No evidence of triclosan-resistant bacteria following long-term use of triclosan-containing toothpaste.

BACKGROUND AND OBJECTIVE: There is a paucity of data in relation to the possible emergence of triclosan (TCS)-resistant bacteria following long-term exposure to TCS toothpaste. Therefore, this study investigated whether long-term continuous exposure to TCS in toothpaste selects for TCS-resistant bacteria within the oral biofilm. MATERIAL AND METHODS: Dental plaque samples were collected from 40 individuals during year 5 of a randomised controlled trial. Participants had been randomly assigned to use TCS (3000 µg/mL TCS) (n = 18) or placebo toothpaste (n = 22). Diluted plaque samples were plated on to Wilkins-Chalgren agar plates containing 5% (v/v) laked sheep red blood cells and TCS (concentrations ranging from 25 to 150 µg/mL) and incubated at 37 °C under microaerophilic and anaerobic conditions for 2-10 d. Selected bacterial isolates were identified by partial 16S rDNA sequencing and TCS minimum inhibitory concentration (MIC) determined for each isolate. RESULTS: At 3000 µg/mL TCS no growth was observed under microaerophilic or anaerobic conditions in either group. The MICs of TCS for all isolates ranged from 125 to 1000 µg/mL in both groups. Species common to both groups had similar MICs. Veillonella parvula and Campylobacter gracilis were the most frequent isolates from both groups, with similar MICs in both groups. CONCLUSION: The use of TCS-containing toothpaste did not appear to lead to an increase in MIC of TCS of oral bacterial isolates.

Nitric oxide production by a murine macrophage cell line (RAW264.7 cells) stimulated with Aggregatibacter actinomycetemcomitans surface-associated material.

Nitric oxide (NO) may play a crucial role in the pathogenesis of periodontal disease and, hence, the aim of the present study was to test the hypothesis that Aggregatibacter actinomycetemcomitans surface-associated material (SAM) stimulates inducible nitric oxide synthase (iNOS) activity and NO production by the murine macrophage cell line RAW264.7. Cells were stimulated with untreated or heat-treated A. actinomycetemcomitans SAM and with or without pre-treatment with L-N(6)-(1-Iminoethyl)-lysine (L-NIL) (an iNOS inhibitor), polymyxin B, interferon-gamma (IFN-γ) and Interleukin-4 (IL-4), IL-10, genistein [a protein tyrosine kinase (PTK) inhibitor], bisindolylmaleimide [a protein kinase C (PKC) inhibitor], bromophenacyl bromide (BPB) [a phospholipase A(2) (PLA2) inhibitor] or wortmannin [phosphatidylinositol 3-kinase (PI-3K) inhibitor]. The iNOS activity and nitrite production in the cell cultures were determined. Untreated but not heat-treated A. actinomycetemcomitans SAM-stimulated both iNOS activity and nitrite production in RAW264.7 cells. L-NIL, IL-4, IL-10, genistein, bisindolylmaleimide, or BPB, suppressed but IFN-γ enhanced both iNOS activity and nitrite production by A. actinomycetemcomitans SAM-stimulated cells. Wortmannin and polymyxin B failed to alter both iNOS activity or nitrite production by A. actinomycetemcomitans SAM treated cells. Therefore, the present study suggests that a heat-sensitive protein constituent(s) of A. actinomycetemcomitans SAM stimulates both iNOS activity and nitrite production by RAW264.7 cells in a cytokine, PTK, PKC, and PLA(2) but not PI-3K-dependent fashion.

Periodontal disease in free-ranging koalas (Phascolarctos cinereus) from the Mount Lofty Ranges, South Australia, and its association with koala retrovirus infection.

