Periodontal ligament-derived mesenchymal stem cells modulate neutrophil responses via paracrine mechanisms.

BACKGROUND: Mesenchymal stem cells differentiate into distinct mesenchymal cell lineages and regulate the immune response. The aim of this study was to determine whether periodontal ligament-derived mesenchymal stem cells (PDLSCs) have the ability to modulate neutrophil responses via paracrine mechanisms. METHODS: CD105-enriched PDLSCs were seeded for 24 h and challenged with Porphyromonas gingivalis total protein extract (PgPE) (0 or 2 ug/mL) for 3 h. Cells were then washed and further cultured for 18 h and the supernatants were collected and stored. Next, neutrophil-differentiated human promyelocytic leukemia HL-60 cells (HL60D) were treated with PDLSCs supernatants and HL-60D activation and functional responses were determined. RESULTS: PgPE treatment induced higher secretion of inflammatory markers and chemokines by PDLSCs, including RANTES, eotaxin, interferon (IFN)-γ- inducible protein 10 (IP-10), monocyte chemoattractant protein-1 (MCP-1), IFN-γ, interleukin (IL)-6, IL-8 and IL-1ra (P < 0.05). HL-60D recruitment rate was increased by 4.7 ± 1.09-fold when exposed to PgPE-treated PDLSCs supernatants. PgPE-treated PDLSCs supernatants promoted a 1.78 ± 1.04-fold increase in the production of intracellular reactive oxygen species (ROS) by PMA-stimulated HL-60D, whereas PgPE-untreated PDLSCs supernatants led to a 16% reduction in intracellular ROS. In sharp contrast, neither PgPE-untreated nor PgPE-treated PDLSCs supernatants altered tumor necrosis factor (TNF)-α and IL-1ß secretion by HL-60D cells. CONCLUSION: Together, these findings suggest an important role of PDLSCs in the recognition of P. gingivalis, paracrine recruitment and activation of antimicrobial mechanisms in innate immune cells, without interfering in cytokine responses.

Does enamel matrix derivative application improve clinical outcomes after semilunar flap surgery? A randomized clinical trial.

OBJECTIVES: To evaluate the treatment of gingival recessions by semilunar coronally positioned flap plus enamel matrix derivative (SCPF + EMD). MATERIALS AND METHODS: Thirty patients with class I localized gingival recession were included. They were randomly allocated in two groups: SCPF + EMD and SCPF. Recession height (RH), recession width (RW), width of keratinized tissue (WKT), thickness of keratinized tissue (TKT), probing depth (PD), and clinical attachment level (CAL) were measured at baseline, 6 and 12 months post-surgery. Patient/professional evaluation of esthetics and root sensitivity was performed. RESULTS: After 12 months, mean root coverage was 1.98 ± 0.33 mm for SCPF + EMD (90.86 ± 14.69%) and 1.85 ± 0.41 mm (79.76 ± 17.44%) for SCPF (p > 0.05). The esthetic evaluation by the patient showed preference for SCPF + EMD. According to the professional evaluation (QCE), the use of EMD decreases the appearance of postoperative scar tissue line. There was a significant reduction in root hypersensitivity with no further complaints by the patients. CONCLUSIONS: The addition of EMD provides significantly better esthetics to SCPF, according to patient and professional assessments. SCPF + EMD is effective but not superior to SCPF for root coverage, after 12 months. CLINICAL RELEVANCE: Previous clinical trials showed that the combination of EMD with coronally advanced flaps may enhance the outcome of root coverage. There is a lack of studies testing the combination of EMD with SCPF. The combination SCPF + EMD provides better esthetics when compared to the SCPF and is effective, but not superior, to SCPF for root coverage, after 12 months. TRIAL REGISTRATION: NCT02459704.

Computational analysis for GNAQ mutations: New insights on the molecular etiology of Sturge-Weber syndrome.

