Periodontal tissue engineering by nano beta-tricalcium phosphate scaffold and fibroblast growth factor-2 in one-wall infrabony defects of dogs.

BACKGROUND AND OBJECTIVE: Nanoparticle bioceramics are being investigated for biomedical applications. We fabricated a regenerative scaffold comprising type I collagen and beta-tricalcium phosphate (ß-TCP) nanoparticles. Fibroblast growth factor-2 (FGF-2) is a bioeffective signaling molecule that stimulates cell proliferation and wound healing. This study examined the effects, on bioactivity, of a nano-ß-TCP/collagen scaffold loaded with FGF-2, particularly on periodontal tissue wound healing. MATERIAL AND METHODS: Beta-tricalcium phosphate was pulverized into nanosize particles (84 nm) and was then dispersed. A nano-ß-TCP scaffold was prepared by coating the surface of a collagen scaffold with a nanosize ß-TCP dispersion. Scaffolds were characterized using scanning electron microscopy, compressive testing, cell seeding and rat subcutaneous implant testing. Then, nano-ß-TCP scaffold, nano-ß-TCP scaffold loaded with FGF-2 and noncoated collagen scaffold were implanted into a dog one-wall infrabony defect model. Histological observations were made at 10 d and 4 wk postsurgery. RESULTS: Scanning electron microscopy images show that TCP nanoparticles were attached to collagen fibers. The nano-ß-TCP scaffold showed higher compressive strength and cytocompatibility compared with the noncoated collagen scaffold. Rat subcutaneous implant tests showed that the DNA contents of infiltrating cells in the nano-ß-TCP scaffold and the FGF-2-loaded scaffold were approximately 2.8-fold and 3.7-fold greater, respectively, than in the collagen scaffold. Histological samples from the periodontal defect model showed about five-fold greater periodontal tissue repair following implantation of the nano-ß-TCP scaffold loaded with FGF-2 compared with the collagen scaffold. CONCLUSION: The ß-TCP nanoparticle coating strongly improved the collagen scaffold bioactivity. Nano-ß-TCP scaffolds containing FGF-2 are anticipated for use in periodontal tissue engineering.

Osteoclastogenesis in Local Alveolar Bone in Early Decortication-Facilitated Orthodontic Tooth Movement.

OBJECTIVE: In the current study, we aimed to investigate the effects of alveolar decortication on local bone remodeling, and to explore the possible mechanism by which decortication facilitates tooth movement. MATERIALS AND METHODS: Forty rabbits were included in the experiment. The left mandible was subjected to decortication-facilitated orthodontics, and the right mandible underwent traditional orthodontics as a control. The animals were sacrificed on the days 1, 3, 5, 7 and 14, after undergoing orthodontic procedures. Tooth movement was measured by Micro-CT, and the local periodontal tissues were investigated using H&E, Masson's trichrome and tartrate-resistant acid phosphatase (TRAP) staining. The mRNA levels of genes related to bone remodeling in the alveolar bone were analyzed using real-time PCR. RESULT: On days 3, 5, 7 and 14, tooth movement was statistically accelerated by decortication (P<0.05) and was accompanied by increased hyperemia. Despite the lack of new bone formation in both groups, more osteoclasts were noted in the decorticated group, with two peak counts (P<0.05). The first peak count was consistent with the maximum values of ctsk and TRAP expression, and the second peak counts accompanied the maximum nfatc1 and jdp2 expression. The increased fra2 expression and the ratio of rankl/opg also accompanied the second peak counts. CONCLUSIONS: Following alveolar decortication, osteoclastogenesis was initially induced to a greater degree than the new bone formation which was thought to have caused a regional acceleratory phenomenon (RAP). The amount of steoclastogenesis in the decorticated alveolar bone was found to have two peaks, perhaps due to attenuated local resistance. The first peak count in osteoclasts may have been due to previously existing osteoclast precursors, whereas the second may represent the differentiation of peripheral blood mononuclear cells which came from circulation as the result of hyperemia.

Periodontal microstructure change and tooth movement pattern under different force magnitudes in ovariectomized rats: an in-vivo microcomputed tomography study.

