RESUMEN
We have formatted an assay to detect Mycobacterium tuberculosis complex infections of non-human primates. Commercially available reagents were used to elicit a specific immune response that was measured by interferon-gamma release. Initial evaluation using blood samples from Rhesus macaques experimentally infected with M tuberculosis distinguished infected versus uninfected animals.
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Ensayo de Inmunoadsorción Enzimática/veterinaria , Ensayos de Liberación de Interferón gamma/veterinaria , Macaca mulatta , Enfermedades de los Monos/diagnóstico , Mycobacterium tuberculosis/aislamiento & purificación , Tuberculosis/diagnóstico , Animales , Ensayo de Inmunoadsorción Enzimática/métodos , Ensayos de Liberación de Interferón gamma/métodosRESUMEN
Follicular regulatory T cells (Tfr) are a unique subset of CD4 T cells that control and impact adaptive immune responses in the lymphoid follicles and germinal centers (GC). Since their relatively recent discovery, several studies have revealed that Tfr interact with other cells within this niche and shape ensuing responses. Recent advances defining the functional and developmental characteristics of Tfr have revealed key characteristics of Tfr differentiation, GC recruitment and retention, and regulatory properties. Further, Tfr shape the GC response and balance tolerance through interactions with Tfh, by modifying Tfh number, diversity and function, as well as with B cells. Mechanisms by which Tfr regulate the GC include cell-to-cell interactions with Tfh and B cells, as well as altering their environment through cytokine production and sequestration. Tfr have been shown to have a diverse T cell receptor (TCR) repertoire and can be specific for immunizing agents, demonstrating a potential role in vaccine development. Due to these important characteristics and functions, Tfr play a major role in immune tolerance, response to infection, and vaccine efficacy.
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Linfocitos B/inmunología , Citocinas/inmunología , Centro Germinal/metabolismo , Tolerancia Inmunológica , Infecciones/inmunología , Linfocitos T Reguladores/inmunología , Animales , Linfocitos B/patología , Centro Germinal/patología , Humanos , Infecciones/patología , Linfocitos T Reguladores/patologíaRESUMEN
Follicular regulatory T (TFR) cells are a subset of CD4+ T cells in secondary lymphoid follicles. TFR cells were previously included in the follicular helper T (TFH) cell subset, which consists of cells that are highly permissive to HIV-1. The permissivity of TFR cells to HIV-1 is unknown. We find that TFR cells are more permissive than TFH cells to R5-tropic HIV-1 ex vivo TFR cells expressed more CCR5 and CD4 and supported higher frequencies of viral fusion. Differences in Ki67 expression correlated with HIV-1 replication. Inhibiting cellular proliferation reduced Ki67 expression and HIV-1 replication. Lymph node cells from untreated HIV-infected individuals revealed that TFR cells harbored the highest concentrations of HIV-1 RNA and highest levels of Ki67 expression. These data demonstrate that TFR cells are highly permissive to R5-tropic HIV-1 both ex vivo and in vivo This is likely related to elevated CCR5 levels combined with a heightened proliferative state and suggests that TFR cells contribute to persistent R5-tropic HIV-1 replication in vivoIMPORTANCE In chronic, untreated HIV-1 infection, viral replication is concentrated in secondary lymphoid follicles. Within secondary lymphoid follicles, follicular helper T (TFH) cells have previously been shown to be highly permissive to HIV-1. Recently, another subset of T cells in secondary lymphoid follicles was described, follicular regulatory T (TFR) cells. These cells share some phenotypic characteristics with TFH cells, and studies that showed that TFH cells are highly permissive to HIV-1 included TFR cells in their definition of TFH cells. The permissivity of TFR cells to HIV-1 has not previously been described. Here, we show that TFR cells are highly permissive to HIV-1 both ex vivo and in vivo The expression of Ki67, a marker of proliferative capacity, is predictive of expression of viral proteins, and downregulating Ki67 leads to concurrent decreases in expression of viral proteins. Our study provides new insight into HIV-1 replication and a potential new cell type to target for future treatment.
