Oral Microbiota Transplant in Dogs with Naturally Occurring Periodontitis.

In periodontitis patients, dysbiosis of the oral microbiota is not only found at clinically diseased periodontal sites but also at clinically healthy periodontal sites, buccal mucosae, tongue, and saliva. The present study evaluated the safety and efficacy of an oral microbiota transplant (OMT) for the treatment of periodontitis in dogs. Eighteen systemically healthy beagle dogs with naturally occurring periodontitis were enrolled in the study and randomly assigned to a test or control group. A 4-y-old, periodontally healthy female beagle dog served as a universal OMT donor. To reduce periodontal inflammation, all dogs received full-mouth mechanical debridement of teeth and mucosae 2 wk before baseline. At baseline, full-mouth mechanical debridement was repeated and followed by adjunctive subgingival and oral irrigation with 0.1% NaOCl. Subsequently, test dogs were inoculated with an OMT from the healthy donor. No daily oral hygiene was performed after OMT transplantation. Adverse events were assessed throughout the observation period. Clinical examinations were performed and whole-mouth oral microbiota samples were collected at week 2, baseline, week 2, and week 12. The composition of oral microbiota samples was analyzed using high-throughput 16S ribosomal RNA gene amplicon sequencing followed by taxonomic assignment and downstream bioinformatic and statistical analyses. Results demonstrated that the intergroup difference in the primary outcome measure, probing pocket depth at week 12, was statistically insignificant. However, the single adjunctive OMT had an additional effect on the oral microbiota composition compared to the full-mouth mechanical and antimicrobial debridement alone. The OMT resulted in an "ecological shift" toward the composition of the donor microbiota, but this was transient in nature and was not observed at week 12. No local or systemic adverse events were observed throughout the study period. The results indicate that OMT may modulate the microbiota composition in dogs with naturally occurring periodontitis and can be applied safely.

Inhibition of in vitro adhesion and virulence of Porphyromonas gingivalis by aqueous extract and polysaccharides from Rhododendron ferrugineum L. A new way for prophylaxis of periodontitis?

The effect of an aqueous extract from the leaves of Rhododendron ferrugineum (RF) was investigated for its capacity of inhibiting the adhesion of Porphyromonas gingivalis cells to epithelial buccal KB cells. RF was characterized by HPLC (12.1% taxifolin-3-O-ß-l-arabinopyranoside, 1.6% hyperoside, 0.9% isoquercitrin, 1.6% chlorogenic acid and a tannin content of 8.7%). Additionally raw polysaccharides (RPS) were obtained from the leaves of R. ferrugineum by aqueous extraction. RF and RPS interacted in a dose-dependent manner (max. 25% reduction at 1mg/ml each) with the adhesion of P. gingivalis by influencing bacterial outer membrane proteins. On protein level a time- and concentration-dependent inhibition of Arg-gingipain activity by RF was observed, while the Lys-gingipain activity remained unaltered. In addition, RF and RPS inhibited the bacterial hemagglutinin. RF affected the P. gingivalis adhesion also by interacting with KB cells in pre-incubation assays of the eukaryotic host cells, leading to reduced bacterial adhesion of about 75%. Gene expression analysis by RT-PCR indicated significant downregulation for arginine-specific gingipain rgpA by RF, while lysin-specific gingipain kgp and fimbrillinA fimA were strongly upregulated. Moreover, pre-incubation with RF abolished the P. gingivalis induced expression of IL-1ß, IL-6, IL-8 and TNFα in KB cells. Results of this study indicate that an aqueous extract from R. ferrugineum combines cytoprotective and antimicrobial effects by both downregulating the expression of pro-inflammatory genes and inhibiting the adhesion of P. gingivalis. Thus RF may be potential candidate for the development of an adjunctive antimicrobial approach in the prevention of periodontal diseases.

Severity of MIH findings at tooth surface level among German school children.

