Mesenchymal Stem Cell Properties of Periodontal Ligament Cells From Deciduous and Permanent Teeth

Background: Human postnatal stem cells have been identified in periodontal ligaments (PDLs). In this study, the in vitro biologic properties of CD105(+) enriched cell subsets from PDLs harvested from deciduous (DePDL) and permanent (PePDL) teeth are comparatively assessed. Methods: PDL tissue was obtained from 12 teeth (six primary and six permanent) from which CD105(+) CD34(-) CD45(-) cells were isolated by magnetic cell sorting. To identify and quantitatively compare the stem cell markers, DePDL and PePDL cells were assessed for CD166 surface antigen expression by flow cytometry, real-time polymerase chain reaction, and immunostaining for Stro-1 and Oct-4, osteogenic and adipogenic differentiation, and proliferation rate by trypan blue method. Results: Magnetic cell sorting isolated cell populations containing 23.87% (+/- 11.98%) and 11.68% (+/- 6.27%) of CD105(+) expressing cells from PePDL and DePDL, respectively. Flow cytometric analysis demonstrated a higher proportion of CD105(+) cells coexpressing CD166 surface antigen in PePDL, whereas immunostaining and real-time polymerase chain reaction analysis demonstrated that both cell subsets expressed Stro-1 and Oct-4. DePDL-CD105(+) subsets were more proliferative compared to PePDL subsets, and both cell populations showed multipotential capabilities to differentiate in vitro to osteoblast/cementoblast- and adipocyte-like cells. However, a higher expression of adipogenic-related genes was observed in DePDL cells, whereas PePDL-CD105(+) cell subset presented a more homogeneous osteoblast/cementoblast response. Conclusion: These findings demonstrate that highly purified mesenchymal progenitor cell subsets can be obtained from the PDLs of both deciduous and permanent teeth, and further indicate phenotype dissimilarities that may have an impact on their clinical applications. J Periodontol 2010;81:1207-1215.

Effects of periodontal therapy on glycemic control and inflammatory markers

Background: Periodontitis, a complication of diabetes mellitus (DM), can induce or perpetuate systemic conditions. This double-masked, placebo-controlled study evaluated the effects of periodontal therapy (scaling and root planing [SRP]) on the serum levels of glycated hemoglobin (HbA1c) and on inflammatory biomarkers. Methods: Thirty subjects with type 2 DM and periodontitis were treated with SRP + placebo (SRP; N = 15) or with SRP + doxycycline (SRP+Doxy; N = 15), 100 mg/day, for 14 days. Clinical and laboratory data were recorded at baseline and at 3 months after treatment. Results: After 3 months, the reduction in probing depth Was 0.8 mm for the SRP group (P <0.01) and 1.1 mm for the SRP+Doxy group (P <0.01) followed by a 0.9% (SRP; P = 0.17) and 1.5% (SRP+Doxy; P<0.01) reduction in HbA1c levels. A significant reduction in interleukin (IL)-6; interferon-inducible protein 10; soluble fas ligand; granulocyte colony-stimulating factor; RANTES; and IL-12 p70 serum levels were also verified (N = 30). To our knowledge, this is the first report on the effects of periodontal therapy on multiple systemic inflammatory markers in DM. Conclusions: Periodontal therapy may influence the systemic conditions of patients with type 2 DM, but no statistical difference was observed with the adjunctive systemic doxycycline therapy. Moreover, it is possible that the observed improvement in glycemic control and in the reduction of inflammatory markers could also be due to diet, which was not controlled in our study. Therefore, a confirmatory study with a larger sample size and controlled diet is necessary.

Morphologic evaluation and expression of matrix metalloproteinases-2 and 9 and nitric oxide during experimental periodontal disease in rat

The immunopathologic and inflammatory mechanisms involved in periodontal disease (PD) include the participation of host resident, inflammatory cells and chemical mediators. Metalloproteinases (MMPs) and nitric oxide (NO) play essential role in extracellular matrix turnover of periodontal tissue destruction. In this study, by means of RT-PCR through semi-quantitative densitometric scanning methods, the expression of MMPs -2 and -9 and inducible NO synthase (iNOS) was temporally and spatially investigated during the destructive mechanisms of experimentally induced PD in rats. Samples from different periods were microscopically analyzed and compared with the contralateral side (control). Our results showed significant expression of MMP-9 and iNOS in tissues affected by PD, as compared with controls, three days after PD induction, simultaneously with the beginning of alveolar bone loss. At 7 days post induction, only the MMP-9 mRNA presented a significantly higher expression, as compared with the respective controls. Thus, in the rat ligature-induced PD, MMP-9 and iNOS might importantly participate in the early stages of the disease, including inflammatory cell migration, tissue destruction and alveolar bone resorption. Also, we may suggest that the exuberant presence of PMNs may be related to the important expression of iNOS and MMP-9 found at 3 days post induction.