Somatic activating mutations in the GNAQ have been recently associated with several congenital genetic disorders and tumors; however, the molecular mechanism/etiology that leads to GNAQ somatic mosaic mutation are unknown. Here, we reported a case of Sturge-Weber Syndrome (SWS) manifesting cutaneous vascular malformations (hemifacial Port-wine stain), cerebral and ocular vascular abnormalities (including epilepsy and glaucoma) and harboring a c.548G>A (p.R183Q) somatic mosaic mutation in GNAQ. Computational modeling studies were performed to assistant with the comprehension of the functional impact of p.R183Q and p.Q209L mutations in GNAQ, which encodes a G protein subunit alpha q (Gαq). The p.R183Q mutation was predicted to abolish hydrogen bonds between R183 residue and GDP molecule, destabilizing the inactive GDP-bound conformation of the Gαq mutants. Furthermore, replacement of R183 by Q183 residue was predicted to promote conformation changes in protein surface features affecting the switch I region, a key region that undergoes conformational changes triggered by receptor binding during signal transduction. In addition, replacement of Q209 by L209 residue was predicted to affect the molecular interaction between Gαq and Gß subunit, impairing formation of the inactive heterotrimeric complex. These findings, in association with PPI network analysis, indicate that p.R183Q and p.Q209L mutations result in the over-activation of different downstream effectors, which in turn will determine the distinct cell responses and phenotype. These findings bring new insights on molecular etiology of vascular malformations associated to SWS and on different mechanisms underlying hyperactivation of downstream pathways to Gαq.

Xenogenous Collagen Matrix and/or Enamel Matrix Derivative for Treatment of Localized Gingival Recessions: A Randomized Clinical Trial. Part II: Patient-Reported Outcomes.

BACKGROUND: Gingival recession (GR) might be associated with patient discomfort due to cervical dentin hypersensitivity (CDH) and esthetic dissatisfaction. The aim is to evaluate the effect of root coverage procedure with a xenogenous collagen matrix (CM) and/or enamel matrix derivative (EMD) in combination with a coronally advanced flap (CAF) on CDH, esthetics, and oral health-related quality of life (OHRQoL) of patients with GR. METHODS: Sixty-eight participants with single Miller Class I/II GRs were treated with CAF (n = 17), CAF + CM (n = 17), CAF + EMD (n = 17), and CAF + CM + EMD (n = 17). CDH was assessed by evaporative stimuli using a visual analog scale (VAS) and a Schiff scale. Esthetics outcome was assessed with VAS and the Questionnaire of Oral Esthetic Satisfaction. Oral Health Impact Profile-14 (OHIP-14) questionnaire was used to assess OHRQoL. All parameters were evaluated at baseline and after 6 months. RESULTS: Intragroup analysis showed statistically significant reduction in CDH and esthetic dissatisfaction with no intergroup significant differences (P >0.05). The impact of oral health on QoL after 6 months was significant for CAF + CM, CAF + EMD, and CAF + CM + EMD (P <0.05). Total OHIP-14 score and psychologic discomfort, psychologic disability, social disability, and handicap dimensions showed negative correlation with esthetics. OHIP-14 physical pain dimension had positive correlation with CDH (P <0.05). OHIP-14 showed no correlation with percentage of root coverage, keratinized tissue width, or keratinized tissue thickness (P >0.05). CONCLUSION: Root coverage procedures improve patient OHRQoL by impacting on a wide range of dimensions, perceived after reduction of CDH and esthetic dissatisfaction of patients with GRs treated with CAF + CM, CAF + EMD, and CAF + CM + EMD.

Viability and Osteogenic Differentiation of Human Periodontal Ligament Progenitor Cells Are Maintained After Incubation With Porphyromonas gingivalis Protein Extract.

