In vitro comparison of zinc-based, chlorhexidine, and essential oil mouth rinses.

BACKGROUND: Chlorhexidine (CHX)-based mouth rinses are frequently prescribed following periodontal surgeries. A more recently available brand of zinc-based mouth rinses advertises one of its mouth rinses as a substitute for chlorhexidine. The purpose of this study was to evaluate, in vitro, the effects of this brand of zinc-based mouth rinses on cell survival, cell motility, and gene expression of human gingival fibroblasts (HGFs). METHODS: HGFs were exposed to essential oil (EO), CHX, and three types of one brand of zinc-based mouth rinses designed to treat breath malodor (ZnA), dry mouth (ZnB), and gingivitis (ZnC). Each mouth rinse was tested over a range of concentrations for its effects on HGF survival and motility. Gene expression of cytokines, interleukins, and growth factors were evaluated via reverse transcriptase-polymerase chain reaction (RT-PCR), as a means to assess potential influences on inflammation and wound healing. RESULTS: Cell survival was significantly decreased for CHX and ZnC at 10% dilutions (p < 0.05). For all time points, cells exposed to ZnC displayed the greatest reduction in cell motility (p < 0.05). The various mouth rinses examined differentially altered the expression of growth factor transcripts. ZnC particularly enhanced the expression of BMP-2 and FGF-2. CONCLUSION: ZnC was more cytotoxic and inhibited cell motility to a greater extent than any of the other mouth rinses. Therefore, using ZnC as an alternative to CHX could potentially have negative effects on wound healing after periodontal surgery. However, further investigation is required to confirm the clinical relevance of these in vitro findings. PLAIN LANGUAGE SUMMARY: One type of zinc-based mouth rinse designed to replace chlorhexidine (often prescribed after oral surgeries) demonstrated the greatest oral cell death and reduction in cell movement when compared to other zinc-based mouth rinses. These zinc-based mouth rinses also reduced the amounts of proteins involved in regulating inflammation, potentially reducing the destruction of bone holding the teeth in place. They also changed the amounts of several molecules involved in tissue healing. It is unknown if this will speed or slow the healing of the soft tissues of the mouth.

Utilization of a periodontal endoscope in nonsurgical periodontal therapy: A randomized, split-mouth clinical trial.

BACKGROUND: The removal of subgingival calculus to obtain gingival health is an integral part of nonsurgical periodontal therapy. The periodontal endoscope is used by some clinicians to help enhance access to effectively remove subgingival calculus; however, longer-term studies on this subject are still lacking. The purpose of this randomized, controlled clinical trial was to compare the clinical outcomes of scaling and root planing (SRP) using a periodontal endoscope versus conventional SRP using loupes for up to 12 months, utilizing a split-mouth design. METHODS: Twenty-five patients were recruited who exhibited generalized stage II or stage III periodontitis. SRP was rendered by the same experienced hygienist using either a periodontal endoscope or conventional SRP using loupes, following random assignment of the left and right halves of the mouth. All periodontal evaluations were done by the same periodontal resident at baseline, and at 1, 3, 6, and 12 months after therapy. RESULTS: Single-rooted teeth interproximal sites displayed a significantly lower percentage of improved sites (P < 0.05) than multirooted teeth for probing depth and clinical attachment level (CAL). Maxillary multirooted interproximal sites favored the use of the periodontal endoscope at the 3- and 6-month time periods (P = 0.017 and 0.019, respectively) in terms of the percentage of sites with improved CAL. Mandibular multirooted interproximal sites showed more sites with improved CAL using conventional SRP than with the periodontal endoscope (P < 0.05). CONCLUSION: Overall, the use of a periodontal endoscope was more beneficial in multirooted sites compared to single-rooted sites, specifically in maxillary multirooted sites.

Effect of different concentrations of commercially available mouthwashes on wound healing following periodontal surgery: a randomized controlled clinical trial.

