Acellular Extrinsic Fiber Cementum Is Invariably Present in the Superficial Layer of Apical Cementum in Mouse Molar.

The cementum is a highly mineralized tissue that covers the tooth root. The regional differences among the types of cementum, especially in the extrinsic fibers that contribute to tooth support, remain controversial. Therefore, this study used second harmonic generation imaging in conjunction with automated collagen extraction and image analysis algorithms to facilitate the quantitative examination of the fiber characteristics and the changes occurring in these fibers over time. Acellular extrinsic fiber cementum (AEFC) was invariably observed in the superficial layer of the apical cementum in mouse molars, indicating that this region of the cementum plays a crucial role in supporting the tooth. The apical AEFC exhibited continuity and fiber characteristics comparable with the cervical AEFC, suggesting a common cellular origin for their formation. The cellular intrinsic fiber cementum present in the inner layer of the apical cementum showed consistent growth in the apical direction without layering. This study highlights the dynamic nature of the cementum in mouse molars and underscores the requirement for re-examining its structure and roles. The findings of the present study elucidate the morphophysiological features of cementum and have broader implications for the maintenance of periodontal tissue health.

Immunohistochemical Evaluation of Periodontal Regeneration Using a Porous Collagen Scaffold.

(1) Aim: To immunohistochemically evaluate the effect of a volume-stable collagen scaffold (VCMX) on periodontal regeneration. (2) Methods: In eight beagle dogs, acute two-wall intrabony defects were treated with open flap debridement either with VCMX (test) or without (control). After 12 weeks, eight defects out of four animals were processed for paraffin histology and immunohistochemistry. (3) Results: All defects (four test + four control) revealed periodontal regeneration with cementum and bone formation. VCMX remnants were integrated in bone, periodontal ligament (PDL), and cementum. No differences in immunohistochemical labeling patterns were observed between test and control sites. New bone and cementum were labeled for bone sialoprotein, while the regenerated PDL was labeled for periostin and collagen type 1. Cytokeratin-positive epithelial cell rests of Malassez were detected in 50% of the defects. The regenerated PDL demonstrated a larger blood vessel area at the test (14.48% ± 3.52%) than at control sites (8.04% ± 1.85%, p = 0.0007). The number of blood vessels was higher in the regenerated PDL (test + control) compared to the pristine one (p = 0.012). The cell proliferative index was not statistically significantly different in pristine and regenerated PDL. (4) Conclusions: The data suggest a positive effect of VCMX on angiogenesis and an equally high cell turnover in the regenerated and pristine PDL. This VCMX supported periodontal regeneration in intrabony defects.

A novel serum-free medium for the isolation, expansion and maintenance of stemness and tissue-specific markers of primary human periodontal ligament cells.

PURPOSE: Periodontal ligament (PDL) cell cultures are classically maintained in serum-containing media. However, unwanted side-effects of these conditions on cellular and molecular characteristics demand a serum-free alternative. Even though these limitations are well known and efforts for the development of adequate serum-free alternatives have been made, these approaches for replacement remained unsuccessful so far. This study aimed at developing a well-defined, serum-free formulation supporting both isolation from tissue samples and efficient expansion of PDL cells. Here, of particular focus was the perpetuation of tissue-characteristic markers detectable in primary tissues and of stemness features. BASIC PROCEDURES: Primary PDL cell cultures from generally healthy human donors (n = 3) were maintained in basal media N2B27 and E6 together with different concentrations of growth and attachment factors. Cell proliferation was recorded via microscopy and WST assay. Gene expression of RUNX2, Periostin, ALP, CD73, CD90, CD105, CD45, SOX10 and SOX2 was compared to primary PDL explants via qRT-PCR. Immunocytochemistry was performed for anti-CD105, SSEA-3, CD271, HNK1. Serum-containing sDMEM medium served as control. MAIN FINDINGS: N2B27 medium substituted with 25 ng/mL EGF, 25 ng/mL IGF1, 0.5 mg/mL Fetuin plus gelatine coating (designated N2B27-PDLsf) emerged as potent serum-free formulation ensuring adequate culture isolation and expansion. Here, PDL primary tissue signature markers RUNX2 and Periostin remained stable in N2B27-PDLsf compared to controls (229.0-fold ±101.0 and 83.2-fold ±9.6 increase). Additionally, stemness markers ALP and CD105 were significantly upregulated on transcriptional, and CD105 and SOX2 on protein level. PRINCIPAL CONCLUSIONS: This investigation identified a novel serum-free medium for the isolation, and expansion of primary human PDL cells with constantly high proliferation rates. Here, purity and stemness properties are maintained. Thus, N2B27-PDLsf represents a valid replacement for serum-containing media in PDL cultures.

