Modulation of Wnt5a Expression by Periodontopathic Bacteria

Wingless proteins, termed Wnt, are involved in embryonic development, blood cell differentiation, and tumorigenesis. In mammalian hematopoiesis, Wnt signaling is essential for stem-cell homeostasis and lymphocyte differentiation. Recent studies have suggested that these molecules are associated with cardiovascular diseases, rheumatoid arthritis, and osteoarthritis. Furthermore, Wnt5a signaling is essential for the general inflammatory response of human macrophages. Periodontitis is a chronic inflammatory disease caused by gram-negative periodontopathic bacteria and the resultant host immune response. Periodontitis is characterized by loss of tooth-supporting structures and alveolar bone resorption. There have been no previous reports on Wnt5a expression in periodontitis tissue, and only few study reported the molecular mechanisms of Wnt5a expression in LPS-stimulated monocytic cells. Using RT-PCR, we demonstrated that Wnt5a mRNA expression was up-regulated in chronic periodontitis tissue as compared to healthy control tissue. P. gingivalis LPS induced Wnt5a mRNA in the human monocytic cell line THP-1 with a peak at 4 hrs after stimulation. P. gingivalis LPS induced higher up-regulation of Wnt5a mRNA than E. coli LPS. The LPS receptors TLR2 and TLR4 were equally expressed on the surface of THP-1 cells. P. gingivalis LPS induced IκBα degradation and was able to increase the NF-κB binding activity to DNA. P. gingivalis LPS-induced Wnt5a expression was inhibited by NF-κB inhibitors, suggesting NF-κB involvement. Furthermore, IFN-γ synergistically enhanced the P. gingivalis LPS-induced production of Wnt5a. Pharmacological investigation and siRNA experiments showed that STAT1 was important for P. gingivalis LPS-induced Wnt5a expression. These results suggest that the modulation of Wnt5a expression by P. gingivalis may play an important role in the periodontal inflammatory process and serve a target for the development of new therapies

Palatal Soft Tissue Thickness on Maxillary Posterior Teeth and Its Relation to Palatal Vault Angle Measured by Cone-Beam Computed Tomography

Objective. Analyzing palatal soft tissue thickness in cone-beam computed tomography (CBCT) images and evaluating the relationship between tissue thickness and palatal vault angulation. Methods. Out of 1,737 CBCT images, fifty-six images met the inclusion criteria and were included in this cross-sectional study. The palatal vault angle on the maxillary first molar was measured and divided the images into 3 groups. The soft tissue thickness between the maxillary first premolar and second molar was measured at a distance of 3, 6, 7, 8, and 9 mm from the cementoenamel junction. All the image measurements were performed using CBCT-viewer software. Result. In this study, 56 CBCT images with full permanent maxillary posterior teeth and absence of light scattering were found. The mean age of the patients was 31.59 ± 13.92 years. The moderate and deep palatal vault angle patterns had the greatest and least prevalence, respectively. The average thickness on shallow, moderate, and deep palatal vault groups was 4.02 ± 0.58, 3.75 ± 0.73, and 3.43 ± 0.38 mm, respectively. Furthermore, the mean palatal mucosal thickness was statistically different between the deep and shallow palatal vault angle groups (p<0.05, power of test 0.8). Based on the Pearson correlation coefficient, there was a negative correlation between the palatal mucosal thickness and palatal vault angle (p<0.05, power of test 0.85). Conclusion. A negative correlation between the palatal mucosal thickness and palatal vault angle was observed. Furthermore, this study suggested that the shape of the palatal vault can be one of the supporting data for evaluating the graft dimensions

Anti-Early Stage of Bacterial Recolonization Effect of Curcuma longa Extract as Photodynamic Adjunctive Treatment

Objective. To evaluate the amount of Fusobacterium nucleatum (F. nucleatum) and Prevotella intermedia (P. intermedia) on subgingival recolonized plaque after mechanical debridement and photodynamic treatment by using blue light-emitting diodes (LEDs) in combination with topical Curcuma longa gel extract. Methods. A total of 12 subjects with stage III grade B periodontitis were recruited for the study. Maxillary posterior teeth with periodontal pocket >4 mm were selected. These teeth were examined for periodontal clinical data at baseline and at 1, 2, 4, and 6 weeks after treatment. All remaining teeth were treated by scaling and root planing (SRP). Then, the teeth were bilaterally divided using randomized split-mouth design with and without photodynamic adjunctive therapy (PDT). Samples of the subgingival microbiota were obtained in each visit. All samples were analyzed by multicolor TaqMan real-time polymerase chain reaction (PCR) for the presence of target bacteria. Results. Throughout the six-week follow-up, long-term improvement of probing depth and bleeding on probing was revealed on the PDT group. The number of subgingival F. nucleatum and P. intermedia also significantly reduced, compared to the baseline. There was a statistically significant recolonization in F. nucleatum and P. intermedia number after 2 and 4 weeks of conventional SRP, respectively. Our quantitative PCR method showed no significant recolonization of those subgingival bacteria on PDT sites throughout the 6-week study duration. Conclusion. The results showed that adjunctive photodynamic treatment by using blue LEDs in combination with topical Curcuma longa gel extract was effective to alter the recolonization patterns of F. nucleatum and P. intermedia after conventional debridement

