Type 2 Diabetes Mellitus and Periodontitis: Bidirectional Association in Population-based 15-year Retrospective Cohorts.

OBJECTIVE: A two-way relationship between periodontitis and diabetes has been proposed. However, bidirectional epidemiological observation is limited and inconsistent. OBJECTIVE: Using the National Health Insurance Research Database of Taiwan (covering over 99% of the entire population), we aimed to estimate the development of diabetes in periodontitis patients or that of periodontitis in patients with type 2 diabetes mellitus (T2DM), respectively. METHODS: A total of 11 011 patients with severe periodontitis were recruited from 2000 to 2015. After matching by age, sex, and index date, 11 011 patients with mild periodontitis and 11 011 non-periodontitis controls were registered. Additionally, 157 798 patients with T2DM and 157 798 non-T2DM controls were enrolled, in whom the development of periodontitis was traced. Cox proportional hazards model was performed. RESULTS: Periodontitis patients tended to have a statistically high risk for T2DM. The adjusted hazard ratio was 1.94 (95% CI, 1.49-2.63, P < .01) and 1.72 (95% CI, 1.24-2.52, P < .01) for severe and mild periodontitis groups, respectively. The patients with severe periodontitis had a higher risk of having T2DM relative to those with mild periodontitis (1.17 [95% CI, 1.04-1.26, P < .001]). Conversely, the risk of periodontitis increased significantly in patients with T2DM (1.99 [95% CI, 1.42-2.48, P < .01]). However, high risk was observed for the outcome of severe periodontitis (2.08 [95% CI, 1.50-2.66, P < .001]), but not for mild periodontitis (0.97 [95% CI, 0.38-1.57, P = .462]). CONCLUSION: We suggest that the bidirectional association is between T2DM and severe but not mild periodontitis.

Is periodontitis a risk factor of benign or malignant colorectal tumor? A population-based cohort study.

OBJECTIVE: To examine the risk of developing benign or malignant colorectal tumors in patients with periodontitis within 15 years using Taiwan's National Health Insurance Database. BACKGROUND: Studies have shown that colorectal carcinoma often develops under inflammatory conditions and changes of microbiota in the gut. Recently, a link between Fusobacterium nucleatum, a periodontal pathogen, and colorectal carcinoma has been proposed. However, whether periodontitis is a risk of developing colorectal tumor remains uncertain. METHODS: In total, 35 124 participants were enrolled from 2000 to 2015 to examine the development risk of benign colorectal tumors, including 11 708 patients with periodontitis who received therapy (group 1), 11 708 patients with periodontitis not receiving periodontal treatment (group 2), and 11 708 non-periodontitis controls after matching for gender, age, and index year. To examine the risk of developing colorectal malignancy, 11 720 participants were assigned to each of the three groups. Cox proportional hazards model and Kaplan-Meier methods were used to compare the risks. Sensitivity analysis was performed, excluding the diagnoses during the first 1 or 5 years. RESULTS: After the follow-up, 177, 154, and 63 participants in group 1, group 2, and control group had benign colorectal tumors. Patients with periodontitis tended to be associated with a greater rate of having a benign colorectal tumor. The adjusted hazard ratios (aHRs) were 3.77 (95% confidence interval [CI] 2.01-4.82, p < .001) and 2.85 (95% CI 1.62-3.74, p < .001) for groups 1 and 2, respectively. Regarding the risk of malignant colorectal tumor, 20, 18, and 14 participants who developed malignant tumors were included in group 1, group 2, and control group; however, no significant increase in malignancy was observed in periodontitis groups (aHR1.92, 95% CI 0.74-2.36, p = .482; aHR 1.50, 95% CI 0.68-1.97, p = .529, for the two periodontitis groups, respectively). CONCLUSIONS: The results of this study suggest that patients with periodontitis may have an increased risk of developing benign, but not malignant, colorectal tumors.

Cyanidin-3-O-glucoside downregulates ligation-activated endoplasmic reticulum stress and alleviates induced periodontal destruction in rats.

