BMP-2 and asporin expression regulate 5-aza-dC-mediated osteoblast/cementoblast differentiation of periodontal dental ligament mesenchymal progenitor cells.

Periodontal dental ligament (PDL) is composed of heterogeneous population of mesenchymal progenitor cells. The mechanisms that regulate the differentiation of these cells towards osteoblast/cementoblast phenotype are not fully understood. Some studies have demonstrated that is possible to change the pattern of cell differentiation via epigenetic mechanisms. The proposal of this study was to investigate whether 5-aza-2'-deoxycytidine (5-aza-dC) treatment would stimulate the osteoblast/cementoblast differentiation of periodontal ligament mesenchymal progenitor cells (PDL-CD105+ enriched cells), characterized as low osteoblast potential, through bone morphogenetic protein-2 (BMP-2) modulation. PDL-CD105+ cells from a single donor were cloned and characterized in two populations as high osteoblast/cementoblast potential (HOP) and low osteoblast/cementoblast potential (LOP) by mineralization in vitro and expression of osteogenic gene markers, such as runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), osteocalcin (OCN), bone morphogenetic protein 2 (BMP-2) and asporin (ASPN). Next, two LOP clones (L1 and L2) were pretreated with 5-aza-dC (10 µM) for 48 h, cultured under osteogenic condition and evaluated for mineralized matrix in vitro, transcription modulation of osteogenic gene markers, methylated and hydroxymethylated DNA levels of BMP-2 and ASPN and intracellular/extracellular expression of BMP-2 protein. LOP clones showed high expression of ASPN transcripts associated with low mRNA levels of BMP-2, RUNX2, ALP, and OCN. 5-aza-dC treatment raised hydroxymethylated DNA levels of BMP-2 and increased the expression of BMP-2 transcripts in both LOP clones. However, BMP-2 protein (intracellular and secreted forms) was detected only in L1 cell clones, in which it was observed an increased expression of osteoblast/cementoblast markers (RUNX2, ALP, OCN) associated with higher mineralization in vitro. In L2 cell clones, 5-aza-dC increased gene expression of ASPN, with no great change in for osteoblast/cementoblast differentiation potential. These data show that 5-aza-dC improves osteoblast/cementoblast differentiation of PDL-CD105+ cells via BMP-2 secretion, and this effect depends on low levels of ASPN expression.

The familial trend of the local inflammatory response in periodontal disease.

OBJECTIVES: The imbalanced host response in front of a dysbiotic biofilm is one of the major aspects of severe periodontitis, which also presents a strong familial aggregation related to the susceptibility factors transmission within family members. This study hypothesized that aggressive periodontitis (GAgP) patients and their descendants could present a similar trend of a local inflammatory response that is different from healthy controls. METHODS: Fifteen GAgP subjects and their children and fifteen healthy subjects and their children were clinically assessed, and the concentration of interferon (IFN)-γ, interleukin (IL)-10, IL-17, IL-1ß, IL-4, IL-6, IL-8, and tumor necrosis factor (TNF)-α was evaluated in the gingival fluid using the multiplexed bead immunoassay. RESULTS: Children from the GAgP group presented lower IL-10 and IFN-γ subgingival concentration than Health children, despite no difference in the clinical parameters. GAgP parents showed a lower IFN-γ, IL-10, and IL-6 than healthy subjects. IL-10/IL-1ß and IFN-γ/IL-4 ratios were reduced in GAgP dyads, suggesting a familial trend in the subgingival cytokine's profile. The cytokines correlated to the clinical data and were predictors of probing depth increase. CONCLUSION: GAgP parents and their children presented a similar cytokine profile and an imbalance in the subgingival response characterized by decreased IFN-γ/IL-4 and IL10/IL-1ß ratios.

Triclosan toothpaste as an adjunct therapy to plaque control in children from periodontitis families: a crossover clinical trial.