BACKGROUND: Porphyromonas gingivalis (Pg) is a major periodontal pathogen that contains immunostimulatory components. Periodontal ligament mesenchymal stem cells (PDLMSCs) are responsible for regeneration of the periodontium that is lost due to periodontitis. Pathologic factors within the microenvironment that impair resident PDLMSCs are not well understood. The present study investigates in vitro the effects of Pg protein extract (PgPE) on biologic properties of CD105-enriched PDL progenitor cell populations (PDL-CD105+). METHODS: Five populations of PDL-CD105+ cells were exposed to PgPE and assessed for cell viability, apoptosis, and proinflammatory gene expression (interleukin-1ß [IL-1ß], tumor necrosis factor-alpha [TNF-α], and IL-6) by quantitative reverse transcription polymerase chain reaction, IL-6 immunostaining, activation of IL-6/signal transducer and activator of transcription (STAT) 3 signaling pathway, and osteogenic differentiation potential. RESULTS: PgPE treatment (2 µg/mL) did not affect cell viability or survival but induced a significant increase in IL-1ß, TNF-α, and IL-6 messenger RNA (mRNA) expression and positive staining for IL-6. A total of 29 genes from the IL-6/STAT3 pathway were upregulated on PgPE stimulation. These genes are related to biologic processes involved in the control of cell survival (B-cell lymphoma 2 [BCL2]), cell proliferation (hepatocytehepatocyte growth factor), cytokine-mediated signaling pathway (suppressor of cytokine signaling 3, C-X-C ligand 8 [CXCL8]), and response to stress (CXCL8, mitogen-activated protein kinase 3, BCL2-associated X protein, and BCL2). Additionally, PgPE treatment caused an increase in alkaline phosphatase mRNA expression in PDL-CD105+ cells after 7 days of osteogenic induction, although mineral nodule formation was comparable to the control group. CONCLUSIONS: These results suggest that the inflammatory profile induced by PgPE treatment in PDL-CD105+ cells did not affect cell viability, apoptosis, or osteogenic differentiation, perhaps due to increased expression of genes involved in the control of cell proliferation and protection against cell death.

Osteogenic potential of periodontal ligament stem cells are unaffected after exposure to lipopolysaccharides.

Periodontitis develops as a result of a continuous interaction between host cells and subgingival pathogenic bacteria. The periodontium has a limited capacity for regeneration, probably due to changes in periodontal ligament stem cells (PDLSCs) phenotype. The aim of this study was to evaluate the effects of lipopolysaccharides from Porphyromonas gingivalis (PgLPS) on mesenchymal phenotype and osteoblast/cementoblast (O/C) potential of PDLSCs. PDLSCs were assessed for Toll-like receptor 2 (TLR2) expression by immunostaining technique. After, cells were exposed to PgLPS, and the following assays were carried out: (i) cell metabolic activity using MTS; (ii) gene expression for IL-1ß, TNF-α and OCT-4 by real-time polymerase chain reaction (RT-qPCR); (iii) flow cytometry for STRO-1 and CD105, and (iv) osteogenic differentiation. PDLSCs were positive for TLR2. PgLPS promoted cell proliferation, produced IL-1ß and TNF-α, and did not affect the expression of stem cell markers, STRO-1, CD105 and OCT-4. Under osteogenic condition, PDLSCs exposed to PgLPS showed a similar potential to differentiate toward osteoblast/cementoblast phenotype compared to control group as revealed by mineralized matrix deposition and levels of transcripts for RUNX2, ALP and OCN. These results provide evidence that PgLPS induces pro-inflammatory cytokines, but does not change the mesenchymal phenotype and osteoblast/cementoblast differentiation potential of PDLSCs.

Exposure of periodontal ligament progenitor cells to lipopolysaccharide from Escherichia coli changes osteoblast differentiation pattern.