OBJECTIVE: Cyanidin-3-O-glucoside (C3G), a dietary anthocyanin, possesses various biological properties, including alleviating endoplasmic reticulum (ER) stress. This study examined the effect of C3G on periodontitis via ER stress in rats. DESIGN: Periodontitis was induced by placing silk sutures around maxillary second molars. C3G (0, 3, or 9 mg/kg) was fed on the day before ligation (10 rats/group). Further, 10 non-ligation control rats received deionized water. On day 8, gingivae were obtained to determine CCAAT/enhancer-binding protein homologous protein (CHOP), c-Jun N-terminal kinase (JNK), phospho-JNK (p-JNK), and nuclear factor-κB (NF-κB) by immunoblotting. Periodontal destruction was evaluated using micro-computed tomography (µCT) and histology. RESULTS: Gingival expression of CHOP, p-JNK/JNK, and NF-κB significantly increased in ligation rats (0 mg/kg C3G) than that in controls. However, protein expression in ligation groups presented a negative association with C3G concentration. By µCT, the distance of cemento-enamel junction to bone significantly increased in ligation groups; however, distances showed a negative association with C3G concentration. In the region of interest, bone volume and trabecular thickness and number significantly decreased in ligation groups but they were positively associated with C3G concentration. In terms of trabecular separation, opposite results were found. Histologically, infiltrated connective tissue (ICT) and periodontal destructions increased in ligation groups; however, they were negatively associated with C3G concentration. Moreover, ICT area is positively correlated with µCT- and histologically measured destructions and protein expression of CHOP, p-JNK/JNK, or NF-κB. CONCLUSION: C3G promotes favorable modulation of ER stress and alleviates destruction of periodontitis, which may imply a new strategy.

Antioxidants protect against gingival overgrowth induced by cyclosporine A.

OBJECTIVE: We investigated the importance of reactive oxygen species (ROS) on developing gingival overgrowth (GO) and then introduced the antioxidant strategy to prevent, or even reduce GO. BACKGROUND: Gingival overgrowth is a common side effect of the patients receiving cyclosporine A (CsA), an immune suppressant. Although it has been broadly investigated, the exact pathogenesis of the induced GO is still uncertain. METHODS: We cultured human primary gingival fibroblasts and used animal model of GO to investigate the ameliorative effects of antioxidants on CsA-induced GO. To examine the CsA-induced oxidative stress, associated genes and protein expression, and the overgrown gingiva of rats by using immunocytochemistry, confocal laser scanning microscopy, real-time PCR, ELISA, gelatin zymography, gingival morphological, and immunohistochemical analysis. RESULTS: We found for the first time that ROS was responsible for the CsA-induced oxidative stress and TGF-ß1 expression in human primary gingival fibroblasts, as well as the GO of rats. The antioxidants (oxidative scavenger of vitamin E and an antioxidative enzyme inducer of hemin) ameliorated CsA-induced pathological and morphological alterations of GO without affected the CsA-suppressed il-2 expression in rats. CsA-induced oxidative stress, HO-1, TGF-ß1, and type II EMT were also rescued by antioxidants treatment. CONCLUSIONS: We concluded that CsA repetitively stimulating the production of ROS is the cause of CsA-GO which is ameliorated by treating antioxidants, including vitamin E and sulforaphane. Furthermore, the immunosuppressive effect of CsA is not interfered by antioxidant treatments in rats. This finding may thus help the clinician devise better prevention strategies in patients susceptible to GO.

Immediate hyperbaric oxygen after tooth extraction ameliorates bisphosphonate-related osteonecrotic lesion in rats.

BACKGROUND: This study aims to assess whether hyperbaric oxygen (HBO) applied immediately after tooth extraction could ameliorate medication-related osteonecrosis of the jaw in rats. METHODS: To evaluate whether osteonecrosis could be successfully induced, healing of extraction maxillary molars was examined in 40 female Sprague Dawley rats received zoledronic acid (7.5 µg/kg) plus dexamethasone (1 mg/kg). Rats were divided into four groups, receiving zero, two, four, or seven injection(s) for 7 days, respectively. Effect of HBO, pressurized to 2.5 atmospheres absolute (ATA) at rate of 0.15 ATA/min with 100% oxygen for 90 minutes, applied immediately after tooth extraction, on the development of osteonecrosis was evaluated. Lesions among groups were compared by size of ulceration, exact area (mm2 ) or relative area (%), and by histology. RESULTS: Unhealed ridge was observed in all nine rats in four and seven injection groups, but none of 10 rats in the control (non-injection) group. Immediate HBO significantly reduced the lesions in rats that received four injections, regardless of the distribution and the total/relative areas of lesions (P <0.01). Histological findings showed the lesions were uncovered epithelium and severe tissue inflammation. CONCLUSION: This is the first in vivo study demonstrating the HBO applied immediately after tooth extraction effectively decreases the development of medication-related osteonecrosis.