Monitoring Biochemical and Structural Changes in Human Periodontal Ligaments during Orthodontic Treatment by Means of Micro-Raman Spectroscopy.

The aim of the study was to examine the biochemical and structural changes occurring in the periodontal ligament (PDL) during orthodontic-force application using micro-Raman spectroscopy ( µ -RS). Adolescent and young patients who needed orthodontic treatment with first premolar extractions were recruited. Before extractions, orthodontic forces were applied using a closed-coil spring that was positioned between the molar and premolar. Patients were randomly divided into three groups, whose extractions were performed after 2, 7, and 14 days of force application. From the extracted premolars, PDL samples were obtained, and a fixation procedure with paraformaldehyde was adopted. Raman spectra were acquired for each PDL sample in the range of 1000-3200 cm - 1 and the more relevant vibrational modes of proteins (Amide I and Amide III bands) and CH 2 and CH 3 modes were shown. Analysis indicated that the protein structure in the PDL samples after different time points of orthodontic-force application was modified. In addition, changes were observed in the CH 2 and CH 3 high wavenumber region due to local hypoxia and mechanical force transduction. The reported results indicated that µ -RS provides a valuable tool for investigating molecular interchain interactions and conformational modifications in periodontal fibers after orthodontic tooth movement, providing quantitative insight of time occurring for PDL molecular readjustment.

Elastic fiber system evaluated in the digestive organ of rats.

According to our previous reports, the intraperiodontal elastic fiber system comprises oxytalan fibers, whereas all types of elastic system fibers are present in the gingiva. Much remains to be elucidated regarding the topographic development of the elastic fiber system that constitutes the walls of the digestive organs. This study aimed to examine the topographic development of the elastic fiber system in the periodontal tissue, oral cavity and digestive tract of rats at light- and electron microscopic levels. At embryonic day 20, in situ hybridization revealed the mRNA expression of tropoelastin in the putative gingival lamina propria but not in the dental follicle. At the postnatal stage, the masticatory mucous membrane of the gingiva and hard palate comprised three different types of elastic system fibers (oxytalan, elaunin and elastic fibers). Conversely, the elastic fiber system comprised elaunin and elastic fibers in other oral mucosae and the lining mucosae of digestive tract organs (the esophagus, stomach and small intestine). The findings of our study suggest that the elastic fiber system is mainly related to tissue resistance in the periodontal ligament and tissue elasticity in the oral mucosae without masticatory mucosae and the overlying mucosa of digestive tracts and both functions in the gingiva and hard palate, respectively. The appearance of elaunin fibers in the periodontium of rats aged 14 weeks suggests the expression of tropoelastin induced by mechanical stressors such as mastication. The intraperiodontal difference in the distribution of elaunin fibers suggests heterogeneity among fibroblasts constituting the periodontium.

Decellularized human periodontal ligament for periodontium regeneration.

