The Porphyromonas gingivalis collagenase-specific serum immunoglobulin A (IgA), IgM, and IgG responses from 20 patients with early-onset periodontitis (EOP), 20 patients with adult periodontitis, (AP), and 20 age- and sex-matched healthy controls were examined by immunoblot analysis. A recombinant collagenase antigen used for the immunoblot analysis was produced by using the plasmid pGEX-2T, which allows the fusion between the collagenase and glutathione S-transferase. There was no significant difference in collagenase-specific IgG antibody detection between samples from the EOP, AP, and control groups. In contrast, 85% of AP and EOP sera had collagenase-specific IgA antibodies, whereas only 20% of control sera showed collagenase-specific IgA reactivity. Plaque samples from all groups were assessed by PCR with primers complementary to the collagenase-encoding gene prtC. The results indicated that 90% of AP and EOP plaque samples and 10% of control samples were positive for P. gingivalis. All patients with collagenase-specific IgA antibodies were PCR positive. The results of the study indicate a nearly complete concordance (k = 0.856) between the presence of collagenase-specific IgA antibodies and PCR detection of P. gingivalis. By using PCR as the "gold standard," the sensitivity and specificity of the IgA immunoblot test were 94.7 and 90.9%, respectively. Therefore, the recombinant collagenase is a potential candidate for use in the serodiagnosis of periodontitis.
The objective of this study was to determine if a relationship exists between antibody reactive with the Actinobacillus actinomycetemcomitans serotype b lipopolysaccharide (LPS) and severity of periodontal disease in generalized early-onset periodontitis (G-EOP). The concentration of antibody reactive with A. actinomycetemcomitans serotype b LPS was determined for 102 G-EOP subjects. Analysis of the relationship between antibody reactive with A. actinomycetemcomitans serotype b LPS and measures of periodontal attachment loss indicated that the patients with the highest concentrations of antibody reactive with A. actinomycetemcomitans serotype b LPS had significantly less attachment loss. These high-responder subjects also had anti-A. actinomycetemcomitans serotype b LPS with significantly higher relative avidity. The results suggest that antibody reactive with A. actinomycetemcomitans serotype b LPS is protective in G-EOP patients.
The present study was performed to estimate the observed frequencies of the immunoglobulin heavy-chain (Gm) and light-chain (Km) allotypes among patients with early-onset periodontitis (EOP) and their effect on the IgG2 subclass responses against Actinobacillus actinomycetemcomitans Y4 and Porphyromonas gingivalis 381, respectively. Sixty-nine EOP patients, including 11 with localized juvenile periodontitis (LJP), 19 who had LJP, 15 with LJP-rapidly progressing periodontitis (RPP), and 24 with RPP, were examined for the Gm and Km allotypes by a hemagglutination inhibition test. Levels of immunoglobulin G2 (IgG2) antibodies against the two organisms were determined by enzyme-linked immunosorbent assay. Fifty race- and age-matched, periodontally healthy subjects were also included as a control group. The observed frequencies of the Gm haplotype afnb and Km(1) were significantly higher in the RPP and LJP groups, respectively. The G2m(n)+ group of those with RPP and the Km(1)+ group of those with LJP had significantly higher levels of IgG2 antibodies to A. actinomycetemcomitans and P. gingivalis, respectively. The results indicate that linkage disequilibrium of the G2m(n) locus in RPP patients or the Km(1) locus in LJP patients may be associated with high IgG2 antibody responses to the respective bacteria. It was reasoned that the IgG2 antibody responses are associated with the immunoglobulin allotypes. The function of IgG2 antibodies in their reaction to different bacterial antigens may be interpreted as either protective or nonprotective in the two different types of EOP (i.e., LJP and RPP).