BACKGROUND: In northern Australian koala populations (Queensland and New South Wales), periodontal disease (gingivitis and periodontitis) is common while koala retrovirus subtype A is endogenous, with other subtypes transmitted exogenously. Koala retrovirus has been hypothesised to cause immune suppression and may predispose koalas to diseases caused by concurrent infections. In southern Australia populations (Victoria and South Australia) periodontal disease has not been investigated, and koala retrovirus is presumably exogenously transmitted. This study described oral health in South Australian koalas and investigated if an association between periodontal disease and koala retrovirus exists. METHODS: Oral health was examined for wild-caught koalas from the Mount Lofty Ranges (n = 75). Koala retrovirus provirus was detected in whole blood using nested PCR and proviral load determined with qPCR. Periodontal disease severity was recorded and used to calculate the Final Oral Health Index (0-normal, 24-severe).Results Periodontal disease was observed in 84% (63/75) of koalas; 77% had gingivitis (58/75) and 65% (49/75) had periodontitis. The average Final Oral Health Index was 5.47 (s.d 3.13). Most cases of periodontal disease were associated with the incisors. Koala retrovirus-infected koalas were more likely to present with periodontitis (p = 0.042) and the Final Oral Health Index was negatively correlated with proviral load (ρ = -0.353, p = 0.017). CONCLUSION: South Australian koalas had a high prevalence of gingivitis and periodontitis. Periodontal disease was more prevalent in the incisors. Exogenous koala retrovirus infection may also facilitate the development of periodontitis by modulation of the immune response to concurrent oral bacterial infections.

Nitric oxide production by a human osteoblast cell line stimulated with Aggregatibacter actinomycetemcomitans lipopolysaccharide.

BACKGROUND/AIM: Human osteoblasts induced by inflammatory stimuli express an inducible nitric oxide synthase (iNOS). The aim of the present study was to test the hypothesis that Aggregatibacter actinomycetemcomitans lipopolysaccharide stimulates the production of nitric oxide (NO) by a human osteoblast-like cell line (HOS cells). METHODS: Cells were stimulated directly with A. actinomycetemcomitans lipopolysaccharide or pretreated with the following l-NIL (an iNOS inhibitor), anti-CD14, Toll-like receptor 2 (TLR2), or TLR4 antibody before stimulation with A. actinomycetemcomitans lipopolysaccharide. The role of the cyclic nucleotides was assessed by pretreating the cells with the following; ODQ (a guanylyl cyclase inhibitor); SQ22536 (an adenylyl cyclase inhibitor); db-cAMP (a cyclic adenosine monophosphate analog); br-cGMP (a cyclic guanosine monophosphate analog); forskolin (an adenylyl cyclase activator), IBMX [a non-specific phosphodiesterase (PDE) inhibitor], or KT5720 [a protein kinase A (PKA) inhibitor]. The cells were also preincubated with genistein [a protein tyrosine kinase (PTK) inhibitor], bisindolylmaleimide [a protein kinase C (PKC) inhibitor], BPB [a phospholipase A2 (PLA2) inhibitor], and NDGA (a lipoxygenase inhibitor). The iNOS activity and nitrite production in the cell cultures were determined spectrophotometrically. RESULTS: The results showed that A. actinomycetemcomitans lipopolysaccharide stimulated both iNOS activity and nitrite production by HOS cells; this was reduced by l-NIL, anti-CD14, or anti-TLR4 antibody, SQ22536, KT5720, genistein, bisindolylmaleimde, BPB, and NDGA, but was enhanced by db-cAMP, IBMX, and forskolin. CONCLUSION: These results therefore suggest that A. actinomycetemcomitans lipopolysaccharide may induce the production of NO by HOS cells via a CD14-TLR4 molecule complex, a cAMP-PKA pathway, as well as by a PTK, PKC, PLA2, and lipoxygenase-dependent mechanism.

Effect of exogenous nitric oxide on murine immune response induced by Aggregatibacter actinomycetemcomitans lipopolysaccharide.