Regenerating the periodontal ligament (PDL) is a crucial factor for periodontal tissue regeneration in the presence of traumatized and periodontally damaged teeth. Various methods have been applied for periodontal regeneration, including tissue substitutes, bioactive materials, and synthetic scaffolds. However, all of these treatments have had limited success in structural and functional periodontal tissue regeneration. To achieve the goal of complete periodontal regeneration, many studies have evaluated the effectiveness of decellularized scaffolds fabricated via tissue engineering. The aim of this study was to fabricate a decellularized periodontal scaffold of human tooth slices and determine its regeneration potential. We evaluated two different protocols applied to tooth slices obtained from human healthy third molars. The extracellular matrix scaffold decellularized using sodium dodecyl sulfate and Triton X-100, which are effective in removing nuclear components, was demonstrated to preserve an intact structure and composition. Furthermore, the decellularized scaffold could support repopulation of PDL stem cells near the cementum and expressed cementum and periodontal-ligament-related genes. These results show that decellularized PDL scaffolds of human teeth are capable of inducing the proliferation and differentiation of mesenchymal stem cells, thus having regeneration potential for use in future periodontal regenerative tissue engineering.

Cellular and collagen reference values of gingival and periodontal ligament tissues in rats: a pilot study.

Reference data are lacking on the periodontal ligament and the gingival tissue of the rat model, which would be useful for studies of new medical or biomaterial periodontal treatments. The objective of the current study was to propose cellular and collagen reference values of gingival and periodontal ligament tissues in rat, using a simple and reliable quantitative method after decalcification. Mandibular samples of ten adult Sprague-Dawley rats were used. Mild decalcification was carried out using ethylenediaminetetraacetic acid (EDTA) to preserve the morphology of tissues. Half of the samples were decalcified and the other half were not. The gingiva and the periodontal ligament were analyzed. Descriptive histology and computer-assisted image analysis were performed. The data showed that qualitatively, cellular and extracellular matrix morphologies were well preserved compared to non-decalcified periodontal soft tissue biopsies. Histomorphometrically, constitutive cellularity and the total amount of native collagen, collagen directionality and collagen anisotropy in both experimental conditions did not significantly differ. Taken together, these results suggested that EDTA decalcification did not negatively affect the studied endpoints. Moreover, this mild decalcification method allowed in situ maintenance of the periodontal soft and hard tissue integrity. The structural and compositional computerized assessment performed in the healthy periodontal soft tissue could provide reference values that will be required for future assessment on the effects of pathological, reparative and regenerative processes in rat periodontal soft tissues.

Sequential application of bFGF and BMP-2 facilitates osteogenic differentiation of human periodontal ligament stem cells.

BACKGROUND AND OBJECTIVES: Basic fibroblast growth factor (bFGF) promotes cells proliferation and chemotaxis and maintains stemness while inhibits mineralized nodule formation. Bone morphogenetic protein 2 (BMP-2) shows great potential in promoting bone formation. However, sequential application of these two growth factors on periodontal ligament stem cells (PDLSCs) has not been explored. In this study, we aimed to identify the optimal concentration and time of bFGF on PDLSCs proliferation, migration and then investigate the sequential delivery of bFGF and BMP-2 on osteogenic differentiation of PDLSCs in vitro. MATERIALS AND METHODS: Periodontal ligament stem cells were isolated by limiting dilution method. Dose-dependent additive effects of bFGF and BMP-2 on PDLSCs were detected. Cell counting assay, cell migration assay, alkaline phosphatase (ALP) activity assay, Alizarin red staining, quantitative real-time polymerase chain reaction (qRT-PCR), and western blot analysis were used to determine different application modalities of bFGF and BMP-2 on proliferation, migration, and osteogenic differentiation of PDLSCs. RESULTS: 50 ng/mL bFGF significantly promoted PDLSCs proliferation and chemotaxis while time-dependently inhibited BMP-2 induced ALP activity. Sequential application of 25 ng/mL bFGF for first 3 days and followed with 50 ng/mL BMP-2 for another 9, 18, and 25 days significantly promoted PDLSCs osteogenic differentiation. Compared with bFGF and BMP-2 simultaneous group, sequential application of bFGF and BMP-2 group significantly enhanced ALP activity, osteogenesis-related genes and proteins expression and mineral deposition. CONCLUSION: Sequential application of bFGF and BMP-2 synergistically promoted osteogenic differentiation of PDLSCs, and this sequential application modality of growth factors would provide a new strategy for periodontal regeneration.