The serotype b-specific carbohydrate antigen (SbAg) of Actinobacillus actinomycetemcomitans Y4 is reported to be the O antigen of lipopolysaccharide, and the highest titers of serum antibody reactive with A. actinomycetemcomitans in early-onset periodontitis (EOP) patients bind SbAg. These high titers of serum antibody reactive with SbAg are associated with a lesser extent and severity of periodontal disease. The aim of this study was to determine if a limited number of genes code for anti-SbAg antibodies as has been shown for immunoglobulin G (IgG) reactive with the type b polysaccharide from Haemophilus influenzae. Serum IgG reactive with the SbAg was prepared from 20 high-titer EOP patients by affinity chromatography. The IgG subclass concentrations were determined, and heterogeneity was analyzed by isoelectric focusing (IEF). IgG2 was the dominant subclass (83% of total IgG) in the anti-SbAg IgG fraction and represented an average of 1.33% of total serum IgG2. The IgG2 reactive with SbAg was isolated from the affinity-purified IgG fraction by affinity chromatography with protein A and subclass-specific monoclonal antibodies. On IEF gels, only 4 to 20 bands were observed in the anti-SbAg IgG fractions, indicating limited heterogeneity. N-terminal amino acid sequence analysis of eight representative anti-SbAg IgG2 preparations indicated that variable heavy and light chains consisted largely of V(H)III and V(kappa)II, respectively. However, a significant fraction of anti-SbAg may use V(H) and V(lambda) genes with blocked N termini. In short, these findings indicate that IgG reactive with SbAg is very much like the antibody reactive with H. influenzae type b polysaccharide. Similarities include IgG2 dominance, limited bands on IEF gels, supporting an oligoclonal response, and use of genes from V(H)III and V(kappa)II regions.
Recent data indicate that smoking is an important risk factor for the development of periodontitis. Smoking is also known to reduce serum immunoglobulin G (IgG) levels. Interestingly, patients with the localized form of early-onset periodontitis (LJP) have elevated levels of serum IgG2, and those who smoke are not clinically different from nonsmoking LJ subjects. In contrast, patients with the generalized form of early-onset periodontitis (G-EOP) who smoke have more extensive destruction than their nonsmoking counterparts. Given the effects of smoking on EOP and the association of IgG2 with less severe disease, we hypothesized that smoking might reduce serum IgG2 and that this might be most apparent in G-EOP. We therefore examined the effects of smoking on serum IgG subclass concentrations in race-matched groups: LJP, G-EOP, and age-matched periodontally healthy controls (NPs). Smoking status was established from serum cotinine levels, and serum IgG subclass concentrations were determined by using radial immunodiffusion. The data indicated that the effects of smoking were remarkably selective with respect to both IgG subclass and race. Smoking did not appear to have any effect on the concentration of IgG1 or IgG3 in either black or white subjects. In contrast, smoking was associated with depressed serum IgG2 concentrations in both white NP and G-EOP subgroups. Serum IgG2 levels in black subjects did not appear to be depressed by smoking, with the single striking exception of the black G-EOP subgroup which also had depressed serum IgG4 levels. The results here confirm that smoking has effects on serum immunoglobulin levels, but the effects were both race and serum IgG subclass specific. Furthermore, the periodontal diagnosis of EOP subjects appeared to be important, as indicated by the fact that IgG2 and IgG4 levels were reduced in smoking black G-EOP subjects whereas the IgG2 and IgG4 levels in black LJP and NP subjects were not reduced by smoking.
The purpose of the present study was to determine the serum immunoglobulin A (IgA), IgM, and IgG reactivities against proteins of Actinobacillus actinomycetemcomitans in patients with periodontitis. Serum samples from 20 patients with early-onset periodontitis, 20 patients with adult periodontitis, and 20 age- and sex-matched healthy controls were assessed by immunoblot analysis. IgG antibody reactivity against a sarcosyl-insoluble 110-kDa protein of A. actinomycetemcomitans was detected in 65 and 45% of patients with early-onset periodontitis and adult peritonitis, respectively, and IgA antibodies against this protein were found in 70 and 55% of these patients, respectively. However, control subjects showed no IgG reactivity, and IgA antibodies against the sarcosyl-insoluble 110-kDa protein were detected in only 5% of the patients (P < 0.05). There was no IgM antibody reactivity against this protein in any of the diseased or healthy subjects. The sensitivity and specificity of serum IgA antibody reactivity against the 110-kDa protein in detecting subgingival A. actinomycetemcomitans infection, as determined by PCR, were 77 and 66%, respectively. The results of the study indicated that the sarcosyl-insoluble 110-kDa protein is a potential candidate for use in the serodiagnosis of periodontal disease.
Early-onset periodontitis (EOP) is characterized by rapidly progressive alveolar bone loss, chemotactic defects of neutrophils, and significant familial aggregation. We found immature myeloid lineage cells, defined as promyelocytes, in the peripheral blood in patients with EOP. A hematological examination of peripheral blood cells showed normal reference values regarding cell proportions. Flow cytometry revealed significantly lower expression of CD16, a glycosylphosphatidylinositol (GPI)-anchored protein, on peripheral neutrophils in patients compared with those in age- and sex-matched healthy controls, whereas the levels of CD11a and CD11b expression were similar. The chemotactic response of neutrophils was lower toward not only formyl-methionyl-leucyl-phenylalanine but also complement fragment C5a than that of healthy controls. The expression of another GPI-anchored protein, CD14, was equally expressed by controls and patients. Therefore, the low level of CD16 expression was not due to the incomplete synthesis of the GPI anchor. GPI anchors of CD16 on neutrophils from controls and patients were both partially resistant to phosphatidylinositol-specific phospholipase C. The presence of promyelocytes in peripheral blood, low expression of CD16, and low chemotactic response of neutrophils suggest that patients with EOP have an abnormal maturation system in myeloid lineage cells in the bone marrow, which may be associated with the onset and course of EOP.
