Dendritic cells, antibodies reactive with oxLDL, and inflammation.

Periodontitis appears to promote chronic inflammatory diseases, including atherosclerosis, but relevant mechanisms need clarification. Oral bacteria induce antibodies that bind not only bacteria, but also oxLDL. Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans induce remarkable IgG responses that are dominated by IgG2, and IgG2 is IFN-γ-dependent and is promoted by dendritic cells (DCs). LDL-reactive antibodies induced by P. gingivalis and A. actinomycetemcomitans include anti-phosphorylcholine (α-PC) and ß2-glycoprotein-1-dependent anticardiolipin (α-CL), and these antibodies may link chronic inflammatory diseases at a mechanistic level. Antibody-mediated uptake of oxLDL or bacteria dramatically enhances DC-IL-12, and DC-IL-12 induces NK-cell-IFN-γ responses that promote Th-1 responses and sustained inflammation. DCs may be derived from monocytes, and this is striking in cultures of aggressive periodontitis (AgP) monocytes, where DC numbers are about double control levels. Moreover, serum α-CL levels in individuals with AgP are frequently elevated, and these antibodies promote atherosclerosis in persons with antiphospholipid syndrome. Elevated serum levels of soluble-intercellular adhesion molecule, soluble-vascular cell adhesion molecule, and soluble-E-selectin are atherosclerosis-associated indicators of vascular inflammation, and these markers are elevated in the subset of AgP patients with high α-CL. We reason that periodontitis patients with elevated antibodies reactive with oxLDL could be a subgroup at high risk for cardiovascular sequelae.

Anti-phosphorylcholine-opsonized low-density lipoprotein promotes rapid production of proinflammatory cytokines by dendritic cells and natural killer cells.

BACKGROUND AND OBJECTIVE: Epidemiological and animal studies suggest that periodontal infections increase atherosclerosis risk. Periodontitis patients have elevated levels of anti-phosphorylcholine (anti-PC) reactive not only with numerous periodontal organisms but also with minimally modified low-density lipoprotein (mmLDL). Dendritic cells (DCs) reside in arterial walls and accumulate in atherosclerotic lesions. The ability of anti-PC to bind mmLDL prompted the hypothesis that opsonized mmLDL would stimulate DCs and enhance the production of proinflammatory cytokines that promote atherogenic plaque development. MATERIAL AND METHODS: Monocyte-derived DCs (mDCs) were generated using granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4, then stimulated with mmLDL or with anti-PC-opsonized mmLDL. The anti-PC effect was determined using flow cytometry, cofocal microscopy and cytokine assays. The production of CD83, IL-12p35 mRNA, IL-12p40 mRNA, IL-12p70 and IL-10 by DCs was monitored. RESULTS: Dendritic cells stimulated with mmLDL expressed little CD83 and produced little IL-12p70. However, anti-PC-opsonized mmLDL enhanced DC maturation, as indicated by upregulated CD83 and rapid (≤ 48 h) production of IL-12p70 if a source of interferon-γ (IFN-γ) was available. In leukocyte cultures, natural killer (NK) cells rapidly produced IFN-γ (≤ 48 h) when interacting with IL-12-producing DCs activated by anti-PC-opsonized mmLDL. Moreover, IFN-γ promoted DC IL-12 responses that were further augmented when mmLDL was opsonized with anti-PC. CONCLUSION: Minimally modified LDL-stimulated DCs and NK cells were mutually stimulatory, with DC IL-12p70 needed by NK cells and with NK cell IFN-γ needed by DCs. Moreover, production of these proinflammatory cytokines was markedly enhanced when LDL was opsonized by anti-PC. In short, the data suggest that the elevated anti-PC levels in periodontitis patients could promote a mechanism that facilitates atherosclerosis.

IL-17 in sera from patients with aggressive periodontitis.