Influence of chemical fixation process on primary mesenchymal stem cells evidenced by Raman spectroscopy.

In investigation of (patho)physiological processes, cells represent frequently used analyte as an exceptional source of information. However, spectroscopic analysis of live cells is still very seldom in clinics, as well as in research studies. Among others, the reasons are long acquisition time during which autolysis process is activated, necessity of specified technical equipment, and inability to perform analysis in a moment of sample preparation. Hence, an optimal method of preserving cells in the existing state is of extreme importance, having in mind that selection of fixative is cell lineage dependent. In this study, two commonly used chemical fixatives, formaldehyde and methanol, are used for preserving primary mesenchymal stem cells extracted from periodontal ligament, which are valuable cell source for reconstructive dentistry. By means of Raman spectroscopy, cell samples were probed and the impact of these fixatives on their Raman response was analyzed and compared. Different chemical mechanisms are the core processes of formaldehyde and methanol fixation and certain Raman bands are shifted and/or of changed intensity when Raman spectra of cells fixed in that manner are compared. In order to get clearer picture, comprehensive statistical analysis was performed.

An unusual disordered alveolar bone material in the upper furcation region of minipig mandibles: A 3D hierarchical structural study.

Teeth are subjected to compressive loads during mastication. Under small loads the soft tissue periodontal ligament (PDL) deforms most. However when the loads increase and the PDL is highly compressed, the tooth and the alveolar bone supporting the tooth, begin to deform. Here we report on the structure of this alveolar bone in the upper furcation region of the first molars of mature minipigs. Using light microscopy and scanning electron microscopy (SEM) of bone cross-sections, we show that this bone is hypermineralized, containing abundant small pores around 1-5 µm in diameter, lacunae around 10-20 µm as well as larger spaces. This bone does not possess the typical lamellar motif or other repeating structures normally found in cortical or trabecular mammalian bone. We also use high resolution focused ion beam scanning electron microscopy (FIB-SEM) in the serial surface mode to image the 3D organization of the demineralized bone matrix. We show that the upper furcation bone matrix has a disordered isotropic structure composed mainly of individual collagen fibrils with no preferred orientation, as well as highly staining material that is probably proteoglycans. Much larger aligned arrays of collagen fibers - presumably Sharpey's fibers - are embedded in this material. This unusual furcation bone material is similar to the disordered material found in human lamellar bone. In the upper furcation region this disordered bone comprises almost all the volume excluding Sharpey's fibers. We surmise that this most unusual bone type functions to resist the repeating compressive loads incurred by molars during mastication.

Periostin, dentin matrix protein 1 and P2rx7 ion channel in human teeth and periodontal ligament.

The periostin is a matricellular protein present in the human periodontal ligament and human dental pulp-derived cells lines, that up-regulates the in vitro expression of some genes involved in the dentin mineralization, such as dentin matrix protein 1 and P2x7-ion channel receptor. Here we investigated the distribution of periostin in human teeth and periodontal ligaments, mapping in parallel the localization of dentin matrix protein 1 and P2x7-ion channel receptor to establish whether or not they are expressed in the same places as periostin. The periodontal ligament and the subodontoblastic layer of the dental pulp displayed strong periostin immunoreactivity, whereas dentin matrix protein 1 was detected in the periodontal ligament co-localized with periostin in the vicinity of the cement. The P2x7 ion channel receptor was regularly absent in both the periodontal ligament and dental tissues, but in some cases, it was observed in the odontoblasts. Present results demonstrate the occurrence of periostin in the healthy adult human tooth without co-localization with proteins involved in tooth mineralization, the expression of which it regulates. These results might serve as a baseline for future studies on pathological conditions.

Laser Photobiomodulation Over Teeth Subjected to Orthodontic Movement.