OBJECTIVES—To study interaction of early onset pauciarticular juvenile chronic arthritis (EOP-JCA) and pregnancy in the Polish population, in particular to confirm the ameliorating effect of pregnancy on disease activity reported by others and to analyse the factors that govern the occurrence of postpartum flare, with emphasis on the potential role of breast feeding.
METHODS—The reproductive outcome and disease status in 39 adult women with history of EOP- JCA was examined by means of a questionnaire and an interview. In all patients the disease onset occurred before the 6th birthday, 19 had persistent pauciarticular JCA (PeEOP-JCA) and 20 had extended pauciarticular JCA (ExEOP-JCA).
RESULTS—23 women had at least one successful pregnancy, seven had unsuccessful pregnancies but all of them had also one or more successful pregnancies. Among those who have never been pregnant (n=16) there was a higher frequency of eye disease and ExEOP-JCA compared with the rest of the group. In almost all cases pregnancy was associated with remission of disease activity, however a postpartum flare appeared after 22 pregnancies (52%). The flares were more frequent in women who had an active disease before pregnancy, had a flare after a previous pregnancy and/or were breast feeding.
CONCLUSIONS—In EOP-JCA patients pregnancy generally has a good outcome and induces amelioration of disease activity. After delivery, however, a flare of disease often appears, especially in women who were breast feeding, had a postparum flare previously or had an active disease before pregnancy. The pattern of interaction between disease and pregnancy found in EOP-JCA makes EOP-JCA similar in this respect to RA, but different from systemic lupus erythematosus and ankylosing spondylitis.
Over-replication of periodontal pathogens in the periodontium induces production of proinflammatory cytokines and C-reactive protein that can stimulate systemic inflammatory status and can initiate atherosclerosis and its consequences. In our pilot study we examined whether periodontal status and serum levels of interleukin-6 and C-reactive protein are associated with the presence of Aggregatibacter actinomycetemcomitans in the periodontium of patients with cardiovascular diseases (CVD).
We randomly selected 38 of 166 outpatients with CVD, of which 21 patients had chronic ischemic heart disease (IHD) only and 17 had both IHD and essential hypertension (HT). The presence of Aggregatibacter actinomycetemcomitans (A.a.) in the periodontium evaluated by PCR was compared with the values of periodontal indices, namely probe depth (PD) and Community Periodontal Index of Treatment Need (CPITN), as well as with interleukin-6 (IL-6) and CRP serum levels.
When comparing A.a.-positive and A.a.-negative groups of patients, no statistically significant differences were noticed as to the age and values of PD and CPITN, respectively. However, the proportion of CRP and IL-6 positive values was significantly higher (p≤0.001) among A.a.-positive than in A.a.-negative patients.
The presence of A.a. in patients with CVD may be associated with significantly higher serum levels of some proinflammatory markers.
cytokines; CRP; periodontal indexes; periodontal diseases
The aims of this study were to determine the immunodominant antigens of Actinobacillus actinomycetemcomitans serotype b (Aab) for the different immunoglobulin (Ig) classes and subclasses and to determine the relative levels of these different Igs in serum. Seropositive early-onset periodontitis patients were sampled, and the Ig classes IgG, IgA, and IgM and subclasses IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2 were studied. Reactivity with Aab antigens was assessed by using the Western blot (immunoblot) in limiting dilution analysis and radioimmunoassay with sera from 13 early-onset periodontitis subjects. A smeared antigen in the upper portion of the immunoblots, typical of high-molecular-weight LPS, was immunodominant for IgG, IgA, IgM, IgG1, IgG2, IgG3, IgA1, and IgA2. This smeared antigen was present in every patient for all of these Igs at the endpoint. A few additional antigens were also present at the endpoint in some patients, but none were present in more than half of the subjects. The distribution of antibody titers by Ig classes reactive with the Aab immunodominant antigen was IgG > IgA > IgM. The distribution of antibody titers by IgG subclass was IgG2 > IgG1 approximately IgG3. Further quantitation by radioimmunoassay revealed that the mean concentration of IgG2 (65.7 micrograms/ml) was significantly greater than that of IgG1 (8.8 micrograms/ml). The IgA subclass distribution was IgA1 >> IgA2, with IgA1 apparently being second only to IgG2. Therefore, the Aab antigen eliciting the highest antibody level in virtually all Ig classes and subclasses appeared to be lipopolysaccharide, and IgG2 was markedly elevated over all other serum Ig classes or subclasses reactive with Aab.
