Probiotics enhance alveolar bone microarchitecture, intestinal morphology and estradiol levels in osteoporotic animals.

BACKGROUND AND OBJECTIVE: Osteoporosis is associated with bone microarchitecture alterations, and the depletion of estrogen during menopause is a major contributing factor to its development. The literature highlights the noteworthy role of gut microbiota in bone metabolism, particularly in the progression of osteoporosis. Periodontal disease leads to alveolar bone loss, which may be influenced by estrogen deficiency, and this mechanism is intricately associated with an imbalance in systemic microbiota. The aim of this study was to evaluate the effects of Bifidobacterium animalis subsp. lactis HN019 (B. lactis HN019) and Lacticaseibacillus casei 01 (L. casei 01) administrations on an osteoporosis animal model. MATERIALS AND METHODS: Thirty-three female rats were randomly divided into three groups: control (C-OVX), C-OVX-HN019 and C-OVX-LC01. All animals were ovariectomized. In groups C-OVX-HN019 and C-OVX-LC01, the probiotics were administered for 4 months. All animals were euthanized after 16 weeks from ovariectomy. Microtomographic, histopathological and immunohistochemical examinations were conducted on periodontal tissues, whereas histomorphometry, histopathological and immunohistochemical analyses were carried out on the intestine. The levels of estradiol were assessed in blood using an immunoenzymatic assay. The data were subjected to statistical analyses (p < .05). RESULTS: The C-OVX-LC01 group exhibited a significant reduction in alveolar bone porosity and an increase in connective tissue density compared to C-OVX (p < .05). The C-OVX-HN019 and C-OVX-LC01 groups presented reduced expression of TRAP and RANKL compared to the C-OVX (p < .05). The C-OVX group presented villi defects, mild neutrophil infiltration, decrease in both villous height and intestinal crypts and reduced expression of intestinal junctional epithelium markers e-cadherin and claudin 01 compared to C-OVX-HN019 and C-OVX-LC01 (p < .05). The C-OVX group had lower estradiol levels than C-OVX-HN019 and C-OVX-LC01 (p < .05). CONCLUSION: The probiotic therapy promoted a reduction in alveolar bone destruction and intestinal permeability as well as an increase in estradiol levels in ovariectomized rats. Specifically, the probiotic strain Lacticaseibacillus casei 01 exhibited greater effectiveness compared to Bifidobacterium animalis subsp. lactis HN019, indicating strain-dependent outcomes.

NOD2 contributes to Porphyromonas gingivalis-induced bone resorption.

The NOD-like receptors are cytoplasmic proteins that sense microbial by-products released by invasive bacteria. Although NOD1 and NOD2 are functionally expressed in cells from oral tissues and play a role triggering immune responses, the role of NOD2 receptor in the bone resorption and in the modulation of osteoclastogenesis is still unclear. We show that in an experimental model of periodontitis with Porphyromonas gingivalis W83, NOD2(-/-) mice showed lower bone resorption when compared to wild type. Quantitative polymerase chain reaction analysis revealed that wild-type infected mice showed an elevated RANKL/OPG ratio when compared to NOD2(-/-) infected mice. Moreover, the expression of 2 osteoclast activity markers-cathepsin K and matrix metalloproteinase 9-was significantly lower in gingival tissue from NOD2(-/-) infected mice compared to WT infected ones. The in vitro study reported an increase in the expression of the NOD2 receptor 24 hr after stimulation of hematopoietic bone marrow cells with M-CSF and RANKL. We also evaluated the effect of direct activation of NOD2 receptor on osteoclastogenesis, by the activation of this receptor in preosteoclasts culture, with different concentrations of muramyl dipeptide. The results show no difference in the number of TRAP-positive cells. Although it did not alter the osteoclasts differentiation, the activation of NOD2 receptor led to a significant increase of cathepsin K expression. We confirm that this enzyme was active, since the osteoclasts resorption capacity was enhanced by muramyl dipeptide stimulation, evaluated in osteoassay plate. These results show that the lack of NOD2 receptor impairs the bone resorption, suggesting that NOD2 receptor could contribute to the progression of bone resorption in experimental model of periodontitis. The stimulation of NOD2 by its agonist, muramyl dipeptide, did not affect osteoclastogenesis, but it does favor the bone resorption capacity identified by increased osteoclast activity.