OBJECTIVES: Studies have demonstrated that children from aggressive periodontitis (AgP) parents presented precocious alterations in their periodontal condition, and the use of chemical agents in association to plaque control could be useful to control these alterations. This study aimed to evaluate the effect of Triclosan toothpaste to modulate the clinical and subgingival condition in children from AgP parents. METHODS: Fifteen children from AgP parents and 15 from periodontally healthy parents were included in this crossover placebo study. Children were randomly allocated into triclosan or placebo therapy, using selected toothpaste for 45 days. After 15 days of wash-out, groups were crossed, changing the used toothpaste. Clinical examination and saliva, crevicular gingival fluid (GCF), and subgingival biofilm collection were performed at baseline and 45 days of each phase. GCF cytokines' levels were analyzed by Luminex/MAGpix platform and subgingival and salivary periodontal pathogens' levels by qPCR. RESULTS: At baseline, AgP group presented higher plaque index (PI), gingival index (GI), and bleeding on probing (BoP), higher Aggregatibacter actinomycetemcomitans (Aa) abundance in saliva and subgingival biofilm, and lower levels of INF-É£, IL-4, and IL-17 in GCF. Placebo therapy only reduced PI in both groups. Triclosan toothpaste reduced PI and GI in both groups. Triclosan promoted reduction of BoP and probing depth (PD), Aa salivary, and IL-1ß levels in AgP group. In health group, triclosan reduced INF-É£ and IL-4 concentration. CONCLUSION: Triclosan toothpaste demonstrated to be more effective than placebo toothpaste to control the periodontal condition in children from AgP parents, by reducing the BoP, PD, salivary Aa, and IL-1ß. CLINICAL RELEVANCE: Triclosan toothpaste can improve oral conditions in higher-risk population for AgP. TRIAL REGISTRATION: This study was registered at ClinicalTrials.gov with the identifier NCT03642353.

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.

Coronally advanced flap with or without porcine collagen matrix for root coverage: a randomized clinical trial.

OBJECTIVES: The objective of this study is to clinically evaluate the outcomes following treatment of single gingival recessions with either coronally advanced flap technique (CAF) alone or combined with a porcine collagen matrix graft (CM). MATERIALS AND METHODS: This is a randomized parallel design clinical trial, including forty patients with single Miller Class I or II gingival recession, with a depth ≥ 2 mm and located at upper canines or premolars. The patients were randomly assigned to receive either CAF or CAF + CM. The primary outcome variable was gingival recession reduction (Rec Red). RESULTS: Baseline recession depth was 3.14 ± 0.51 mm for CAF group and 3.16 ± 0.65 mm for CAF + CM group (p > 0.05). Both groups showed significant Rec Red (p < 0.05), up to 6 months. Rec Red for CAF + CM was 2.41 ± 0.73 mm and was 2.25 ± 0.50 mm for CAF alone (p > 0.05). Root coverage was 77.2 % in the CAF + CM group and 72.1 % in the CAF group (p > 0.05). Complete root coverage (CRC) was found in 40 % of the cases in the CAF + CM group and in 35 % of the sites treated with CAF. Keratinized tissue thickness (KTT) was 0.26 mm higher in CAF + CM group (p < 0.05). CONCLUSIONS: It can be concluded that CAF + CM does not provide a superior recession reduction when compared to CAF; however, it may offer a small gain in KTT after 6 months. CLINICAL RELEVANCE: CAF + CM can be suggested as a valid therapeutic option to achieve root coverage and some increase in soft tissue thickness after 6 months.

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.

Autologous periodontal ligament cells in the treatment of class II furcation defects: a study in dogs.

AIM: The goal of this study was to histologically investigate the use of periodontal ligament cells (PDL cells) in tissue engineering to regenerate class II furcation defects. MATERIAL AND METHODS: PDL cells were obtained from the mandibular tooth extracted from each dog (seven), cultured in vitro and phenotypically characterized with regard to their biological properties. Following, bilateral class II furcation lesions were created at maxillary 3rd premolars and were randomly assigned to the test group [PDL cells+guided tissue regeneration (GTR)] or the control group (GTR). After 3 months, the animals were euthanized to evaluate the histometric parameters. RESULTS: In vitro, PDL cells were able to promote mineral nodule formation and to express bone sialoprotein, type I collagen and alkaline phosphatase. Histometrically, data analysis demonstrated that the cell-treated group presented a superior length of new cementum (6.00 ± 1.50 and 8.08 ± 1.08 mm), a greater extension of periodontal regeneration (3.94 ± 1.20 and 7.28 ± 1.00 mm), a lower formation of connective tissue/epithelium (2.15 ± 1.92 and 0.60 ± 0.99 mm), a larger area of new bone (7.01 ± 0.61 and 9.02 ± 2.30 mm(2)) and a smaller area of connective tissue/epithelium (5.90 ± 1.67 and 4.22 ± 0.95 mm(2)), when compared with control group. CONCLUSION: PDL cells in association with GTR may significantly promote periodontal regeneration in class II furcation defects in dog.