OBJECTIVES: The purpose of this study was to evaluate the effect of chlorhexidine and essential oils containing mouth rinses on oral wound healing after periodontal flap surgery. MATERIALS AND METHODS: Eighty subjects participated in the study and were randomly assigned to use water, 0.12% chlorhexidine (CHX), essential oils (EO), 5% CHX, and 10% EO. Subjects were examined at 1, 2, and 3 weeks postoperatively. Plaque index (PI) and the modified gingival index (GI) were recorded, while wound epithelialization was measured to evaluate the healing process. Numerical data were analyzed with parametric test for multiple comparisons (ANOVA) with Bonferroni correction. Categorical data were analyzed using Chi-square test/fisher exact test. RESULTS: All groups demonstrated a gradual GI reduction from first to third visit. Patients in the CHX group presented statistically significant lower PI scores than patients in the water group at the all-time points of the study. Wound epithelialization analysis demonstrated that 100% of the sites in the CHX group were healing by secondary intention at visit 1. This finding was statistically significant. CONCLUSION: Full strength concentrations of CHX and EO did not show any detrimental effects on healing after traditional periodontal surgery at the end of the observation period. CLINICAL RELEVANCE: The use of chlorhexidine and EO containing mouthwashes does not appear to delay wound healing. Diluting these commercial mouthwashes may present an approach that could possibly reduce the adverse effects (such as tooth staining) associated with their use, while maintaining their antibacterial properties.

Cytotoxic effects of silver diamine fluoride.

PURPOSE: To investigate the effect of silver diamine fluoride (SDF) and fluoride varnish (FV) on human gingival fibroblasts (HGF) and bacteria. METHODS: HGF cell viability was assessed after exposure to various dilutions of SDF or FV. Hydroxyapatite (HA) discs treated with SDF, FV, or saline were rinsed in artificial saliva for 84 days. HGF were exposed to treated discs and viability assessed fluorescently. Oral bacteria were exposed to treated discs and survival quantified. RESULTS: At 0.01%, SDF was almost 100% cytotoxic to HGF. SDF and FV treated HA discs, induced near-complete cell death after 24 hours of contact. After rinsing FV discs for 21 days, cell survival exceeded 95%. SDF treated discs were toxic to HGF and bacteria after 9 weeks of rinsing. CLINICAL SIGNIFICANCE: SDF and FV can induce cell death. FV lost its cytotoxicity within 3 weeks, while SDF remained cytotoxic even after 9 weeks of rinsing. This research confirms that SDF has long lasting antimicrobial effects at very low concentrations although it does raise concerns regarding cytotoxicity. However, HGF cells are exposed to other cytotoxic substances in dentistry with little, if any, long-term effects.

The Effects of Irrigants on the Survival of Human Stem Cells of the Apical Papilla, Including Endocyn.

INTRODUCTION: Endocyn, a pH-neutral solution of hypochlorous acid and hypochlorite has been developed for use as an endodontic irrigant. The purpose of this study was to evaluate the effect of Endocyn on human periodontal ligament (PDL) fibroblasts, rat osteosarcoma cells (UMR-106), and stem cells of the apical papilla (SCAP) compared with other commonly used endodontic irrigants. METHODS: To determine cytotoxicity, cells were exposed to various concentrations of Endocyn, 6% sodium hypochlorite (NaOCl), 17% EDTA, and 2% chlorhexidine for 10 minutes, 1 hour, or 24 hours. Cell survival was measured fluorescently using calcein AM. Endocyn also was tested for its ability to inhibit SCAP proliferation and alkaline phosphatase activity. Finally, SCAP transcript expression was examined via reverse-transcriptase polymerase chain reaction. RESULTS: Endocyn was no more toxic to PDL and UMR cells than water for up to 24 hours. Endocyn concentrations of 50% were toxic to SCAP after 1 hour of exposure. Endocyn concentrations of >20% inhibited SCAP proliferation, whereas concentrations of ≥10% inhibited alkaline phosphatase activity. Exposure of SCAP to 10% Endocyn for 3 days did not alter most transcript expression, but did significantly reduce the expression of alkaline phosphatase, fibromodulin, and osteomodulin. CONCLUSION: Endocyn was significantly less cytotoxic to PDL, UMR-106, and SCAP cells compared with other commonly used endodontic irrigants. High concentrations of Endocyn did inhibit some transcript expression and alkaline phosphatase activity, indicating a potential reduction in the osteogenic potential of stems cells exposed to Endocyn.

Greater Sensitivity of Oral Fibroblasts to Smoked Versus Smokeless Tobacco.