UNLABELLED: Periodontal ligament mesenchymal stem cells (PDLMSCs) are an important alternative source of adult stem cells and may be applied for periodontal tissue regeneration, neuroregenerative medicine, and heart valve tissue engineering. However, little is known about the impact of bacterial toxins on the biological properties of PDLSMSCs, including self-renewal, differentiation, and synthesis of extracellular matrix. OBJECTIVE: This study investigated whether proliferation, expression of pro-inflammatory cytokines, and osteogenic differentiation of CD105-enriched PDL progenitor cell populations (PDL-CD105(+) cells) would be affected by exposure to bacterial lipopolysaccharide from Escherichia coli (EcLPS). MATERIAL AND METHODS: Toll-like receptor 4 (TLR4) expression was assessed in PDL-CD105(+) cells by the immunostaining technique and confirmed using Western blotting assay. Afterwards, these cells were exposed to EcLPS, and the following assays were carried out: (i) cell viability using MTS; (ii) expression of the interleukin-1 beta (IL-1ß), interleukin-6 (IL-6), interleukin-8 (IL-8), and tumor necrosis factor alpha (TNF-α) genes; (iii) osteoblast differentiation assessed by mineralization in vitro, and by mRNA levels of run-related transcription factor-2 (RUNX2), alkaline phosphatase (ALP) and osteocalcin (OCN) determined by quantitative PCR. RESULTS: PDL-CD105+ cells were identified as positive for TLR4. EcLPS did not affect cell viability, but induced a significant increase of transcripts for IL-6 and IL-8. Under osteogenic condition, PDL-CD105+ cells exposed to EcLPS presented an increase of mineralized matrix deposition and higher RUNX2 and ALP mRNA levels when compared to the control group. CONCLUSIONS: These results provide evidence that CD105-enriched PDL progenitor cells are able to adapt to continuous Escherichia coli endotoxin challenge, leading to an upregulation of osteogenic activities.

Intermittent parathyroid hormone administration improves periodontal healing in rats.

BACKGROUND: Intermittent administration of parathyroid hormone (PTH) promotes new bone formation in patients with osteoporosis and bone fractures. It was shown previously that PTH also reduces periodontitis-related bone loss. The aim of this study is to evaluate the effect of treatment with PTH on periodontal healing in rats. METHODS: Fenestration defects were created at the buccal surface of the distal root of the mandibular first molars, and both periodontal ligament (PDL) and cementum were removed. Animals were then assigned to two groups (eight animals per group): group 1: control, placebo administration; and group 2: test, human PTH (hPTH) 1-34 administration at a concentration of 40 µg/kg. For both groups, the animals were injected every 2 days, and the animals were sacrificed at 14 and 21 days after surgery. Specimens were harvested and processed for routine decalcified histologic sections. The following parameters were assessed: 1) remaining bone defect extension (RBDE); 2) newly formed bone density (NFBD); 3) total callus area (TCA); 4) osteoclast number (ON) in the callus region; and 5) newly formed dental cementum-like tissue (NFC). Birefringence of root PDL reattachment was also evaluated. RESULTS: Birefringence analysis showed root PDL reattachment for both groups 21 days after treatment. Intermittent hPTH 1-34 administration decreased RBDE (P <0.01) and increased NFBD (P <0.01), TCA (P <0.01), area of NFC (P <0.01), and ON in the callus region (P <0.01). CONCLUSION: Within the limits of the present study, intermittent administration of hPTH 1-34 led to an enhanced periodontal healing process compared with non-treated animals.

One stage, full-mouth, ultrasonic debridement in the treatment of severe chronic periodontitis in smokers: a preliminary, blind and randomized clinical trial.