BACKGROUND AND OBJECTIVE: Elevated nitric oxide (NO) has been associated with destructive periodontal disease. The aim of the present study was to test the hypothesis that exogenous NO may inhibit a protective immune response to Aggregatibacter actinomycetemcomitans lipopolysaccharide (LPS) in a murine model. MATERIAL AND METHODS: Mice of the BALB/c strain were sham immunized, immunized with A. actinomycetemcomitans LPS, treated with S-nitroso-N-acetyl penicillamine (SNAP; a NO donor) and immunized with A. actinomycetemcomitans LPS or treated with SNAP plus 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO) and immunized with A. actinomycetemcomitans LPS. All animals were then challenged subcutaneously with viable A. actinomycetemcomitans. The serum-specific immunoglobulin G (IgG) subclasses and both interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) as well as splenic inducible nitric oxide synthase (iNOS) activity before and after bacterial challenge were assessed. The diameter of skin lesions was determined. Groups of mice were treated with l-N(6)-(1-iminoethyl)-lysine (l-NIL), an iNOS inhibitor, or 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one (ODQ), a guanylyl cyclase inhibitor, prior to injections with SNAP and/or A. actinomycetemcomitans LPS, and the skin lesions were assessed. RESULTS: Treatment with SNAP increased the iNOS activity, suppressed both serum-specific IgG2a and IFN-gamma levels, and delayed the healing of the lesions. These SNAP-induced immune alterations were restored by treatment with carboxy-PTIO. Pretreatment with l-NIL resulted in partial healing, whereas pretreatment with ODQ induced a delayed healing of the lesions. CONCLUSION: The present study suggests that exogenous NO may suppress a protective T helper 1-like murine immune response to A. actinomycetemcomitans LPS by an endogenous NO-independent but a cyclic GMP-dependent mechanism.

Effect of exogenous nitric oxide on murine splenic immune response induced by Aggregatibacter actinomycetemcomitans lipopolysaccharide.

The aim of this study was to determine the effect of exogenous nitric oxide (NO) on the induction of murine splenic immune response to Aggregatibacter actinomycetemcomitans lipopolysaccharide (LPS) in vitro. BALB/c mice were immunized with A. actinomycetemcomitans LPS and a control group was sham-immunized. Spleen cells were obtained, cultured and stimulated with A. actinomycetemcomitans LPS with or without the presence of S-nitroso acetyl-penicillamine (SNAP), a NO donor, and carboxy-PTIO, an NO scavenger. Culture supernatants were assessed for inducible nitric oxide synthase (iNOS) activity, specific IgG subclass levels, and both IFN-gamma and IL-4 levels. The results showed that in A. actinomycetemcomitans LPS-stimulated cells, SNAP enhances iNOS activity but inhibits the levels of specific IgG2a and IFN-gamma suggesting a Th1 response. The effect of SNAP on these immune parameters was ablated by carboxy-PTIO. These results suggest that exogenous NO may suppress the Th1-like immune response of A. actinomycetemcomitans LPS-stimulated murine spleen cells.

Malocclusions in the koala (Phascolarctos cinereus).

Malocclusions are a misalignment or incorrect positioning of the teeth when the upper and lower jaws close. These are poorly described in the koala and can result in irregular mastication which can have lifelong effects on body condition and oral health. A total of 370 koalas from two populations in Queensland (295) and one in South Australia (75) were examined for malocclusions. The prevalence of malocclusions in South Australian free-ranging koalas, captive Queensland koalas and Queensland free-ranging koalas was 39% (44), 30% (29) and 22% (29) respectively. Four types of malocclusion were identified based on severity of misalignment of the incisor/canine region, types 1, 2, 3 and 4. Maxillary overbite measurements of the molariform teeth were determined and these anisognathic values were then used to describe malocclusions within familial relationships in captive colonies. Captive koalas with a malocclusion had narrower mandibular width that ranged between 0.5 and 1% less than the normal measurements. The specific malocclusions reported in this study affected individuals by leading to tooth rotation, mobility and erosion with inefficient mastication of food and vegetation compaction. These changes increased the oral cavity pathology, by placing animals at risk of periodontal disease. There was evidence of familial links to malocclusion types in captive animals. Therefore captive breeding recommendations should consider known koala malocclusion traits to minimise their effect on future generations.