AIM: This study was to investigate the distribution and clinical characteristics of teeth diagnosed with MIH at surface and defect type level in a cohort of German children. METHODS: The study cohort included 242 children diagnosed with MIH which had been recorded during the compulsory dental school examinations of 20 German primary schools. The subjects had been enrolled by cluster sampling. All children attended the second to fourth grade (age 7-10 years, mean 8.1 ± 0.8). The children were examined by five calibrated examiners (kappa = 0.9) after tooth brushing. The recording comprised teeth, surfaces, type and severity of MIH defects and was conducted using a portable light, mirrors and cotton rolls. MIH was registered according to the EAPD criteria. Defects <1 mm were not recorded. Statistical analysis included descriptive statistics and Spearman's correlation. RESULTS: Most affected teeth were first permanent molars (71.4 %) followed by the maxillary central incisors (15.6 %). The most common defects were demarcated opacities (82.2 %), while the remaining 17.8 % of the affected teeth exhibited severe enamel defects. The most frequently affected surface in molars was the occlusal surface (72.4 %); in incisors, it was the buccal surface (73.5 %). There were no atypical restorations in the affected incisors. Different types of MIH defects at various surfaces of the same tooth were common. The number of affected tooth surfaces was positively correlated with the severity of MIH at child (p < 0.001). CONCLUSIONS: The study demonstrates severe enamel defects involving in almost one-fifth of all MIH teeth. The knowledge of the intra-oral distribution and severity of MIH findings at the enamel surface level is important for assessing the treatment needs.

[Periodontal diseases--a review on the association between maternal periodontitis and adverse pregnancy outcome]. / Parodontale Erkrankungen--Eine Übersicht über die Beziehung mütterlicher Parodontitis und negativer Schwangerschaftsergebnisse.

Several prospective clinical trials have indicated an association between maternal periodontal status and adverse pregnancy outcome, e.g., low birth weight, pre-term birth and pre-eclampsia. However, the translation of these findings into clinical care and decision making is still a matter of debate. Gynecologists and obstetricians are usually not very familiar with periodontal diseases and do not always consider this pathology in routine preconception counselling. This article outlines the clinical pictures of the most common periodontal diseases and thus helps the gynecologists to identify patients with periodontal diseases.

Detection of nine microorganisms from the initial carious root lesions using a TaqMan-based real-time PCR.

OBJECTIVE: The purpose of this study was to quantify nine selected cariogenic bacteria in plaque from sound root surfaces and initial carious root lesions using TaqMan PCR and to analyse a putative dependence on the kind of initial periodontal treatment. MATERIAL AND METHODS: Fifty-four subjects with generalized chronic periodontitis were randomly allocated to one of the following initial periodontal therapies: full-mouth disinfection, full-mouth scaling and root planing or scaling and root planing within 7 days. Plaque samples were taken before and after periodontal treatment and analysed by TaqMan PCR. RESULTS: The quantity of the cariogenic bacteria Actinomyces spp., Streptococcus mutans, Streptococcus sobrinus, Lactobacilllus spp., Rothia dentocariosa, Parvimonas micra, Propionibacterium acnes and Neisseria mucosa were significantly higher, while the quantity of Veillonella parvula was significantly lower on initial carious lesions than on the sound surfaces both before and after periodontal therapy. No significant differences could be found in any of the tested bacteria except P. micra on initial carious lesions and sound surfaces for both examinations between the groups. CONCLUSION: All the nine species analysed were found to be present in initial carious root lesions as well as sound root surfaces but in different quantities, independent of the different periodontal therapies.

Irreversible inflammation is associated with decreased levels of the alpha1-, beta1-, and alpha2-subunits of sGC in human odontoblasts.