Interleukin-17 (IL-17), the prototype cytokine produced by the Th17 subset of T-helper cells, plays a role in inflammatory responses, autoimmunity, and antimicrobial responses in a variety of infectious and inflammatory diseases. In view of the inflammatory nature and severity of aggressive periodontitis, we hypothesized that IL-17 might be detected in sera from patients with aggressive periodontitis. We used ELISA to measure IL-17 serum concentrations from 67 periodontally healthy (NP) individuals and from 53 patients with localized (LAgP) and 49 patients with generalized (GAgP) aggressive periodontitis. IL-17 was barely detectable in sera from periodontally healthy individuals (1.9 +/- 2.0 pg/mL), but was present at significantly higher concentrations in sera from those with LAgP (7.6 +/- 2.2 pg/mL) and GAgP (17.1 +/- 2.3 pg/mL). Multivariate analyses demonstrated associations of IL-17 concentrations with periodontal attachment loss, but not with current smoking. Therefore, Th17 responses may be characteristic of AgP, and IL-17 may play a role in the pathogenesis of aggressive periodontitis.

Atherogenic lipoprotein parameters in patients with aggressive periodontitis.

BACKGROUND AND OBJECTIVE: Certain types of chronic infection increase the plasma level of very-low-density lipoprotein, leading to formation of the particularly atherogenic low-density lipoprotein subclass, small dense low-density lipoprotein. In the present study, we examined whether aggressive forms of periodontitis are associated with these atherogenic lipoprotein parameters. MATERIAL AND METHODS: Twelve healthy control subjects without periodontitis, 12 subjects with localized aggressive periodontitis and 12 subjects with generalized aggressive periodontitis were studied. Lipoprotein subclass levels were determined using nuclear magnetic resonance methodology. RESULTS: Healthy control subjects, localized aggressive periodontitis subjects and generalized aggressive periodontitis subjects had progressively higher plasma levels of very-low-density lipoprotein and progressively smaller average low-density lipoprotein size (p < 0.05, one-way analysis of variance). In pairwise comparisons, differences were only significant between healthy controls and generalized aggressive periodontitis subjects (p < 0.05, Tukey's post test). After adjustment for body mass index, the mean periodontal pocket depth correlated positively with plasma very-low-density lipoprotein levels (p = 0.047). Very-low-density lipoprotein concentrations correlated positively with small dense low-density lipoprotein levels and negatively with average low-density lipoprotein size. Prevalence of the atherogenic lipoprotein pattern-B in healthy controls, localized aggressive periodontitis subjects and generalized aggressive periodontitis subjects was 8.3%, 33.3% and 66.6%, respectively. CONCLUSION: These results indicate that periodontal infection is associated with elevated plasma levels of atherogenic lipoprotein species. This association may account for the increased risk of periodontitis patients for cardiovascular disease.

Differential platelet-activating factor synthesis by monocytes and polymorphonuclear leukocytes from subjects with localized aggressive periodontitis.

