Clinical and microbiological effects of minocycline-loaded microcapsules in adult periodontitis.

Clinical and microbiological effects of subgingival delivery of 10% minocycline-loaded (MC), bioabsorbable microcapsules were examined in 15 adult periodontitis patients. Patients received oral hygiene instruction 2 weeks prior to the study. At baseline (day 0) all teeth received supragingival scaling (SC); 2 quadrants received no further treatment and 1 quadrant received subgingival scaling and root planning (SRP). In the fourth quadrant, the tooth with the deepest probing sites (at least 1 site > or = 5 mm) was treated with minocycline microcapsules. The sites were evaluated at baseline and weeks 1, 2, 4, and 6. Clinical indices included bleeding on probing (BOP), probing depths (PD), and attachment loss (AL). Microbiological evaluations included percent morphotypes by phase-contrast microscopy; cultivable anaerobic, aerobic, and black-pigmented Bacteroides (BPB); and percent Porphyromonas gingivalis, Prevotella intermedia, Eikenella corrodens, and Actinomyces viscosus by indirect immunofluorescence. In the SC + MC group, BOP, PD, and AL were significantly reduced from baseline for weeks 1 to 6. BOP in the SC + MC group was significantly reduced compared to the SRP group from weeks 2 to 6. In the SC + MC group the percent of spirochetes and motile rods decreased and the percent of cocci increased after 1 week. The increased cocci and decreased motile rods were statistically greater at weeks 4 and 6 in the SC + MC group compared to the SRP group. This study demonstrates that local subgingival delivery of 10% minocycline-loaded microcapsules as an adjunct to scaling results in reduction in the percent sites bleeding on probing greater than scaling and root planning alone and induces a microbial response more favorable for periodontal health than scaling and root planing.

Clinical and microbiological status of osseointegrated implants.

Peri-implantitis, an inflammatory response around implants, has a poorly defined etiology and pathogenesis. To better understand the role of specific microorganisms in this disease process, clinical and microbiological parameters were examined in 24 patients with 98 osseointegrated implants. Sites were evaluated for probing depth (PD), plaque/calculus index (PI), gingival bleeding index (GBI), mobility, and crevicular fluid flow rate (CFFR). Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Prevotella intermedia in subgingival plaque were identified by latex agglutination assays. Clinically, a statistically significant correlation (P < 0.001) was observed between probing depth and the length of time an implant was present. Mobility was also significantly greater (P < 0.001) in the maxillary than in the mandibular implants. Subgingival sites harboring one of the three microorganisms had significantly greater PD, GBI, and CFFR than non-colonized sites. Implants in partially edentulous patients more frequently were colonized with P. gingivalis/P. intermedia than edentulous patients. The incidence of these microorganisms also correlated with fixture longevity. Implants present for 3 to 4 years had a significantly greater frequency of test microorganisms than implants present for 1 to 2 years. These findings suggest that microbial pathogens associated with periodontitis occur more commonly around implants exhibiting gingival inflammation (GBI) and may contribute to peri-implantitis.

Development of a rapid latex agglutination test for periodontal pathogens.

The studies reported here describe the development, characterization, and initial application of latex agglutination assays for periodontal pathogens. Latex reagents were prepared by sensitization of latex microspheres with rabbit IgG antibodies to either Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, or Prevotella intermedia. The protein concentration utilized for sensitization and microsphere size were optimized. One reagent was prepared to A. actinomycetemcomitans and a second combination reagent was prepared by mixing latex sensitized with antibodies to P. gingivalis and latex sensitized with antibodies to P. intermedia. The sensitivity of both latex reagents in the traditional wet and a dried format was evaluated. In addition, sensitivity and specificity with homologous and heterologous bacterial suspensions were evaluated. The reagents were found to demonstrate both specificity and sensitivity. Initial studies with subgingival human plaque demonstrated the ability of these reagents to detect the specific organisms in plaque.

Humoral antibody responses in periodontal disease.