The nitric oxide (NO) receptor enzyme soluble guanylate cyclase (sGC) contains one prosthetic heme group as an αß heterodimer, and two heterodimer isoforms (α(1)ß(1), α(2)ß(1)) were characterized to have enzyme activity. To test the irreversible inflammation-dependent regulation of sGC in odontoblasts, we incubated decalcified frozen sections of healthy and inflamed human third molars with antibodies against ß-actin, nitrotyrosine, inducible nitric oxide synthase (iNOS), α(1)-, ß(1)-, and α(2)-subunits of sGC and analyzed them at protein levels by quantitative immunohistochemistry. The irreversible inflammation induced an increase in the signal intensities for nitrotyrosine and iNOS and a decrease for the α(1)-, ß(1)-, and α(2)-subunits of sGC in odontoblasts. Inflammatory mediators, reactive oxygen, and nitrogen species may impair the expression of the α(1)-, ß(1)-, and α(2)-subunits in odontoblasts. The decrease of sGC at the protein level in inflamed odontoblasts is compatible with a critical role for sGC to mediate biological effects of NO in health.

Sequence variations in rgpA and rgpB of Porphyromonas gingivalis in periodontitis.

OBJECTIVE: The aim of the present study was to determine sequence variations in the active centre of the Arg-X-specific protease encoding genes rgpA and rgpB of clinical Porphyromonas gingivalis isolates and to analyse their prevalence in periodontitis patients before and 3 months after mechanical periodontal therapy. BACKGROUND: Genetic diversity at nucleotides 281, 283, 286 and 331 has been shown to result in amino acid substitutions in the catalytic domain of RgpA and RgpB that affect the substrate specificity and thus may influence the efficacy of Arg-X-protease specific inhibitors. METHODS: Sequence analysis of rgpA and rgpB genes in clinical P. gingivalis strains isolated from subgingival plaque samples of 82 periodontitis patients before and 3 months after mechanical supra- and subgingival debridement was performed. RESULTS: No specific variation within the rgpA sequence was observed. However, the rgpB sequence in the region of the active centre showed five different rgpB genotypes, which were named NYPN, NSSN, NSSK, NYPK and DYPN according to the derived amino acid substitution. Porphyromonas gingivalis genotype NYPN was detected in 27 patients (32.9%) before and in 8 patients (9.8%) after therapy, NSSN in 26 (31.7%) and 10 (12.2%), NSSK in 22 (26.8%) and 2 (2.4%), NYPK in 5 (6.2%) and 1 (1.2%), and DYPN in 1 patient (1.2%) and 0 patients (0%), respectively. Only one patient (1.2%) harboured two P. gingivalis rgpB genotypes (NSSK/NYPN) before treatment; these were no longer detected after therapy. CONCLUSION: The results indicate that five rgpB genotypes are maintained in natural populations of P. gingivalis. These data may be of importance with regard to the development of specific rgpB inhibitors.

Sequence analysis of kgp in Porphyromonas gingivalis isolates from periodontitis patients.

The aim of the present study was to determine sequence variation in the Lys-x-specific protease (Kgp) encoding gene kgp of Porphyromonas gingivalis and to analyze its association with periodontal disease severity. Pooled subgingival plaque samples were obtained from the six most severely affected sites of 102 patients with periodontitis. Sequence analysis of the kgp gene in 23 clinical P. gingivalis isolates resulted in the identification of two distinct kgp types (kgp-I and kgp-II) according to sequence differences in the region encoding the catalytic domain. Restriction analysis revealed that 59 of the 102 patients were colonized by kgp-I and 43 by kgp-II. Patients harboring kgp-I or kgp-II showed no significant difference in the severity of periodontal disease as assessed by pocket probing depth and bleeding on probing following adjustment for smoking habit and age. Moreover, no differences in proteolytic activity of Kgp-I and Kgp-II were detected. The results indicated that two kgp types are maintained in natural populations of P. gingivalis.

Serum antibody reactivity against recombinant PrtC of Porphyromonas gingivalis following periodontal therapy.