BACKGROUND AND OBJECTIVE: Platelet-activating factor is elevated in localized aggressive periodontitis. We previously demonstrated that the elevated level of platelet-activating factor in localized aggressive periodontitis is at least partially attributable to low levels of platelet-activating factor acetylhydrolase, the enzyme that catabolizes platelet-activating factor. The objective of this study was to determine if platelet-activating factor synthesis was also elevated in localized aggressive periodontitis. To test this, platelet-activating factor synthesis was quantified in the monocytes and polymorphonuclear neutrophils of periodontally healthy patients and of subjects with localized aggressive periodontitis. MATERIAL AND METHODS: Cells were labeled with [(3)H]acetate and treated with vehicle or stimulated with calcium ionophore A23187. Platelet-activating factor was extracted and quantified by scintillation counting. RESULTS: For both subject groups, resting monocytes and polymorphonuclear neutrophils produced platelet-activating factor, and calcium ionophore A23187 stimulated platelet-activating factor production in both cell types. However, calcium ionophore A23187-activated monocytes from subjects with localized aggressive periodontitis produced less platelet-activating factor than did activated periodontally healthy monocytes (p < 0.0001), suggesting an aberrant calcium ionophore A23187 response in monocytes from subjects with localized aggressive periodontitis. Indeed, when the data were expressed as fold induction of platelet-activating factor synthesis in response to calcium ionophore A23187, monocytes from subjects with localized aggressive periodontitis exhibited only a fourfold increase in platelet-activating factor synthesis, whereas calcium ionophore A23187-stimulated monocytes from periodontally healthy, chronic periodontitis and generalized aggressive periodontitis subjects produced approximately 12 times more platelet-activating factor than did resting monocytes. In contrast, both resting and activated localized aggressive periodontitis polymorphonuclear neutrophils synthesized more platelet-activating factor than did periodontally healthy polymorphonuclear neutrophils. CONCLUSION: These data suggest that high levels of platelet-activating factor in subjects with localized aggressive periodontitis result from both increased synthesis and reduced catabolism. While localized aggressive periodontitis polymorphonuclear neutrophils contribute to increased platelet-activating factor mass through synthesis, the contribution of monocytes is probably the result of reduced catabolism by platelet-activating factor acetylhydrolase.

Follicular dendritic cells stimulated by collagen type I develop dendrites and networks in vitro.

Follicular dendritic cells (FDCs) reside in germinal centers in which their dendrites interdigitate and form non-mobile networks. FDC purification requires the use of collagenase and selection columns and leaves FDCs without detectable dendrites when examined by light microscopy. We have reasoned that isolated FDCs might reattach to a collagen matrix, extend their processes, and form immobile networks in vitro. As a test for this, cells were plated on collagen type I, laminin, biglycan, and hyaluronan. After 12 h, 80%-90% of FDCs adhered to all tested matrices but not to plastic. Within 2 weeks, FDCs adhering to type I collagen had spread out and had begun to acquire processes with occasional interconnections. By day 30, most FDCs had fine processes that formed networks through interdigitation with neighboring cells. FDC identity was confirmed by FDC-M1 labeling, immune complex trapping, and retention by FDCs in the networks. Scanning electron microscopy confirmed that groups of FDCs were in networks composed of convolutions and branching dendrites emanating from FDC cell bodies. In vivo, collagen type I was co-localized with FDCs, 5 h after challenge of immune mice with antigen. However, 2 days later, the collagen type I fibers were largely found at the periphery of the active follicles. Flow cytometry established the expression of CD29 and CD44 on FDCs; this may have partly mediated FDC-collagen interactions. Thus, we report, for the first time, that FDCs attach to collagen type I in vitro and regenerate their processes and networks with features in common with networks present in vivo.

Rapid tissue factor induction by oral streptococci and monocyte-IL-1beta.

The ability of pro-inflammatory cytokines to promote coagulation prompted the hypothesis that pro-inflammatory cytokines induced by oral streptococci might play a role in the pathogenesis of viridans endocarditis. We used supernatant fluids from peripheral blood mononuclear monocyte (PBMC) cultures, stimulated for just 4-6 hrs with representative streptococcal isolates, to study cytokines that promoted endothelial tissue factor (TF) activity. Neutralizing antibodies demonstrated that interleukin-1beta (IL-1beta) was a major early endothelial TF inducer, and that recombinant IL-1beta was comparable with the supernatant fluid in activity. IL-1beta-rich supernatant fluids from oral streptococci-stimulated or lipopolysaccharide-stimulated PBMC cultures up-regulated the expression of endothelial ICAM-1 and E-selectin. These molecules could help trap TF-producing monocytes or dendritic cells bearing streptococci at the site. Thus, the rapid IL-1beta-inducing capacity of oral streptococci could facilitate the early deposition of bacteria in fibrin clots and promote endocarditis.

Influence of proinflammatory cytokines on Actinobacillus actinomycetemcomitans specific IgG responses.