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Infecciones por VIH/inmunología , VIH-1/fisiología , Linfocitos T Colaboradores-Inductores/virología , Linfocitos T Reguladores/virología , Tropismo Viral , Adulto , Anciano , Células Cultivadas , Niño , Femenino , Células HEK293 , Humanos , Antígeno Ki-67/metabolismo , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/virología , Masculino , Persona de Mediana Edad , Tonsila Palatina/citología , Tonsila Palatina/virología , Replicación ViralRESUMEN
Targeting of myeloid-dendritic cell receptor DC-SIGN by numerous chronic infectious agents, including Porphyromonas gingivalis, is shown to drive-differentiation of monocytes into dysfunctional mDCs. These mDCs exhibit alterations of their fine-tuned homeostatic function and contribute to dysregulated immune-responses. Here, we utilize P. gingivalis mutant strains to show that pathogen-differentiated mDCs from primary human-monocytes display anti-apoptotic profile, exhibited by elevated phosphorylated-Foxo1, phosphorylated-Akt1, and decreased Bim-expression. This results in an overall inhibition of DC-apoptosis. Direct stimulation of complex component CD40 on DCs leads to activation of Akt1, suggesting CD40 involvement in anti-apoptotic effects observed. Further, these DCs drove dampened CD8+ T-cell and Th1/Th17 effector-responses while inducing CD25+Foxp3+CD127- Tregs. In vitro Treg induction was mediated by DC expression of indoleamine 2,3-dioxygenase, and was confirmed in IDO-KO mouse model. Pathogen-infected &CMFDA-labeled MoDCs long-lasting survival was confirmed in a huMoDC reconstituted humanized mice. In conclusion, our data implicate PDDCs as an important target for resolution of chronic infection.
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Células Dendríticas/inmunología , Porphyromonas gingivalis/patogenicidad , Animales , Apoptosis , Proteína 11 Similar a Bcl2/metabolismo , Antígenos CD40/metabolismo , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Diferenciación Celular , Citocinas/metabolismo , Células Dendríticas/metabolismo , Células Dendríticas/microbiología , Proteína Forkhead Box O1/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenasa/deficiencia , Indolamina-Pirrol 2,3,-Dioxigenasa/genética , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Monocitos/metabolismo , Porphyromonas gingivalis/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Linfocitos T Reguladores/citología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Células TH1/inmunología , Células TH1/metabolismo , Células Th17/citología , Células Th17/inmunología , Células Th17/metabolismoRESUMEN
During chronic HIV infection, viral replication is concentrated in secondary lymphoid follicles. Cytotoxic CD8 T cells control HIV replication in extrafollicular regions, but not in the follicle. Here, we show CXCR5hiCD44hiCD8 T cells are a regulatory subset differing from conventional CD8 T cells, and constitute the majority of CD8 T cells in the follicle. This subset, CD8 follicular regulatory T cells (CD8 TFR), expand in chronic SIV infection, exhibit enhanced expression of Tim-3 and IL-10, and express less perforin compared to conventional CD8 T cells. CD8 TFR modestly limit HIV replication in follicular helper T cells (TFH), impair TFH IL-21 production via Tim-3, and inhibit IgG production by B cells during ex vivo HIV infection. CD8 TFR induce TFH apoptosis through HLA-E, but induce less apoptosis than conventional CD8 T cells. These data demonstrate that a unique regulatory CD8 population exists in follicles that impairs GC function in HIV infection.
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Linfocitos T CD8-positivos/inmunología , Centro Germinal/inmunología , Infecciones por VIH/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Subgrupos de Linfocitos T/inmunología , Animales , Técnicas de Cocultivo , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Humanos , Macaca mulatta , Tonsila Palatina/inmunologíaRESUMEN
During untreated disease, HIV replication is concentrated within T follicular helper cells (TFH). Heightened permissiveness, the presence of highly infectious virions on follicular dendritic cells (FDCs), low frequencies of virus-specific cytotoxic T lymphocytes (CTLs) in B cell follicles, expansions in TFH, and TFH dysfunction, all likely promote replication in TFH. Limited data suggest that memory TFH play a role in the latent or subclinical reservoir of HIV during antiretroviral therapy (ART), potentially for many of the same reasons. A better understanding of the role of memory TFH and FDC-bound virions in promoting recrudescent viremia in the setting of ART cessation is essential. Studies that target follicular virus reservoirs are needed to determine their role in HIV latency and to suggest successful cure strategies.