Effectiveness of an oral function intervention for older Taiwanese people.

OBJECTIVE: Oral function disorders occur often in older people with increasing age. Oral function disorders affect bodily function and self-esteem, which are related to quality of life (QOL). The aim of this study was to evaluate the effectiveness of an oral function intervention programme on the oral function of older Taiwanese people. MATERIALS AND METHODS: A one-group pretest-post-test study design was used. A total of 529 older Taiwanese people (women, 68.2%; men, 31.8%; average age, 75.07 ± 5.95 years) participated in this study. The oral function intervention programme consisted of a brief oral health education programme and oral function exercises. The total test period was 8 months. The oral condition and function examination comprised two questionnaires (self-reported symptoms of oral function disturbance and the Geriatric/General Oral Health Assessment Index [GOHAI]) and three oral function assessments (Repetitive Saliva Swallowing Test [RSST], Oral Diadochokinesia Test [ODT] and Cheek Expanding Test [CET]). RESULTS: After the oral function intervention, the self-reported symptoms on the oral function questionnaire and GOHAI showed significant improvement (P < 0.05). Additionally, RSST, ODT and CET showed differences between pretest and post-test measurements (P < 0.05). CONCLUSION: The oral function intervention programme was effective in maintaining their feeding, swallowing and articulatory functions of older Taiwanese people. Significant improvements in self-reported symptoms of oral function and GOHAI scores indicated that the oral function intervention programme might improve the QOL of older Taiwanese people.

Ameliorative effect of hesperidin on ligation-induced periodontitis in rats.

BACKGROUND: This study evaluated the ameliorative effect of hesperidin (HES), an anti-inflammatory flavanone, in rats with ligation (Lig)-induced periodontitis. METHODS: A total of 48 rats were randomly divided into non-ligation group (NL), Lig group, and two ligation-plus-HES groups (L+H). HES was administered immediately after ligature placement at a dose of 75 or 150 mg/kg by intragastric feeding. Destruction of the ligated maxillary second and mandibular first molars were evaluated by dental radiography, microcomputed tomography (micro-CT), and histometry performed after sacrificing the rats on the seventh day. The expression levels of interleukin (IL)-1ß, IL-6, and inducible nitric oxide synthase (iNOS) messenger (m)RNAs in the gingiva were determined by reverse-transcription polymerase chain reaction. The expression of iNOS was examined by immunohistochemistry. RESULTS: The dental radiography and micro-CT findings revealed significantly increased alveolar bone loss in the Lig group, which was significantly prevented by HES. The histometry results revealed less gingival inflammation and connective tissue loss in the L+H groups compared with that in the Lig group. The mRNA expression levels of IL-6, IL-1 ß, and iNOS were significantly increased in the Lig group but were reduced in the L+H groups. The immunostaining results showed that the ligation-induced iNOS expression was also decreased by HES. CONCLUSIONS: Oral administration of HES promotes an ameliorative effect against the ligation-induced alveolar bone loss and effectively inhibits the production of proinflammatory mediators in rats with experimentally induced periodontitis. Therefore, HES may be a good candidate for modulating oral inflammatory diseases.

Comparison of oral malodors before and after nonsurgical periodontal therapy in chronic periodontitis patients.