Antimicrobial activity of copaíba (Copaifera langsdorffii) oleoresin on bacteria of clinical significance in cutaneous wounds / Atividade antimicrobiana do óleo-resina de copaíba (Copaifera langsdorffii) em bactérias de significância clínica em úlceras cutâneas

The present study was designed to evaluate the in vitro antimicrobial activity of Copaifera langsdorffii oleoresin, which has been used in folk medicine as an anti-inflammatory, antibacterial, healing among others. The oleoresin was tested against Gram-positive (Staphylococcus aureus, Streptococcus pyogenes, Enterococcus faecalis) and Gram-negative (Pseudomonas aeruginosa and Escherichia coli) bacteria related to infections in cutaneous wounds. Antimicrobial activity was determined by the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assays. Copaiba oleoresin showed antimicrobial activity only against the Gram-positive bacteria with MIC of 200 µg/mL, 400 µg/mL and 1100 µg/mL for S. aureus, S. pyogenes and E. faecalis, respectively. MBC values were the same as MIC for S. aureus and S. pyogenes and for E. faecalis it was 1200 µg/mL. Considering that infection significantly impairs the wound healing process, we believe that the use of copaiba oleoresin as a component of a topical formulation could be a valuable adjunct in the treatment of infected wounds, mainly in the case of wounds infected by Gram-positive microorganisms.

Este trabalho avaliou a atividade antimicrobiana in vitro do óleo-resina da Copaifera langsdorffii, o qual vem sendo utilizado há muitos anos na medicina tradicional popular, principalmente devido às suas propriedades antiinflamatórias, antibacterianas, cicatrizante entre outras. O óleo-resina foi testado em bactérias Gram-positivas (Staphylococcus aureus, Streptococcus pyogenes, Enterococcus faecalis) e Gram-negativas (Pseudomonas aeruginosa e Escherichia coli) relacionadas com infecções de úlceras cutâneas. A atividade antimicrobiana foi determinada pelos testes da Concentração Inibitória Mínima (CIM) e Concentração Bactericida Mínima (CBM). O óleo-resina apresentou atividade antimicrobiana in vitro apenas para as bactérias Gram-positivas, com valores de CIM de 200 µg/mL, 400 µg/mL e 1100 µg/mL para S. aureus, S. pyogenes e E. faecalis, respectivamente. Os valores de CBM foram os mesmos que os valores de MIC para S. aureus e S. pyogenes. O valor de CBM para o microrganismo E. faecalis foi de 1200 µg/mL. Considerando que a presença de infecção significativamente impede o processo normal de cicatrização de úlceras cutâneas, acreditamos que o óleo-resina de copaíba, utilizado como componente de formulações tópicas, poderia ser um adjunto importante no tratamento de úlceras cutâneas infectadas, principalmente nos casos de infecção por microrganismos Gram-positivos.

Halitosis and periodontal disease in subjects with mental disabilities.

OBJECTIVE: The aim of this study was to investigate the relationship between halitosis, presence of N-benzoyl-DL-arginine-2-naphthylamide (BANA) positive activity (indicative of Treponema denticola, Porphyromonas gingivalis, Tannerella forsythensis) and periodontal conditions in subjects with mental disabilities. MATERIALS AND METHODS: The population consisted of 17 Down syndrome (DS), 17 mentally retarded (MR) and 17 mentally healthy subjects (MH) - control group. A portable sulfide monitor was used to measure the concentration of volatile sulfur compounds (VSC) found in the mouth. Clinical parameters [Plaque Index (PlI), Probing Depth (PD), Bleeding on Probing (BOP)] were obtained from six reference teeth. Subgingival plaque samples for BANA Test were taken from the same six teeth and dorsum of the tongue. RESULTS: PlI and BOP were higher in DS than in MR and MH, respectively (P < 0.05). Mean PD was similar between DS and healthy groups but it was higher than the MR group. No significant differences were found among the three groups regarding the presence of BANA positive species, however the VSC levels were significantly lower in DS (97.23 ppb) than in MR (203.23 ppb) and MH individuals (180.53 ppb). CONCLUSIONS: Even though the presence of BANA positive activity was similar among DS, MR and MH subjects, halitosis seemed to be lower in the DS group.

Subgingival utilization of a 1% chlorhexidine collagen gel for the treatment of periodontal pockets. A clinical and microbiological study.