Sorption, solubility and cytotoxicity of novel antibacterial nanofilled dental adhesive resins.

Dental adhesives hydrolyze in the mouth. This study investigated the water sorption (SOR), solubility (SOL) and cytotoxicity (CYTO) of experimental adhesives containing nitrogen-doped titanium dioxide nanoparticles (N_TiO2). Specimens (n = 15/group [SOR, SOL]; n = 10/group [CYTO]) of unaltered Clearfil SE Protect (CSP), OptiBond Solo Plus (OSP), Adper Scotchbond (ASB) and experimental adhesives (OSP + 25% or 30% of N_TiO2) were fabricated, desiccated (37 °C) and tested for SOR and SOL according to ISO Specification 4049 (2009). CYTO specimens were UV-sterilized (8 J/cm2) and monomer extracted in growth medium (1, 3 or 7 days). Human pulp cells were isolated and seeded (0.5 × 104) for MTT assay. SOR and SOL data was analyzed using GLM and SNK (α = 0.05) and CYTO data was analyzed with Kruskal-Wallis and SNK tests (α = 0.05). SOR and SOL values ranged from 25.80 µg/mm3 (30% N_TiO2) to 28.01 µg/mm3 (OSP) and 23.88 µg/mm3 (30% N_TiO2) to 25.39 µg/mm3 (25% N_TiO2). CYTO results indicated that pulp cells exposed to experimental materials displayed comparable viabilities (p > 0.05) to those of OSP. Experimental materials displayed comparable SOR, SOL and CYTO values (p > 0.05) when compared to unaltered materials. N_TiO2 incorporation have not adversely impacted SOR, SOL and CYTO properties of unaltered adhesives.

Transcriptome profile of highly osteoblastic/cementoblastic periodontal ligament cell clones.

BACKGROUND: Heterogeneous cell populations of osteo/cementoblastic (O/C) or fibroblastic phenotypes constitute the periodontal dental ligament (PDL). A better understanding of these PDL cell subpopulations is essential to propose regenerative approaches based on a sound biological rationale. OBJECTIVE: Our study aimed to clarify the differential transcriptome profile of PDL cells poised to differentiate into the O/C cell lineage. METHODOLOGY: To characterize periodontal-derived cells with distinct differentiation capacities, single-cell-derived clones were isolated from adult human PDL progenitor cells and their potential to differentiate into osteo/cementoblastic (O/C) phenotype (C-O clones) or fibroblastic phenotype (C-F clones) was assessed in vitro. The transcriptome profile of the clonal cell lines in standard medium cultivation was evaluated using next-generation sequencing technology (RNA-seq). Over 230 differentially expressed genes (DEG) were identified, in which C-O clones showed a higher number of upregulated genes (193) and 42 downregulated genes. RESULTS: The upregulated genes were associated with the Cadherin and Wnt signaling pathways as well as annotated biological processes, including "anatomical structure development" and "cell adhesion." Both transcriptome and RT-qPCR showed up-regulation of WNT2, WNT16, and WIF1 in C-O clones. CONCLUSIONS: This comprehensive transcriptomic assessment of human PDL progenitor cells revealed that expression of transcripts related to the biological process "anatomical structure development," Cadherin signaling, and Wnt signaling can identify PDL cells with a higher potential to commit to the O/C phenotype. A better understanding of these pathways and their function in O/C differentiation will help to improve protocols for periodontal regenerative therapies.

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.