Down-regulation of expression of osteoblast and osteocyte markers in periodontal tissues associated with the spontaneous alveolar bone loss of interleukin-10 knockout mice

The aim of this study was to unravel the mechanisms by which interleukin (IL)-10, a potent pleiotropic cytokine, modulates alveolar bone homeostasis in C57BL/6 wild-type (WT) and IL-10 knockout (IL-10 KO) mice, evaluated at 8, 24, and 48 wk of age. Interleukin-10 KO mice presented significant alveolar bone loss when compared with WT mice, and this was not associated with changes in leukocyte counts or bacterial load. The levels of expression of messenger RNA (mRNA) for tumor necrosis factor-alpha (TNF-alpha), IL-1 beta, IL-6, transforming growth factor-beta (TGF-beta), receptor activator of nuclear factor kappa B ligand (RANKL), osteoprotegerin (OPG), and matrix metalloproteinase 13 (MMP13) were similar between both strains, whereas a significant decrease of tissue inhibitor of metalloproteinase 1 (TIMP1) mRNA expression was found at 48 wk in IL-10 KO mice. The osteoblast markers core binding factor alpha1 (CBFA1) and type I collagen (COL-I) were expressed at similar levels in both strains, whereas the levels of alkaline phosphatase (ALP) and osteocalcin (OCN), and those of the osteocyte markers phosphate-regulating gene endopeptidases (PHEX) and dentin matrix protein 1 (DMP1) were significantly lower in IL-10 KO mice. Our results demonstrate that the alveolar bone loss in the absence of IL-10 was associated with a reduced expression of osteoblast and osteocyte markers, an effect independent of microbial, inflammatory or bone-resorptive pathways.

PPAR-gamma agonist rosiglitazone prevents inflammatory periodontal bone loss by inhibiting osteoclastogenesis

Rosiglitazone (RGZ), an oral anti-hyperglycemic agent used for non-insulin-dependent diabetes mellitus, is a high-affinity synthetic agonist for peroxisome proliferator-activated receptor-gamma (PPAR-gamma). Both in vitro and in vivo experiments have also revealed that RGZ possesses anti-inflammatory properties. Therefore, in the present study, we investigated the anti-inflammatory effects of RGZ in a rat model of periodontal disease induced by ligature placed around the mandible first molars of each animal. Male Wister rats were divided into four groups: 1) animals without ligature placement receiving administration of empty vehicle (control); 2) animals with ligature receiving administration of empty vehicle; 3) animals with ligature receiving administration with oral RGZ (10 mg/kg/day); and 4) animals with ligature receiving administration of subcutaneous RGZ (10 mg/kg/day). Thirty days after induction of periodontal disease, the animals were sacrificed, and mandibles and gingival tissues were removed for further analysis. An in vitro assay was also employed to test the inhibitory effects of RGZ on osteoclastogenesis. Histomorphological and immunohistochemical analyses of periodontal tissue demonstrated that RGZ-treated animals presented decreased bone resorption, along with reduced RANKL expression, compared to those animals with ligature, but treated with empty vehicle. Corresponding to such results obtained from in vivo experiments, RGZ also suppressed in vitro osteoclast differentiation in the presence of RANKL in MOCP-5 osteoclast precursor cells, along with the down-regulation of the expression of RANKL-induced TRAP mRNA. These data indicated that RGZ may suppress the bone resorption by inhibiting RANKL-mediated osteoclastogenesis elicited during the course of experimental periodontitis in rats. (C) 2009 Elsevier B.V. All rights reserved.