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Infecciones por VIH/virología , VIH-1/fisiología , Linfocitos T Colaboradores-Inductores/virología , Animales , Infecciones por VIH/sangre , Infecciones por VIH/inmunología , Infecciones por VIH/terapia , Humanos , Linfocitos T Colaboradores-Inductores/inmunología , Latencia del Virus/inmunología , Latencia del Virus/fisiología , Replicación Viral/fisiologíaRESUMEN
HIV-1 replication is concentrated within CD4(+) T cells in B cell follicles of secondary lymphoid tissues during asymptomatic disease. Limited data suggest that a subset of T follicular helper cells (TFH) within germinal centers (GC) is highly permissive to HIV-1. Whether GC TFH are the major HIV-1 virus-producing cells in vivo has not been established. In this study, we investigated TFH permissivity to HIV-1 ex vivo by spinoculating and culturing tonsil cells with HIV-1 GFP reporter viruses. Using flow cytometry, higher percentages of GC TFH (CXCR5(high)PD-1(high)) and CXCR5(+)programmed cell death-1 (PD-1)(low) cells were GFP(+) than non-GC TFH (CXCR5(+)PD-1(intermediate)) or extrafollicular (EF) (CXCR5(-)) cells. When sorted prior to spinoculation, however, GC TFH were substantially more permissive than CXCR5(+)PD-1(low) or EF cells, suggesting that many GC TFH transition to a CXCR5(+)PD-1(low) phenotype during productive infection. In situ hybridization on inguinal lymph node sections from untreated HIV-1-infected individuals without AIDS revealed higher frequencies of HIV-1 RNA(+) cells in GC than non-GC regions of follicle or EF regions. Superinfection of HIV-1-infected individuals' lymph node cells with GFP reporter virus confirmed the permissivity of follicular cells ex vivo. Lymph node immunostaining revealed 96% of CXCR5(+)CD4(+) cells were located in follicles. Within sorted lymph node cells from four HIV-infected individuals, CXCR5(+) subsets harbored 11-66-fold more HIV-1 RNA than CXCR5(-) subsets, as determined by RT PCR. Thus, GC TFH are highly permissive to HIV-1, but downregulate PD-1 and, to a lesser extent, CXCR5 during HIV-1 replication. These data further implicate GC TFH as the major HIV-1-producing cells in chronic asymptomatic HIV-1 infection.
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Centro Germinal/inmunología , Infecciones por VIH/inmunología , VIH-1/fisiología , Linfocitos T Colaboradores-Inductores/inmunología , Enfermedades Asintomáticas , Diferenciación Celular , Células Cultivadas , Infecciones por VIH/virología , Especificidad del Huésped , Humanos , Tonsila Palatina/patología , Fenotipo , Receptor de Muerte Celular Programada 1/metabolismo , Receptores CXCR5/metabolismo , Sobreinfección , Linfocitos T Colaboradores-Inductores/virología , Replicación ViralRESUMEN
T follicular helper cells (TFH) are a specialized subset of CD4 T cells that reside in B cell follicles and promote B cell maturation into plasma cells and long-lived memory B cells. During chronic infection prior to the development of AIDS, HIV-1 (HIV) replication is largely concentrated in TFH. Paradoxically, TFH numbers are increased in early and midstages of disease, thereby promoting HIV replication and disease progression. Despite increased TFH numbers, numerous defects in humoral immunity are detected in HIV-infected individuals, including dysregulation of B cell maturation, impaired somatic hypermutation, and low quality of antibody production despite hypergammaglobulinemia. Clinically, these defects are manifested by increased vulnerability to bacterial infections and impaired vaccine responses, neither of which is fully reversed by antiretroviral therapy (ART). Deficits in TFH function, including reduced HIV-specific IL-21 production and low levels of co-stimulatory receptor expression, have been linked to these immune impairments. Impairments in TFH likely contribute as well to the ability of HIV to persist and evade humoral immunity, particularly the inability to develop broadly neutralizing antibodies. In addition to direct infection of TFH, other mechanisms that have been linked to TFH deficits in HIV infection include upregulation of PD-L1 on germinal center B cells and augmented follicular regulatory T cell responses. Challenges to development of strategies to enhance TFH function in HIV infection include lack of an established phenotype for memory TFH as well as limited understanding of the relationship between peripheral TFH and lymphoid tissue TFH. Interventions to augment TFH function in HIV-infected individuals could enhance immune reconstitution during ART and potentially augment cure strategies.