Validation of reported GLT6D1 (rs1537415), IL10 (rs6667202), and ANRIL (rs1333048) single nucleotide polymorphisms for aggressive periodontitis in a Brazilian population.

BACKGROUND: Aggressive periodontitis (AgP) is influenced by genetic factors. Recently, the single nucleotide polymorphisms (SNPs) rs1537415 (GLT6D1), rs6667202 (IL10), and rs1333048 (ANRIL) were associated with AgP in different European populations. However, these specific SNPs have not yet been determined in Brazilians. Therefore, this study investigated whether these SNPs previously associated with AgP could be replicated among Brazilians. METHODS: The SNPs rs1537415, rs6667202, and rs1333048 were genotyped using 5'-nuclease allelic discrimination assay in AgP (n = 200), chronic periodontitis (CP, n = 190), and healthy patients (H, n = 196). Differences in allele and genotype frequencies were analyzed using chi-square tests and stepwise logistic regression. RESULTS: The minor C allele of rs6667202 was less frequently detected in AgP patients (23.5%) when compared to non-AgP groups (H = 34.2% and CP = 30.3%; p < 0.01), making the SNP protective against AgP occurrence. Moreover, the final logistic model for AgP diagnosis included gender (p = 0.001) and the SNP rs6667202 (p < 0.001) as significant variables. The SNPs rs1537415 and rs1333048 did not show associations with AgP. CONCLUSION: Only the SNP rs6667202 was associated with AgP in a Brazilian population, being the minor C allele protective against AgP. Moreover, SNPs rs1333048 and rs1537415, previously associated with AgP in other population, was not validated to Brazilian population.

Influence of aging on biological properties of periodontal ligament cells.

The majority of patients eligible for periodontal regenerative therapies are aged subjects. Since periodontal ligament cells (PDLC) are essential for periodontal regeneration, the aim of the present study was to determine the effect of cellular aging on PDLC, including genes associated with extracellular matrix metabolism and growth-associated factors. PDLC cultures were obtained from subjects aged 15 to 20 years and subjects aged more than 60 years. Proliferation, cell viability, mineralization assays, and mRNA levels were assessed for type I and III collagen, platelet-derived growth factor (PDGF)-1, basic fibroblast growth factor (bFGF), metalloproteinase (MMP)-2 and-8, and tissue inhibitor of metalloproteinases (TIMP)-1 and-2. Data analysis demonstrated that aging negatively influenced cell proliferation and mineral nodule formation (p < 0.05). Gene expression analysis further showed that mRNA levels for bFGF, PDGF-1, and TIMP-2 were not affected by aging (p > 0.05). In addition, mRNA levels for type I and III collagen were significantly lower in aged cells (p < 0.05), whereas MMP-2 and-8 and TIMP-1 mRNA levels were higher (p < 0.05). Within the limits of the present study, data analysis suggests that aging modulates important biological properties of periodontal ligament cells, diminishes the potential for mineral nodule formation, and favors extracellular matrix degradation.

Stem cells: potential therapeutics for periodontal regeneration.

This paper is concerned about dental-derived stem cells and their characterization in vitro and in vivo. Additionally, since conventional periodontal techniques remain insufficient to attain complete and reliable periodontal regeneration, the potential of dental-derived stem cells in promoting periodontal tissue regeneration is also reviewed.

Physiological features of periodontal regeneration and approaches for periodontal tissue engineering utilizing periodontal ligament cells.

Experimental studies have shown that the potential of periodontal regeneration seems to be limited by the regenerative capacity of the cells involved. The regeneration of damaged periodontal tissues is mediated by various periodontal cells and is regulated by a vast array of extracellular matrix informational molecules that induce both selective and nonselective responses in different cell lineages and their precursors. In this paper, we first review periodontal ligament tissue and its different cell subpopulations including fibroblasts and paravascular stem cells, and their functions during the development and homeostasis of periodontal tissues. Because conventional periodontal regeneration methods remain insufficient to obtain a complete and reliable periodontal regeneration, the concept of periodontal tissue engineering has been based on the generation of the conditions necessary to improve the healing of periodontal tissues. Additionally, the potential of periodontal ligament cells for use in periodontal tissue engineering to overcome the limitations of conventional periodontal regenerative therapies is discussed, followed by an update of the recent progress and future directions of research utilizing periodontal ligament cells for predictable periodontal regeneration.