Evidence of the presence of T helper type 17 cells in chronic lesions of human periodontal disease

Periodontal disease is a chronic inflammation of the attachment structures of the teeth, triggered by potentially hazardous microorganisms and the consequent immune-inflammatory responses. In humans, the T helper type 17 (Th17) lineage, characterized by interleukin-17 (IL-17) production, develops under transforming growth factor-beta (TGF-beta), IL-1 beta, and IL-6 signaling, while its pool is maintained by IL-23. Although this subset of cells has been implicated in various autoimmune, inflammatory, and bone-destructive conditions, the exact role of T lymphocytes in chronic periodontitis is still controversial. Therefore, in this study we investigated the presence of Th17 cells in human periodontal disease. Gingival and alveolar bone samples from healthy patients and patients with chronic periodontitis were collected and used for the subsequent assays. The messenger RNA expression for the cytokines IL-17, TGF-beta, IL-1 beta, IL-6, and IL-23 in gingiva or IL-17 and receptor activator for nuclear factor-kappa B ligand in alveolar bone was evaluated by real-time polymerase chain reaction. The production of IL-17, TGF-beta, IL-1 beta, IL-6, and IL-23 proteins was evaluated by immunohistochemistry and the presence of Th17 cells in the inflamed gingiva was confirmed by immunofluorescence confocal microscopy for CD4 and IL-17 colocalization. Our data demonstrated elevated levels of IL-17, TGF-beta, IL-1 beta, IL-6, and IL-23 messenger RNA and protein in diseased tissues as well as the presence of Th17 cells in gingiva from patients with periodontitis. Moreover, IL-17 and the bone resorption factor RANKL were abundantly expressed in the alveolar bone of diseased patients, in contrast to low detection in controls. These results provided strong evidence for the presence of Th17 cells in the sites of chronic inflammation in human periodontal disease.

Histological and histomorphometrical alterations of the periodontal ligament in gerbils submitted to teeth extraction

This study verified the effect of unilateral teeth extraction on the periodontal ligament in gerbils (Meriones unguiculatus). Ten adult male gerbils weighing about 50 g had induced occlusal alterations by upper left molar extractions while the other ten animals, only submitted to surgical stress, were considered as controls. The periodontal ligament was characterized by qualitative and quantitative analysis, histological description and histomorphometric quantification. Significant alterations were observed on the left side of the experimental group (P < 0.05), the hypofunctional region, when it was compared with the contralateral side and the corresponding region of the control group. Two months after occlusal alterations induced by unilateral teeth extraction, atrophic histological alterations and a decrease in the periodontal space on the ipsilateral side characterized the periodontal ligament. In this study it was possible to conclude that the gerbil can be used in experimental models attempting to correlate the periodontium`s biological response to various mechanical stresses, as the periodontal ligament was shown to be highly sensitive to occlusal alterations.

Short communication: Identification of subclinical cow mastitis pathogens in milk by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Subclinical mastitis is a common and easily disseminated disease in dairy herds. Its routine diagnosis via bacterial culture and biochemical identification is a difficult and time-consuming process. In this work, we show that matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) allows bacterial identification with high confidence and speed (1 d for bacterial growth and analysis). With the use of MALDI-TOF MS, 33 bacterial culture isolates from milk of different dairy cows from several farms were analyzed, and the results were compared with those obtained by classical biochemical methods. This proof-of-concept case demonstrates the reliability of MALDI-TOF MS bacterial identification, and its increased selectivity as illustrated by the additional identification of coagulase-negative Staphylococcus species and mixed bacterial cultures. Matrix-assisted laser desorption-ionization mass spectrometry considerably accelerates the diagnosis of mastitis pathogens, especially in cases of subclinical mastitis. More immediate and efficient animal management strategies for mastitis and milk quality control in the dairy industry can therefore be applied.

Wild canids, domestic dogs and their pathogens in Southeast Brazil: disease threats for canid conservation

Wild canids are under many pressures, including habitat loss, fragmentation and disease. The current lack of information on the status of wildlife health may hamper conservation efforts in Brazil. In this paper, we examined the prevalence of canine pathogens in 21 free-ranging wild canids, comprising 12 Cerdocyon thous (crab-eating fox), 7 Chrysocyon brachyurus (maned wolf), 2 Lycalopex vetulus (hoary fox), and 70 non-vaccinated domestic dogs from the Serra do Cip National Park area, Southeast Brazil. For wild canids, seroprevalence of antibodies to canine parvovirus, canine adenovirus, canine coronavirus and Toxoplasma gondii was 100 (21/21), 33 (7/21), 5 (1/19) and 68 (13/19) percent, respectively. Antibodies against canine distemper virus, Neospora caninum or Babesia spp. were not found. We tested domestic dogs for antibodies to canine parvovirus, canine distemper virus and Babesia spp., and seroprevalences were 59 (41/70), 66 (46/70), and 42 (40/70) percent, respectively, with significantly higher prevalence in domestic dogs for CDV (P < 0.001) and Babesia spp. (P = 0.002), and in wild canids for CPV (P < 0.001). We report for the first time evidence of exposure to canine coronavirus in wild hoary foxes, and Platynossomun sp. infection in wild maned wolves. Maned wolves are more exposed to helminths than crab-eating foxes, with a higher prevalence of Trichuridae and Ancylostomidae in the area. The most common ectoparasites were Amblyomma cajennense, A. tigrinum, and Pulex irritans. Such data is useful information on infectious diseases of Brazilian wild canids, revealing pathogens as a threat to wild canids in the area. Control measures are discussed.