BACKGROUND/PURPOSE: Periodontal diseases have been considered as a source of oral malodor or halitosis. Improvement of oral malodor in chronic periodontitis patients has recently been observed after nonsurgical periodontal therapy in combination with tongue cleaning and/or chlorhexidine mouth rinsing. The present study, however, evaluated the impact of nonsurgical periodontal therapy alone on the oral malodor in chronic periodontitis patients by comparing the intraoral concentrations of volatile sulfur compounds (VSCs) before and after nonsurgical therapy. MATERIALS AND METHODS: Using a sulfide monitor, the total VSCs in exhaled breath were measured in 80 patients with chronic periodontitis prior to and 1 month after nonsurgical periodontal therapy (re-evaluation phase). Malodor was defined as a VSC score > 75 parts per billion (ppb) and > 110 ppb, respectively. RESULTS: Significantly lower level of VSCs was recorded at periodontal re-evaluation (55 ± 9.7 ppb) than before treatment (89 ± 16.3 ppb). Before treatment, 27 (34%) patients were considered to have malodor, defined as VSCs > 75 ppb. After treatment, 16 patients (20%) had VSC scores > 75 ppb, including 10 of 27 patients with baseline VSC scores > 75 ppb and six of 53 patients with baseline scores ≤ 75 ppb. The risk of malodor differed significantly before and after treatment (P = 0.035, McNemar's test). However, when malodor was defined as VSCs > 110 ppb, the difference in risk showed only borderline significance (P = 0.077). CONCLUSION: On the basis of our findings, we suggest that nonsurgical periodontal therapy has a mild impact on oral malodor.

Crosstalk Between Human Monocytic U937 Cells and Gingival Fibroblasts in Coculturally Enhanced Matrix Metalloproteinase-2 Expression.

BACKGROUND: Matrix metalloproteinases (MMPs) play a key role in inflammatory periodontal disease. Synergistically enhanced MMP-2 expression in a coculture of human gingival fibroblasts (HGFs) and human monocytic U937 cells was observed. Crosstalk between these two cells via the extracellular matrix metalloproteinase inducer (EMMPRIN) was demonstrated. METHODS: Enzyme levels of MMP-2 in HGFs and direct coculture with U937 were examined by zymography. MMP-2 and EMMPRIN expressions of HGFs and U937 were determined in coculture and conditioned cultures (using supernatants from HGF- or U937-conditioned medium). The crosstalk was evaluated by EMMPRIN extrasupplement and EMMPRIN inhibition, through pretreatment of U937 with cyclosporine-A. RESULTS: Direct coculturing of HGFs and U937 enhanced MMP-2 enzyme level and mRNA expression. Coculturing also increased membranous EMMPRIN expression of U937, but not from HGFs. In conditioned cultures, mRNA expression of MMP-2 increased in HGFs which received U937-conditioned medium. Increased MMP-2 was not observed in U937 with HGF-conditioned medium, although mRNA expression of EMMPRIN increased. Enhanced MMP-2 was observed after administration of exogenous EMMPRIN in HGFs; however, reduced MMP-2 enzyme level was noted if EMMPRIN of cocultured U937 was inhibited. CONCLUSIONS: In the coculture of HGFs and U937, upregulated EMMPRIN expression in U937, which may be triggered by HGFs, can enhance MMP-2 expression in HGFs. Crosstalk between HGFs and U937 involving MMP-2 from HGFs was proposed; EMMPRIN from U937 may play a particular role.

Effects of Salvia miltiorrhiza ethanolic extract on lipopolysaccharide-induced dental alveolar bone resorption in rats.

BACKGROUND/PURPOSE: Salvia miltiorrhiza (SM) Bunge (Labiatae/Lamiaceae; common name danshen) is a Chinese medicine that improves blood circulation and inhibits inflammatory response. Thus, it is used for the treatment of cardiac diseases and inflammation. In this study, we aimed to evaluate the effect of an ethanolic extract of SM (SME) on the dental alveolar bone resorption induced by bacterial lipopolysaccharide (LPS) in rats. MATERIALS AND METHODS: An ethanolic extract was prepared from roots of SM. The major constituents of this extract were determined by high-performance liquid chromatography. The activity of the extract was evaluated in a rat model in which the dental alveolar bone resorption was induced by injection of bacterial LPS into the palatal gingiva around the maxillary molar teeth. The effect of SME on the bone resorption was studied by histologic and histomorphometric analysis. RESULTS: The number of osteoclasts and the percentage of osteoclasts covering the alveolar bone surfaces were significantly increased in the LPS group compared with those in the phosphate-buffered saline (PBS) group. The number and percentage of the osteoclasts on the bony surfaces were significantly reduced in the SME group in comparison with the LPS group, although it was still higher than the numbers observed in the PBS group. CONCLUSION: Because SME reduced bone resorption caused by the injections of bacterial LPS in rats, we suggest that SME might have a protective effect on dental alveolar bone resorption in periodontitis.