Effect of inhibition of inducible nitric oxide synthase (iNOS) on the murine splenic immune response induced by Aggregatibacter (Actinobacillus) actinomycetemcomitans lipopolysaccharide.

Animal studies suggest that inducible nitric oxide synthase (iNOS) may be associated with destructive periodontal disease. l-N(6)-(1-Iminoethyl)-lysine (L-NIL) has been shown to inhibit iNOS in a selective manner, and hence the aim of the present study was to test the hypothesis that treatment with l-NIL may induce a T-cell helper 1 (Th1)-like immune response by Aggregatibacter (Actinobacillus) actinomycetemcomitans lipopolysaccharide (LPS)-stimulated murine spleen cells in vitro. BALB/c mice were either sham-immunized or immunized with A. actinomycetemcomitans LPS. Spleen cells were stimulated with A. actinomycetemcomitans LPS in the presence or absence of L-NIL. Nitric oxide (NO), iNOS activity, specific IgG subclass antibodies, interferon-gamma (IFN-gamma), and interleukin-4 (IL-4) levels and cell proliferation were determined. The results showed that treatment with L-NIL suppressed both NO production and iNOS activity but enhanced specific IgG2a, IFN-gamma levels, and increased cell proliferation following stimulation with A. actinomycetemcomitans LPS-stimulated cells. The results of the present study suggest that inhibition of iNOS activity by L-NIL may skew the A. actinomycetemcomitans LPS-stimulated murine splenic immune response towards the Th1-like immune profile in vitro.

An abattoir survey of equine dental abnormalities in Queensland, Australia.

OBJECTIVE: A cadaver study to estimate the prevalence of dental disorders in horses presented at an abattoir in Queensland, Australia. METHODS: Cadaver heads at a Queensland abattoir were examined for the presence of dental abnormalities and categorised into age groups. The prevalence of abnormalities was analysed by binomial observation of observed proportion, Pearson's Chi-square test or Fisher's exact correlation test. Strength of association was evaluated using Cramer's V test. RESULTS: Heads from horses (n=400) estimated to be between 1 and 30 years of age were placed into four age groups. The most common abnormalities were sharp enamel points (55.3%) and hooks (43%). The highest frequency of dental diseases and abnormalities were in horses 11-15 years old (97.5%). CONCLUSIONS: Common abnormalities were found in all groups and the prevalence increased with age. This study suggests that all horses should have regular complete dental examinations to detect and treat dental disorders in order to limit more severe dental pathologies later in life.

Microbiological and serological investigations of oral lesions in Papillon-Lefèvre syndrome.

Microbiological and serological (enzyme linked immunosorbent assay) investigations were carried out, including karyotyping, on two Asian children with Papillon-Lefèvre syndrome. In case 1, a girl aged four years, the most prevalent putative periodontopathogens were Eikenella corrodens, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia (deciduous dentition) and Bacteroides gracilis, E corrodens and F nucleatum (permanent dentition). In case 2, a boy aged nine years, they were F nucleatum, P intermedia and P loeschii and E corrodens. Serum from case 2 showed a raised specific IgG antibody response to Actinomyces actino-mycetemcomitans serotype b. Thus, a wider range of species than hitherto reported may be associated with Papillon-Lefèvre syndrome, including A actino-mycetemcomitans and F nucleatum.

A microbiological study of Papillon-Lefévre syndrome in two patients.