Serosurvey Of Selected Avian Pathogens In Brazilian Commercial Rheas (Rhea americana) And Ostriches (Struthio camelus)

Ratite farming of has expanded worldwide. Due to the intensive farming methods used by ratite producers, preventive medicine practices should be established. In this context, the surveillance and control of some avian pathogens are essential for the success of the ratite industry; however, little is known on the health status of ratites in Brazil. Therefore, the prevalence of antibodies against Newcastle Disease virus, Chlamydophila psittaci, Mycoplasma gallisepticum, Mycoplasma synoviae, and Salmonella Pullorum were evaluated in 100 serum samples collected from commercial ostriches and in 80 serum samples from commercial rheas reared in Brazil. All sampled animals were clinically healthy. The results showed that all ostriches and rheas were serologically negative to Newcastle disease virus, Chlamydophila psittaci, Myco plasma gallisepticum, and Myco plasma synoviae. Positive antibody responses against Salmonella Pullorum antigen were not detected in ostrich sera, but were detected in two rhea serum samples. These results can be considered as a warning as to the presence of Salmonella spp. in ratite farms. Therefore, the implementation of good health management and surveillance programs in ratite farms may contribute to improve not only animal production, but also public health conditions.

Periodontal attachment loss in an untreated isolated population of Brazil

Background: The aim of this study was to assess the prevalence, extent, and severity of clinical attachment loss (CAL) and to investigate the association of demographic, socioeconomic, and behavioral risk indicators with CAL in an untreated isolated population in Brazil. Methods: All subjects aged >= 12 years were identified by a census. Participants were submitted to a full-mouth clinical examination of six sites per tooth and were interviewed using a structured, written questionnaire. Results: Among the 214 subjects who were interviewed and examined clinically, CAL >= 5 mm in at least one site was observed in 8% of the 12- to 19-year-olds and in all dentate subjects >= 50 years of age; the age-dependent prevalence of CAL >= 7 mm in at least one site ranged from 5% among 12- to 19-year-olds to 83% among subjects >= 50 years old. Multivariate analysis identified plaque (odds ratio [OR] = 2.8), supragingival calculus (OR = 2.9 to 10.6), age >= 30 years (OR = 11.4), and smoking (OR = 2.4) as risk indicators for CAL >= 5 mm and smoking (OR = 8.2) as a risk indicator for CAL >= 7 mm. Conclusions: CAL is highly prevalent in this isolated population. The high occurrence of CAL in young age groups and the confirmation of traditional risk indicators for CAL in this study suggest that other factors, such as host susceptibility, may be needed to explain the high levels of CAL found. Age and behavioral factors were risk indicators associated significantly with the CAL found in this population and may be useful indicators of high-risk subjects for periodontal diseases.

Periodontal disease in gestational and type 1 diabetes mellitus pregnant women

Objective: The present study evaluated the relationship between periodontal disease and its clinical variables in Brazilian non-diabetic pregnant women (C), gestational diabetes mellitus (GDM), or type 1 diabetes mellitus (T1DM). Subjects and methods: A periodontal exam was performed in one hundred and sixty-one pregnant women (GDM:80; T1DM:31; C:50) by a single-blinded calibrated examiner who recorded plaque index (PI), gingival index (GI), bleeding index (BI), gingival margin location (GM), probing depth (PD), clinical attachment level (CAL), bleeding on probing (BOP), and tooth mobility index (MI). The medical variables were age, pregestational body mass index (pre-BMI), fasting plasma glucose (FPG), and glycated hemoglobin (HbA(1c)). Results: The GI, GM, PD, CAL, BOP, and MI were significantly higher (P < 0.01) among GDM and T1DM than for C. The PI was higher in GDM and similar between C and T1DM. The Adjusted Final Model for medical variables to evaluate the effects of groups on periodontal parameters confirmed these results. Conclusions: The presence of periodontal disease was significantly higher in Brazilian diabetic pregnancies (GDM and T1DM) when compared to non-diabetic pregnant women (C). The degree of periodontal disease was similar between the GDM and T1DM groups. Age, pregestational BMI, and HbA(1c) were factors related to CAL development in these two types of diabetes mellitus.