Preparation and Evaluation of Metronidazole-Loaded Pectin Films for Potentially Targeting a Microbial Infection Associated with Periodontal Disease

The objective of this study was to develop the metronidazole loaded high and low methoxyl pectin films (HM-G-MZ and LM-G-MZ) for the treatment of periodontal disease. The films were prepared by pectin 3% w/v, glycerin 40% w/v, and metronidazole 5% w/v. The developed films were characterized by scanning electron microscope and evaluated for thickness, weight variation, and elasticity. The developed films showing optimal mechanical properties were selected to evaluate radial swelling properties, in vitro release of metronidazole and the antimicrobial activity against Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans by the disc diffusion method. The results demonstrated that LM-MZ and HM-G-MZ films were colorless and yellowish color, respectively, with the film thickness around 0.36&ndash;0.38 mm. Furthermore, both films exhibited good elasticity with low puncture strength (1.63 &plusmn; 0.37 and 0.84 &plusmn; 0.03 N/mm2, respectively) and also showed slight increase in radial swelling, so that they could be easily inserted and fitted into the periodontal pocket during a clinical use. However, HM-G-MZ showed a decrease in radial swelling after 1 h due to the film erosion. The in vitro release study of LM-G-MZ showed a burst release that was initially followed by a slow release rate profile, capable to maintain the therapeutic level in periodontal pocket for seven days, whereas HM-G-MZ showed an immediate release profile. The cumulative percentage of metronidazole release from HM-G-MZ was less than LM-G-MZ during the first 5 min as metronidazole was in a crystalline form inside HM-G-MZ film. For antimicrobial activity test, both films showed the inhibitory effect against P. gingivalis and A. actinomycetemcomitans, and there was no difference in the inhibition zone between LM-G-MZ and HM-G-MZ. The present study showed, for the first time, that low methoxyl pectin film containing glycerin and metronidazole could be potentially considered as a promising clinical tool for the drug delivery via intra-periodontal pocket to target an oral disease that is associated with polymicrobial infection

Inhibitory Effects of Erythrosine/Curcumin Derivatives/Nano-Titanium Dioxide-Mediated Photodynamic Therapy on Candida albicans

This study focuses on the role of photosensitizers in photodynamic therapy. The photosensitizers were prepared in combinations of 110/220 µM erythrosine and/or 10/20 µM demethoxy/bisdemethoxy curcumin with/without 10% (w/w) nano-titanium dioxide. Irradiation was performed with a dental blue light in the 395–480 nm wavelength range, with a power density of 3200 mW/cm2 and yield of 72 J/cm2. The production of ROS and hydroxyl radical was investigated using an electron paramagnetic resonance spectrometer for each individual photosensitizer or in photosensitizer combinations. Subsequently, a PrestoBlue® toxicity test of the gingival fibroblast cells was performed at 6 and 24 h on the eight highest ROS-generating photosensitizers containing curcumin derivatives and erythrosine 220 µM. Finally, the antifungal ability of 22 test photosensitizers, Candida albicans (ATCC 10231), were cultured in biofilm form at 37 °C for 48 h, then the colonies were counted in colony-forming units (CFU/mL) via the drop plate technique, and then the log reduction was calculated. The results showed that at 48 h the test photosensitizers could simultaneously produce both ROS types. All test photosensitizers demonstrated no toxicity on the fibroblast cells. In total, 18 test photosensitizers were able to inhibit Candida albicans similarly to nystatin. Conclusively, 20 µM bisdemethoxy curcumin + 220 µM erythrosine + 10% (w/w) nano-titanium dioxide exerted the highest inhibitory effect on Candida albicans