OBJECTIVE: High levels of serum anti-Actinobacillus actinomycetemcomitans immunoglobulin G (IgG) correlate with reduced extent and severity of periodontal disease and the present study was undertaken to begin testing the hypothesis that proinflammatory cytokines are important in the induction of optimal anti-A. actinomycetemcomitans IgG responses. BACKGROUND: Studies with pokeweed mitogen indicate that interleukin-1alpha (IL-1alpha) and IL-1beta are necessary for optimal IgG1 and IgG2 production and that prostaglandin E(2) (PGE(2)) and interferon-gamma (IFN-gamma) selectively promote IgG2, which is a major component of the anti-A. actinomycetemcomitans response in vivo. The pokeweed mitogen results suggest that these proinflammatory cytokines would also be necessary for optimal production of IgG specific for A. actinomycetemcomitans. METHODS: Peripheral blood mononuclear cells from A. actinomycetemcomitans-seropositive subjects with localized aggressive periodontitis were stimulated with A. actinomycetemcomitans in immune complexes capable of binding follicular dendritic cells that participate in the induction of recall responses in vivo. Cultures were manipulated with anti-IL-1alpha, anti-IL-1beta, anti-IFN-gamma, anti-IL-12, anti-CD21, indomethacin, and PGE(2). Actinobacillus actinomycetemcomitans specific IgG production was monitored by enzyme-linked immunosorbent assay (ELISA). RESULTS: Addition of follicular dendritic cells to peripheral blood mononuclear cells cultures resulted in follicular dendritic cell-lymphocyte clusters and increased anti-A. actinomycetemcomitans IgG responses (3-40-fold increases) compared with controls lacking follicular dendritic cells. Anti-IL-1alpha, anti-IL-1beta, anti-IFN-gamma, anti-IL-12, anti-CD21 and indomethacin suppressed anti-A. actinomycetemcomitans IgG production by half or more. PGE(2) restored IgG responses suppressed by indomethacin. CONCLUSIONS: The cytokines IL-1alpha, IL-1beta, IFN-gamma, IL-12, and PGE(2) were all necessary for optimal production of human anti-A. actinomycetemcomitans and the need for proinflammatory cytokines including the T helper 1 (Th1) cytokines is consistent with a response with a significant IgG2 component.

Dendritic-NK cell interactions in P. gingivalis-specific responses.

Patients with localized aggressive periodontitis have type-1 cytokines in gingival crevicular fluid and high titers of IFN-gamma-dependent IgG2 reactive with P. gingivalis in gingival crevicular fluid and serum. Localized aggressive periodontitis monocytes spontaneously differentiate into dendritic cells that can stimulate IFN-gamma production by NK cells. These relationships prompted the hypothesis that P. gingivalis-dendritic cell-NK cell interactions might promote type-1 cytokine responses. Although P. gingivalis is not a potent inducer of Th1 responses, it stimulated strong IL-12 responses by monocyte-derived dendritic cells in the presence of IFN-gamma, and IFN-gamma was produced by NK cells within 24 hrs in the presence of dendritic cells. Anti-P. gingivalis IgG2 responses were enhanced by dendritic cells, and removal of NK cells reduced IFN-gamma- and P. gingivalis-specific IgG2. Thus, P. gingivalis-dendritic cell-NK cell interactions apparently resulted in reciprocal stimulation and increased type-1 cytokine production by both dendritic cells and NK cells, and increased P. gingivalis-specific IgG2.

Dendritic cells stimulated with Actinobacillus actinomycetemcomitans elicit rapid gamma interferon responses by natural killer cells.