RESUMEN
Human and simian immunodeficiency viruses (HIV and SIV) exploit follicular lymphoid regions by establishing high levels of viral replication and dysregulating humoral immunity. Follicular regulatory T cells (TFR) are a recently characterized subset of lymphocytes that influence the germinal centre response through interactions with follicular helper T cells (TFH). Here, utilizing both human and rhesus macaque models, we show the impact of HIV and SIV infection on TFR number and function. We find that TFR proportionately and numerically expand during infection through mechanisms involving viral entry and replication, TGF-ß signalling, low apoptosis rates and the presence of regulatory dendritic cells. Further, TFR exhibit elevated regulatory phenotypes and impair TFH functions during HIV infection. Thus, TFR contribute to inefficient germinal centre responses and inhibit HIV and SIV clearance.
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Infecciones por VIH/inmunología , Ganglios Linfáticos/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Linfocitos T Colaboradores-Inductores/fisiología , Linfocitos T Reguladores/fisiología , Adulto , Anciano , Animales , Células Cultivadas , Enfermedad Crónica , Femenino , Humanos , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Macaca mulatta , Masculino , Persona de Mediana Edad , Tonsila Palatina/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Adulto JovenRESUMEN
Signaling via pattern recognition receptors (PRRs) expressed on professional antigen presenting cells, such as dendritic cells (DCs), is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs) and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs). We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs.
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Autofagia/inmunología , Moléculas de Adhesión Celular/metabolismo , Células Dendríticas/metabolismo , Lectinas Tipo C/metabolismo , Células Mieloides/metabolismo , Porphyromonas gingivalis/metabolismo , Receptores de Superficie Celular/metabolismo , Receptor Toll-Like 2/metabolismo , Dendritas/ultraestructura , Células Dendríticas/inmunología , Células Dendríticas/ultraestructura , Fimbrias Bacterianas , Humanos , Espacio Intracelular/inmunología , Espacio Intracelular/metabolismo , Monocitos/inmunología , Monocitos/ultraestructura , Células Mieloides/inmunología , Receptor Toll-Like 2/inmunologíaRESUMEN
BACKGROUND: The oral cavity is colonized by >10(9) bacteria, many of which can increase heart disease risk when seeded into the bloodstream. Most dentoalveolar surgeries require the use of surgical sutures. Suture placement and removal can increase the risk of postoperative infection and bacteremia. The aim of this study is to evaluate the antimicrobial activity of a novel quaternary ammonium compound, K21, when coated on different suture materials. METHODS: The periodontal pathogen Porphyromonas gingivalis and the endodontic species Enterococcus faecalis were grown to early log phase and inoculated on enriched Brucella blood agar, on which were placed identical lengths of surgical suture (chromic gut, polyester suture, silk, and nylon suture) and control unwaxed dental floss impregnated with K21 at 5%, 10%, 20%, and 25% volume/volume in ethanol vehicle. Controls included the following: 1) sutures treated with vehicle; 2) untreated sutures; and 3) unwaxed floss. Zones of inhibition in millimeters were measured at five randomized sites per suture/floss for each concentration and material used. Mean ± SD of zones of inhibition were calculated, and analysis of variance (P <0.05) was used to determine whether differences were statistically significant. RESULTS: The results indicate that K21-coated suture at concentrations ranging from 5% to 25%, depending on the type of suture, have antimicrobial activity for P. gingivalis and E. faecalis. Nylon suture coated with K21 at 5%, 10%, 20%, and 25% resulted in zones ranging from 3 to 11 mm. Polyester suture was more effective at lower K21 concentrations with 5% (P = 0.0031), 10% (P = 0.0011), and 20% (P = 0.0002), yielding 7.5, 8.3, and 10.5 mm zones of inhibition. K21-coated silk suture yielded significant zones of inhibition at 25% (P <0.0001), whereas chromic gut was effective at K21 concentrations of 5% (P = 0.0081) and 25% (P <0.0001). CONCLUSION: It is concluded that K21-coated surgical sutures have antimicrobial activity for bacterial species of direct relevance to postoperative infection and bacteremia.