Background: Orthodontics of the 21st century requires aesthetic, painless, predictable, and quick treatments. This demand for faster results generated orthodontic movement acceleration protocols (OMAPs); among other OMAPs we present low-level laser (LLL) as a candidate. Objective: To evaluate levels of interleukin (IL)-1, IL-10, and type 1 collagen in the periodontal ligament of first molars of rats subjected to orthodontic traction with and without LLL irradiation, compared with untreated controls (CO), and to evaluate whether the dose of LLL used in this work is eligible as an OMAP. Materials and methods: A total of 35 male Wistar rats were distributed into three groups: group 1 NI (nonirradiated) n = 15, group 2 IR (laser irradiated using 5 J, 177 J/cm2, and 100 mW applied in contact to the vestibular mesial, vestibular distal, and palatal faces of gum tissue around molar region for 50 sec each point, for 3 consecutive days, immediately 24 and 48 h after orthodontic device placement.) n = 15, and group 3 CO n = 5; groups 1 and 2 were subjected to orthodontic force and each group was divided into three subgroups that were sacrificed after 3, 5, and 7 days, IL-1/10 and COL-1 levels were analyzed. Results: In the IR group, levels of IL-1/10 and COL-1 showed peak anticipation after LLL irradiation compared with those in the NI and CO groups. Conclusions: These results can also infer that this dose of LLL can be used as an OMAP.

Extracellular Matrix from Periodontal Ligament Cells Could Induce the Differentiation of Induced Pluripotent Stem Cells to Periodontal Ligament Stem Cell-Like Cells.

The periodontal ligament (PDL) plays an important role in anchoring teeth in the bone socket. Damage to the PDL, such as after severe inflammation, can be treated with a therapeutic strategy that uses stem cells derived from PDL tissue (PDLSCs), a strategy that has received intense scrutiny over the past decade. However, there is an insufficient number of PDLSCs within the PDL for treating such damage. Therefore, we sought to induce the differentiation of induced pluripotent stem (iPS) cells into PDLSCs as an initial step toward PDL therapy. To this end, we first induced iPS cells into neural crest (NC)-like cells. We then captured the p75 neurotrophic receptor-positive cells (iPS-NC cells) and cultured them on an extracellular matrix (ECM) produced by human PDL cells (iPS-NC-PDL cells). These iPS-NC-PDL cells showed reduced expression of embryonic stem cell and NC cell markers as compared with iPS and iPS-NC cells, and enrichment of mesenchymal stem cell markers. The cells also had a higher proliferative capacity, multipotency, and elevated expression of PDL-related markers than iPS-NC cells cultured on fibronectin and laminin (iPS-NC-FL cells) or ECM produced by human skin fibroblast cells (iPS-NC-SF cells). Overall, we present a culture method to produce high number of PDLSC-like cells from iPS cells as a first step toward a strategy for PDL regeneration.

Scaffold requirements for periodontal regeneration with enamel matrix derivative proteins.

Periodontitis affects the attachment of natural teeth, and infection or inflammation associated with periodontitis may affect peri-implant tissues. Enamel matrix derivative (EMD) proteins provide stimulation for self-regeneration of the damaged tissue when applied to wide intrabony defects as part of a mixture with bone graft material. As a first step of the process enhancing cell proliferation and ligament formation, we demonstrated that EMD protein precipitation depends strongly on the physical and chemical characteristics of the bone grafts used in the mixture. To guarantee optimum protein-stimulated self-regulation, the pH of the initial EMD formulation must therefore be adjusted between 3.9 and 4.2 in order to compensate the change in pH induced by the bone graft. Moreover, the interaction between the two components resulted in precipitates of different shape and size differently covering the grafts. This outcome might potentially have clinical implications on cell attachment and periodontal ligament extension, which deserve further in vitro and in vivo tests.

Quest for Cells Responsible for Age-related Increase of Salivary Glycine and Proline.