Human immunoglobulin G2 (IgG2) serum concentrations and the IgG2 antibody response to Actinobacillus actinomycetemcomitans can be influenced by genes, by environmental factors such as smoking, and by periodontal disease status. Examination of the IgG2 response to phosphorylcholine (PC), a response thought to be mainly induced by the C polysaccharide of Streptococcus pneumoniae, suggested that periodontal disease status was also associated with this response. This prompted the hypothesis that PC is an important oral antigen associated with organisms in the periodontal flora and that anti-PC antibody is elevated as a consequence of periodontal disease. Subjects in various periodontal disease diagnostic categories in which attachment loss is exhibited were tested for anti-PC in serum. Those with adult periodontitis, localized juvenile periodontitis, generalized early-onset periodontitis, and gingival recession all had similar levels of anti-PC IgG2 serum antibody which were significantly greater than in the group of subjects with no attachment loss. Analysis of plaque samples from subgingival and supragingival sites in all diseases categories for reactivity with the anti-PC specific monoclonal antibody TEPC-15 revealed that a substantial proportion of the bacteria in dental plaque (30 to 40%) bear PC antigen; this antigen was not restricted to morphotypes resembling only cocci but was also present on rods and branched filamentous organisms. We found that S. mitis, S. oralis, and S. sanguis, as well as oral actinomycetes, including A. viscosus, A. odontolyticus, and A. israelii, incorporated substantial amounts of [3H]choline from culture media. Further analysis of antigens derived from these organisms by Western blot indicated that S. oralis, S. sanguis, A. viscosus, A. odontolyticus, and A. israelii contained TEPC-15-reactive antigens. The data show that many commonly occurring bacterial species found in dental plaque contain PC antigen and that immunization with plaque-derived PC antigens as a consequence of inflammation and periodontal attachment loss may influence systemic anti-PC antibody concentrations.
A powerful, cost-effective new method for studying single-nucleotide polymorphisms (SNPs) is described. This method is based on the use of hairpin-shaped primers (HP), which give a sensitive and specific PCR amplification of each specific allele, without the use of costly fluorophore-labeled probes and any post-PCR manipulation. The amplification is monitored in real-time using SYBR Green I dye and takes only 2 h to yield results. The HP assay has a simple design and utilizes a conventional real-time PCR apparatus. The −44 C→G transversion in the DEFB1 gene (which encodes human β-defensin 1) has been previously associated with Candida carriage in oral epithelia. In this study, we analyzed the association between early-onset periodontal disease (EOP) and the −44 SNP. We used an HP assay to study the distribution of the −44 SNP in 264 human DNAs obtained from two cohorts of EOP patients and healthy controls from different ethnic backgrounds. The results indicate that the −44 SNP has a similar distribution between EOP and healthy patients, suggesting that it is not associated with the disease.
OBJECTIVE—Although the influence of age on clinical and laboratory features has been widely demonstrated in many arthropathies, studies on elderly onset (> 60 years) psoriatic arthritis (EOPsA) are rare. This study compares manifestations at onset and two year outcome of EOPsA with those of younger onset PsA (YOPsA).
PATIENTS AND METHODS—Sixty six consecutive PsA patients with disease duration < 1 year, 16 EOPsA (>60 years) and 50 YOPsA (⩽60 years) were admitted to a prospective study. Clinical, laboratory, and radiographic assessment were carried out at admission and after two years. HLA class I and bone scintigraphy were also recorded. In 10 patients with EOPsA and 24 with YOPsA it was possible to obtain synovial fluid, which was subsequently analysed for local inflammatory indices, including interleukin (IL) 1β, IL6, and IL8.