BACKGROUND: Smokers have an increased incidence and severity of periodontal disease. Although cigarette smoke contains >4,000 chemical components that could affect periodontal tissues, less is understood about the effect of smokeless tobacco. Therefore, this study compares the effects of cigarette smoke extract (CSE) and smokeless tobacco extract (STE) on cell survival and motility of periodontal ligament (PDL) and gingival fibroblasts in vitro. METHODS: PDL and gingival fibroblasts were exposed to various concentrations of CSE, STE, or nicotine alone. Viable cells were labeled with calcein acetoxymethyl, visualized using fluorescent microscopy, and quantified using a fluorescence multi-well plate reader. In vitro wounding and collagen gel contraction assays were used to assess cell motility. RESULTS: Both gingival and PDL fibroblasts displayed reduced cell viability with increasing concentrations of CSE and STE. Based on relative nicotine content, CSE was significantly more cytotoxic than STE. PDL fibroblasts were also more sensitive to both CSE and STE compared with gingival fibroblasts. Finally, sublethal doses of CSE reduced cell motility and gel contraction, whereas STE had less effect. Nicotine alone ≤0.5 mM had little to no effect in any of these assays. CONCLUSIONS: Many of the underlying effects of tobacco products on periodontal tissues may be due to direct inhibition of normal fibroblast function. CSE is found to be more deleterious to the function of both PDL and gingival fibroblasts than STE. PDL fibroblasts appear to be more sensitive to CSE and STE than gingival fibroblasts. Therefore, cigarette smoke may have more profound effects than smokeless tobacco.

Synthesis, Antifungal Activity, and Biocompatibility of Novel 1,4-Diazabicyclo[2.2.2]Octane (DABCO) Compounds and DABCO-Containing Denture Base Resins.

The fungal pathogen Candida albicans causes a variety of oral infections, including denture stomatitis, which is characterized by inflammation of the oral mucosa in direct contact with dentures and affects a significant number of otherwise healthy denture wearers. While antifungal treatment reduces symptoms, infections are often recurrent. One strategy to address this problem is to incorporate compounds with fungicidal activities into denture materials to prevent colonization. Our laboratory synthesized novel derivatives of 1,4-diazabicyclo[2.2.2]octane (DABCO), which is an organic compound typically used as a catalyst in polymerization reactions. DABCO derivatives with different aliphatic chain lengths (DC16, DC16F, DC18, and C6DC16), as well as methacrylate monomers conjugated to DABCO compounds (DC11MAF and C2DC11MAF), were synthesized and tested for antimicrobial activity. All the compounds exhibited fungicidal activity against several Candida species at concentrations ranging between 2 and 4 µg/ml. Moreover, acrylic denture base resins fabricated to contain 1, 2, or 4 wt% DABCO compounds inhibited surface C. albicans biofilm formation, as well as fungal growth, in disc diffusion assays. Remarkably, discs (4 wt%) aged for 2 months also exhibited approximately 100% growth-inhibitory activity. While some DABCO compounds exerted intermediate to high cytotoxicity against mammalian oral cell types, DC11MAF and denture base resin discs containing 2 or 4 wt% C2DC11MAF exhibited relatively low cytotoxicity against periodontal ligament (PDL) cell and gingival fibroblast (GF) lines, as well as primary oral epithelial cells. These studies demonstrate that DABCO derivatives can be incorporated into denture materials and exert fungicidal activity with minimal cytotoxicity to mammalian cells. DC11MAF and C2DC11MAF are considered strong candidates as therapeutic or preventive alternatives against Candida-associated denture stomatitis.

Resistance to cigarette smoke is increased in periodontal ligament cells by attachment to collagen and fibronectin.

BACKGROUND: The toxic effects of cigarette smoke often presents in smokers as increased incidence and severity of periodontal disease. These patients demonstrate symptomatic inflammation, increased probing depth, and tooth loss likely attributable to the direct effects of cigarette smoke on periodontal ligament (PDL) fibroblasts. The goal of this in vitro study is to investigate the direct effects of smoking on PDL fibroblasts, focusing on cell-extracellular matrix (ECM) interactions and cell survival. METHODS: PDL cells were plated for various times on tissue culture plastic, PDL-derived ECMs, collagen Type I, or fibronectin. Cells were exposed to various concentrations of cigarette smoke extract (CSE) at different times during the cell attachment process. Subsequently, cell survival was quantified using calcein-acetoxymethyl ester compound and a fluorescent plate reader. RESULTS: After exposure to CSE, PDL cell survival increased with increased cell attachment time to plastic. These observations were independent of soluble factors present in PDL cell-conditioned media. PDL-derived ECMs and collagen Type I-pretreated plates promoted increased cell survival after 1 day of cell attachment. Fibronectin-pretreated plates demonstrated increased cell survival after 3 days of cell attachment. CONCLUSIONS: Cell-ECM interactions increase survival of PDL cells exposed to CSE. It is suggested that the increased survival is attributable to PDL cells altering their ECM, potentially by depositing collagen and fibronectin. This may imply that cells embedded in an ECM would be more resistant to the toxic effects of cigarette smoke, leading to increased cell death near the exposed edges of a wound.