Bifid mandibular canals and their cortex thicknesses: A comparison study on images obtained from cone-beam and multislice computed tomography.

BACKGROUND/PURPOSE: High prevalence of bifid mandibular canals has been visualized with various types of computerized tomography (CT). Along the canals, a various ranged corticalization was recently reported. The depiction of the fine anatomic structures on multislice and cone-beam CT images was compared. MATERIAL AND METHODS: The presence or absence of the bifid canal was assessed on 327 images obtained by multislice CT (MSCT; n = 173) or by cone-beam CT (CBCT; n = 154), according to the configuration. The cortex thickness and distribution were also assessed. RESULTS: The prevalence of bifid canal detected by CBCT was significantly greater than that detected by MSCT (42.2% vs. 18.7% for hemi-mandibles and 58.4% vs. 30.6% for patients). Cortical thickness recorded by CBCT was significantly thinner than that recorded by MSCT (0.48 mm vs. 0.65 mm, P < 0.001); however, the distributions of corticalization detected by the two tomography methods were similar. There was a significant association of cortex thickness with CT type and corticalization degree (R 2 = 0.530, P < 0.001). CONCLUSION: Thinner cortices, but greater prevalence of bifid canals recorded by CBCT, compared to MSCT, suggests that clinicians should be cautious when using CT to interpret this fine anatomic structure.

Association between periodontal disease and osteoporosis by gender: a nationwide population-based cohort study.

Periodontitis and osteoporosis are primary concerns in public health and clinical management. The aim of this study was to investigate the association between periodontitis and osteoporosis by gender.Data were retrieved from the National Health Insurance Research Database, Taiwan. A diagnosis of periodontitis was defined on the basis of subgingival curettage, periodontal flap operation, and gingivectomy (excluding those with restorative or aesthetic indications). Multiple logistic regression was used for analysis. After adjusting for age, sex, income, and geographical region, there was a significant association between periodontitis and osteoporosis among women (odds ratio: 1.96; 95% confidence interval 1.17-3.26). The association between periodontitis and osteoporosis was significant among women.

Role of Shh and TGF in cyclosporine-enhanced expression of collagen and α-SMA by gingival fibroblast.

OBJECTIVE: Cyclosporine-A (CsA)-induced gingival overgrowth may arise from an alteration in stoma matrix homeostasis. Sonic hedgehog (Shh) plays a key role during embryogenic development and fibrotic progression, and may be involved in CsA-altered gingival matrix homeostasis. METHODS: Using the reverse transcription-polymerase chain reaction and Western blot analysis, we investigated the mRNA and protein expressions of Shh, type 1 collagen (COL1), alpha-smooth muscle actin (α-SMA) and transforming growth factor-beta (TGF-ß) in human gingival fibroblasts after CsA treatments. The effect of Shh on CsA-induced alterations was further evaluated by the extra-supplement or inhibition of Shh or TGF-ß. RESULTS: Cyclosporine-A enhanced COL1, α-SMA, Shh and TGF-ß expressions in human gingival fibroblasts. The exogenous Shh/TGF-ß augmented the expression of COL1 and α-SMA, and the Shh/TGF-ß inhibition suppressed the CsA-enhanced COL1 and α-SMA expressions. Moreover, Shh mRNA and protein expressions increased if extra-supplementing the exogenous TGF-ß, whereas the CsA-upregulated Shh was mitigated by the TGF-ß pathway inhibitor. However, neither exogenous Shh nor the Shh pathway inhibitor alters TGF-ß expression or CsA-up-regulated TGF-ß expression. CONCLUSIONS: Shh, regulated by TGF-ß, mediates CsA-altered gingival matrix homeostasis.

Gelatinases and extracellular matrix metalloproteinase inducer are associated with cyclosporin-A-induced attenuation of periodontal degradation in rats.