Several forms of periodontal disease are considered to be infectious diseases with associated specific bacteria. This study examined the humoral antibody levels as assayed by ELISA to Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Prevotella intermedia in adult periodontitis (AP), localized juvenile periodontitis (LJP), rapidly progressive periodontitis (RPP), and in periodontally healthy subjects (HS). Sixty-two of the 64 (96.9%) patients had significantly elevated antibody levels to at least one of the three organisms. Elevated levels of antibodies to P. gingivalis occurred in 82.8% of the RPP, LJP, and AP patients with all 3 disease groups showing greater responses than HS controls. Antibodies to A. actinomycetemcomitans were found in 59.4% of the RPP, LJP, and AP patients and were significantly higher in both LJP and RPP patients. Only 21.9% of the RPP, LJP, and AP patients showed elevated levels to P. intermedia with only significantly higher levels in the RPP and LJP groups. Antibodies to A. actinomycetemcomitans and P. intermedia were rarely found alone (only 5.1% and 2.6% of the patients respectively) but were usually accompanied by P. gingivalis. These results suggest that one or more combinations of these 3 bacteria may play a role in the pathogenesis of these forms of periodontal disease.

Investigation of the PMN chemotaxis in periodontal disease.

The chemotactic responses of peripheral polymorphonuclear leukocytes in 36 patients with rapidly progressive periodontitis (RPP) were compared to responses in 15 patients with localized juvenile periodontitis (LJP) and 10 patients with adult periodontitis (AP). In the RPP group, 13 (36%) patients exhibited reduced chemotactic activity and 7 (19%) exhibited activity. Compared to this, 12 (80%) of the LJP patients displayed reduced chemotactic activity. Among the AP-patients 4 (40%) showed altered chemotactic activity including 1 (10%) with reduced chemotactic activity and 3 (30%) with increased activity. These results suggest that depressed chemotaxis is greatest in LJP, intermediate in RPP and least in AP.

Actinobacillus actinomycetemcomitans serotypes and leukotoxicity in Korean localized juvenile periodontitis.

Actinobacillus actinomycetemcomitans is though to play an important role in the pathogenesis of localized juvenile periodontitis (LJP). Preliminary data suggested that the serotype distribution of A. actinomycetemcomitans in Korea and the United States differ. This study evaluated A. actinomycetemcomitans prevalence, serotype distribution, and leukotoxicity in Korean LJP patients by culture, enzyme-linked immunosorbent assay, indirect immunofluorescence, and lactate dehydrogenase release from polymorphonuclear leukocytes exposed to A. actinomycetemcomitans. A. actinomycetemcomitans occurred in 75% of LJP lesions and 6% of normal sites with approximately equal distribution of serotype a, b, and c. Single serotypes were isolated from nine patients while three patients harbored two serotypes either in the same or different disease sites. A. actinomycetemcomitans leukotoxicity occurred in 22% isolates with a 69% prevalence. Individual sites harbored both leukotoxic and non-leukotoxic strains with no serotype association. The distribution of serotypes and leukotoxic strains of A. actinomycetemcomitans in Korean LJP patients differed from those reported in the United States. This suggests that serotype b may not be more important in the pathogenesis of LJP.

The treatment of desquamative gingival lesions.

The treatment of desquamative gingival lesions is dependent on the correct diagnosis of the underlying disease, which is frequently dermatologic. This is determined by clinical observations, as well as by histologic and immunologic examination of gingival biopsy specimens. The recommended treatment of desquamative lesions caused by lichen planus, cicatricial pemphigoid (benign mucous membrane pemphigoid), pemphigus, and psoriasis, the most common dermatoses causing gingival lesions, is summarized on the basis of 62 cases and reports in the literature.

The presence of bacteria in the oral epithelium in periodontal disease. II. Immunohistochemical identification of bacteria.