INTRODUCTION: The purpose of this study was to evaluate the dynamic changes in the periodontal microstructure and the molar displacement pattern during orthodontic tooth movement in ovariectomized rats. METHODS: Twenty ovariectomized rats received either 100 or 30 g of orthodontic force to induce mesial movement of the maxillary left first molars over 14 days. Ten healthy rats underwent sham operations as controls. Periodontal ligament thickness, alveolar bone microstructural properties, and displacement of the molar were measured with 6 in-vivo microcomputed tomography scans for each sample. RESULTS: The ovariectomized rats that received 100 g of orthodontic force had obvious changes in periodontal ligament thickness at day 1 and poor periodontal ligament thickness recovery from days 5 through 14. The bone volume fraction increased and the trabecular separation decreased significantly in this group at day 3, and obvious bone loss was observed at day 14. Molar linear and angular movements were also higher in this group than in the other 2 groups. CONCLUSIONS: Relatively heavier force applications in ovariectomized rats resulted in poor periodontal ligament thickness recovery and local alveolar bone overcompression, and consequently induced undermining resorption and obvious alveolar bone loss; these led to high rates of tooth movement and molar inclination.

An experimental study on periodontal regeneration after subcutaneous transplantation of rat molar with and without cryopreservation: an in vivo study.

This study analysed the effects of cryopreservation on periodontal regeneration of autotransplanted rat molars. First and second maxillary molars (n=92) of 24 four-week-old Wistar rats were gently extracted and autotransplanted into the abdominal tissue immediately (control group n=44) or after cryopreservation in liquid nitrogen for 7 days (experimental group n=48). At 1, 2, 4 and 10 weeks after transplantation, the transplanted molars were excised and regeneration of the periodontal tissues was analysed on histological sections stained with routine H&E and Goldner method. Different tissue responses were scored on a tooth basis: inflammation, regeneration of the periodontal ligament, resorption/apposition of cementum, and alveolar bone formation. Sixty-two teeth were available for histological evaluation, including 30 experimental and 32 control samples. One week after transplantation, both control and test teeth were surrounded by granulation tissue and some areas of root resorption could be seen. After 2 weeks, signs of regeneration of the periodontal ligament, cementum apposition, and new bone formation roughly coincided in both groups, however markedly retarded in the experimental group. After 4 weeks, regeneration progressed equally in both groups, presenting fewer areas of cementum apposition in experimental samples. Finally, 10 weeks after baseline transplantation, no significant differences between both groups could be observed. Cryopreservation followed by autotransplantation of extracted teeth in rats appears to have minimal detrimental effects on regeneration of periodontal tissues after integration periods of 1-10 weeks. However, the present findings indicated that the regeneration process in general is retarded for cryopreserved teeth, as compared to their immediately transplanted homologues.

Adhesion and biologic behavior of human periodontal fibroblast cells to resin ionomer Geristore: a comparative analysis.

AIM: The resin ionomer Geristore, originally designed for restorative procedures, has been used extensively in treating subgingival defects (such as root resorption and perforations) and as a retrofilling material. The purpose of this study was to evaluate the cell adhesion as well as in vitro biocompatibility of human periodontal fibroblast cells with resin ionomer Geristore in comparison with mineral trioxide aggregate (MTA) and glass ionomer cement (GIC). MATERIAL AND METHOD: Adhesion, growth, and morphology of human periodontal fibroblasts over test materials were evaluated by scanning electron microscopy (SEM). Biocompatibility was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide salt (MTT) assay. RESULTS: Compared to glass coverslips, cells grew and spread qualitatively better over the surface of Geristore in comparison with the other test materials. In vitro interpretation indicates that Geristore is significantly less cytotoxic to human periodontal ligament cells. Results of statistical analysis revealed that material extracts had significant effect on cell proliferation at both 24 h (F = 547.62, P < 0.05) and at 48 h (F = 6048.18, P < 0.05). CONCLUSION: Our study supports that Geristore has enhanced biologic behavior to human periodontal ligament cells and superior biocompatibility in comparison with MTA and GIC, so it can be suggested as a material of choice in root resorption, perforations, and root-end filling.

Ultra-structure characterization of self-etching treated cementum surfaces

Objectives: to evaluate the effect of different conditioning treatments on surface roughness and topography ofdental cementum. Study Design: Extracted human canines were used for the present study. The mesial surfacefrom the cervical third of the roots were ground flat with wet 600-grit silicon carbide paper. They were polished(up to 1/4 μm diamond paste) and treated as follows: 1) No treatment, 2) 35% H3PO4 during 15 s, 3) Clearfil SEBond primer (SEB), 4) One-Up Bond F (OUB). The adhesive systems were applied following manufacturer’s instructions.SEB primer and OUB were removed from surfaces by washing and ultrasonic agitation with ascendingethanol solutions. Digital images of treated surfaces (5x5 and 15x15 μm) were obtained by means of an atomicforce microscope (AFM) analysis. The average surface roughness (Ra nanometers) of the scanned areas was assessed.Data were analyzed by ANOVA and SNK multiple comparisons tests (p<0.05). Results: phosphoric acidtreatment produced the highest mean roughness value, at all scan sizes. At 5x5 μm AFM images, for self-etchadhesive systems no differences in roughness were detected. At 15x15 μm, when One-Up Bond F was employedthe lowest value was obtained. Conclusions: When phosphoric acid treatment was applied, cementum surfaceroughness increased and a strong demineralization with exposed collagen fibers could be observed (AU)

Periodontal alteration of the microcirculation and hypercholesterolemia: a possible correlation?