Introducing evidence-based dentistry to dental students using histology.

The expansion of evidence-based dentistry (EBD) is essential to the continued growth and development of the dental profession. Expanding EBD requires increased emphasis on critical thinking skills during dental education, as noted in the American Dental Education Association's Competencies for the New General Dentist. In order to achieve this goal, educational exercises must be introduced to increase the use of critical thinking skills early in the dental curriculum, with continued reinforcement as students progress through subsequent years. Described in this article is one approach to increasing student exposure to critical thinking during the early basic science curriculum-specifically, within the confines of a traditional histology course. A method of utilizing the medical and dental research literature to reinforce and enliven the concepts taught in histology is described, along with an approach for using peer-to-peer presentations to demonstrate the tools needed to critically evaluate research studies and their presentation in published articles. This approach, which could be applied to any basic science course, will result in a stronger foundation on which students can build their EBD and critical thinking skills.

A novel GuttaFlow sealer supports cell survival and attachment.

INTRODUCTION: The purpose of this in vitro study was to compare the biocompatibility of a novel formulation of a silicone-based endodontic sealer GuttaFlow 2 (GF2; Coltène/Whaledent, Langenau, Germany) with the original (GFO) and fast-set (GFF) formulations of GuttaFlow and with an epoxy resin sealer, AHPlus Jet (AH+J; Dentsply, York, PA). METHODS: Sealers were set into 3 × 5.5 mm discs. Cell culture media was used to extract leachable products at 24 hours and 1, 2, and 4 weeks. Primary human periodontal ligament fibroblasts were incubated with sealer elutes for 24 hours and evaluated using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay and the calcein AM assay. Cell attachment was evaluated on set sealer that was either rinsed or unrinsed with cell media for 1 week. Statistical analysis was performed using the Student t test. RESULTS: Both calcein and 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assays revealed that periodontal ligament cell viability was reduced on AH+J at 1, 2, and 4 weeks compared with all GuttaFlow sealers. There were no differences in cell viability between the GuttaFlow samples, and all displayed high rates of cell survival at all time periods. After 2 hours, cell attachment to the rinsed GFO and GFF samples exceeded the control, and at 24 hours cell attachment on all GuttaFlow samples exceeded the control. AH+J sealers supported significantly less cell attachment when compared with all GuttaFlow sealers. Cell attachment to set sealers showed better cell attachment when rinsed compared with unrinsed. CONCLUSIONS: GuttaFlow sealers were more biocompatible than AHJ in vitro. The novel GF2 displayed comparable biocompatibility with GFF and GFO.

Electrochemical dissolution of nickel-titanium endodontic files induces periodontal ligament cell death.

INTRODUCTION: Fractured endodontic files present a major problem. A novel method has been proposed to retrieve fractured nickel-titanium (NiTi) endodontic files by using electrochemical dissolution. However, the effect of file dissolution on adjacent soft tissues such as the periodontal ligament (PDL) has not been investigated. The aim of this study was to determine the effects of the dissolution products on PDL fibroblasts. METHODS: Endodontic files were dissolved in sodium fluoride (NaF) by passing a 50-mA current through the NiTi files while immersed in the NaF solution. NaF/NiTi solutions were diluted with minimal essential medium-α media containing 10% serum. PDL cells were treated for up to 24 hours, and cell viability was quantified by using calcein AM to label live cells and ethidium homodimer to label dead cells. This was repeated by using artificial saliva (AS) as an alternative to NaF. RESULTS: NaF solution reduced PDL cell survival, and the NaF/NiTi solution further reduced PDL cell survival. AS alone did not reduce cell survival, whereas AS/NiTi solution reduced PDL cell survival. Particles that resulted from the electrochemical dissolution of NiTi files were highly cytotoxic. CONCLUSIONS: Electrochemically dissolving NiTi files in NaF results in solutions that are cytotoxic to PDL fibroblasts. AS may be a less toxic alternative for dissolving NiTi files.