AIM: To analyse the microflora of subgingival plaque from patients with Papillon-Lefévre syndrome (PLS), which is a very rare disease characterised by palmar-plantar hyperkeratosis with precocious periodontal destruction. METHODS: Bacterial isolates were identified using a combination of commercial identification kits, traditional laboratory tests, and gas liquid chromatography. Some isolates were also subjected to partial 16S rDNA sequencing. Plaque samples were also assayed for the presence of Porphyromonas gingivalis, Prevotella intermedia, and Actinobacillus actinomycetemcomitans in a quantitative enzyme linked immunosorbent assay (ELISA) using monoclonal antibodies. RESULTS: The culture results showed that most isolates were capnophilic and facultatively anaerobic species-mainly Capnocytophaga spp and Streptococcus spp. The latter included S. constellatus, S. oralis, and S. sanguis. Other facultative bacteria belonged to the genera gemella, kingella, leuconostoc, and stomatococcus. The aerobic bacteria isolated were species of neisseria and bacillus. Anaerobic species included Prevotella intermedia, P. melaninogenica, and P. nigrescens, as well as Peptostreptococcus spp. ELISA detected P gingivalis in one patient in all sites sampled, whereas A. actinomycetemcomitans was detected in only one site from the other patient. Prevotella intermedia was present in low numbers. CONCLUSIONS: Patients with PLS have a very complex subgingival flora including recognised periodontal pathogens. However, no particular periodontopathogen is invariably associated with PLS.

A longitudinal study of Streptococcus mutans colonization in infants after tooth eruption.

We previously reported that, before tooth eruption, over one-half of infants aged 6 mos were already infected with Streptococcus mutans. The aim of this investigation was to determine the colonization of S. mutans after tooth eruption in the same cohort of 111 infants (35 pre-term, 76 full-term). Our results showed that S. mutans colonization increased with increasing age, so that by 24 mos of age, 84% harbored the bacteria (p < 0.01). The mean and median ages of S. mutans colonization in dentate infants were 15.7 mos and 16.0 mos, respectively. Factors associated with S. mutans colonization were sweetened fluids taken to bed (p < 0.01), frequent sugar exposure (p < 0.03) and snacking (p < 0.03), sharing of foods with adults (p < 0.03), and maternal S. mutans levels of > 10(5) CFU/mL (p < 0.02). In contrast, non-colonization of S. mutans was associated with toothbrushing (p < 0.03) and multiple courses of antibiotics (p < 0.001). Analysis of our data establishes the timing of S. mutans colonization in children from birth to 24 mos of age.

Oral colonization of Streptococcus mutans in six-month-old predentate infants.

We hypothesize that S. mutans colonization occurs more frequently in pre-term children due to their relative immaturity. In this study of 172 predentate, six-month-old infants, we found that 50% of pre-term and 60% of full-term children harbored S. mutans. The colonization was confirmed by repeat sampling. Although there were minor differences, factors associated with S. mutans infection in pre-term and full-term infants were generally similar. In both groups, increased frequency of sugar was ranked the most important factor (p < 0.001), followed by breast-feeding (p < 0.001), and habits which allowed saliva transfer from mother to infant (p < 0.01). By contrast, non-colonization of S. mutans was associated with multiple courses of antibiotics (p < 0.001). Compared with pre-term children, there were higher percentages of full-term who had night feedings and consumed sugar during sleep times. Mothers with infected infants had S. mutans levels > 5 x 10(5) CFU/mL saliva (p < 0.001), poorer oral hygiene, more periodontal disease, and lower socio-economic status (p < 0.02) and snacked frequently (p < 0.001), compared with mothers with non-infected infants.

The effects of interleukin-10 depletion in vivo on the immune response to Porphyromonas gingivalis in a murine model.