RESULTS—Presenting manifestations of EOPsA differed from YOPsA in number of active joints (mean (SD)) (12.2 (6.3) v 6.7 (4.6), p<0.001), foot bone erosions (2.7 (1.2) v 1.1 (1.1), p<0.001), erythrocyte sedimentation rate (64.2 (35.3) v 30.5 (30.0) mm 1st h, p<0.001), C reactive protein (3.9 (2.0) v 1.3 (1.3) mg/dl, p<0.001) and synovial fluid IL1β (8.0 (4.7) v 3.0 (3.0) pg/ml, p<0.001) and IL6 (828.2 (492.6) v 469.3 (201.4) pg/ml, p<0.005). No differences were found in the number of subjects with dactylitis, pitting oedema, HLA-B27, or signs of sacroiliac and sternoclavicular joint involvement at bone scintigraphy. After two years, progression was more evident in EOPsA than in YOPsA, as the number of new erosions in the hands and also the C reactive protein were higher in EOPsA patients.
CONCLUSION—PsA has a more severe onset and a more destructive outcome in elderly people (onset >60 years) than in younger subjects. This behaviour may be influenced by immune changes associated with aging, as suggested by the higher concentrations of IL1β and IL6 found in the synovial fluid of EOPsA than in YOPsA.
Keywords: elderly onset arthritis; psoriatic arthritis; synovial fluid; interleukins
Actinobacillus actinomycetemcomitans is a facultative gram-negative bacterium which has been associated with severe oral and nonoral infections. This study examined its occurrence in the oral cavities of 10 normal juveniles, 11 normal adults, 10 juvenile periodontitis patients, and 12 adult periodontitis patients. Four deep periodontal pockets and two normal periodontal sites were sampled in the diseased patients, and six normal periodontal sites were sampled in the healthy individuals. In all subjects samples were obtained from the cheek, tongue, and saliva. Samples from a total of 172 normal periodontal sites, 83 deep periodontal pockets, 42 cheek mucosae, 42 tongue dorsa, and 42 salivas were examined. Isolation was performed by using a medium for selective isolation of A. actinomycetemcomitans (Trypticase soy agar [BBL Microbiology Systems] supplemented with 10% serum and 75 μg of bacitracin per ml). The carrier rates were 20% for normal juveniles, 36% for normal adults, 50% for adult periodontitis patients, and 90% for juvenile periodontitis patients. A. actinomycetemcomitans was on average recovered in about fivefold-higher numbers from infected deep periodontal pockets than from infected normal subgingival areas. Samples of periodontal pockets generally contained 100-fold-more cells of A. actinomycetemcomitans than did samples of the cheek, tongue, and saliva. A. actinomycetemcomitans is commonly isolated from patients with juvenile periodontitis, often isolated from patients with adult periodontitis, and occasionally isolated from normal juveniles and adults. Its primary oral ecological niche appears to be dental plaque and periodontal pockets.
Mutations in the PTEN‐induced kinase 1 (PINK1) gene have been identified in recessively inherited and sporadic early‐onset parkinsonism (EOP).
A total of 131 Norwegian patients diagnosed with Parkinson's disease were included. Of them, 89 participants had EOP (onset ⩽50 years); the remaining had familial late‐onset disease (mean age at onset 64 years). PINK1 analysis included sequencing and gene dose assessment. Mutations were examined in 350 controls .
Heterozygous missense mutations in PINK1 were found in 3 of 131 patients; none of the patients carried homozygous or compound heterozygous mutations. One of these three patients had a father affected by Parkinson's disease, and he carried the mutation. Three new and seven known polymorphic variants were identified, although none seemed to be associated with disease risk.
PINK1 mutations are rare in Norwegian patients with EOP and familial Parkinson's disease. However, the data suggest that some heterozygous mutations might increase the risk of developing Parkinson's disease.
Actinobacillus actinomycetemcomitans is the etiologic agent of localized aggressive periodontitis, a rapidly progressing oral disease that occurs in adolescents. A. actinomycetemcomitans can also cause systemic disease, including infective endocarditis. In early work on A. actinomycetemcomitans workers concluded that this bacterium is not beta-hemolytic. More recent reports have suggested that A. actinomycetemcomitans does have the potential to be beta-hemolytic. While growing A. actinomycetemcomitans on several types of growth media, we noticed a beta-hemolytic reaction on media from one manufacturer. Beta-hemolysis occurred on Columbia agar from Accumedia with either sheep or horse blood, but not on similar media from other manufacturers. A surprising result was that mutants of A. actinomycetemcomitans defective for production of leukotoxin, a toxin that is reportedly highly specific for only human and primate white blood cells, are not beta-hemolytic. Purified leukotoxin was able to lyse sheep and human erythrocytes in vitro. This work showed that in contrast to the accepted view, A. actinomycetemcomitans leukotoxin can indeed destroy erythrocytes and that the production of this toxin results in beta-hemolytic colonies on solid medium. In light of these results, the diagnostic criteria for clinical identification of A. actinomycetemcomitans and potentially related bacteria should be reevaluated. Furthermore, in studies on A. actinomycetemcomitans leukotoxin workers should now consider this toxin's ability to destroy red blood cells.