OBJECTIVES: In 34 patients with chronic periodontitis, the presence of IgA, IgG, and IgG subclass serum antibodies against recombinant PrtC (rPrtC) of Porphyromonas gingivalis was assessed by immunoblot analysis 24 months after therapy. METHODS: rPrtC was produced from P. gingivalis ATTC 33277 using the plasmid pGEX-2T. In addition, intraoral colonization with P. gingivalis was detected by PCR in subgingival plaque and swab samples from buccal mucosae, tonsils and tongue at baseline, 10 d, and 3, 6, 9, 12, 18, and 24 months. RESULTS: All patients were found to harbor P. gingivalis in the oral cavity at least once during the observation period. The identified antibody responses against the rPrtC of P. gingivalis were IgA (97%, i.e. 33/34 patients) and IgG (100%, i.e. 34/34), with an IgG subclass distribution of IgG2 (65%, i.e. 22/34 patients) > IgG3 (47%, i.e. 16/34) > IgG1 (38%, i.e. 13/34) > IgG4 (29%, i.e. 10/34). Anti-rPrtC IgA and IgG antibody reactivity was found in all but one patients (anti-rPrtC IgA negative), who tested negative for P. gingivalis at all of the assessed intraoral sites for at least 6 months before sera collection. There was no association between IgG subclass reactivity against the rPrtC of P. gingivalis and progression of periodontal attachment loss. CONCLUSION: The results indicated that anti-rPrtC IgA and IgG antibodies may serve as an indicator for past or present intraoral colonization with P. gingivalis.

Eruption cyst formation associated with cyclosporin A.

BACKGROUND: Cyclosporin A (CyA) is a potent immunomodulatory agent with a wide range of applications. Despite its therapeutic value, multiple adverse effects of CyA have been identified. This case report describes eruption cyst formation as a possible adverse effect of CyA administration during tooth eruption in a boy treated with CyA as a consequence of a cardiac transplantation. The clinical diagnosis of eruption cyst was confirmed by histopathological examination. TREATMENT: The periodontal treatment consisted of supragingival and subgingival scaling, followed by surgical removal of the tissues overlying the crowns of the teeth associated with eruption cysts, and flap surgery in the region of gingival overgrowth. The patient was then placed on quarterly periodontal supportive therapy and his immunosuppressive medication was switched from CyA to tacrolimus. RESULTS: Twenty months after therapy, neither new cyst formation nor recurrence of gingival overgrowth was registered. CONCLUSION: Formation of an eruption cyst may be an adverse effect of CyA in children with erupting teeth.

In-dental-office screening for diabetes mellitus using gingival crevicular blood.

BACKGROUND: Diabetes mellitus (DM) is undiagnosed in approximately 1/2 of the patients actually suffering from the disease. In addition, the prevalence of DM is more than 2x as high in patients with periodontitis when compared to periodontally healthy subjects. Thus, a high number of patients with periodontitis may have undiagnosed DM. AIM: The purpose of this pilot study was to evaluate, whether blood oozing from gingival tissues during routine periodontal examination can be used for determining glucose levels. 32 non-diabetic and 13 diabetic patients with moderate to severe periodontitis were enrolled and subjected to routine clinical periodontal examination. Periodontal pocket probing was performed using a standard force. Blood oozing from gingival tissues of anterior teeth following periodontal pocket probing was collected with the stick of a glucose self-monitoring device (Elite(R) 2000, Bayer Diagnostics GmbH, Munich). As control, fingerstick capillary blood was taken. Statistical analysis was performed by Pearson's correlation coefficient. RESULTS: The patient blood glucose levels ranged from 3.57 mmol/l to 18.01 mmol/l and the values of blood samples taken from gingiva or finger tip showed a very high intrapatient correlation (r=0.98; p<0.0001). CONCLUSION: The results suggested that blood oozing during routine periodontal examination may be used for diabetes mellitus screening in a dental office setting.

Interleukin-3 and interleukin-6 stimulate bovine adrenal cortisol secretion through different pathways.