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Antibacterianos/farmacología , Materiales Biocompatibles Revestidos/farmacología , Enterococcus faecalis/efectos de los fármacos , Porphyromonas gingivalis/efectos de los fármacos , Compuestos de Amonio Cuaternario/farmacología , Suturas , Técnicas Bacteriológicas , Catgut , Relación Dosis-Respuesta a Droga , Etanol/química , Humanos , Ensayo de Materiales , Pruebas de Sensibilidad Microbiana , Nylons , Vehículos Farmacéuticos/química , Poliésteres , SedaRESUMEN
We previously demonstrated that HIV replication is concentrated in lymph node B cell follicles during chronic infection and that HIV-specific CTL fail to accumulate in large numbers at those sites. It is unknown whether these observations can be generalized to other secondary lymphoid tissues or whether virus compartmentalization occurs in the absence of CTL. We evaluated these questions in SIVmac239-infected rhesus macaques by quantifying SIV RNA(+) cells and SIV-specific CTL in situ in spleen, lymph nodes, and intestinal tissues obtained at several stages of infection. During chronic asymptomatic infection prior to simian AIDS, SIV-producing cells were more concentrated in follicular (F) compared with extrafollicular (EF) regions of secondary lymphoid tissues. At day 14 of infection, when CTL have minimal impact on virus replication, there was no compartmentalization of SIV-producing cells. Virus compartmentalization was diminished in animals with simian AIDS, which often have low-frequency CTL responses. SIV-specific CTL were consistently more concentrated within EF regions of lymph node and spleen in chronically infected animals regardless of epitope specificity. Frequencies of SIV-specific CTL within F and EF compartments predicted SIV RNA(+) cells within these compartments in a mixed model. Few SIV-specific CTL expressed the F homing molecule CXCR5 in the absence of the EF retention molecule CCR7, possibly accounting for the paucity of F CTL. These findings bolster the hypothesis that B cell follicles are immune privileged sites and suggest that strategies to augment CTL in B cell follicles could lead to improved viral control and possibly a functional cure for HIV infection.
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Ganglios Linfáticos/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Virus de la Inmunodeficiencia de los Simios/fisiología , Bazo/inmunología , Linfocitos T Citotóxicos/inmunología , Animales , Antígenos Virales/inmunología , Movimiento Celular , Células Cultivadas , Progresión de la Enfermedad , Macaca mulatta , ARN Viral/análisis , Receptores CCR7/metabolismo , Receptores CXCR5/metabolismo , Linfocitos T Citotóxicos/virología , Replicación ViralRESUMEN
The majority of risk factors for chronic inflammatory diseases are unknown. This makes personalized medicine for assessment, prognosis, and choice of therapy very difficult. It is becoming increasingly clear, however, that low-grade subclinical infections may be an underlying cause of many chronic inflammatory diseases and thus may contribute to secondary outcomes (e.g., cancer). Many diseases are now categorized as inflammatory-mediated diseases that stem from a dysregulation in host immunity. There is a growing need to study the links between low-grade infections, the immune responses they elicit, and how this impacts overall health. One such link explored in detail here is the extreme sensitivity of myeloid dendritic cells (mDCs) in peripheral blood to chronic low-grade infections and the role that these mDCs play in arbitrating the resulting immune responses. We find that emerging evidence supports a role for pathogen-induced mDCs in chronic inflammation leading to increased risk of secondary clinical disease. The mDCs that are elevated in the blood as a result of low-grade bacteremia often do not trigger a productive immune response, but can disseminate the pathogen throughout the host. This aberrant trafficking of mDCs can accelerate systemic inflammatory disease progression. Conversely, restoration of dendritic cell homeostasis may aid in pathogen elimination and minimize dissemination. Thus it would seem prudent when assessing chronic inflammatory disease risk to consider blood mDC numbers, and the microbial content (microbiome) and activation state of these mDCs. These may provide important clues ("the canary in the coal mine") of high inflammatory disease risk. This will facilitate development of novel immunotherapies to eliminate such smoldering infections in atherosclerosis, cancer, rheumatoid arthritis, and pre-eclampsia.