BACKGROUND: The immune response to Porphyromonas gingivalis in the mouse abscess model is known to be dependent upon CD4 T-cell activation and the regulatory role of cytokines. The role of interleukin-10 (IL-10) in this mouse model was examined in vivo. METHODS: One-week-old, female BALB/c mice were divided into 4 groups. Groups 1 and 2 were given intraperitoneal (i.p.) injections of phosphate buffered saline (PBS) weekly for 5 weeks. Group 3 was given an i.p. injection of rat immunoglobulin. Group 4 was injected with rat anti-IL-10 antibodies. At week 6, group 1 was sham-immunized with PBS, and groups 2, 3, and 4 were injected with P. gingivalis lipopolysaccharide (Pg-LPS) weekly for 2 weeks. One week after the final immunization, delayed-type hypersensitivity (DTH) was assessed by footpad swelling to Pg-LPS. The level of serum antibodies to Pg-LPS and IFN-gamma (IFN-gamma) was determined by enzyme-linked immunosorbent assay. Dorsal abscess formation induced by the injection of viable P. gingivalis was examined daily for 30 days. RESULTS: The footpad swelling of the anti-IL-10-treated group (group 4) was significantly higher than that of groups 1 to 3. Similarly, the serum IFN-gamma level in group 4 was much higher than that of the other experimental groups. There was no significant difference in serum IgG antibodies to Pg-LPS in any of the experimental groups. However, the level of IgM antibodies in group 4 mice was significantly lower than that in groups 2 and 3. In addition, serum IgG1 was suppressed in group 4 mice, while IgG2a antibodies were raised. However, there was no difference observed between the levels of IgG2b and IgG3 antibodies in any group of mice. The lesions in sham-immunized mice (group 1) persisted for 30 days, and those in group 2 and 3 were undetected by day 18 and 20, respectively. In sharp contrast, lesions in group 4 had healed completely by day 13. CONCLUSIONS: This study has shown that IL-10 depletion in vivo in P. gingivalis LPS-induced immune response in mice led to an elevated DTH response, an increase in serum IFN-gamma levels, and raised levels of IgG and IgG2a antibodies. Treatment with anti-IL-10 antibodies resulted in suppressed IgG1 and IgM responses and a more rapid healing of abscesses than in non-IL-10-depleted mice. These results suggest that IL-10 depletion in Pg-LPS-induced immune response in mice may lead to a Th1-like immune response and provide strong protection against a subsequent challenge with live P. gingivalis in an abscess model.

Cytokine profiles of lesional and splenic T cells in Porphyromonas gingivalis infection in a murine model.

T cell cytokine profiles in the spleens and Porphyromonas gingivalis-induced lesions of P. gingivalis-immunized mice were examined. BALB/c mice were immunized with P. gingivalis outer membrane (OM) antigens/mouse weekly for 3 weeks followed by challenge with live organisms 2 weeks after the final immunization. Control mice were immunized with PBS. Spleens were excised at 0 and 4 days and lesions at 1, 4, and 7 days after challenge. Splenic and lesional CD4 and CD8 cells were stained for intracytoplasmic interleukin (IL)-4, interferon (IFN)-gamma, and IL-10. More than 50% of the T cells in the spleens of immunized mice were IFN-gamma positive at day 0 which was significantly higher than for IL-4 or IL-10, these levels decreasing significantly 4 days after challenge. Less than 6% of the T cells in sham immunized mice were cytokine positive at day 0, although at day 4, there was a significant increase in the percent IL-10 positive CD4 cells and IL-4 and IL-10 positive CD8 cells. There were no differences in the percent IL-4, IFN-gamma, or IL-10 positive T cells in the lesions of immunized mice, but there was a dramatic decrease at day 7 to very low levels in control mice. In conclusion, the results of the present study show a predominant Th1 response in the spleens of BALB/c mice after immunization with P. gingivalis OM antigens, suggesting that a protective immune response to P. gingivalis may involve a strong IFN-gamma response.

Mucosal induction of systemic T cell tolerance by Fusobacterium nucleatum.