Periodontal disease in adults with untreated congenital growth hormone deficiency: a case-control study

P>Aim The aim of this study was to investigate the possible associations between isolated growth hormone deficiency (IGHD) and periodontal attachment loss (PAL) in adults affected by congenital IGHD. Materials and methods Forty-five previously identified IGHD subjects were eligible for this study. The final study sample comprised 32 cases (gender:20M/12F; age:44.8 +/- 17.5) matched for age, gender, diabetes, smoking status and income to 32 controls (non-IGHD subjects). Participants were submitted to a full-mouth clinical examination of six sites per tooth and were interviewed using a structured, written questionnaire. Periodontitis was defined as proximal PAL >= 5 mm affecting >= 30% of teeth. Results No significant differences were observed in the percentage of sites with visible plaque between IGHD and non-IGHD subjects (59.4% versus 46.9%, p=0.32). IGHD subjects had significant less supragingival calculus (31.3% versus 59.4%, p=0.02) and more bleeding on probing (71.9% versus 18.8%, p < 0.01) than controls. PAL >= 5 mm was significantly more prevalent (100% versus 71.9%, p < 0.01) and affected more teeth (30.5% versus 6.7%, p < 0.01) in cases than in controls. After adjusting for supragingival calculus, IGHD cases had a higher likelihood of having periodontitis than controls (OR=17.4-17.8, 95% CI=2.3-134.9, p=0.004-0.005). Conclusion Congenital IGHD subjects have a greater chance of having PAL.

Release of cytokines by stimulated peripheral blood mononuclear cells in chronic periodontitis

Objective: The emergence of periodontal medicine increased interest in defining the behaviour of peripheral blood cells in periodontitis subjects in comparison with healthy group. The aim of this study was to evaluate the levels of interleukin (IL)-8, tumour necrosis factor-alpha (TNF-alpha), IL-6 and IL-10 released by Escherichia coli lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMC) obtained from the peripheral blood of chronic periodontitis subjects. Design: PBMC samples were isolated from 19 systemically healthy donors, divided into generalized chronic periodontitis (n = 10) and healthy (n = 9) subjects. Cells were incubated for 24-48 h in 500 mu L wells containing RPM! 1640 and stimulated with 1.0 ng/mL of E. coli LPS. Supernatants were used to quantify the amounts of IL-8, TNF-alpha, IL-6 and IL-10 released using enzyme-linked immunosorbent assay (ELISA). Results: PBMC cells from periodontitis subjects released higher levels of TNF-alpha and IL-6 than those from healthy subjects (P < 0.05). Conversely, the supernatants of the stimulated PBMC cells obtained from healthy subjects presented higher amounts of IL-8 than those from periodontitis (P < 0.05). No differences were observed in the levels of IL-10 (P > 0.05) between groups. Conclusion: In conclusion, the results of the present study showed that E. coli LPS-stimulated PBMC from subjects with periodontitis present a different pattern of cytokine release when compared to PBMC from healthy subjects. This phenomenon could have implications locally, in periodontitis, as well as in systemic diseases. (C) 2010 Elsevier Ltd. All rights reserved.

Herpes viruses in periodontal compromised sites: comparison between HIV-positive and -negative patients

Aims: The objective of this study was to compare the frequency of herpes simplex virus type 1 (HSV-1), Epstein-Barr virus (EBV) and human cytomegalovirus (HCMV) in subgingival plaque, saliva and peripheral blood of HIV-positive and-negative patients with periodontal disease. Materials and Methods: Fifty HIV-positive subjects (23 with gingivitis, 27 with periodontitis) and 50 healthy HIV-negative patients with chronic periodontitis were included in the study. Parameters of probing depth (PD), clinical attachment level (CAL), gingival index and plaque index were recorded. The samples were processed for viral identification by the nested polymerase chain reaction technique. Results: HCMV was the most prevalent virus in HIV-positive (82%) and-negative patients (84%), and the detection in the three samples was similar (p > 0.05). HSV-1 was the least prevalent virus in both groups, being detected in similar frequencies in oral sites and in peripheral blood. EBV-1 was found more frequently in saliva and subgingival plaque of HIV-positive patients than in HIV-negative patients (p <= 0.05). Conclusions: EBV-1 was more frequently recovered in oral sites of HIV-positive patients than in HIV-negative patients.