BACKGROUND: The present study aims to examine the inhibitory effect of cyclosporin-A (CsA) on periodontal breakdown and to further explore the correlations of CsA-induced attenuation of periodontal bone loss with the expressions of gelatinases (i.e., matrix metalloproteinase [MMP]-2 and MMP-9) and extracellular matrix metalloproteinase inducer (EMMPRIN). METHODS: Forty Sprague-Dawley rats were randomly divided into four groups: 1) control; 2) CsA; 3) ligature (Lig); and 4) ligature plus CsA (Lig + CsA). The CsA group received 10 mg ⋅ Kg(-1) ⋅ d(-1) CsA for 8 days. The Lig group received silk ligature on selected molars. The Lig + CsA group received silk ligature and CsA treatment. The inhibitory effects of CsA on the ligature-induced periodontal breakdown was examined with microcomputed tomography (micro-CT) and histometric analyses to analyze the amount of attachment loss, crestal bone loss, connective tissue attachment, and the surface area with inflammatory cell infiltration. The effects of CsA on ligature-induced expressions of gelatinases and EMMPRIN in gingival tissues were examined with Western blotting and zymography, respectively. RESULTS: By micro-CT and histology, the Lig + CsA group had significantly more periodontal breakdown than the control and CsA groups but less periodontal breakdown than the Lig group. Consistent results were found for the expressions of gelatinases and EMMPRIN among the groups demonstrating that the Lig + CsA group had significantly less gingival protein expression of gelatinases and EMMPRIN than the Lig group. CONCLUSIONS: CsA inhibited the expressions of gelatinase MMPs and EMMPRIN and partially prevented the periodontal breakdown in ligature-induced experimental periodontitis. The CsA-induced attenuation of periodontal bone loss was strongly correlated positively with the expressions of MMP-2, MMP-9, and EMMPRIN in gingiva.

Role of transforming growth factor-beta1 in cyclosporine-induced epithelial-to-mesenchymal transition in gingival epithelium.

BACKGROUND: It has been proposed that cyclosporin A (CsA) may induce epithelial-to-mesenchymal transition (EMT) in gingiva. The aims of the present study are to confirm the notion that EMT occurs in human gingival epithelial (hGE) cells after CsA treatment and to investigate the role of transforming growth factor beta1 (TGF-ß1) on this CsA-induced EMT. METHODS: The effects of CsA, with and without TGF-ß1 inhibitor, on the morphologic changes of primary culture of hGE cells were examined in vitro. The changes of protein and messenger RNA (mRNA) expressions of two EMT markers (E-cadherin and alpha-smooth muscle actin) in the hGE cells after CsA treatment with and without TGF-ß1 inhibitor were evaluated with immunocytochemistry and real-time polymerase chain reaction. RESULTS: The epithelial cells became spindle-like, elongated, and disassociated from neighboring cells and lost their original cobblestone monolayer pattern when CsA was added. However, the epithelial cells stayed in their original cobblestone morphology with treatment of TGF-ß1 inhibitor on top of the CsA treatment. When CsA was given, the protein and mRNA expressions of E-cadherin and α-SMA were significantly altered, and these alterations were significantly reversed with pretreatment of TGF-ß1 inhibitor. CONCLUSIONS: CsA could induce Type 2 EMT in gingiva by changing the morphology of epithelial cells and altering the EMT markers/effectors. The CsA-induced gingival EMT is dependent or at least partially dependent on TGF-ß1.

Cyclosporine A enhances gingival ß-catenin stability via Wnt signaling.

BACKGROUND: Cyclosporine A (CsA) increases ß-catenin messenger RNA (mRNA) and protein expression. The present study demonstrates that Wnt/ß-catenin signaling inhibits ß-catenin degradation in the gingiva. METHODS: Forty 5-week-old male Sprague-Dawley rats were assigned to two study groups after healing from right maxillary molar extractions. The rats in the experimental group were fed 30 mg/kg CsA daily for 4 weeks, whereas the control rats were fed mineral oil. At the end of the study, all rats were sacrificed, and the gingivae were obtained. The gingival morphology after CsA treatment was evaluated by histology, and the genes related to Wnt/ß-catenin signaling were initially screened by microarray. Polymerase chain reaction, Western blotting, and immunohistochemistry were used to examine the mRNA and protein expression of proliferating cell nuclear antigen, cyclin D1, E-cadherin, ß-catenin, Dvl-1, glycogen synthase kinase-3ß, axin-1, and adenomatous polyposis coli (APC). Phosphoserine and ubiquitinylated ß-catenin were detected after immunoprecipitation. RESULTS: In rats treated with CsA, overgrowth of gingivae was observed, and altered expression of genes related to Wnt/ß-catenin signaling was detected by the microarray. The gingival mRNA and protein expression profiles for genes associated with Wnt/ß-catenin signaling further confirmed the effect of CsA: ß-catenin and Dvl-1 expression increased, but APC and axin-1 expression decreased. Western blotting and immunohistochemistry showed decreases in ß-catenin serine phosphorylation (33/37) and ubiquitinylation in the gingivae of CsA-treated rats. CONCLUSION: CsA-enhanced gingival ß-catenin stability may be involved in gene upregulation or ß-catenin degradation via the Wnt/ß-catenin pathway.