RESUMEN
Several intracellular pathogens, including a key etiological agent of chronic periodontitis, Porphyromonas gingivalis, infect blood myeloid dendritic cells (mDCs). This infection results in pathogen dissemination to distant inflammatory sites (i.e., pathogen trafficking). The alteration in chemokine-chemokine receptor expression that contributes to this pathogen trafficking function, particularly toward sites of neovascularization in humans, is unclear. To investigate this, we utilized human monocyte-derived DCs (MoDCs) and primary endothelial cells in vitro, combined with ex vivo-isolated blood mDCs and serum from chronic periodontitis subjects and healthy controls. Our results, using conditional fimbria mutants of P. gingivalis, show that P. gingivalis infection of MoDCs induces an angiogenic migratory profile. This profile is enhanced by expression of DC-SIGN on MoDCs and minor mfa-1 fimbriae on P. gingivalis and is evidenced by robust upregulation of CXCR4, but not secondary lymphoid organ (SLO)-homing CCR7. This disruption of SLO-homing capacity in response to respective chemokines closely matches surface expression of CXCR4 and CCR7 and is consistent with directed MoDC migration through an endothelial monolayer. Ex vivo-isolated mDCs from the blood of chronic periodontitis subjects, but not healthy controls, expressed a similar migratory profile; moreover, sera from chronic periodontitis subjects expressed elevated levels of CXCL12. Overall, we conclude that P. gingivalis actively "commandeers" DCs by reprogramming the chemokine receptor profile, thus disrupting SLO homing, while driving migration toward inflammatory vascular sites.
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Infecciones por Bacteroidaceae/metabolismo , Movimiento Celular/fisiología , Periodontitis Crónica/metabolismo , Células Dendríticas/microbiología , Células Mieloides/microbiología , Porphyromonas gingivalis/fisiología , Receptores de Quimiocina/metabolismo , Infecciones por Bacteroidaceae/inmunología , Infecciones por Bacteroidaceae/microbiología , Estudios de Casos y Controles , Moléculas de Adhesión Celular/metabolismo , Quimiocina CXCL12/metabolismo , Quimiotaxis/fisiología , Periodontitis Crónica/inmunología , Periodontitis Crónica/microbiología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Células Endoteliales/metabolismo , Fimbrias Bacterianas/genética , Fimbrias Bacterianas/fisiología , Humanos , Lectinas Tipo C/metabolismo , Lipopolisacáridos/farmacología , Células Mieloides/efectos de los fármacos , Células Mieloides/inmunología , Células Mieloides/metabolismo , Neovascularización Patológica/microbiología , Fenotipo , Receptores CCR7/metabolismo , Receptores CXCR4/metabolismo , Receptores de Superficie Celular/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Maintenance of blood DC homeostasis is essential to preventing autoimmunity while controlling chronic infection. However, the ability of bacteremic pathogens to directly regulate blood DC homeostasis has not been defined. One such bacteremic pathogen, Porphyromonas gingivalis, is shown by our group to survive within mDCs under aerobic conditions and therein, metastasize from its oral mucosal niche. This is accompanied by expansion of the blood mDC pool in vivo, independently of canonical DC poietins. We presently know little of how this bacteremic pathogen causes blood DC expansion and the pathophysiological significance. This work shows that optimum differentiation of MoDCs from primary human monocytes, with or without GM-CSF/IL-4, is dependent on infection with P. gingivalis strains expressing the DC-SIGN ligand mfa-1. DC differentiation is lost when DC-SIGN is blocked with its ligand HIV gp120 or knocked out by siRNA gene silencing. Thus, we have identified a novel, noncanonical pathway of DC differentiation. We term these PDDCs and show that PDDCs are bona fide DCs, based on phenotype and phagocytic activity when immature and the ability to up-regulate accessory molecules and stimulate allo-CD4(+) T cell proliferation when matured. The latter is dependent on the P. gingivalis strain used to initially "educate" PDDCs. Moreover, we show that P. gingivalis-infected, conventional MoDCs become resistant to apoptosis and inflammatory pyroptosis, as determined by levels of Annexin V and caspase-8, -3/7, and -1. Taken together, we provide new insights into how a relatively asymptomatic bacteremia may influence immune homeostasis and promote chronic inflammation.
