado/microbiologia , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Gravidez , Infecções Estreptocócicas/complicações
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