OBJECTIVE: The aim of this clinical trial was to assess the performance of a full-mouth ultrasonic debridement protocol in the treatment of severe chronic periodontitis in comparison with scaling and root planing in a quadrant-wise procedure in smokers. MATERIALS AND METHODS: The trial consisted of 30 participants presenting with periodontitis divided into 3 groups: Group FMUD - full-mouth ultrasonic debridement, i.e., one session of 45 minutes of ultrasonic instrumentation for smokers (n = 10), Group SRP- scaling and root planing performed in a quadrant-wise manner for smokers (n = 10), and Group Control - SRP for nonsmokers (n = 10), treated following the same protocol as the SRP group. The parameters evaluated were: plaque/bleeding on probing indices, probing pocket depth, relative recession, and relative probing attachment level at baseline, 45, 90 and 180 days after therapy. RESULTS: Full-mouth ultrasonic debridement and scaling and root planing resulted in comparable gain of attachment 6 months after therapy. Both groups exhibited probing pocket depth reduction at all experimental periods as compared to baseline. Smokers, however, had less probing pocket depth reduction and relative probing attachment level gain compared to non-smokers, despite the mechanical protocol used (p < 0.05). Moreover, at 180 days, nonsmokers presented with fewer sites requiring re-treatment (probing pocket depth > 5 mm and bleeding on probing) than smokers (p < 0.05). CONCLUSIONS: Full-mouth ultrasonic debridement and scaling and root planing result in comparable clinical outcomes for the treatment of smokers with severe chronic periodontitis. Despite the non-surgical technique used, smokers had a less favorable clinical response than non-smokers.

Healing patterns after subgingival placement of a resin-modified glass-ionomer restoration: a histometric study in dogs.

The aim of this study was to evaluate, clinically and histometrically, the effects of subgingival placement of a resin-modified glass-ionomer restoration during flap surgery. Nine dogs were included in this study. The mandibular canines were randomly assigned to receive either a transgingival resin-modified glass-ionomer restoration (test group) or no restoration (control group). The apical margins of the restorations in the test group and a reference notch on those in the control group were placed at the level of the bone crest. Clinical parameters were recorded 7 days before sacrifice. The dogs were sacrificed after 107 days, and undecalcified sections were obtained for histologic evaluation. Clinically, both groups presented significant clinical attachment loss and an increase in probing depth, but differences between groups were not statistically significant (P > .05). Histologically, a significant difference between groups was observed for length of epithelium (test, 4.05 ± 0.57 mm; control, 3.36 ± 0.63 mm; P = .01). The test group showed more bone resorption (2.02 ± 1.47 mm) when compared with the control group (0.74 ± 0.37 mm) (P = .048). It can be concluded that even with the claimed favorable properties of resin-modified glass ionomer, the presence of the restoration within the biologic width causes increased migration of the apical epithelium and bone resorption.

Parathyroid hormone (1-34) compensates the negative effect of smoking around implants.

OBJECTIVES: This study aimed to investigate the effect of Recombinant Human Parathyroid Hormone (PTH 1-34) on attenuating the influence of cigarette smoke on bone around titanium implants. MATERIAL AND METHODS: Forty-eight female Wistar rats were used. At the beginning of the study, 15 animals were randomly assigned to Group 1 (control) and received subcutaneous injections of saline solution, three-times/week, after implant placement. The other animals received intermittent cigarette smoke inhalation (CSI), 60 days prior and 60 days after implant placement ( Al 2 O 3 -blasted titanium implants - 4.0 × 2.2 mm). After surgery, these animals were randomly assigned to: Group 2 - subcutaneous injections of saline solution, three-times/week (n = 16) and Group 3 - intermittent doses of PTH (1-34) (40 µg/Kg), three-times/week (n = 17). Animals were sacrificed 60 days after surgery, and degree of bone-to-implant contact (BIC), bone area (BA) within the limits of the threads and proportion of mineralized tissue (PMT) adjacent to the implants (500 µm wide zone) were separately obtained in cortical and cancellous bone. RESULTS: Data analysis confirmed that CSI negatively affects bone around implants, as observed for BIC in cortical zone (Cohen's d (d) = -1.26) and for PMT in both zones (d = -6.09 and d = -4.46 for cortical and cancellous zones, respectively). In addition, in the presence of CSI, PTH (1-34) promoted the highest BIC in both regions and BA and PMT in cancellous bone (P < 0.05). The histometric parameter that was not influenced by both PTH and CSI (1-34) was BA in cortical bone (P > 0.05). CONCLUSION: In the presence of cigarette smoke, a factor related to poor bone healing and low bone density, PTH (1-34) increased bone volume around implants.

