sodium-dodecyl-sulfate and Aggressive-Periodontitis

Immunological studies examining the homogeneity of the major antigenic components of P. gingivalis have suggested 3 serotypes and have indicated a limited distribution of the serotypes in an individual patient. These studies prompted us to define the immunodominant antigens and distribution of immune responses to P. gingivalis serotypes. Serum IgG antibody levels in periodontitis patients in the present study were most frequently elevated above the normal subjects when tested against P. gingivalis serotype A (i.e., 33277). Nearly 1/3 of the patients showed significantly elevated antibody to multiple serotypes of the P. gingivalis apparently resulting from cross-reacting antigens. We determined distinctive differences among outer envelope protein and antigen patterns obtained from the three serotypes. Moreover, the results identified considerable similarities in the qualitative and quantitative antigen response patterns among patients to a particular serotype. There was a strong positive correlation between IgG antibody levels (ELISA) and the total level of reactivity determined in the immunoblots, as well as a positive correlation to the proportion of antibody to particular antigens. These findings suggest that responses to these antigens comprised a major portion of the response to the intact microorganism. Additionally, the detection of antibody to particular antigen bands was indicative of early responses to each of the P. gingivalis serotypes. The results of our study indicate that a subpopulation of periodontitis patients develop an extensive serum antibody response often to multiple serotypes of P. gingivalis and may define a patient population with a P. gingivalis disease. Finally, our results indicate a more consistent antigenic composition for P. gingivalis which may enhance the potential for strategies to immunologically interfere with disease caused by this microorganism.

Topics: Adolescent; Adult; Aggressive Periodontitis; Antibodies, Bacterial; Antigens, Bacterial; Bacterial Outer Membrane Proteins; Blotting, Western; Cross Reactions; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunoglobulin G; Male; Middle Aged; Periodontitis; Periodontium; Porphyromonas gingivalis; Serotyping; Sodium Dodecyl Sulfate

The aim of the present study was to characterize the eventual presence and molecular forms of gelatinase/type IV collagenase activities in gingival crevicular fluid (GCF) and saliva in different forms of periodontitis; patients with clinically healthy periodontium served as controls. Enzyme activities were monitored electrophoretically by zymography using gelatin and type IV collagen as substrates and analyzed visually and/or densitometrically. Both saliva and GCF collected from adult periodontitis, localized juvenile periodontitis and type II diabetes mellitus periodontitis patients contained species moving identically with gelatinase isolated from human neutrophils or MMP-9 (mean 98 kD), and species with mobility similar to gelatinase in fibroblast cell culture supernatants or MMP-2 (mean 76 kD). Hitherto, undescribed high molecular weight forms (mean 128 kD), were found, possibly representing polymerized or complexed enzyme active/activated in situ in the gel matrix. Small molecular forms of gelatinases (mean 51 kD and 46 kD), unable to cleave type IV collagen, were also found, most likely representing in vivo proteolytically activated, truncated enzymes. Although multiple forms of gelatinases/type IV collagenases in saliva and GCF may take part in the tissue destruction in periodontitis, their profile judged according to molecular weights does not differentiate between different forms of periodontitis.

Topics: Adult; Aggressive Periodontitis; Collagenases; Diabetes Mellitus, Type 2; Electrophoresis, Polyacrylamide Gel; Female; Gelatinases; Gingival Crevicular Fluid; Humans; Male; Matrix Metalloproteinase 9; Middle Aged; Molecular Weight; Periodontal Pocket; Periodontitis; Periodontium; Saliva; Sodium Dodecyl Sulfate

Levels of serum IgG antibody to the 29 kilodalton outer membrane protein of A. actinomycetemcomitans Y4 in sera of periodontally healthy subjects and localized juvenile periodontitis patients were determined using an enzyme-linked immunosorbent assay (ELISA). The 29 kDa protein was isolated by solubilization of an octylglucoside-NaCl insoluble fraction by incubation at ambient temperature in 20 mM sodium phosphate, pH 7.5, containing 1% sodium dodecyl sulfate. The isolated protein migrated on SDS-polyacrylamide gels with an apparent molecular mass of 29 kDa following incubation in sample buffer at ambient temperature. However, the protein migrated with an apparent molecular mass of 34 kDa following incubation in sample buffer at 100 degrees C for 10 minutes. Geometric mean IgG antibody titers to the 29 kDa protein were significantly higher in sera from LJP subjects than in sera obtained from periodontally healthy subjects. Twenty-two of 35 LJP sera (63%) had antibody titers greater than 2 standard deviations from the mean titer of the periodontally healthy group. Among LJP subjects, elevated antibody titers to the 29 kDa protein were found primarily in subjects greater than or equal to 18 years of age, with the highest titers seen in patients 18 to 21 years of age. The results of this study indicate that the humoral response of LJP subjects to A. actinomycetemcomitans includes the production of IgG antibodies which recognize the major outer membrane proteins of this organism.

Topics: Actinobacillus; Adolescent; Adult; Age Factors; Aggressive Periodontitis; Antibodies, Bacterial; Antibody Specificity; Bacterial Proteins; Child; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Haemophilus; Humans; Immunoblotting; Immunoglobulin G; Membrane Proteins; Middle Aged; Molecular Weight; Periodontium; Sodium Dodecyl Sulfate

Localized juvenile periodontitis (LJP) is a progressively destructive infection of the supporting tissues of the teeth, primarily affecting adolescents. In this disease, patients' polymorphonuclear leukocytes (PMN) exhibit decreased chemotaxis (CTX) and decreased binding of the chemotactic peptide N-formyl-l-methionyl-l-leucyl-l-phenyl-alanine (FMLP) to specific receptors on the PMN surface. Since the FMLP receptor is involved in the activation of the PMN, and its subsequent response to chemotactic stimuli, a decrease in the chemotactic peptide receptor, as seen in LJP patients, is suspected to be a predisposing factor for this disease. To define differences in the FMLP receptor between CTX defective LJP patients and CTX normal donors, a battery of monoclonal antibodies reactive against the FMLP receptor was prepared. The FMLP receptor was affinity-purified, and was found to be comprised of two components, one of 68 kDa, and the other of 94 kDa. Only the 68 kDa component specifically bound a radioiodinated FMLP analogue in a photoaffinity experiment. Seven monoclonal antibodies were selected on the basis of their reactivity with the 68 kDa receptor component, and of these, 5 showed reduced binding against PMN from CTX defective LJP donors when compared to their reactivity against PMN from CTX normal subjects. Two of the 7 anti-68 kDa antibodies reacted with PMN from both sets of subjects at a comparable extent. Furthermore, the presence of 20 nmol of FMLP inhibited the binding of 5 of the anti-receptor antibodies to whole PMN, including one that showed no difference in binding between CTX normal and defective PMN, and 4 of the 5 that did show such difference.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adolescent; Adult; Affinity Labels; Aggressive Periodontitis; Amino Acids; Antibodies; Antibodies, Monoclonal; Antibody Affinity; Autoradiography; Blotting, Western; Cell Membrane; Chemotaxis, Leukocyte; Child; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Humans; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Receptors, Formyl Peptide; Receptors, Immunologic; Sodium Dodecyl Sulfate