This study evaluates the effect of subgingival irrigation with a 1% chlorhexidine collagen gel in periodontal pockets as an adjunct procedure to scaling and root planing (SRP). Thirty-seven sites with probing depth (PD) of 5-7 mm and BANA positive in 6 patients with chronic periodontal disease were selected. Sites were assigned to different treatment groups consisting of SRP only (group 1), SRP + irrigation with collagen gel (group 2), or SRP + irrigation with collagen gel containing 1% chlorhexidine (group 3). Subgingival irrigation was performed after initial SRP and at 7, 14 and 21 days. Clinical measurements including PD, plaque index (PI), gingival index (GI), gingival recession (GI), bleeding on probing (BOP) and clinical attachment level (CAL) were performed at the selected sites at baseline, 60 and 90 days and the BANA test was performed on plaque samples from the same sites at baseline and 90 days. There was an improvement in clinical parameters in all groups with a significantly greater decrease in GI and bleeding in the chlorhexidine group. There was a greater reduction of BANA positive sites in groups 2 and 3. The authors concluded that 1% chlorhexidine collagen gel is a promising adjunct to SRP in the treatment of adult periodontitis.

[Effect of sodium fluoride mouth rinses containing xylitol and sorbitol on the number of Streptococcus mutans from human saliva]. / Efeito de soluções fluoretadas contendo xilitol e sorbitol no número de estreptococos do grupo mutans na saliva de seres humanos.

The objective of this study was to assess the effect of 0.05% sodium fluoride solutions containing 2.5% or 12.5% xylitol on the number of Streptococcus mutans in the human mouth. Fifty boys between 8 and 16 years of age participated in this double-blind crossover study. Of the original 50 boys, 33 finished the study. Participants were randomly divided into four groups. The following solutions were employed: placebo solution; 0.05% sodium fluoride solution; 0.05% sodium fluoride + 2.5% xylitol + 2% sorbitol; 0.05% sodium fluoride + 12.5% xylitol + 2% sorbitol. Each solution was used for a 28-day period (20 mL/day, twice a day), with a 10-day washout period between solutions. There were no significant differences (P = 0.32) between the two xylitol-containing solutions (2.5% vs. 12.5%) concerning the number of Streptococcus mutans. However, there was a significant difference between these two xylitol-containing solutions and the sodium fluoride and placebo solutions (P < 0.001). Our results suggest that the 0.05% sodium fluoride solutions containing either 2.5% or 12.5% xylitol caused a significant reduction in the number of Streptococcus mutans.

Relationship between the presence or absence of gingival bleeding and the enzymatic BANA test.

When periodontal disease started to be considered a bacterial infection mainly mediated by subgingival plaque, the basic problem faced by periodontists was the identification and/or quantification of periodontopathogenic bacteria. However, clinical methods continue to be of great value for the diagnosis of periodontal disease. In the present study we show a significant correlation between an index widely used in clinical practice, the Gingival Index of Löe (1967), based on the presence or absence of bleeding on probing, and the methodology of the BANA test for the detection of the specific enzymatic activity of microorganisms involved in periodontal disease. More sensitive and specific clinical parameters, taken together with other microbiologic methods, will be useful in daily clinical practice even before periodontal treatment.

The relationship between place BANA reactivity and clinical parameters in subjects with mental disabilities.

The purpose of the present investigation was to determine whether subjects institutionalized with mental retardation have a relationship between periodontal clinical parameters and the presence of the BANA-positive periodontal pathogens Porphyromonas gingivalls, Treponema denticola, and Bacteroides forsythus in their subgingival plaques. Fifty institutionalized subjects (25 patients with Down syndrome and 25 subjects with mental retardation) were matched with respect to age and sex. Periodontal clinical parameters (Bleeding on Probing, BOP; Papillary Bleeding Score, PBS; and Probing Depth, PD) were obtained from 6 reference teeth (3, 8, 14, 19, 24, 30). In addition, subgingival plaque samples taken from the same 6 teeth were analyzed for the presence of the BANA-positive species, by means of the chairside BANA test. In both the patients with Down syndrome and the group with mental retardation, the presence of BANA-positive plaques was significantly associated with bleeding on probing (p < 0.05) and increased probing depth (p < 0.01, Chi-square). Analysis of these data indicated that the BANA test could be used in combination with clinical criteria to diagnose a periodontopathy anaerobic infection in institutionalized subjects.

Correlation between the CPITN score and anaerobic periodontal infections assessed by BANA assay.

In the present investigation the ability of subgingival plaque to hydrolyze BANA (Perioscan) was correlated with CPITN scores. Among 281 sites investigated, 136 had a CPITN equal to 2 with a highly significant positive BANA value (107 sites). A CPITN equal to 3 was also significantly BANA positive (90 sites). These findings clearly demonstrate the relationship between CPITN and anaerobic microorganisms (BANA positive).

Relationship between clinical probing depth and reactivity to the BANA test of samples of subgingival microbiota from patients with periodontitis.