Autologous periodontal ligament cells in the treatment of Class III furcation defects: a study in dogs.

AIM: This study aimed to evaluate, histomorphometrically, the use of periodontal ligament cells (PDL cells) in the treatment of class III furcation defects. MATERIAL AND METHODS: PDL cells were obtained from the mandibular tooth extracted from each dog (7), cultured in vitro and phenotypically characterized. Bilateral class III furcation defects were created at mandibular 3rd and 4th premolars and were randomly assigned to: CONTROL GROUP: coronally positioned flap, GTR Group: GTR, Sponge Group: carrier + GTR, Cell Group: carrier + PDL cells + GTR. RESULTS: After 3 months of healing, data analysis demonstrated that the Cell Group presented a superior length of new cementum (4.82 ± 0.61 mm; 3.66 ± 0.95 mm; 2.87 ± 0.74 mm and 1.70 ± 0.60 mm, p < 0.001), a greater extension of periodontal regeneration (3.43 ± 1.44 mm; 2.33 ± 0.95 mm; 1.52 ± 0.39 mm and 0.69 ± 0.59 mm, p = 0.001) and a larger area of new bone (5.45 ± 1.58 mm(2) ; 3.94 ± 1.52 mm(2) ; 2.91 ± 0.56 mm(2) and 1.89 ± 0.95 mm(2) , p = 0.0012), when compared with Sponge, GTR and CONTROL GROUP, respectively. CONCLUSION: The PDL cells in association with GTR may significantly promote periodontal regeneration in class III furcation defects surgically created in dogs.

Steroidal and non-steroidal cyclooxygenase-2 inhibitor anti-inflammatory drugs as pre-emptive medication in patients undergoing periodontal surgery.

The aim of the present study was to compare the pre-emptive use of a cyclooxygenase-2 (COX-2) inhibitor with a well established steroidal anti-inflammatory drug for pain and edema relief following periodontal surgery for crown lengthening. Thirty patients requiring periodontal surgery were randomly assigned to receive one of the following medications: selective COX-2 inhibitor or steroidal anti-inflammatory drug, 60 min before the surgical procedure. To examine patient anxiety, a Corah's dental anxiety scale was applied before surgery. Using a visual analog scale, the extent of pain/discomfort during the trans-operative period and immediately after the surgery was measured. Additionally, intensity of pain/discomfort and edema were examined 4, 8, 12 and 24 h postoperatively. With regard to anxiety, no statistical differences between the groups were observed (p>0.05). With respect to the extent of pain/discomfort during the trans-operative, immediate and late postoperative period, data demonstrated no significant differences (p>0.05) between the COX-2 inhibitor and steroidal groups. With regard to edema, intragroup analysis did not reveal any statistically significant difference (p>0.05) during the 24 h following surgery in either group. In conclusion, both anti-inflammatory drugs presented a similar potential for pain and edema relief following periodontal surgery.

TLR2 and TLR4 gene promoter methylation status during chronic periodontitis.

AIM: The objective of this study was to analyse the status of DNA methylation in the promoter region of the toll-like receptor (TLR)2 and TLR4 genes in gingival tissue samples from healthy subjects, smokers and non-smokers affected by chronic periodontitis. MATERIAL AND METHODS: Genomic DNA and total RNA were purified from gingival tissue using the TRIZOL reagent protocol. Genomic DNA was then digested by methylation-sensitive restriction enzymes, amplified by polymerase chain reaction (PCR), electrophoresed on a 10% polyacrylamide gel and stained using SYBR Gold. Real-time PCR was also performed to verify the transcript levels. RESULTS: The CpG dinucleotides analysed were observed to be unmethylated in the majority of DNA samples of the three groups and statistical differences were not found among groups (p>0.05). However, a trend towards methylation was observed in the TLR2 HhaI site in the samples of the periodontitis non-smoker groups. In fact, the analysis of all CpG sites together shows which complete methylation is observed in the shortest level in the samples of periodontitis non-smoker group. The analysis of transcript levels demonstrated no difference among groups (p>0.05). CONCLUSION: The results demonstrated major unmethylation of the TLR4 gene promoter in all groups. However, the results for the TLR2 gene promoter are inconclusive; this gene was found as a mosaic of methylated and unmethylated DNA in the majority of samples of the three groups and we also observed a trend towards the DNA methylation of CpG sites recognized by the HhaI enzyme.