We have previously reported that salivary glycine and proline levels are increased to nearly butanoate level in elderly people. In order to identify the source of glycine and proline, we performed high-performance liquid chromatography analysis of amino acid production to a total of seven oral cells before and after stimulation with inflammation inducers. We found that production of amino acids (per a given number of cells) by normal oral mesenchymal cells (gingival fibroblast, pulp cell, periodontal ligament fibroblast) was approximately three-fold that of oral squamous cell carcinoma cell lines (HSC-2, HSC-3, HSC-4, Ca9-22), and that production of glycine and especially proline by all these seven cells was much lower than that of glutamine and glutamic acid. Treatment of three oral mesenchymal cells with interleukin (IL)-1ß or lipopoly-saccharide (LPS) reproducibly increased the production of glutamic acid and glutamine, but not that of glycine and proline. Glycine and proline only marginally stimulated the IL-8 production by IL-1ß-stimulated gingival fibroblast, whereas glycine dose-dependently inhibited the nitric oxide production by lipopolysaccharide-stimulated mouse macrophage-like RAW264.7 cells. These data demonstrated that normal oral mesenchymal cells are not the major source of glycine and proline that accumulates in the saliva of aged people, suggesting the involvement of the deregulation of collagen metabolism during aging.

Expression of Wnt3a, Wnt10b, ß-catenin and DKK1 in periodontium during orthodontic tooth movement in rats.

OBJECTIVE: To investigate the expression of Wnt3a, Wnt10b, ß-catenin and DKK1 in the periodontal ligament (PDL) during orthodontic tooth movement (OTM) in rats. MATERIALS AND METHODS: Nickel-titanium closed-coil springs were used to deliver an initial 50 g mesial force to the left maxillary first molars in 30 rats. The force was kept constant for 1, 3, 5, 7, 10 and 14 days until the animals were sacrificed. The right maxillary molars without force application served as control. Paraffin-embedded sections of the upper jaws were prepared for histological and immunohistochemical analyses to detect Wnt3a, Wnt10b, ß-catenin and DKK1 expression in PDL. RESULTS: Wnt3a, Wnt10b, ß-catenin and DKK1 were expressed on both the ipsilateral and contralateral sides of PDL in each group. After the application of orthodontic force, the expression of ß-catenin and DKK1 was initially increased and then decreased on both sides, with maximal levels of expression at day 7 and day 10, respectively. On the compression side, Wnt3a and Wnt10b levels started to increase at day 5, while on the tension side, these two molecules began to increase at day 1. Furthermore, the expression levels of Wnt3a, Wnt10b, and ß-catenin were much stronger on the tension side than on the compression side at any of the observation points, while DKK1 level was much higher on the compression side. CONCLUSION: Wnt3a, Wnt10b, ß-catenin and DKK1 expression may be related to the periodontal tissue remodeling following the application of an orthodontic force in rats. These observations suggest that the Wnt/ß-catenin signaling pathway may play a crucial role in periodontal tissue remodeling during OTM.

Expression analysis of α-smooth muscle actin and tenascin-C in the periodontal ligament under orthodontic loading or in vitro culture.

α-smooth muscle actin (α-SMA) and tenascin-C are stress-induced phenotypic features of myofibroblasts. The expression levels of these two proteins closely correlate with the extracellular mechanical microenvironment. We investigated how the expression of α-SMA and tenascin-C was altered in the periodontal ligament (PDL) under orthodontic loading to indirectly reveal the intrinsic mechanical microenvironment in the PDL. In this study, we demonstrated the synergistic effects of transforming growth factor-ß1 (TGF-ß1) and mechanical tensile or compressive stress on myofibroblast differentiation from human periodontal ligament cells (hPDLCs). The hPDLCs under higher tensile or compressive stress significantly increased their levels of α-SMA and tenascin-C compared with those under lower tensile or compressive stress. A similar trend was observed in the tension and compression areas of the PDL under continuous light or heavy orthodontic load in rats. During the time-course analysis of expression, we observed that an increase in α-SMA levels was matched by an increase in tenascin-C levels in the PDL under orthodontic load in vivo. The time-dependent variation of α-SMA and tenascin-C expression in the PDL may indicate the time-dependent variation of intrinsic stress under constant extrinsic loading.