The present study was undertaken to determine if mucosal presentation of the periodontopathic bacterium Fusobacterium nucleatum could induce systemic tolerance. Two separate protocols of mucosal priming were carried out. In the first, mice were gastrically intubated on 2 consecutive days; this was repeated 5 days later. In the second protocol, mice were similarly primed but received another priming dose after a further 7 days. Positive control mice were similarly primed with sheep red blood cells (SRBC) while negative control animals were sham-primed with saline. Following mucosal priming, mice were systemically sensitized with the respective antigen and then subsequently challenged in the left hind footpad. The right footpad was challenged with saline and served as a negative control. Serum antibody levels were measured using enzyme-linked immunoabsorbent assay (ELISA) and haemagglutination assays. Mucosal priming with F. nucleatum was found to suppress the local delayed type hypersensitivity reaction as determined by footpad measurements. Sham-priming did not suppress the local response. On the other hand, the levels of serum antibodies were not influenced by mucosal priming. These results suggest that under the experimental conditions used, mucosal presentation of F. nucleatum can induce a degree of split tolerance in which T cell responses are suppressed while B cell responses remain intact. The implication of this finding to human periodontal disease is yet to be determined.

The influence of genetic variation on the splenic T cell cytokine and specific serum antibody responses to Porphyromonas gingivalis in mice.

BACKGROUND: T cell cytokine profiles in the spleens and anti-Porphyromonas gingivalis antibodies in the sera of P. gingivalis-immunized BALB/c (H-2d), CBA/CaH (H-2k), C57BL6 (H-2b), and DBA/2J (H-2d, C5 deficient) mice were examined. METHODS: Mice were immunized either by intraperitoneal injections of P. gingivalis outer membrane antigens and Freund's incomplete adjuvant weekly for 3 weeks or sham-immunized with PBS and adjuvant, followed by subcutaneous challenge with live organisms 1 week after the final immunization. Spleens were excised and blood samples collected by heart puncture at 0 and 7 days after challenge. Splenic CD4 and CD8 cells were stained for intracytoplasmic interleukin (IL)-4, interferon (IF)-gamma, and IL-10 and levels of anti-P. gingivalis antibodies in the serum samples determined by ELISA. RESULTS: Lesion sizes in immunized BALB/c mice remained stable for the 7-day experimental period. Immunized CBA/CaH and C57BL6 mice exhibited large lesions at day 1 reducing by day 7 particularly in the latter strain. Lesions in immunized DBA/2J mice were still larger than the other strains at day 7. With the exception of DBA/2J mice, sham-immunized mice demonstrated lesions which did not show signs of healing by day 7. T cell cytokine responses in sham-immunized mice at day 0 were low, increasing to a variable degree by day 7 after challenge in the 4 strains. Immunized BALB/c mice demonstrated intermediate T cell responses while generally exhibiting a stronger IFN-gamma response than IL-4 or IL-10. Immunized CBA/CaH and C57BL6 mice showed weak T cell cytokine responses while immunized DBA/2J displayed the strongest T cell responses particularly in regard to IL-4 positive cells. Sham-immunized mice had low levels of serum anti-P. gingivalis antibody levels at day 0 with levels increasing significantly by day 7 after challenge. Antibody levels in immunized mice seemed to correlate with lesion sizes. Immunized C57BL6 mice had the highest antibody levels followed by CBA/CaH, BALB/c with DBA/2J exhibiting low levels. The T cell and B cell antibody responses in each strain appeared to exhibit an inverse relationship. CONCLUSIONS: This study has shown that genetic differences at the level of H-2 haplotype induce variations in the local and T and B cell responses to P. gingivalis antigens. The responses of DBA/2J mice which have the same haplotype as BALB/c mice suggest that factors other than H-2 haplotype such as the C5 deficiency may influence this immune response. The significance of the specific antibody and T cell responses and of their inverse relationship to susceptibility to periodontal disease remains to be determined.

Protective immunity to Porphyromonas gingivalis infection in a murine model.