OBJECTIVE: We evaluated the morphological and parametric characteristics of the periodontal microcirculation in patients diagnosed as having hypercholesterolemia and high levels of low-density lipoprotein (LDL). METHODS: Forty patients were recruited, 20 of whom were affected by hypercholesterolemia and 20 of whom were considered healthy. A videocapillaroscopic examination was carried out on the periodontal mucosa in the proximity of the frenulum (II, V sextant). RESULTS: The difference between the parameters of the hypercholesterolemia group and the control group was evaluated with the Mann-Whitney U-test for non-parametric ordinal data; the level of significance being P<0.05. The videocapillaroscopy documented extremely significant differences between the two groups, regarding the following parameters: total diameter of the loop (P=0.0017), diameter of the afferent loops (P=0.0004), diameter of the efferent loops (P=0.00008) and periodontal density (P=0.0001). CONCLUSIONS: The capillaroscopic examination revealed a morphological alteration of the periodontal microcirculation in patients with hypercholesterolemia, which is an expression of peripheral vascular phlogosis.

Hormonal influence on the physiology and pathology of the dental gingival unit: the preliminary results of a pilot study.

AIM: The effects of steroid hormones on the periodontium are most prominent at certain stages of a woman's life especially during the menstrual cycle when there is an increase in the secretion of sex hormones or a significant fluctuation in their concentration. The deterioration of existing periodontal conditions can be attributed to a fluctuation in the steroid hormones in circulation. By contributing to our understanding of periodontal changes caused by variation in hormone concentrations, this study aims to encourage the implementation in dental practice of the most suitable forms of treatment for hormone-related pathologies. METHODS: Tartar was removed from the teeth of five young women and four biopsies and blood tests were carried out on them at regular intervals. The information gathered was used to monitor periodontal changes arising from variation in hormone concentrations. RESULTS: The histological analysis of the test samples under an optical microscope did not reveal signs of inflammation, hyperaemia or oedema at any stage of the menstrual cycle in the patients examined. The extent of gingival Keratinization was found to be comparable to that present in the follicular phase. CONCLUSION: The occurrence of ovulation could not be shown in the pilot study. The histological analysis and the analysis of hormone concentrations show primarily the absence of surges in estradiol and LH which normally accompany ovulation; the levels recorded are similar to those found in the follicular phase.

Periodontal tissue engineering and regeneration: current approaches and expanding opportunities.

The management of periodontal tissue defects that result from periodontitis represents a medical and socioeconomic challenge. Concerted efforts have been and still are being made to accelerate and augment periodontal tissue and bone regeneration, including a range of regenerative surgical procedures, the development of a variety of grafting materials, and the use of recombinant growth factors. More recently, tissue-engineering strategies, including new cell- and/or matrix-based dimensions, are also being developed, analyzed, and employed for periodontal regenerative therapies. Tissue engineering in periodontology applies the principles of engineering and life sciences toward the development of biological techniques that can restore lost alveolar bone, periodontal ligament, and root cementum. It is based on an understanding of the role of periodontal formation and aims to grow new functional tissues rather than to build new replacements of periodontium. Although tissue engineering has merged to create more opportunities for predictable and optimal periodontal tissue regeneration, the technique and design for preclinical and clinical studies remain in their early stages. To date, the reconstruction of small- to moderate-sized periodontal bone defects using engineered cell-scaffold constructs is technically feasible, and some of the currently developed concepts may represent alternatives for certain ideal clinical scenarios. However, the predictable reconstruction of the normal structure and functionality of a tooth-supporting apparatus remains challenging. This review summarizes current regenerative procedures for periodontal healing and regeneration and explores their progress and difficulties in clinical practice, with particular emphasis placed upon current challenges and future possibilities associated with tissue-engineering strategies in periodontal regenerative medicine.

Ultrastructural modifications at the level of marginal periodontium in the case of incorrect dental reconstruction.

Our study focused on the evolution of the marginal periodontium inflammatory process caused by an incorrect dental reconstruction. Our research studied a control group and a group of patients having traumatic and inflammatory lesions in different stages of evolution. A pronounced rarefaction of the junction desmosome structures as well as an inflammatory process pointed out by the presence of macrophages, neutrophils, Langerhans' cells, and mastocytes. The presence of altered fibroblasts and collagen fibers in the electron microscopic sections of vascular lesions represents microscopic signs of the inflammation and support the theory of local immunoglobulin synthesis.