Serial histological sections of gingiva obtained from each of six advanced adult periodontitis, two localized juvenile periodontitis and two periodontally healthy patients were used for specific identification of bacteria within the oral epithelium and adjacent connective tissue. Healthy gingival biopsies served as controls. Sections from patients and control biopsies were Gram-stained and also screened with antibacterial sera associated with the peroxidase immunocytochemical technique for specific bacterial identification. The "Pop-off" electron microscopic technique was also used to further demonstrate the bacterial nature of peroxidase-stained material. In addition, the possible correlation between bacteria and areas of possible reduced keratinization was investigated. The results showed that sections of orthokeratinized healthy gingiva did not contain bacteria. Gram-stained sections from diseased sites contained large numbers of bacteria in the oral epithelium and adjacent connective tissue. Bacteroides gingivalis and to a lesser extent Capnocytophaga gingivalis were found in periodontitis, and Actinobacillus actinomycetemcomitans was found in juvenile periodontitis when the immunoperoxidase technique was used. The bacterial nature of peroxidase-stained material was confirmed by the "pop-off" technique. In the disease biopsies, bacterial presence was correlated with areas of reduced amounts of keratin suggesting that the oral epithelium may be a portal of entry for bacteria into gingival tissues.

Antibodies to Actinobacillus actinomycetemcomitans in a Korean population.

Previous studies demonstrated that serum antibodies to Actinobacillus actinomycetemcomitans strain Y4 are significantly elevated in sera from localized juvenile periodontitis (LJP) and postlocalized juvenile periodontitis (P-LJP) patients compared to normal controls in United States populations. This study examined the age of subjects in relation to the antibody levels to A. actinomycetemcomitans in a Korean population. Seven groups were investigated including sera from newborns, infants, children, periodontally normal puberty and adult groups and LJP and P-LJP groups. Antibody levels were determined by an enzyme-linked immunosorbent assay (ELISA) to A. actinomycetemcomitans strain SNUDC 10-1 (serotype C) isolated from a Korean LJP patient. In the healthy non-LJP and non-P-LJP subjects, IgG levels decreased from the newborn group to the 5-month-old group and then gradually increased through the adult group. The IgM levels in these groups continuously increased from birth until a plateau was reached in the 2- to 6-year group. Serum IgA levels to strain SNUDC 10-1 were too low to be measured by this assay.

Gingival basement membrane antigens in desquamative lesions of the gingiva.

Desquamative gingival lesions are a clinical manifestation of several entities where histologic alterations in the gingival basement membrane are frequent. Normal and lesional gingival specimens from six patients with desquamative lesions of the gingiva associated with cicatricial pemphigoid or lichen planus and from one normal subject were examined by indirect immunofluorescence for immunologic alterations in basement membrane antigens. In four out of five desquamative gingival lesions from patients with lichen planus, type IV collagen and EBA basement membrane antigens were interrupted, thickened, or partially or completely absent as compared to normal gingiva. One specimen showed multiplication and duplication of the basement membrane. In the two desquamative gingival lesions from patients with cicatricial pemphigoid, type IV collagen was either partially absent or associated with the connective tissue side of the subepithelial bullae. These results suggest structural alterations in antigens within the basement membrane of patients with desquamative lesions of the gingiva.

The alteration in gingival basement membrane antigens in chronic periodontitis.

The gingival basement membrane antigens, Type IV collagen, bullous pemphigoid antigen and epidermolysis bullosa acquisita antigen were studied by indirect immunofluorescence in 11 gingival specimens from patients with periodontitis and 2 normal gingival specimens. In the normal control gingival specimens, the antigens were all present and stained with a continuous linear pattern. In periodontitis, alterations occurred in the gingival basement membrane antigens in the apical portion of the pockets. These included thinning, interruptions, partial or complete absence involving one or more rete pegs and fragmentation. These alterations may result form the disease process or play a role in the pathogenesis.

The regeneration of gingival basement membrane antigens during secondary wound healing.