Periodontitis and cardiovascular diseases: The link and relevant mechanisms

This paper reviews the association between periodontitis and CVD. In addition, the potential mechanisms of any association between periodontitis and CVD as well as the effects of periodontal treatment on CVD are herein discussed. Among the studies carried out by this group and others on coronary artery diseases, peripheral arterial diseases, abdominal aortic aneurysm, and Buerger's disease, periodontopathic bacteria were frequently detected in the diseased blood vessels, thus suggesting an association between periodontitis and CVD. The potential mechanisms of the association between periodontitis and CVD are not fully elucidated. However, inflammation and some autoimmune mechanisms, including molecular mimicry between the periodontopathic bacteria and host molecules, are suggested. The effects of periodontal treatment on CVD might thus vary among the different treatment modalities, and full-mouth mechanical debridement might induce strong transient systemic inflammatory responses in comparison to the quadrant-wise mechanical debridement

Wnt5a was specifically up-regulated in THP-1 cells by <i>P. gingivalis</i> LPS.

<p>(A) HGF-1 and THP-1 cells were stimulated with <i>A. actinomycetemcomitans</i> sonicated extract, <i>P. gingivalis</i> sonicated extract, <i>P. gingivalis</i> LPS, and TNF-α for 4 hrs, and the expression of Wnt5a mRNA was detected by RT-PCR. PCR products were electrophoresed on a 1.5% agarose gel and visualized by UV illumination. GAPDH served as the internal control. (B) THP-1 cells were stimulated with 1 µg/ml of <i>P. gingivalis</i> LPS for 0.5, 2, 4, 12, or 24 hrs, and the expression of Wnt5a mRNA was detected by real-time PCR. Relative expression levels of Wnt5a are shown. (C) THP-1 cells were stimulated with 0.01–10 µg/ml of <i>P. gingivalis</i> LPS (black bars) or <i>E. coli</i> LPS (gray bars) for 4 hrs, and then the expression of Wnt5a mRNA was detected by real-time PCR. Relative expression levels of Wnt5a are presented. (D) THP-1 cells were stimulated with <i>E. coli</i> 055:B5 LPS (middle and right upper panels) or <i>P. gingivalis</i> LPS (middle and right lower panels) for 30 min or 4 hrs, and then the expression of surface TLR2 and TLR4 protein was determined by flow cytometry. Left upper panel shows no-staining condition, and left lower panel shows un-stimulated condition with staining. (E, F, G, H) The expression of Wnt5a mRNA was detected by real-time PCR. Relative expression levels of Wnt5a mRNA are shown. (E) THP-1 cells were stimulated with 10⁴–10⁷ cells/ml of live <i>P. gingivalis</i> for 4 hrs. (F, G) Primary human gingival fibroblasts (HGF) and human monocytes were stimulated with 1 µg/ml of <i>P. gingivalis</i> LPS for 4 hrs. Monocytes were pretreated with the NF-κB inhibitor MG132 for 1 hr. Here we describe typical dates of three samples. (H) THP-1 cells were stimulated with <i>P. gingivalis</i> LPS for 4 hrs after being transfected with TLR2 siRNA, TLR4 siRNA or control siRNA for 72 hrs. *p<0.05.</p

Induction of Wnt5a expression is partly JAK/STAT dependent.

<p>(A–D) The expression of Wnt5a mRNA was detected by real-time PCR. Relative expression levels of Wnt5a are presented. (A, B) THP-1 cells were pre-treated with AG490 (A) or fludarabine (B) for 1 hr and then stimulated with <i>P. gingivalis</i> LPS for 4 hrs. DMSO is a solvent control for both inhibitors. (C) THP-1 cells were stimulated with <i>P. gingivalis</i> LPS for 4 hrs after transfection with STAT1 siRNA or control siRNA for 18 hrs. (D) THP-1 cells were pre-treated with STA21 for 1 hr and stimulated with <i>P. gingivalis</i> LPS for 4 hrs. *p<0.05.</p

<i>P. gingivalis</i> LPS-induced Wnt5a expression was inhibited by wedelolactone or dnIκBα.

<p>(A, B) THP-1 cells were pre-treated with the IKK inhibitor wedelolactone or the IKK inhibitor VII for 1 hr. The expression of Wnt5a mRNA was detected by real-time PCR. Relative expression levels of Wnt5a are presented. DMSO is a solvent control for both inhibitors. (C) THP-1 cells were transfected with a dominant-negative form of IκBα (dnIκBα) or the parent plasmid (pUSE) for 12 hrs. Fold induction of Wnt5a mRNA expression is shown. *p<0.05.</p