Cigarette smoke inhalation modulates gene expression in sites of bone healing: a study in rats.

OBJECTIVE: The aim of this study was to assess the effect of cigarette smoke inhalation (CSI) on gene expression in alveolar bone healing sites. STUDY DESIGN: Wistar rats were randomly assigned to the groups: control [animals not exposed to CSI (n = 20)] and test [animals exposed to CSI, starting 3 days before teeth extraction and maintained until killing them (n = 20)]. First mandibular molars were bilaterally extracted, and the expression of alkaline phosphatase, bone morphogenetic protein (BMP) 2 and 7, receptor activator of nuclear factor κB ligand, osteoprotegerin, and d2 isoform of vacuolar adenosine triphosphatase V(0) domain were assessed by quantitative polymerase chain reaction in the newly formed tissue in the sockets. RESULTS: Overall, data analysis demonstrated that CSI significantly affected the expression pattern of all of the studied genes except BMP-7. CONCLUSION: The expression of key genes for bone healing may be affected by CSI in tooth extraction sites.

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.

Effect of aluminum oxide-blasted implant surface on the bone healing around implants in rats submitted to continuous administration of selective cyclooxygenase-2 inhibitors.

PURPOSE: The continual use of selective cyclooxygenase-2 (COX-2) inhibitors may have a negative impact on bone repair around titanium implants. Because modified implant surfaces could be considered an important strategy to increase success rates in some conditions that interfere in bone healing, the aim of this study was to investigate whether an aluminum oxide (Al2O3)-blasted implant surface could reduce the negative action promoted by the continuous administration of selective COX-2 inhibitors on bone healing around implants. MATERIALS AND METHODS: Thirty Wistar rats received one titanium implant (machined or Al2O3-blasted surface) in each tibia and were randomly assigned to one of the following groups: saline (n = 14) or meloxicam (n = 16); each was administered daily for 60 days. Bone-to-implant contact (BIC), bone area (BA) within the limits of threads, and bone density (BD) in a zone lateral to the implant were examined in undecalcified sections. RESULTS: The Al2O3-blasted surface resulted in significantly increased BIC in both groups, and meloxicam significantly reduced bone healing around implants (P < .05). For the machined surface, significant differences were observed for BIC (39.48 +/- 10.18; 25.23 +/- 9.29), BA (60.62 +/- 4.09; 42.94 +/- 8.12), and BD (56.31 +/- 3.64; 49.30 +/- 3.15) in the saline and meloxicam groups, respectively. For the Al2O3-blasted surface, data analysis also demonstrated significant differences for BIC (45.92 +/- 11.34; 33.30 +/- 7.56), BA (61.04 +/- 4.39; 44.89 +/- 7.11), and BD (58.77 +/- 2.93; 50.04 +/- 3.94) for the saline and meloxicam groups, respectively. CONCLUSIONS: The Al2O3-blasted surface may increase BIC; however, it does not reverse the negative effects promoted by a selective COX-2 inhibitor on bone healing around implants.

Inflammatory and bone-related genes are modulated by aging in human periodontal ligament cells.