The mouse abscess model has been used extensively to demonstrate protection after challenge with periodontopathic organisms. In the present study, an outer membrane (OM) preparation of P. gingivalis ATCC 33277 was used to immunize BALB/c mice prior to challenge with live P. gingivalis organisms. This OM preparation, particularly at the highest dose level of 100 micrograms/immunization, was able to induce high levels of specific antibody and subsequent protective immunity. Protection in all immunized mice was noted by the rapid healing of the primary lesions, a low incidence of secondary lesions, and, in the highest dose group, an absence of septicemia. Non-immunized animals demonstrated a slower development as well as healing of primary lesions, with higher numbers and larger sizes of secondary lesions. Weight loss and behavior patterns such as hunched bodies, ruffled hair, and stiffness of the hind legs were particularly noted in this group. Depletion of CD4 T cells in mice prior to immunization with 100 micrograms P. gingivalis OM resulted in significantly depressed serum levels of anti-P. gingivalis antibody and an increase in the physical signs of disease compared with both the immunized and control groups. Western blot analysis demonstrated three antigen bands (63.3, 50.1, and 45.1) recognized by all immunized groups and also the control non-immunized group, although the latter recognition occurred only after challenge. A further antigen band of 36.1 kDa was recognized by sera from the highest dose group only. This study has demonstrated the ability of P. gingivalis OM to provide protection against challenge with live P. gingivalis organisms. The increased physical signs of disease seen in the CD4 depleted animals compared with the control group not only illustrate the protective role of serum antibody, but also suggest a possible role for T cell mechanisms in control of the lesion locally. The ability of specific OM antigens to provide similar protective immunity remains to be ascertained.

IgG antibody subclass response to Porphyromonas gingivalis outer membrane antigens in gingivitis and adult periodontitis.

Porphyromonas gingivalis is an important oral pathogen with a strong association with adult periodontitis. Significant titers of specific IgG antibodies to P. gingivalis can be found in the sera of both gingivitis and periodontitis patients. Since IgG subclasses have different biological characteristics, the present study dealt with the serum IgG subclass response to outer membrane antigens of P. gingivalis. Western blot analysis of P. gingivalis outer membrane was carried out using 20 adult periodontitis and 20 age- and sex-matched gingivitis patients. Antibodies in sera of both adult periodontitis and gingivitis patients recognized 38 antigen bands, ranging in molecular mass from 11.1 to 161 kDa. IgG2 was the predominant antibody subclass response in both patient groups in terms of the numbers of outer membrane antigens recognized, followed by IgG3, IgG1, and IgG4. More antigens in all IgG subclasses except IgG4 were recognized in adult periodontitis cases. Of the 23 antigens identified by IgG2 antibodies, 9 were recognized predominantly in adult periodontitis and 3 in the gingivitis group. In the IgG1 subclass, 4 antigens were recognized predominantly in the adult periodontitis group while only 1 antigen was recognized significantly more in the gingivitis group. The IgG3 response identified 14 antigens ranging in molecular mass from 11.1 to 61.2 kDa in both groups. Ten antigens were recognized significantly by the adult periodontitis group. The lowest response was seen by IgG4 antibodies, with only 3 antigens of molecular mass 61.2, 52.3, and 38.8 kDa recognized, the latter two significantly in the adult periodontitis group.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-1 and interleukin-1 inhibitor production by human adherent cells stimulated with periodontopathic bacteria.

This study examined the effect of the putative periodontopathic bacteria Bacteroides gingivalis and Fusobacterium nucleatum on the production of interleukin-1 (IL-1) and IL-1 inhibitors by human plastic-adherent mononuclear cells from normal donors. Fusobacterium mortiferum was used as a non-oral, non-pathogenic control organism. Unstimulated adherent cells spontaneously secreted an IL-1 inhibitor, whereas stimulation with B. gingivalis induced the synthesis and secretion of IL-1. With both fusobacteria IL-1 was present in the intracellular environment, whereas the predominant secretory product was either IL-1 or an IL-1 inhibitor. These results suggest that bacteria are capable of modulating cytokine production by monocytes and may thereby alter the local immune response.