Effect of paeonol on tissue destruction in experimental periodontitis of rats.

We evaluated the effects of paeonol, a phenolic compound of Moutan Cortex, on the tissue inflammation and destruction in experimental periodontitis of rats. The maxillary palatal bony surfaces of 18 rats received injections of lipopolysaccharide (LPS, 5 mg/mL), PBS or LPS-plus-paeonol (40 mg/kg, intra-peritoneal injection) for three days. Five days later, the osteoclasts were examined and compared after tartrate-resistant acid phosphatase staining. In another 36 rats, the experimental periodontitis was induced by placing the ligatures around the maxillary second and mandibular first molars. Seven days later, the periodontal destruction and inflammation in rats with paeonol (40 mg/kg or 80 mg/kg) and those who had no ligature or without paeonol were compared by dental radiography, micro-computerized tomography (micro-CT), and histology. Gingival mRNA expressions of pre-inflammatory cytokines, including IL-1ß' IL-6 and TNF-α were also examined. Compared to the effect of the LPS positive control, the paeonol injection significantly reduced the induced osteoclast formation. In ligature-induced periodontitis, the periodontal bone supporting ratio was significantly higher in the ligature-plus-paeonol groups compared to that of the ligature group, although they were still less than those in the non-ligature group. By micro-CT and by histology/histometry, a consistent anti-destructive effect was observed when paeonol was added. Moreover, less amount of inflammatory cell-infiltrated connective tissue area, connective tissue attachment, and mRNA expressions of pro-inflammatory cytokines were presented in the ligature-plus-paeonol groups than those in the ligature group. These results suggested that paeonol might have a protective potential on gingival tissue inflammation and alveolar bone loss during the process of periodontitis by inhibiting pro-inflammatory cytokines.

Effects of enamel matrix derivative on the proliferation and osteogenic differentiation of human gingival mesenchymal stem cells.

INTRODUCTION: Gingiva-derived mesenchymal stem cells (GMSCs) have recently been harvested and applied for rebuilding lost periodontal tissue. Enamel matrix derivative (EMD) has been used for periodontal regeneration and the formation of new cementum with inserting collagen fibers; however, alveolar bone formation is minimal. Recently, EMD has been shown to enhance the proliferation and mineralization of human bone marrow mesenchymal stem cells. Because the gingival flap is the major component to cover the surgical wound, the effects of EMD on the proliferation and mineralization of GMSCs were evaluated in the present study. METHODS: After single cell suspension, the GMSCs were isolated from the connective tissues of human gingiva. The colony forming unit assay of the isolated GMSCs was measured. The expression of stem cell markers was examined by flow cytometry. The cellular telomerase activity was identified by polymerase chain reaction (PCR). The osteogenic, adipogenic and neural differentiations of the GMSCs were further examined. The cell proliferation was determined by MTS assay, while the expression of mRNA and protein for mineralization (including core binding factor alpha, cbfα-1; alkaline phosphatase, ALP; and osteocalcin, OC; ameloblastin, AMBN) were analyzed by real time-PCR, enzyme activity and confocal laser scanning microscopy. RESULTS: The cell colonies could be easily identified and the colony forming rates and the telomerase activities increased after passaging. The GMSCs expressed high levels of surface markers for CD73, CD90, and CD105, but showed low expression of STRO-1. Osteogenic, adipogenic and neural differentiations were successfully induced. The proliferation of GMSCs was increased after EMD treatment. ALP mRNA was significantly augmented by treating with EMD for 3 hours, whereas AMBN mRNA was significantly increased at 6 hours after EMD treatment. The gene expression of OC was enhanced at the dose of 100 µg/ml EMD at day 3. Increased protein expression for cbfα-1 at day 3, for ALP at day 5 and 7, and for OC at week 4 after the EMD treatments were observed. CONCLUSIONS: Human GMSCs could be successfully isolated and identified. EMD treatments not only induced the proliferation of GMSCs but also enhanced their osteogenic differentiation after induction.