Previously published work has shown that sera from healthy sickle cell disease (SCD) patients inhibits normal lymphocyte response to phytohemagglutinin (PHA) in vitro. The objective of the current study is to ascertain what the combined effects of SCD sera plus penicillin have on normal lymphocyte cytokine production and mitogenic response to PHA. Steady state sera from 20 SCD patients not on penicillin prophylaxis and 20 comparable healthy controls were used in all experiments. Four normal healthy individuals were used as donors for obtaining peripheral blood mononuclear cells (PBMC), by density gradient. PBMC with or without penicillin were PHA stimulated by standard in vitro culture for mitogenic response and cytokine production. Supernatant cytokine levels for interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha and interleukin (IL)2 were quantified by ELISA technique. Results revealed suppression of mitogenic response in the SCD group with or without penicillin, compared to control sera (P < .001). Cytokine production in the SCD sera group showed increased production of IFN-gamma and TNF-alpha in the absence of penicillin, but suppression at all doses of penicillin. The control group results were as follows: no significant difference in IFN-gamma production with or without penicillin, mean TNF-alpha levels were the opposite of SCD sera with lower levels in the absence of penicillin. IL-2 production demonstrated a similar pattern for both groups of sera. IL-2 production was low without penicillin, but there was increased production with penicillin, which appeared dose related. The data suggests that sera from healthy SCD patients and in vitro added penicillin may have a combined suppressive effect on normal lymphocyte in vitro production of IFN-gamma and TNF-alpha. The current study results suggest that penicillin has the beneficial effect of decreasing TNF-alpha production and increasing IL-2 production when combined with SCD steady state sera. However, this in vitro benefit must be weighed against suppression of IFN-gamma production and ultimately, perhaps the long-term utility of penicillin prophylaxis in patients with SCD.
Periodontitis, a prime cause of tooth loss in humans, is implicated in the increased risk of systemic diseases such as heart failure, stroke, and bacterial pneumonia. The mechanisms by which periodontitis and antibacterial immunity lead to alveolar bone and tooth loss are poorly understood. To study the human immune response to specific periodontal infections, we transplanted human peripheral blood lymphocytes (HuPBLs) from periodontitis patients into NOD/SCID mice. Oral challenge of HuPBL-NOD/SCID mice with Actinobacillus actinomycetemcomitans, a well-known Gram-negative anaerobic microorganism that causes human periodontitis, activates human CD4+ T cells in the periodontium and triggers local alveolar bone destruction. Human CD4+ T cells, but not CD8+ T cells or B cells, are identified as essential mediators of alveolar bone destruction. Stimulation of CD4+ T cells by A. actinomycetemcomitans induces production of osteoprotegerin ligand (OPG-L), a key modulator of osteoclastogenesis and osteoclast activation. In vivo inhibition of OPG-L function with the decoy receptor OPG diminishes alveolar bone destruction and reduces the number of periodontal osteoclasts after microbial challenge. These data imply that the molecular explanation for alveolar bone destruction observed in periodontal infections is mediated by microorganism-triggered induction of OPG-L expression on CD4+ T cells and the consequent activation of osteoclasts. Inhibition of OPG-L may thus have therapeutic value to prevent alveolar bone and/or tooth loss in human periodontitis.
This article may have been published online in advance of the print edition. The date of publication is available from the JCI website, http://www.jci.org. J. Clin. Invest. 106:R59–R67 (2000).
The aim of this study was to determine whether the increased serum cell-free fetal DNA (cffDNA) level of gravidas developed into early-onset preeclampsia (EOPE) subsequently in the early second trimesters is related to prenatal screening markers. Serum was collected from 1011 gravidas. The level of cffDNA and prenatal screening markers were analyzed in 20 cases with EOPE and 20 controls. All fetuses were male. The maternal serum cffDNA level was assessed by amplification of the Y chromosome specific gene. Correlations between the variables were examined. (Logged) cffDNA in EOPE (median, 3.08; interquartile range, 2.93–3.68) was higher than controls (median, 1.79; interquartile range, 1.46–2.53). The increased level of (logged) cffDNA was correlated significantly with the increased human chorionic gonadotropin (HCG) level (r = 0.628, p < 0.001). Significant reciprocal correlations between cffDNA and babies’ birth weight as well as gestation weeks at delivery were noted (r = −0.516, p = 0.001; r = −0.623, p < 0.001, respectively). The sensitivity and specificity of cffDNA to discriminate between the EOPE cases and the controls were 90% and 85%, respectively. CffDNA is a potential marker for EOPE, which had a significant reciprocal correlation with babies’ birth weight and gestation weeks at delivery. Moreover, it may help in indicating the underlying hypoxic condition in the placenta.