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.

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.

Influence of rs6667202 SNP on Interleukin-10 levels in the gingival fluid of patients with periodontitis grade C

Grade C periodontitis in youngers is characterized by a severe form of periodontitis, and IL10 rs6667202 single nucleotide polymorphism (SNP) has been described as an important feature in this disease etiology. Aim: This study aimed to evaluate, in vivo, the functionality of IL10 rs6667202 SNP on IL-10 gingival fluid levels. Methods: Thirty patients with Perio4C were selected, 15 with the IL10 AA genotype (rs6667202) and 15 with AC/CC genotypes. The gingival fluid was collected from two sites with probing depth ≥ 7 mm and bleeding on probing, and two healthy sites. The IL-10 concentration was determined by Luminex/MAGpix platform. Results: In deep pockets, the IL10 AA genotype presented a lower concentration of IL-10 when compared with AC or CC genotypes (p<0.05). In shallow pockets, no difference between groups was seen (p>0.05). Conclusion: IL10 rs6667202 SNP decreases the production of IL-10 in crevicular fluid, potentially affecting this disease progression

Tooth-derived stem cells: Update and perspectives.

Tissue engineering is an emerging field of science that focuses on creating suitable conditions for the regeneration of tissues. The basic components for tissue engineering involve an interactive triad of scaffolds, signaling molecules, and cells. In this context, stem cells (SCs) present the characteristics of self-renewal and differentiation capacity, which make them promising candidates for tissue engineering. Although they present some common markers, such as cluster of differentiation (CD)105, CD146 and STRO-1, SCs derived from various tissues have different patterns in relation to proliferation, clonogenicity, and differentiation abilities in vitro and in vivo. Tooth-derived tissues have been proposed as an accessible source to obtain SCs with limited morbidity, and various tooth-derived SCs (TDSCs) have been isolated and characterized, such as dental pulp SCs, SCs from human exfoliated deciduous teeth, periodontal ligament SCs, dental follicle progenitor cells, SCs from apical papilla, and periodontal ligament of deciduous teeth SCs. However, heterogeneity among these populations has been observed, and the best method to select the most appropriate TDSCs for regeneration approaches has not yet been established. The objective of this review is to outline the current knowledge concerning the various types of TDSCs, and discuss the perspectives for their use in regenerative approaches.

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.

Transcriptome profile of highly osteoblastic/cementoblastic periodontal ligament cell clones

Abstract Heterogeneous cell populations of osteo/cementoblastic (O/C) or fibroblastic phenotypes constitute the periodontal dental ligament (PDL). A better understanding of these PDL cell subpopulations is essential to propose regenerative approaches based on a sound biological rationale. Objective Our study aimed to clarify the differential transcriptome profile of PDL cells poised to differentiate into the O/C cell lineage. Methodology To characterize periodontal-derived cells with distinct differentiation capacities, single-cell-derived clones were isolated from adult human PDL progenitor cells and their potential to differentiate into osteo/cementoblastic (O/C) phenotype (C-O clones) or fibroblastic phenotype (C-F clones) was assessed in vitro. The transcriptome profile of the clonal cell lines in standard medium cultivation was evaluated using next-generation sequencing technology (RNA-seq). Over 230 differentially expressed genes (DEG) were identified, in which C-O clones showed a higher number of upregulated genes (193) and 42 downregulated genes. Results The upregulated genes were associated with the Cadherin and Wnt signaling pathways as well as annotated biological processes, including "anatomical structure development" and "cell adhesion." Both transcriptome and RT-qPCR showed up-regulation of WNT2, WNT16, and WIF1 in C-O clones. Conclusions This comprehensive transcriptomic assessment of human PDL progenitor cells revealed that expression of transcripts related to the biological process "anatomical structure development," Cadherin signaling, and Wnt signaling can identify PDL cells with a higher potential to commit to the O/C phenotype. A better understanding of these pathways and their function in O/C differentiation will help to improve protocols for periodontal regenerative therapies.