Several interleukins have been reported to play a major role in the regulation of steroid secretion at all three levels of the hypothalamic-pituitary-adrenal axis. The objective of this study was to investigate the effect of interleukin-3 (IL-3) and interleukin-6 (IL-6) on cortisol secretion of bovine adrenocortical cells in primary culture under serum-free conditions. Both IL-3 and IL-6 stimulated basal cortisol secretion dose-dependently to a similar extent at a similar time course. After incubation with IL-3 or IL-6 at concentrations of 100 microg/l, a maximum 4.1-fold increase of the cortisol secretion was reached after 12 h (P<0.01). Coincubation of IL-3 and IL-6 (100 microg/l) revealed no significant synergism. To elucidate a possible involvement of arachidonic acid metabolites in the signal transduction, we coincubated IL-3 or IL-6 together with the cyclo-oxygenase inhibitor indometacin or the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA). Coincubation with indometacin completely abolished the stimulatory effect of IL-6 but had no effect on IL-3 stimulated cortisol secretion. In contrast, specific inhibition of the lipoxygenase system by nordihydroguaiaretic acid blocked IL-3 stimulated steroidogenesis while the effect of IL-6 was not affected. Neither IL-3 nor IL-6 altered cAMP levels significantly, whereas ACTH significantly induced cAMP levels in parallel to its steroidogenic effect. In conclusion, our data indicate that IL-3 and IL-6 stimulate the steroid secretion of bovine adrenocortical cells to a similar extent and with a similar time course. However, the effects of IL-3 and IL-6 are mediated through different, cAMP-independent pathways. While the stimulatory effect of IL-3 seems to be dependent on the lipoxygenase pathway, the effect of IL-6 on adrenocortical cortisol secretion is mediated through the cyclo-oxygenase pathway.

Protective effect of serum antibodies against a 110-kilodalton protein of Actinobacillus actinomycetemcomitans following periodontal therapy.

Thirty-four adult patients with untreated periodontitis were randomly assigned to receive full mouth scaling alone or scaling with an adjunctive antimicrobial therapy, both followed by supportive periodontal therapy. At 24 months, specific serum immunoglobulin A (IgA), IgG and IgG subclass antibody reactivities against a 110-kDa protein of Actinobacillus actinomycetemcomitans were assessed by Western blot. In patients harboring A. actinomycetemcomitans intraorally, the IgG4 antibody reactivity against the 110-kDa protein of A. actinomycetemcomitans was associated with significantly increased survival rates of teeth and of sites not exhibiting 2 mm or more of probing attachment loss. The same trend was found for IgG3 and IgG2 antibody reactivities, but it was statistically insignificant. No association with clinical treatment outcome was observed for IgA, IgG and IgG1 antibody reactivities. The results indicated that systemic IgG4 antibody reactivity against the 110-kDa protein of A. actinomycetemcomitans may have a protective effect against periodontal disease progression in patients harboring A. actinomycetemcomitans and receiving periodontal therapy.

Clonal infection with Actinobacillus actinomycetemcomitans following periodontal therapy.

Mechanical debridement results in a shift of the bacterial composition in the periodontal pocket on the species level. It is unknown, however, whether a clonal change within a species could lead to the emergence of strains with different levels of virulence. Therefore, in the present study, the genetic variability of Actinobacillus actinomycetemcomitans was assessed and strains identified which were associated with periodontal disease progression following periodontal therapy, i.e., refractory periodontitis. Twenty adult patients with untreated periodontitis and subgingival colonization of A. actinomycetemcomitans were randomly assigned to receive full-mouth scaling alone or scaling with an adjunctive antimicrobial therapy. Both groups received supportive periodontal therapy at 3, 6, 9, 12, 18, and 24 months. Subgingival plaque samples were taken at every visit; venous blood was obtained at 24 months only. A. actinomycetemcomitans isolates were typed by the RAPD method, and antibody reactivity against outer membrane proteins was assessed by immunoblot analysis. Eleven distinct RAPD patterns were found in 18 patients completing the study. All patients harbored only one A. actinomycetemcomitans genotype, and within each patient this genotype persisted throughout the 24-month observation period. No differences in the expression of antibody reactivity against outer membrane proteins were found between strains isolated at baseline and at 24 months. Three genotypes were associated with reduced survival rates of teeth without probing attachment loss of 2 mm or more. The results indicated that (i) most patients harbored only one A. actinomycetemcomitans genotype; (ii) the genotype persisted following therapy; and (iii) only some genotypes were associated with refractory periodontitis.