Human immunoglobulin G2 (IgG2) responses are gamma interferon (IFN-gamma) dependent, and monocyte-derived dendritic cells (mDCs) promote IgG2 production. DCs spontaneously emerge from monocytes in cultures prepared from localized aggressive periodontitis (LagP) patients, and these patients have high levels of IgG2 that is reactive with Actinobacillus actinomycetemcomitans. These results prompted the hypothesis that an interaction between mDCs and A. actinomycetemcomitans promotes IFN-gamma production, and IFN-gamma is known to promote both immunopathology and protective IgG2. A. actinomycetemcomitans induced mDCs to produce interleukin-12 (IL-12), and the addition of A. actinomycetemcomitans and DCs to cultured peripheral blood lymphocytes elicited high levels of IFN-gamma within just 24 h. In contrast, IL-4 was not detectable although DC-derived IL-10 production was apparent. A. actinomycetemcomitans-stimulated macrophages prepared from the same monocytes lacked the ability to induce IL-12 or IFN-gamma responses. NK cells of the innate immune system were the primary source of this early IFN-gamma, although CD8 T cells also contributed some. The NK cell-derived IFN-gamma was IL-12 dependent, and A. actinomycetemcomitans-DC interactions were Toll-like receptor 4 dependent. A. actinomycetemcomitans and A. actinomycetemcomitans lipopolysaccharide (LPS) were more potent than Escherichia coli and E. coli LPS in the ability to induce DC IL-12 and IFN-gamma. The ability of A. actinomycetemcomitans-stimulated DCs to induce NK cells to rapidly produce IFN-gamma in the absence of detectable IL-4 suggests their potential for skewing responses toward Th1. This may help explain the presence of Th1-associated cytokines in gingival crevicular fluid (GCF) from LagP patients and the high levels of IgG2 in their serum and GCF that is reactive with A. actinomycetemcomitans.

Anti-cardiolipin antibodies in sera from patients with periodontitis.

Antiphospholipid antibodies are commonly found in patients with systemic lupus erythematosus or the antiphospholipid syndrome, and a subset of such antibodies is associated with prothrombotic events such as stroke and with adverse pregnancy outcomes and fetal loss. We examined sera from 411 patients who were clinically characterized as to their periodontal disease status for serum levels of beta2-glycoprotein I-dependent anti-cardiolipin autoantibodies (anti-CL). The prevalence of patients with chronic periodontitis (CP) and generalized aggressive periodontitis (GAgP) positive for anti-CL (16.2% and 19.3%, respectively) was greater than that in healthy controls (NP) and localized aggressive periodontitis (LAgP) patients (6.8% and 3.2%). Patients with these autoantibodies demonstrated increased pocket depth and attachment loss compared with patients lacking the antibodies. Analysis of the data indicates that patients with generalized periodontitis have elevated levels of autoantibodies reactive with phospholipids. These antibodies could be involved in elevated risk for stroke, atherosclerosis, or pre-term birth in periodontitis patients.

Evidence of a substantial genetic basis for IgG2 levels in families with aggressive periodontitis.

IgG2 is elevated in localized but not in generalized aggressive periodontitis (AgP). Exposure to pathogenic bacteria is essential for disease. Immune responses are dominated by IgG2 reactive with bacterial surface carbohydrates. We used variance component analyses to assess IgG2 heritability and determine whether genes that influence IgG2 are the same genes that influence disease susceptibility. We studied 17 Caucasian and 43 African American families with two or more localized or generalized AgP-affected members (274 subjects with IgG2 measurements). Only 16% of the variance in IgG2 was attributable to age, race, and smoking. Even with the addition of localized AgP, the model still explained only 19% of IgG2 variance. By contrast, heritability of IgG2 levels was estimated to be 38% and highly significant (P = 0.0006), demonstrating a substantial genetic basis. Bi-trait variance component analyses of IgG2 and quantitative measures of AgP indicate that different genes appear to control IgG2 levels and disease susceptibility.

Non-redundant roles for interleukin-1 alpha and interleukin-1 beta in regulating human IgG2.