The reformation of basement membrane antigens was examined during healing following gingivectomy procedures. Three antigens, Type IV collagen, bullous pemphigoid antigen and epidermolysis bullosa acquisita (EBA) antigen, were identified by indirect immunofluorescent tests. While all three antigens could be identified in the healing wound within 2 days, the antigens regenerated at different rates. The bullous pemphigoid antigen reformed the earliest and extended furthest along the healing front of epithelium. The Type IV collagen reformed at an intermediate rate and the EBA antigen formed the slowest. These results confirm that these three antigens are different and suggest that the gingival basement membrane is a heterogeneous structure which contains elements that form at different rates during secondary wound healing.

Streptococcus mutans-induced nephritis in rabbits.

Intravenous administration of disrupted Streptococcus mutans into rabbits over 23-76 weeks led to severe nephritis involving glomeruli, tubules, and interstitium. Light-microscopic observation of glomeruli documented diffuse endocapillary proliferative glomerulonephritis accompanied often (65%) by epithelial crescents. Electron-microscopic observation revealed humps in glomeruli of 70% of kidney specimens. In the glomeruli of some rabbits, extensive fibrin deposits and sclerosis were evident. Immunofluorescence showed linear, granular, often ribbonlike or patchy immune deposits encompassing, in order of decreasing frequency, C3, IgG, streptococcal antigen, IgA, and IgM. The histopathologic and immunohistologic features of the nephritis seen in rabbits given S mutans thus shows many features of Streptococcus-associated nephritides in man, in particular, the diffuse glomerular nephritis encountered in subacute bacterial endocarditis. Further, analysis of nephritis induced by administration of S mutans may have implications for the evaluation and purification of dental caries vaccines.

Binding of Streptococcus mutans antigens to heart and kidney basement membranes.

Using indirect immunofluorescence, alkali-extracted components of Streptococcus mutans were found to bind in vitro to capillary walls and sarcolemmal sheaths of monkey cardiac muscle and to glomerular and tubular basement membranes of monkey kidney. Adsorption of S. mutans components to tissue fragments was also detected by indirect radioimmunoassay and immunoblotting on nitrocellulose paper. Antibodies did not bind to untreated, control tissues in these experiments, proving that antigens shared by S. mutans and tissue components were not involved. Rabbit and monkey heart and kidney components bound S. mutans antigens of 24,000, 35,000, and 65,000 Mr. Monkey heart also bound molecules of 90,000 and 120,000 Mr. Rabbits immunized by intravenous injection of disrupted S. mutans cells developed severe nephritis that was characterized by the deposition of immunoglobulins, complement component C3, and S. mutans antigens in the glomeruli. Immunoglobulin G eluted from nephritic kidneys reacted in immunoblots with the 24,000, 35,000, and 65,000 Mr components of S. mutans extract, indicating that the antigens that bound to tissue in vitro also bound in vivo and reacted with antibodies in situ. Antibodies to other S. mutans antigens were not detected in the kidney eluate, although they were present in the serum of the same rabbit.

Basement membrane reactivity of antisera to type IV collagen and sera from patients with bullous pemphigoid and epidermolysis bullosa acquisita.

The basement membrane antigenic specificities of antibodies to Type IV collagen were compared to naturally occurring antibodies in sera from patients with bullous pemphigoid and epidermolysis bullosa acquisita (EBA) by indirect immunofluorescence, mixed immunofluorescence and immunoabsorption. Results suggested that the three sera reacted with three different basement membrane antigens. In addition, absorption with Types I, II, III, or IV collagen failed to reduce the basement membrane reactivities of bullous pemphigoid or EBA sera. The antibodies to the basement membrane components should be useful in studying skin and mucous membrane diseases including periodontal diseases.

Serology and tissue lesions in rabbits immunized with Streptococcus mutans.