Pedialyte promotes periodontal ligament cell survival and motility.

INTRODUCTION: The search still continues to find the best storage media for avulsed teeth. Unfortunately, some of the recommended storage solutions are not commonly found in households or do not preserve the periodontal ligament (PDL) cells long-term. The purpose of the present study was to determine whether Pedialyte is a viable alternative storage solution for avulsed teeth by assessing its ability to preserve human PDL cell viability. METHODS: Human PDL cells were exposed to 6 different storage solutions (minimal essential medium [MEMα], Hank's balanced salt solution [HBSS], non-fat milk, coconut water, Pedialyte, or tap water) for 2, 6, 24, or 48 hours at 4°C or 25°C. Cell viability was quantified immediately or 1 week after exposure. The effects of these storage solutions on PDL cell motility and bacterial proliferation were also examined. The results were statistically analyzed by analysis of variance. RESULTS: Pedialyte at 4°C and 25°C showed significantly (P < .001) higher cell survival compared with water after all time intervals. No significant difference was noted between control (MEMα), HBSS, coconut water, and Pedialyte at 4°C after 2 hours. Cells stored in Pedialyte for 24 hours at 25°C and assayed 1 week later showed significantly higher cell survivability compared with milk. Pedialyte supported significantly less bacterial growth compared with non-fat milk and coconut water. No difference in cell motility was observed for cells stored for 24 hours in Pedialyte, MEMα, HBSS, milk, or coconut water. CONCLUSIONS: Pedialyte is a viable alternative as a storage solution for avulsed teeth.

Effect of essential oil and chlorhexidine mouthwashes on gingival fibroblast survival and migration.

BACKGROUND: Chemical plaque control is the most commonly recommended means of oral hygiene after periodontal surgery. Commercially available mouthwashes contain a variety of active ingredients that have bactericidal properties but may potentially be toxic to the host cells. The goal of this in vitro study is to investigate the effect of commercially available mouthwashes on the survival and migratory capacity of human fibroblasts. METHODS: Human gingival and periodontal ligament (PDL) fibroblasts were treated with commercially available mouthwashes that contained either chlorhexidine (CHX) or essential oils (EO) as the active ingredient. Each mouthwash was tested over a range of concentrations for its ability to affect fibroblast survival and migration, as well as long-term effects on cell viability. RESULTS: Undiluted mouthwashes induced near-complete cell death 24 hours after only a 60-second treatment. Dilutions of 15% to 20% for both CHX and EO mouthwashes resulted in 50% cell death. When diluted to 10% to 15%, EO did not reduce cell migration, whereas similar dilutions of CHX resulted in reduced cell migration. Concentrations of 10% of both EO and CHX mouthwashes retained most of their antibacterial capacity. Treatment with EO did not result in gingival fibroblast death, whereas 5% CHX resulted in near-complete gingival fibroblast death 7 days after exposure. CONCLUSIONS: The results of this in vitro study indicate that diluted EO displayed no detectable detrimental effects on human gingival and PDL fibroblasts, whereas diluted CHX reduced both cell migration and long-term survival. Both solutions retained their antimicrobial activity in lower concentrations.

Synthesis and Characterization of New Antibacterial Fluoride-Releasing Monomer and Dental Composite.

A new dimethacrylate chelating monomer containing a BisGMA-like backbone structure and a bis(carboxymethyl)-L-lysine chelating group and its ternary zirconium-fluoride complex (antibacterial fluoride-releasing monomer) have been synthesized. The monomer structures were confirmed by (1)H-NMR, (13)C-NMR, and ES-MS analysis. Several experimental fluoride-releasing dental composites containing different quantities of the new antibacterial fluoride-releasing monomer were formulated and tested for fluoride release, fluoride recharge, compressive and flexural strengths, water sorption and solubility. These composites displayed high fluoride release and recharge capabilities, as well as good physical and mechanical properties.

Altered cell motility and attachment with titanium surface modifications.