BACKGROUND: Serum concentrations of immunoglobulin G2 (IgG2) are elevated in localized aggressive periodontitis (LAgP) patients, and secretory products of monocytes from LAgP patients enhance IgG2 responses of lymphocytes from healthy subjects. Furthermore, genes regulating production of interleukin (IL)-1 influence the risk for both aggressive periodontitis (AgP) and chronic periodontitis. These observations, and the fact that IgG2 dominates responses to carbohydrates from Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis, prompted the hypothesis that IL-1 alpha, IL-1 beta, and IL-RA may help regulate human IgG2 responses. METHODS: Human peripheral blood leukocytes (PBL) were stimulated in culture with pokeweed mitogen (PWM); the levels of available IL-1 gene products were manipulated; and the effect on IgG2 production was monitored. Manipulations of IL-1 were accomplished by adding specific neutralizing monoclonal antibodies or recombinant IL-1RA, IL-1 alpha, or IL-1 beta. RESULTS: Blocking the IL-1 receptor with IL-1RA or neutralizing IL-1 alpha or IL-1 beta with specific antibody dramatically suppressed IgG2 production (50% to 70%). Additionally IL-1 alpha did not compensate for neutralized IL-1 beta, and additional IL-1 beta did not compensate for neutralized IL-1 alpha, suggesting the 2 monokines have separate roles in promoting IgG2. Furthermore, combinations of anti-IL-1 alpha and anti-IL-1 beta were more inhibitory than either antibody alone, and IL-1 alpha and IL-1 beta in combination appeared to work additively in promoting IgG2. Moreover, PBL cultures from a group of LAgP patients with high IgG2 levels had elevated levels of IL-1 beta. CONCLUSION: IL-1 alpha and IL-1 beta appear to have critical and non-redundant roles in the generation and regulation of potent IgG2 responses, which appear to be important in human responses to carbohydrate-bearing bacteria.

Phosphorylcholine-dependent cross-reactivity between dental plaque bacteria and oxidized low-density lipoproteins.

Antibodies reactive with phosphorylcholine (PC) are ubiquitous in human sera, but the antigens stimulating their production and their function are not clear. Previous studies have shown that a significant proportion of dental plaque bacteria contain PC as determined by reactivity with PC-specific mouse myeloma proteins and monoclonal antibodies. Additionally, serum antibody concentrations of immunoglobulin (IgG) G anti-PC are higher in sera of individuals who have experienced periodontal attachment loss than those who are periodontally healthy. These data implicate the oral microflora as a source of antigen-stimulating anti-PC responses. Recent data also indicate that antibodies with specificity for PC are elevated in ApoE-deficient mice, a model for studies of athersclerosis, and that such antibodies bound oxidized low-density lipoproteins (LDL) (oxLDL) in atherosclerotic plaques. These data prompted the hypothesis that human anti-PC could bind to both oral bacteria and human oxLDL, and that these antigens are cross-reactive. We therefore examined the ability of human anti-PC to bind to PC-bearing strains of oral bacteria using enzyme-linked immunosorbent inhibition assays and by assessment of direct binding of affinity-purified human anti-PC to PC-bearing Actinobacillus actinomycetemcomitans. Our results indicated that PC-bearing strains of Streptococcus oralis, Streptococcus sanguis, Haemophilus aphrophilus, Actinomyces naeslundii, Fusobacterium nucleatum, and A. actinomycetemcomitans, as well as a strain of Streptococcus pneumoniae, absorbed up to 80% of anti-PC IgG antibody from human sera. Furthermore, purified anti-PC bound to a PC-bearing strain of A. actinomycetemcomitans but only poorly to a PC-negative strain. OxLDL also absorbed anti-PC from human sera, and oxLDL but not LDL reacted with up to 80% of the anti-PC in human sera. Furthermore, purified anti-PC bound directly to oxLDL but not to LDL. The data indicate that PC-containing antigens on a variety of common oral bacteria are cross-reactive with neoantigens expressed in oxLDL. We propose that PC-bearing dental plaque microorganisms may induce an antibody response to PC that could influence the inflammatory response associated with atherosclerosis.