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Bacteriemia/inmunología , Proteínas Bacterianas/inmunología , Moléculas de Adhesión Celular/inmunología , Células Dendríticas/patología , Proteínas Fimbrias/inmunología , Lectinas Tipo C/inmunología , Porphyromonas gingivalis/fisiología , Receptores de Superficie Celular/inmunología , Aerobiosis , Anexina A5/inmunología , Apoptosis , Linfocitos T CD4-Positivos/inmunología , Caspasas/fisiología , Moléculas de Adhesión Celular/antagonistas & inhibidores , Moléculas de Adhesión Celular/genética , Diferenciación Celular , Supervivencia Celular , Células Cultivadas/inmunología , Células Cultivadas/patología , Células Dendríticas/inmunología , Células Dendríticas/microbiología , Proteínas Fimbrias/deficiencia , Proteínas Fimbrias/genética , Fimbrias Bacterianas/inmunología , Proteína gp120 de Envoltorio del VIH/farmacología , Homeostasis , Interacciones Huésped-Patógeno/inmunología , Humanos , Lectinas Tipo C/antagonistas & inhibidores , Lectinas Tipo C/genética , Monocitos/citología , Mucosa Bucal/inmunología , Mucosa Bucal/microbiología , Fagocitosis , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/inmunología , Interferencia de ARN , ARN Interferente Pequeño/farmacología , Receptores de Superficie Celular/antagonistas & inhibidores , Receptores de Superficie Celular/genéticaRESUMEN
The low-grade oral infection chronic periodontitis (CP) has been implicated in coronary artery disease risk, but the mechanisms are unclear. In this study, a pathophysiological role for blood dendritic cells (DCs) in systemic dissemination of oral mucosal pathogens to atherosclerotic plaques was investigated in humans. The frequency and microbiome of CD19(-)BDCA-1(+)DC-SIGN(+) blood myeloid DCs (mDCs) were analyzed in CP subjects with or without existing acute coronary syndrome and in healthy controls. FACS analysis revealed a significant increase in blood mDCs in the following order: healthy controls < CP < acute coronary syndrome/CP. Analysis of the blood mDC microbiome by 16S rDNA sequencing showed Porphyromonas gingivalis and other species, including (cultivable) Burkholderia cepacia. The mDC carriage rate with P. gingivalis correlated with oral carriage rate and with serologic exposure to P. gingivalis in CP subjects. Intervention (local debridement) to elicit a bacteremia increased the mDC carriage rate and frequency in vivo. In vitro studies established that P. gingivalis enhanced by 28% the differentiation of monocytes into immature mDCs; moreover, mDCs secreted high levels of matrix metalloproteinase-9 and upregulated C1q, heat shock protein 60, heat shock protein 70, CCR2, and CXCL16 transcripts in response to P. gingivalis in a fimbriae-dependent manner. Moreover, the survival of the anaerobe P. gingivalis under aerobic conditions was enhanced when within mDCs. Immunofluorescence analysis of oral mucosa and atherosclerotic plaques demonstrate infiltration with mDCs, colocalized with P. gingivalis. Our results suggest a role for blood mDCs in harboring and disseminating pathogens from oral mucosa to atherosclerosis plaques, which may provide key signals for mDC differentiation and atherogenic conversion.