Dose/response curves of lipoic acid R-and S-forms in the working rat heart during reoxygenation: superiority of the R-enantiomer in enhancement of aortic flow.

Micromolar concentrations of lipoic acid racemate added to a working rat heart during hypoxia have been previously found to improve aortic flow during subsequent reoxygenation. Since the R-form represents the naturally occurring form of lipoic acid, and the S-form does not reveal a positive influence on ATP synthesis in isolated mitoplasts, a dose/response curve of both enantiomers has been performed in working rat hearts. After the end of perfusion mitochondria were isolated and further analysed. At a concentration of 0.05-0.1 mumol of the R-enantiomer, aortic flow rises precipitously during reoxygenation, reaching over 70% of normoxic values compared to 50% of the controls. By contrast, with the S-enantiomer a value of about 60% is attained at 1 mumol, only. Accordingly, ATPase activity in mitochondria isolated from rat hearts previously treated with 0.05-0.1 mumol of the R- or S-enantiomer was significantly decreased or increased respectively. Consequently, whereas mitochondrial ATP synthesis was increased when the R-enantiomer was previously added to the working heart at 0.05-0.1 mumol concentration, with the S-enantiomer ATP synthesis remained within the control range. Mitochondrial membrane fluidity, measured with diphenylhexatriene, revealed a trend towards increase with the R- and decrease with the S-enantiomer. The total amount of thiol added at 0.1 mumol concentration is consistent with a value of 2 nmol/mg mitochondrial protein. This value has previously been found to be connected with -SH groups which add oligomycin-sensitivity to the ATPase complex. It is suggested that oligomycin-sensitive mitochondrial -SH groups contribute to the overall efficiency of low concentrations of lipoic acid R-enantiomer to enhance aortic flow.

Identification and characterization of insulin-like growth factor I (IGF-I) and IGF-II/mannose-6-phosphate (IGF-II/M6P) receptors in bovine adrenal cells.

We have identified and characterized insulin-like growth factor I (IGF-I) and IGF-II/mannose-6-phosphate (IGF-II/M6P) receptors in bovine adrenal cells. Iodine-125-labeled IGF-I ([125I]IGF-I) binding was characteristic of the IGF-I receptor, and binding kinetics as well as receptor densities were similar in cortical and medullary membranes. Scatchard analysis of [125I]IGF-I binding to cultured adrenocortical cells showed a single class of high-affinity binding sites with a Kd of 1.4 nmol/l and an average of 150,000 binding sites/cell. Affinity cross-linking experiments displayed a band at an apparent molecular weight of 135 kD, corresponding to the size of the alpha-subunit of the IGF-I receptor. In analogy, the binding of [125I]IGF-II to bovine adrenal membranes was characteristic of the IGF-II/M6P receptor and no differences between cortical and medullary membrane fractions were found. Scatchard analysis revealed a single class of high-affinity binding sites in adrenocortical cells with a Kd of 1.1 nmol/l and an average of 280,000 binding sites/cell. The identity of the IGF-II/M6P receptor was confirmed by western blotting of adrenocortical membranes with an anti-IGF-II/M6P receptor antibody and by affinity cross-linking of adrenocortical cells with labeled IGF-II. In conclusion, we have identified and characterized IGF-I and IGF-II/M6P receptors in bovine adrenocortical as well as medullary cells. In both regions of the bovine adrenal gland the IGF-II/M6P receptor is much more abundant than the IGF-I receptor.