Follicular dendritic cells: beyond the necessity of T-cell help.

Follicular dendritic cells (FDCs) are potent accessory cells for B cells, but the molecular basis of their activity is not understood. Several important molecules involved in FDC-B-cell interactions are indicated by blocking the ligands and receptors on FDCs and/or B cells. The engagement of CD21 in the B-cell coreceptor complex by complement-derived CD21 ligand on FDCs delivers a crucial signal that dramatically augments the stimulation delivered by the binding of antigen to the B-cell receptor (BCR). The engagement of Fc gamma receptor IIB (FcgammaRIIB) by the Ig crystallizable fragment (Fc) in antigen-antibody complexes held on FDCs decreases the activation of immunoreceptor tyrosine-based inhibition motifs (ITIMs), mediated by the crosslinking of BCR and FcgammaRIIB. Thus, FDCs minimize a negative B-cell signal. In short, these ligand-receptor interactions help to signal to B cells and meet a requirement for B-cell stimulation that goes beyond the necessity of T-cell help.

Suppression of IgE responses in CD23-transgenic animals is due to expression of CD23 on nonlymphoid cells.

Serum IgE is suppressed in CD23-transgenic (Tg) mice where B cells and some T cells express high levels of CD23, suggesting that CD23 on B and T cells may cause this suppression. However, when Tg B lymphocytes were compared with controls in B cell proliferation and IgE synthesis assays, the two were indistinguishable. Similarly, studies of lymphokine production suggested that T cell function in the Tg animals was normal. However, adoptive transfer studies indicated that suppression was seen when normal lymphocytes were used to reconstitute Tg mice, whereas reconstitution of controls with Tg lymphocytes resulted in normal IgE responses, suggesting that critical CD23-bearing cells are irradiation-resistant, nonlymphoid cells. Follicular dendritic cells (FDC) are irradiation resistant, express surface CD23, and deliver iccosomal Ag to B cells, prompting us to reason that Tg FDC may be a critical cell. High levels of transgene expression were observed in germinal centers rich in FDC and B cells, and IgE production was inhibited when Tg FDCs were cultured with normal B cells. In short, suppressed IgE production in CD23-Tg mice appears to be associated with a population of radioresistant nonlymphoid cells. FDCs that interface with B cells in the germinal center are a candidate for explaining this CD23-mediated IgE suppression.

Ontogeny of the follicular dendritic cell phenotype and function in the postnatal murine spleen.

Follicular dendritic cells (FDCs) represent a unique cell population of antigen trapping cells restricted to follicles within the secondary lymphoid tissues. FDCs appear to be involved in the formation of primary follicles during the ontogeny of lymphoid tissue. We sought to determine the kinetics and tissue distribution of cells in the spleen of newborn mice expressing various differentiation antigens restricted to FDCs using immunohistochemistry with monoclonal antibodies (mAb) against FDCs and in vivo immune complex binding and retention. The earliest FDC-specific marker displayed was the antigenic determinant recognized by the FDC-M1 mAb, which was detectable by Day 3 prior to follicle formation on cells located around the peripheral part of the developing white pulp. The appearance of CD21/35 (complement receptor Type 2 and 1, CR1.2) was observed at the end of the first week, revealing a focal pattern in B-cell-rich areas. In addition, at that time there were some FDC-M1-positive cells in the nonfollicular part of the periarteriolar region. The administration of anti-horseradish peroxidase antibody followed by soluble antigen HRP into 7-day-old newborn mice resulted in the trapping and retention of immune complexes onto FDCs even in the absence of Fcgamma receptors. The appearance of another FDC-specific marker, FDC-M2, was observed during the second week after birth and was restricted on the cells located in the same area as CR1.2 cells. The Fcgamma receptor Type II appeared on FDCs after the second postnatal week. The above sequence of phenotypic maturation could also be observed in newborns after lethal irradiation at Day 3. This indicates that not only mature FDCs but also their precursors are highly radioresistant, and their phenotypic maturation follows a programmed path that requires only a small number of mature B cells.