The enzymatic test BANA (N-benzoyl-DL-arginine-naphthylamide) was used to analyze the subgingival microbiota of 28 patients aged 26 to 55 years old with a diagnosis of adult periodontitis. Samples were collected with periodontal curettes at 513 sites, with a mean number of sites of 18.3 +/- 8.8 per patient. The results of the BANA test were correlated with the initial measurements of pocket depth. The data showed a statistically significant correlation between increasing probing depth and a positive BANA test. BANA test detected the presence of BANA-positive microorganisms at sites of < 3 mm probing depth in a statistically significant proportion. Negative (BANA 1) and weakly positive (BANA 2) BANA tests were inversely correlated with increasing pocket depth, and positive (BANA 3) BANA tests were directly correlated with increasing pocket depth. On the basis of the presents results, we consider the BANA test to be of practical applicability in periodontal clinical practice and to represent an important auxiliary diagnostic tool for patients with adult periodontitis.

Similarities of periodontal clinical and microbiological parameters in mother-child pairs.

The aim of this study was to verify the association between children and their mothers (N = 28) for periodontal clinical and microbiological measures. Periodontal clinical parameters (probing depth and bleeding on probing) were obtained from six reference teeth from each mother-child pair. In addition, subgingival plaque samples taken from the same reference teeth were collected and placed on the Perioscan test for the detection of Treponema denticola, Porphyromonas gingivalis and Bacteroides forsythus. There were statistically significant differences between children and mothers regarding probing depth; there was an overall tendency for mothers to exhibit average probing depths greater than their children. In contrast, if a child had a bleeding site, the respective site on the mother usually also bled on probing, implying that there was an association in terms of bleeding between the mother-child pairs. Similar findings were observed for the Perioscan test. It is concluded that the occurrence of bleeding and periodontal anaerobic infections (as determined by the Perioscan test) were similar in reference teeth of mother-child pairs. These data suggest that parents with periodontal disease may serve as a reservoir of periodontopathic organisms for their children.

Antimicrobial action of AXOL on periodontopathic bacteria.

The antimicrobial action of AXOL was tested against a panel of periodontopathic bacteria, which included Treponema denticola, Treponema vincentii, Treponema sp., Porphyromonas gingivalis, Prevotella intermedia, Prevotella melaninogenica, and Fusobacterium nucleatum. The AXOL commercial solution (undiluted) was effective in inhibiting some of the bacteria but not all. The rational for the use of antimicrobials is discussed.

Enzyme profiles of oral spirochetes in RapID-ANA system.

Enzyme profiles of oral Treponema species were determined by using RapID-ANA (Innovative Diagnostic System, Atlanta, Ga.), a 4-h test system which detects 18 enzymatic reactions, including aminopeptidases and glycosidases. Seventy-two clinical isolates of Treponema denticola, four reference strains of T. denticola (ATCC 35404, ATCC 35405, ATCC 35520, and ATCC 33521), one strain of T. vincentii (ATCC 35580), and two strains of T. socranskii subspecies (T. socranskii subsp. buccale ATCC 35534 and T. socranskii subsp. socranskii ATCC 35536) were used in this study. All T. denticola strains produced indole and a variety of aminopeptidases and glycosidases. These organisms could be differentiated into two groups on the basis of tetrazolium reductase and serine, phenylalanine, and glycine aminopeptidase activities. T. vincentii produced N-acetylglucosaminidase and arginine aminopeptidase, which facilitated the differentiation of this organism from T. socranskii subspecies and the T. denticola group. T. socranskii subspecies gave positive reactions for alkaline phosphatase only. These findings suggest that the RapID-ANA system is useful for enzymatic characterization and differentiation of oral spirochetes.

Comparison of three dispersion procedures for quantitative recovery of cultivable species of subgingival spirochetes.

Spirochetes are usually the predominant organisms observed microscopically in subgingival plaques removed from tooth sites associated with periodontitis, but these organisms are rarely isolated by cultural means, presumably because the media do not support their growth and/or because these fragile organisms are disrupted by the various procedures used to disperse plaque samples. In the present investigation, three dispersal procedures, sonification, mechanical mixing, and homogenization, were compared for their ability to permit the isolation of Treponema denticola, Treponema vincentii, Treponema socranskii, and Treponema pectinovorum from plaque samples on media that support the growth of these species. Plaque samples in which the spirochetes averaged 50% of the microscopic count were chosen. The highest viable recoveries of spirochetes were observed when the plaques were dispersed with a Tekmar homogenizer, and the lowest occurred with sonification. The highest recoveries averaged only about 1% of the total cultivable counts, indicating either that the sought-after species were minor members of the flora or that the dispersal procedures were still too harsh. A total of 91% of the isolates were T. denticola, 5% were T. socranskii, and 4% were a heretofore-undescribed species.