preeclampsia; cell-free fetal DNA; HCG
This study was designed to (i) delineate the characteristics of serum antibody responses to Actinobacillus actinomycetemcomitans in patients with periodontitis who are infected with A. actinomycetemcomitans; irrespective of disease classification; (ii) assess the relationship of the elevated antibody levels to colonization of the oral cavity by A. actinomycetemcomitans; and (iii) describe the serotype distribution of A. actinomycetemcomitans and antibodies to the microorganism in infected patients with various clinical classifications. To compare the levels of various isotype-specific antibodies to the different antigens, studies were performed that allowed quantitation of each isotype-specific antibody in a human reference standard. By using this reference standard, it was shown that the levels of immunoglobulin G (IgG), IgM, and IgA responses to A. actinomycetemcomitans were similar among the infected patients, irrespective of disease classification. Also, we demonstrated that the serum antibody response to serotype b was quantitatively greater in all isotypes. Our findings indicate that b was the most frequent A. actinomycetemcomitans serotype detected in the patients and appears to be capable of initiating a substantial serum IgG antibody response that may contain cross-reactive antibodies to other serotypes of A. actinomycetemcomitans. Generally, in cases in which the response to a single serotype was elevated, only that type of A. actinomycetemcomitans was detected in the plaque. Individuals exhibiting elevated antibodies to multiple serotypes were most consistently colonized by the serotype b microorganism. This study represents the first report detailing the distribution of IgG subclass antibodies to A. actinomycetemcomitans in periodontal disease. The results demonstrated that the primary responses of patients with periodontitis to A. actinomycetemcomitans were of the IgG1 and IgG3 subclasses, which is consistent with elicited responses to protein antigens. In contrast, the primary subclass response in normal subjects was limited to the IgG2 subclass and may represent broader cross-reactivity to polysaccharide antigens-lipopolysaccharide from the bacteria.
We developed an assay to detect antibodies spontaneously secreted in vitro by peripheral blood mononuclear cells (PBMC) against Brucella spp. High levels of anti-Brucella immunoglobulin G (IgG) and/or IgM and/or IgA antibodies were detected in the cell supernatant solution of PBMC cultures for 12 patients suffering from acute or focalized brucellosis and for 5 patients recently vaccinated against brucellosis. This spontaneous in vitro antibody production disappeared 5 to 20 months after onset of clinical signs and 20 to 27 days after vaccination. The transient character of this anti-Brucella antibody production by PBMC is consistent with a temporary in vivo stimulation of the immune system by Brucella antigens. Detection of this secretion could improve the diagnosis of evolutive brucellosis.
In a study of members of a large family with a high prevalence of early-onset periodontitis, we sampled the subgingival microflora and characterized 40 isolates from each sample. We surveyed serum samples by enzyme-linked immunosorbent assay for antibodies reacting with any of a panel of 21 periodontal bacteria. The mother and 7 of her 13 children had early-onset periodontitis. Bacteroides gingivalis was not detected in the subgingival flora of any affected or unaffected family member, and Actinobacillus actinomycetemcomitans was isolated from only one affected child. Capnocytophaga ochracea was isolated from five of seven affected children and from none of their normal siblings. We found no significant differences among the floras from family members who had rapidly progressive, juvenile, and prepubertal forms of periodontitis. Elevated levels of serum antibody reacting with one or more of the bacteria tested were found in all family members with disease, but in only one periodontally normal family member. Both children with prepubertal periodontitis had antibodies reacting with C. sputigena, a species not found in their subgingival floras, but with none of the other bacteria tested. All remaining affected family members had antibodies to one or more serotypes of A. actinomycetemcomitans, and four had antibodies reacting with additional bacteria, including C. sputigena, Eikenella corrodens, Fusobacterium nucleatum, and Haemophilus aphrophilus. Sera from patients contained antibodies specific for putative periodontal pathogens not found in their pocket flora, and conversely, putative periodontal pathogens for which no serum antibodies were found frequently comprised a large proportion (10% or more) of the pocket flora. In no case were both the bacterium and its antibody found. These observations are suggestive of sequential infection in the early-onset forms of periodontitis and of induction of protective immunity against reinfection by the same microorganism.