Periodontal ligament cells (PDLC) play a major role in periodontal tissues homeostasis and destruction. Most age-associated diseases seem to be closely related to an underlying chronic inflammatory state. Thus, the present study aimed at evaluating in PDLC the effect of aging on the basal levels of inflammatory and bone-related genes. Primary PDLC cultures were obtained from subjects aged 15-20 years (control- n=5), and subjects aged more than 60 years (test- n=5). Proliferation, cell viability and total secreted protein assays were performed, and mRNA levels were quantitatively assessed for interleukin (IL)-1beta, IL-4, IL-6 and IL-8, and for receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) by real time PCR. Data analysis demonstrated that aging negatively influenced cell proliferation, whereas cell viability and total secreted protein were not affected (p>0.05). Gene expression analysis showed that mRNA levels for RANKL and IL-8 were not affected by aging (p>0.05) whereas, mRNA levels for IL-4 was significantly lower in aged cells (p<0.05) and OPG, IL-1beta and IL-6 mRNA levels were higher (p<0.05). Data analysis suggests that aging decreased the ability of PDLC to proliferate and modulated the expression of important inflammatory and bone-related genes in periodontal ligament cells, favoring a proinflammatory and an antiresorptive profile.

Effect of anti-infective mechanical therapy on clinical parameters and cytokine levels in human peri-implant diseases.

BACKGROUND: The objectives of this study were to clinically and immunologically assess the effects of mechanical anti-infective therapies for mucositis and peri-implantitis and to compare the levels of cytokines in untreated and treated peri-implant diseased sites to healthy ones. METHODS: Titanium dental implants were assigned to one of the following groups: healthy (n = 10) = control; mucositis (n = 10) = mechanical debridement using abrasive sodium carbonate air-powder and resin curets; and peri-implantitis (n = 20) = open surgical debridement using abrasive sodium carbonate air-powder and resin curets. Visible plaque accumulation, marginal bleeding, bleeding on probing, suppuration, and probing depth were assessed at baseline for all groups and at 3 months after therapies for diseased groups. At these times, the total amounts of interleukin (IL)-4, -10, and -12, tumor necrosis factor-alpha (TNF-alpha), receptor activator of nuclear factor-kappa B ligand (RANKL), and osteoprotegerin (OPG) in the peri-implant crevicular fluid (PICF) were measured by enzyme-linked immunosorbent assay. RESULTS: At 3 months, the anti-infective treatments resulted in a significant improvement in all clinical parameters for mucositis and peri-implantitis (P <0.05). Moreover, the total amounts of TNF-alpha in PICF were significantly higher in untreated diseased implants compared to healthy ones, and the OPG/RANKL ratio was higher for healthy implants than for untreated peri-implantitis (P <0.05). TNF-alpha levels were significantly reduced for both diseased groups (P <0.05), achieving the same level as the healthy group at 3 months after therapies (P >0.05). CONCLUSION: The proposed anti-infective therapies may locally modulate the levels of TNF-alpha and the OPG/RANKL ratio and improve clinical parameters around peri-implant tissues.

Cannabis sativa smoke inhalation decreases bone filling around titanium implants: a histomorphometric study in rats.

PURPOSE: Although the harmful effect of tobacco smoking on titanium implants has been documented, no studies have investigated the effects of cannabis sativa (marijuana) smoking. Thus, this study investigated whether marijuana smoke influences bone healing around titanium implants. MATERIALS: Thirty Wistar rats were used. After anesthesia, the tibiae surface was exposed and 1 screw-shaped titanium implant was placed bilaterally. The animals were randomly assigned to one of the following groups: control (n = 15) and marijuana smoke inhalation (MSI) 8 min/d (n = 15). Urine samples were obtained to detect the presence of tetra-hidro-cannabinoid. After 60 days, the animals were killed. The degree of bone-to-implant contact and the bone area within the limits of the threads of the implant were measured in the cortical (zone A) and cancellous bone (zone B). RESULTS: Tetra-hidro-cannabinoid in urine was positive only for the rats of MSI group. Intergroup analysis did not indicate differences in zone A-cortical bone (P > 0.01), however, a negative effect of marijuana smoke (MSI group) was observed in zone B-cancellous bone for bone-to-implant contact and bone area (Student's t test, P < 0.01) values. CONCLUSIONS: Considering the limitations of the present study, the deleterious impact of cannabis sativa smoke on bone healing may represent a new concern for implant success/failure.