BACKGROUND: Titanium implants are widely used in dentistry to replace lost teeth. Various surface modifications have been used to improve implant retention and osseointegration. This study is designed to compare the ability of three titanium surfaces to promote cell attachment and cell motility of cells relevant to periodontal tissues. METHODS: Three clinically relevant surfaces were tested: 1) machined titanium; 2) a titanium surface roughened through acid etching (dual thermal-etched titanium [DTET]); and 3) a titanium surface roughened with nanometer-scale calcium phosphate deposition (nanoscale calcium phosphate-impregnated titanium [NCPIT]). Cell attachment and migration were examined for four cell types: rat osteosarcoma cells, human osteoblasts, and gingival and periodontal ligament (PDL) fibroblasts. RESULTS: All four cell types attached to each of the three titanium surfaces equally by 2 hours, and the PDL and gingival fibroblasts generally displayed less attachment than the osteosarcoma cells and osteoblasts. The cells displayed differential motility and long-term attachment to each of the titanium surfaces. Osteosarcoma cells displayed preferential motility on NCPIT, whereas PDL fibroblasts were more motile on machined titanium, and gingival fibroblasts moved more rapidly on both DTET and NCPIT. Osteoblasts displayed little motility on any of the titanium surfaces and lost viability on NCPIT after 24 hours. Gingival fibroblasts lost attachment to machined titanium. CONCLUSIONS: Periodontal cells displayed differential motility and long-term attachment to titanium surfaces. Selective modification of titanium surface properties in various regions of an implant may be useful in guiding specific cell populations to specific locations where they might best aid in osseointegration and soft tissue remodeling.

Cigarette smoke extract induces select matrix metalloproteinases and integrin expression in periodontal ligament fibroblasts.

BACKGROUND: The periodontal ligament (PDL) is the connective tissue that anchors the cementum of the teeth to the alveolar bone. PDL fibroblasts are responsible for the production of collagen and remodeling of the PDL. Periodontal disease is increased among smokers in both incidence and severity. This study examines the direct effect of smoking on PDL fibroblasts and their production of various matrix components and remodeling enzymes. METHODS: PDL cells were plated for 1 day and then treated with various concentrations of cigarette smoke extract (CSE). Survival of PDL cells was quantified after exposure to CSE, and their ability to contract three-dimensional collagen gels was examined. Changes in transcript expression after CSE treatment was compared using reverse transcription-polymerase chain reaction analysis for matrix metalloproteinases (MMPs), collagens, and integrins. RESULTS: Treatment with CSE-induced cell death at concentrations of ≥5%. PDL-cell-induced collagen gel contraction was reduced at concentrations of 1.5% CSE. Treatment with CSE selectively increased the expression of collagen Vα3 and decreased collagen XIα1. CSE increased the expression of MMP1 and MMP3 and, to a lesser extent, MMP2 and MMP8. CSE also increased the expression of integrins α1, α2, and α10 (collagen receptors) and α9 (a tenascin receptor). CONCLUSIONS: This study shows that cigarette smoking has local effects on the cells of the PDL. CSE reduced survival of PDL cells and their ability to contract collagen matrices. CSE also altered the expression of molecules known to provide the structural integrity of the ligament by altering collagen synthesis and remodeling as well as cell adhesion.

Mechanical tension alters semaphorin expression in the periodontium.

BACKGROUND: Periodontal remodeling requires coordinated cell movement. Semaphorins are cell-surface signals that regulate cell migration and may be differentially regulated by periodontal cells. Mechanical tension can regulate periodontal ligament (PDL) remodeling. We predicted that mechanical tension alters the expression of the subset of semaphorins in the periodontium likely to be most involved with regulating the remodeling of this tissue. METHODS: PDL and gingival cells were exposed to mechanical tension, and their attachment and movement on collagen matrices were evaluated. Alterations in extracellular matrix and semaphorin transcript expression were monitored by semiquantitative reverse transcription-polymerase chain reaction. RESULTS: Mechanical tension induced osteoclast regulatory transcripts in the PDL cells to a greater extent than gingival fibroblasts, increasing the expression of osteoprotegerin and decreasing receptor activator of nuclear factor-kappa B ligand. These mechanical forces reduced PDL cell mingling, without altering cell attachment or motility. Concurrently, these forces induced dynamic changes in several semaphorin molecules in PDL cells, increasing semaphorin 3D and 5B and decreasing semaphorin 7A. In addition, plexin transcript expression was altered, decreasing plexin A1 and increasing plexin C1. These changes were different than those observed in gingival fibroblasts. CONCLUSIONS: These data suggest that a subset of semaphorins and plexins are dynamically regulated in the PDL. Because these molecules may be involved in cell guidance, changes in semaphorins may play a pivotal role in periodontal remodeling, affecting angiogenesis or PDL cell invasion into sites of injury.