A geographically homogeneous population of 83 subjects, from 21 families with localized juvenile periodontitis (LJP), and 35 healthy control subjects was monitored, over a 5-year period, for the presence of the periodontal pathogen Actinobacillus actinomycetemcomitans. Restriction fragment length polymorphism (RFLP) analysis was used to monitor the distribution of genetic variants of this bacterium in LJP-susceptible subjects that converted from a healthy to a diseased periodontal status. A. actinomycetemcomitans was cultured from 57% of the LJP family members accessioned into the study. Nine of 36 LJP-susceptible subjects, in seven families, developed signs of periodontal destruction. All but one of these conversion subjects harbored A. actinomycetemcomitans. Bacterial variants representative of a single RFLP group (II) showed the strongest correlation with conversion (P < 0.002). Six of nine conversion subjects were infected with A. actinomycetemcomitans from this group. RFLP group II variants also prevailed in 8 of 22 probands but were absent in the 35 healthy control subjects. In contrast to the selective distribution of group II variants is diseased individuals, variants belonging to RFLP groups XIII and XIV were found exclusively in the control subjects. Thus, the use of RFLP to type clinical isolates of A. actinomycetemcomitans has resulted in the identification of genetic variants that predominate in LJP and health. These results indicate that studies concerned with the pathogenicity of this bacterium in LJP should be focused on the group II variants.
By using a sensitive enzyme-linked immunosorbent assay, 200 randomly selected sera from Red Cross blood donors were screened for immunoglobulin G (IgG), IgA, and IgM levels against Actinobacillus actinomycetemcomitans, Bacteroides gingivalis, and Bacteroides intermedius. A subgroup of 79 blood donors was clinically examined for type and extent of periodontal destruction, and serological and clinical data were subjected in all possible dual combinations to correlation analyses. The results revealed that the majority of the blood donors suffered from moderate to severe adult periodontitis, often coupled with severe gingival inflammation. No cases of localized juvenile periodontitis or rapidly progressive periodontitis were observed. The extent of periodontal destruction proved to be significantly correlated only to the IgG response levels against B. gingivalis. Corresponding correlation tests assessing the relationships of loss of attachment, bone loss, pocket depth, and papillary bleeding index with the IgG responses to A. actinomycetemcomitans were of marginal significance, while the IgG responses to B. intermedius revealed no relationship to the periodontal health status. The specific IgM responses proved to be unrelated to the clinical parameters, but interestingly, they were found to be highly correlated with each other. Specific IgA levels were frequently too low for enzyme-linked immunosorbent assay testing and, therefore, had to be exempted from statistical analyses. Assessments of the serotype specificity of strongly elevated IgG responses to A. actinomycetemcomitans disclosed no evidence for an association of a particular serotype-specific IgG response with the occurrence of adult periodontal destruction. In contrast to results of earlier studies, a number of sera were found to contain strongly elevated IgG levels against two or even all three serotypes. Although derived by an alternative approach, the reported results largely corroborate earlier observations linking only the occurrence of elevated anti-B. gingivalis IgG responses to the presence of marked periodontal lesions in adults.
Background. Cell signaling via Toll-like receptors (TLRs) leads to synovial inflammation in rheumatoid arthritis (RA). We aimed to assess effects of TLR2 and TLR4 stimulation on proinflammatory cytokine production by peripheral blood mononuclear cells (PBMCs) from patients with recent-onset RA, osteoarthrosis (OA), and healthy control (HC).
Methods. PBMCs were stimulated with LPS, biglycan and cytokine mix. Cytokines were analyzed in supernatants with ELISA. Expression of toll-like receptors mRNA in leukocytes was analyzed using real-time qPCR.
Results. PBMCs from RA patients spontaneously produced less IL-6 and TNFα than cells from OA and HC subjects.
LPS increased cytokines' production in all groups. In RA patients increase was dramatic (30 to 48-fold and 17 to 31-fold, for respective cytokines) compared to moderate (2 to 8-fold) in other groups. LPS induced 15-HETE generation in PBMCs from RA (mean 251%) and OA patients (mean 43%), although only in OA group, the increase was significant. TLR2 and TLR4 gene expressions decreased in response to cytokine mix, while LPS enhanced TLR2 expression in HC and depressed TLR4 expression in OA patients.
Conclusion. PBMCs from recent-onset RA patients are overresponsive to stimulation with bacterial lipopolysaccharide.
TLR expression is differentially regulated in healthy and arthritic subjects.