Rabbits were immunized i.v. or i.d. with sterile suspensions of disrupted Streptococcus mutans strain MT703 or K1R. Indirect immunofluorescence assays indicated that sera from four of 10 rabbits immunized i.d. contained antibodies reactive with monkey and human heart and kidney components; 19 of 24 rabbits immunized i.v. had antibodies reactive with these tissues. Heart-reactive antibodies were also detected by immunoelectrophoresis and indirect radioimmunoassay. These antibodies were absorbed well by cytoplasmic membranes, a whole cell extract, and an alkali extract of S. mutans but only weakly by intact bacteria. Between 6 and 8 weeks after the first i.v. administration of S. mutans vaccines, rabbits developed proteinuria and hematuria with subsequent weight loss and lethargy. Approximately 25% of the animals died from illness between the fifth and sixth month of immunization. In 13 of 15 rabbits, immune deposits of C3 and IgG, IgM, or IgA and fibrinogen were seen in kidneys within the glomeruli, basement membranes of the peritubular capillaries, and in the interstitium. In the heart, deposits were seen along the capillaries of the myocardium. In 8 of 14 rabbits, focal deposits of S. mutans antigen were detected in glomeruli and in the kidney interstitium. The kidneys showed gross pathologic and histopathologic changes. Most kidneys were pale and enlarged. Microscopic examination revealed hypercellularity of the glomeruli, presence of neutrophils, thickening of glomerular and tubular basement membranes, tubular atrophy, edema, and fibrosis of the interstitium. The kidney disease presented features of poststreptococcal glomerulonephritis. Microscopic examination of heart sections revealed mild perivascular infiltration by polymorphonuclear leukocytes and plasma cells in some of the rabbits.

Bacterial IgG and IgM antibody titers in acute necrotizing ulcerative gingivitis.

IgG AND IgM ANTIBODY TITERS to eight bacterial isolates were measured by indirect immunofluorescence and ELISA in sera from acute necrotizing ulcerative gingivitis (ANUG) patients during the acute phase, from ANUG patients during the convalescent phase, from patients with gingivitis and from subjects with normal gingiva. Subjects were matched with respect to age and sex. Compared to the gingivitis and healthy groups, the ANUG groups exhibited significantly higher IgG and IgM titers to intermediate-sized spirochetes and higher IgG titers to Bacteroides melaninogenicus subsp intermedius. The findings support recent studies showing that these organisms are major bacterial components in ANUG lesions. They also suggest that these bacteria proliferate above-normal levels several weeks or months prior to the clinical onset of ANUG.

Immunologic cross-reactivity between Streptococcus mutans and mammalian tissues.

The presence of cross-reacting antigens shared by oral streptococci and human and monkey tissue was investigated by indirect immunofluorescence. Rabbit antisera prepared against two strains of each of the seven serotypes of Streptococcus mutans were incubated on sections of cardiac muscle, skeletal muscle, kidney, brain, liver, and skin. Antisera to S. mutans, serotype g, strain OMZ-65 reacted with cardiac muscle components. This reactivity could be reduced by absorption with bacterial cell membranes. Antisera to S. mutans serotype g, strain K-1 reacted with both cardiac and skeletal muscle but could not be reduced by absorption with bacterial cells, cell walls, or cell membranes. Antisera to S. mutans serotype g, strain OMZ-65 reacted with kidney glomeruli and could be completely absorbed by a cell wall preparation. S. mutans, serotype e, strain MT703 also reacted with kidney glomeruli but could not be reduced by absorption with bacterial antigens.

Dermatitis herpetiformis: IgA deposits in gingiva, buccal mucosa, and skin.

Fourteen patients who had dermatitis herpetiformis of the skin with no gingival lesions were studied. In each patient biopsy specimens of normal gingiva, buccal mucosa, and perilesional skin were examined for IgA and IgG deposits. Depending on the method of handling biopsy specimens for immunofluorescence, IgA deposits were detected in 21 to 25 percent of gingival specimens, 27 to 46 percent of buccal mucosal specimens, and 64 to 76 percent of skin specimens. The occurrence of IgA deposits in the gingival papillae, a site where lesions of DH have not been reported in the past or observed in this study, suggests that these IgA deposits alone may not play a primary role in the pathogenesis of this disease. Similar immunofluorescent findings of IgA deposits were observed in both quick-frozen biopsy specimens and those placed in holding solution, indicating the suitability of the holding solution for transporting suggested DH biopsy material.