Bifid mandibular canals and the factors associated with their presence: a medical computed tomography evaluation in a Taiwanese population.

OBJECTIVES: To examine distribution of bifid mandibular canals in a Taiwanese population and to evaluate factors contributing to the phenomenon. MATERIAL AND METHODS: Computed tomographic images from 173 subjects (97 females and 76 males) were obtained using a 64-slice multidetector computerized tomography system, and the presence of bifid mandibular canals, as well as their widths and lengths, was examined. Association of length of bifid canals with possible contributing factors, including gender, age, and side of presentation, as well as size of cross-sectional bony area of mandible along the long axis of mandibular canal, was evaluated. RESULTS: Bifid mandibular canals, with mean values of 10.1 and 0.9 mm in length and width, were found in 53 (30.6%) of 173 patients and 64 (18.5%) of 346 hemi-mandibles. Bifid canals appeared more frequently and tend to penetrate mandible with greater lengths in males if compared with those in females. When males were compared with females and when mandibles with bifid canals were compared with ones without, the former tend to present with larger bony area at corresponding levels of cross-sectional plane than the later, respectively. By regression analysis, significant association was found between length of bifid canals and gender, side of hemi-mandible, and bony area at mid-zone of mandibular canal. CONCLUSIONS: Bifid canals were observed in 30.6% of subjects and 18.5% of hemi-mandibles. Significant association between length of bifid canals and gender, side of hemi-mandible, and cross-sectional bony area of mandible was observed.

Berberine's effect on periodontal tissue degradation by matrix metalloproteinases: an in vitro and in vivo experiment.

Periodontal disease involves tissue destruction caused by interactions among bacterial antigens and inflammatory mediators including matrix metalloproteinases (MMPs). Berberine, an isoquinoline alkaloid isolated from medicinal herbs, can inhibit the degradative action of extracellular MMPs. The effect of berberine on the periodontal expression of MMPs was examined in vitro and in vivo. Gelatinolytic activity of pro-MMP-2, MMP-2, and MMP-9 in the human gingival fibroblast and/or U-937 was compared after treatment with Porphyromonas gingivalis lipopolysaccharide (P.g. LPS) in four medias containing 0, 1, 10 and 100µM of berberine each. Twelve animals were divided into three groups for the study: (A) non-ligation, (B) ligation, and (C) ligation-plus-berberine (75mg/kg berberine by gastric lavage daily); and the effect of berberine on periodontal destruction was evaluated in the ligature-induced periodontitis in rats for 8 days by micro computerized tomography (micro-CT), histology and immunohistochemistry (IHC). An enhancing effect of P.g. LPS on MMP activities was identified, with a greater effect on fibroblasts/U937 co-culture than on either culture alone. When berberine was added to the LPS-treated cultures, the activities of MMPs were significantly reduced in dose-dependent manner. In the animals, the trends of the following parameters were compared. 1. Micro-CT distances between cemento-enamel junction (CEJ) and dental alveolar bone crest: B>C>A. 2. Histometrically measured crest bone levels: B>C>A. 3. Amount of collagen deposited in tissue areas: A>C>B. 4. Attachment loss: B>C≈A. 5. Connective tissue (CT) attachment: B>either A or C. 6. Expression of cells stained positive for MMP-2 and -9 by IHC: B>C>A. In conclusion, berberine demonstrated in vitro an inhibitory effect on P.g. LPS-enhanced MMP activities of HGF and U937 macrophages, reducing in vivo gingival tissue degradation in periodontitic rats. We thus propose that berberine may slow periodontal degradation through the regulation of MMPs in periodontitis induced by bacterial plaque.