Upregulated Leptin in Periodontitis Promotes Inflammatory Cytokine Expression in Periodontal Ligament Cells.

BACKGROUND: Imbalance or disruption in the expression of inflammatory mediators contributes greatly to the breakdown of the periodontal supporting tissues. Leptin, through binding to its receptor (obesity-related leptin and leptin receptor [OBR]), has potent effects on immunity and inflammation. However, to date, researchers only indicated a role of leptin in periodontitis. No direct or valid evidence exists about how leptin and its receptor are regulated by local inflammation, what effects they have, and the underlying mechanisms. METHODS: Experimental periodontitis was induced by ligation of mandibular second molars in beagle dogs. The expression of leptin, OBR, and interleukin (IL)-1ß was examined by immunohistochemistry. Meanwhile, recombinant human IL-1ß was used to stimulate human periodontal ligament cells (hPDLCs) in vitro, and mRNA and protein levels of leptin were measured using real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Then, mRNA and protein levels of IL-6 and IL-8 were measured using real-time PCR and ELISA, after stimulation with various concentrations of leptin, knocking down all or only the long form of OBR (OBRb) by small interfering RNA and incubation with multiple intracellular signaling pathway inhibitors, respectively. RESULTS: Leptin and OBR increased substantially in inflammatory periodontal tissues, which correlated well with the extent of inflammatory infiltration, and was a result of the upregulation in resident cells themselves. A high dose of leptin could induce the expression of mRNA and protein of IL-6 and IL-8 in hPDLCs through binding with OBRb and activating different intracellular signaling pathways. CONCLUSION: Upregulated leptin and OBR in periodontitis stimulated proinflammatory cytokine expression in PDL cells to additionally promote local inflammation.

Effects of electroacupuncture on experimental periodontitis in rats.

BACKGROUND: Acupuncture has shown the capability of modulating the immuno-inflammatory response of the host. This study aims to evaluate the effects of electroacupuncture (EA) on ligature-induced periodontitis in rats. METHODS: Thirty-two animals were divided into four groups: 1) control; 2) experimental periodontitis (EP); 3) sham-treated (EP/EA-sham); and 4) treated with EA (EP/EA). For the EP groups, a ligature was placed around the right mandibular first molars at day 1. Sessions of EA or EA-sham were assigned every other day. For EA treatment, large intestine meridian points LI4 and LI11 and stomach meridian points ST36 and ST44 were used. EA-sham was performed in off-meridian points. Animals were euthanized at day 11. Histomorphometric and microtomographic analyses were performed. Immunolabeling patterns for the receptor activator of nuclear factor κB ligand (RANKL), osteoprotegerin (OPG), and tartrate-resistant acid phosphatase (TRAP) were assessed. Expressions of interleukin (IL)-1ß, matrix metalloproteinase (MMP)-8, IL-6, and cyclooxygenase (COX)-2 messenger RNAs (mRNAs) were evaluated by quantitative reverse transcription-polymerase chain reaction. Data were analyzed statistically (P <0.05, analysis of variance). RESULTS: Histomorphometric and microtomographic analyses demonstrated that group EP/EA presented reduced alveolar bone loss when compared to group EP (P <0.05). Reduced RANKL immunolabeling and fewer TRAP-positive multinucleated cells were observed in the EA-treated group in relation to group EP. No differences were observed in OPG expression among groups. EA treatment decreased the genic expression of IL-1ß and MMP-8 (P <0.05), increased the mRNA expression of IL-6 (P <0.05), and did not modify the genic expression of COX-2 in animals with EP (P >0.05). CONCLUSION: It can be concluded that EA reduced periodontal tissue breakdown and the expression of some proinflammatory mediators and a proresorptive factor in EP in rats.