Persistence of infectious HIV on follicular dendritic cells.

Follicular dendritic cells (FDCs) trap Ags and retain them in their native state for many months. Shortly after infection, HIV particles are trapped on FDCs and can be observed until the follicular network is destroyed. We sought to determine whether FDCs could maintain trapped virus in an infectious state for long periods of time. Because virus replication would replenish the HIV reservoir and thus falsely prolong recovery of infectious virus, we used a nonpermissive murine model to examine maintenance of HIV infectivity in vivo. We also examined human FDCs in vitro to determine whether they could maintain HIV infectivity. FDC-trapped virus remained infectious in vivo at all time points examined over a 9-mo period. Remarkably, as few as 100 FDCs were sufficient to transmit infection throughout the 9-mo period. Human FDCs maintained HIV infectivity for at least 25 days in vitro, whereas virus without FDCs lost infectivity after only a few days. These data indicate that HIV retained on FDCs can be long lived even in the absence of viral replication and suggest that FDCs stabilize and protect HIV, thus providing a long-term reservoir of infectious virus. These trapped stores of HIV may be replenished with replicating virus that persists even under highly active antiretroviral therapy and would likely be capable of causing infection on cessation of drug therapy.

Cytokine induction by Streptococcus mutans and pulpal pathogenesis.

Chronic pulpal inflammation under caries appears to be elicited by bacterial antigens that diffuse into the pulp through dentinal tubules. This prompted the hypothesis that cytokines elicited by antigens from Streptococcus mutans, which frequently dominates shallow lesions, could play a major role in eliciting the initial T-cell response in the pulp. To test this, we examined the ability of S. mutans to stimulate T cells and elicit cytokines and used Lactobacillus casei, which often predominates in deep carious lesions where B cells and plasma cells predominate, as a control. In addition, the presence of cytokines in the pulp was analyzed at the mRNA level. S. mutans elicited potent gamma interferon (IFN-gamma) responses in peripheral blood mononuclear cell cultures and reduced the CD4/CD8 ratio by promoting CD8(+) T cells. Multiple inflammatory cytokine mRNAs (IFN-gamma, interleukin 4 [IL-4], and IL-10) were detected in human dental pulp. A higher prevalence of IFN-gamma (67%) than IL-4 (19%) or IL-10 (29%) was obtained in shallow caries, suggesting a type 1 cytokine mechanism in early pulpitis where S. mutans predominates. In contrast, in deep caries no differences in cytokine frequency were observed. Furthermore, the presence of IFN-gamma in the pulp correlated with the presence of S. mutans. The extraordinary induction of type 1 cytokines and the preferential activation of CD8(+) T cells by S. mutans offers an explanation for the etiology of the CD8(+) T-cell-dominant lesion in early pulpitis and suggests that S. mutans may have a major impact on the initial lesion and pulpal pathology.

Invasion of human vascular endothelial cells by Actinobacillus actinomycetemcomitans via the receptor for platelet-activating factor.

Strains of the periodontal pathogen Actinobacillus actinomycetemcomitans are variable with respect to display of phosphorylcholine (PC)-bearing antigens. We have examined strains of A. actinomycetemcomitans with and without PC to assess their ability to invade endothelial cells via the receptor for platelet-activating factor (PAF). Results of antibiotic protection assays indicate that PC-bearing A. actinomycetemcomitans invade human vascular endothelial cells by a mechanism inhibitable by CV3988, a PAF receptor antagonist, and by PAF itself. The invasive phenotype was verified by transmission electron microscopy. A PC-deficient strain of this organism was not invasive. This property, in addition to the established ability of A. actinomycetemcomitans to invade epithelial cells, may provide this organism with access to the systemic circulation. The ability of PC-bearing oral bacteria to access the circulation may also explain the elevated levels of anti-PC antibody in serum found in patients with periodontitis.