A simple cell motility assay demonstrates differential motility of human periodontal ligament fibroblasts, gingival fibroblasts, and pre-osteoblasts.

During periodontal regeneration, multiple cell types can invade the wound site, thereby leading to repair. Cell motility requires interactions mediated by integrin receptors for the extracellular matrix (ECM), which might be useful in guiding specific cell populations into the periodontal defect. Our data demonstrate that fibroblasts exhibit differential motility when grown on ECM proteins. Specifically, gingival fibroblasts are twice as motile as periodontal ligament fibroblasts, whereas osteoblasts are essentially non-motile. Collagens promote the greatest motility of gingival fibroblasts in the following order: collagen III>collagen V>collagen I. Differences in motility do not correlate with cell proliferation or integrin expression. Osteoblasts display greater attachment to collagens than does either fibroblast population, but lower motility. Gingival fibroblast motility on collagen I is generally mediated by alpha2 integrins, whereas motility on collagen III involves alpha1 integrins. Other integrins (alpha10 or alpha11) may also contribute to gingival fibroblast motility. Thus, ECM proteins do indeed differentially promote the cell motility of periodontal cells. Because of their greater motility, gingival fibroblasts have more of a potential to invade periodontal wound sites and to contribute to regeneration. This finding may explain the formation of disorganized connective tissue masses rather than the occurrence of the true regeneration of the periodontium.

Use of microarrays to find novel regulators of periodontal ligament fibroblast differentiation.

Periodontal regeneration requires the coordinated movement and differentiation of several cell types in order to re-establish the cementum, periodontal ligament (PDL), and alveolar bone. Cells in culture are often used as model systems for mature tissues, although they may represent expanded progenitor cell populations. Comparison of transcript expression between fresh PDL tissue and PDL cell isolates by MicroArray analysis has revealed numerous molecular differences. Several transcripts (including alkaline phosphatase, bone sialoprotein, periostin, and fibromodulin) are expressed at higher levels in fresh PDL than in cultured PDL cells. In contrast, PDL cells in culture selectively express a variety of growth factors. Several of these growth factors alter PDL fibroblast behavior. Two members of the transforming growth factor beta family of growth factors, namely, bone morphogenic protein-7 (BMP7) and growth differentiation factor-5 (GDF5), reduce cell proliferation and Stro-1 expression (a bone marrow stromal stem cell marker), whereas only BMP7 induces alkaline phosphatase activity. In contrast, fibroblast growth factor-5 induces enhanced cell proliferation and Stro-1 expression, while repressing alkaline phosphatase activity. The stimulation of PDL cells to differentiate (either by BMP7 or GDF5) inhibits cell motility. Thus, PDL cells in culture are regulated by several factors that differentially stimulate a mineralized (cementoblast-like) fate, a non-mineralized fate (mature fibroblasts), or the propagation of a more naive phenotype (potential progenitors).

Transcript profiling of periodontal fibroblasts and osteoblasts.

BACKGROUND: Fibroblasts are critical to the establishment and maintenance of the periodontal attachment apparatus (cementum, periodontal ligament [PDL], and bone). In order to characterize the cellular changes that accompany periodontal regeneration, better tools are necessary to distinguish periodontal ligament fibroblasts (PDLF), gingival fibroblasts, and osteoblasts. Our goal is to identify gene markers to better characterize and identify these cell types. METHODS: We chose to examine and compare the expression of numerous gene transcripts by semiquantitative reverse transcriptase-polymerase chain reaction using primers specific for 44 different gene transcripts in order to better characterize the identity of these cells. RESULTS: Several transcripts were cell-type specific. Specifically, fibromodulin was expressed only in PDL fibroblasts, while osteopontin was expressed only in dermal fibroblasts. In addition, lumican was expressed by all three types of fibroblasts (PDL, gingival, and dermal), while alkaline phosphatase was expressed by osteoblasts as well as PDL and gingival fibroblasts. CONCLUSIONS: Our results indicate that PDL fibroblasts are distinct from either gingival or dermal fibroblasts or osteoblasts. In general, PDL and gingival fibroblasts displayed greater similarity to each other than either displayed toward dermal fibroblasts. Furthermore, both gingival and PDL fibroblasts displayed greater similarity to osteoblasts than to dermal fibroblasts, possibly reflecting their common origin (the neural crest).