Preserved microstructure and mineral distribution in tooth and periodontal tissues in early fossil hominin material from Koobi Fora, Kenya.

The aim of this study was to explore further the preservation of tissues and the mineral distribution in 1.6 million-year-old fossil hominin material from Koobi Fora, Kenya attributed to Paranthropus boisei (KNM-ER 1817). Bone, dentine and cementum microstructure were well preserved. Electron microprobe analysis of dentine and bone revealed an F-bearing apatite. Calcite now filled the original soft tissue spaces. The average Ca/P atomic ratio was 1.93, as compared to 1.67 in biological hydroxyapatite, indicating that the Ca-content had increased during fossilization. Analytical sums for mineral content were approximately 90 wt%. Some of the remaining 10 wt% may be preserved organic material. Demineralized dentine fragments showed irregularly distributed tubules encircled with a fibrous-like electron-dense material. A similar material was observed in demineralized dentine. Within this, structures resembling bacteria were seen. In demineralized bone an electron-dense material with a fibrous appearance and a banding pattern that repeated every 64 nm, similar to that of collagen, was noted. SEM of an enamel fragment (KNM-ER 6081) showed signs of demineralization/remineralization. Retzius lines, Hunter-Schreger bands and prism cross-striations spaced 3.7-7.1.microm apart were noted. Prisms were arranged in a pattern 3 configuration and deeper areas containing aprismatic enamel were occasionally observed. We conclude that a great deal of informative microstructure and ultrastructure remains preserved in this fossil material. We also hypothesize that the high mineral content of the tissues may 'protect' parts of the organic matrix from degradation, since our findings indicate that some organic matrix may still be present.

Recapitulating development: a template for periodontal tissue engineering.

The induction of bone formation by the soluble osteogenic molecular signals of the transforming growth factor-beta (TGF-beta) superfamily is a critical issue to periodontologists, molecular biologists, and tissue engineers alike, because preclinical studies in primates and clinical trials have demonstrated the bone induction capacity of bone morphogenetic and osteogenic proteins (BMPs/OPs) in clinical context. BMPs/OPs, pleiotropic members of the TGF-beta superfamily, induce de novo endochondral bone formation as a recapitulation of embryonic development and act as soluble signals for tissue morphogenesis sculpting the multicellular mineralized structures of the periodontal tissues with functionally oriented periodontal ligament fibers inserting into newly formed cementum. This paper reviews the induction of the complex tissue morphologies of the periodontal tissues in the nonhuman primate Papio ursinus with furcation defects treated with doses of naturally derived and recombinantly produced human BMPs/OPs. Periodontal tissue regeneration develops as a mosaic structure in which the OPs of the TGF-beta superfamily singly, synergistically, and synchronously initiate and maintain tissue induction and morphogenesis.

[Effect of intermitent hypoxia on ultrastructure and prooxidant-antioxidant balance in the soft tissue of the periodontium in acute immobilization stress].

The changes of prooxidant-antioxidant balance and morphofunctional state of soft tissue of paradontum under the acute immobilization stress, intermittent hypoxia and its complex influence were investigated. It was shown, that intermittent hypoxia can stimulate own endogenous mechanisms of nonspecific resistance, such as antioxidant protection in soft tissue of paradontum. Normalization of the prooxidant-antioxidant balance under immobilization stress after intermittent hypoxia essentially improved the morphofunctional state of tissue investigated with decrease of edema and improvement of mitochondria apparatus ultrastructure.

Low frequency noise effects on the periodontium of the Wistar rat - a light microscopy study

Long-term low-frequency noise (LFN)(m500Hz, infrasound included) exposure is known to cause extracellular matrix proliferation with fibrosis, in the absence of inflammatory signs. The aim of this work was to study the morphological alterations to the periodontium of Wistar rats exposed to LFN. 10 rats were exposed to LFN for 2184 consecutive hours and another 10 were kept in silence. The mandibles were removed, kept in 10%buffered formalin, sectioned sagitally, stained with haematoxylin-eosin (HE) and Masson¡¯s trichromic solution (TCM) and observed with light microscopy (LM).The results revealed a disappearance of thecementum, irregular erosion of surface alveolarbone, and signs of bone necrosis, with detached bone particles. The periodontal ligament was disorganized and had deficient anchorage of the fibers.These findings may be due to a direct effectof noise and vibration impinging on the structures,to stress, to vascular alterations or to a combination of these factors. They may also explain the alterations in alveolar bone, reported by other authors (AU)

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A comparative study of systemic subantimicrobial and topical treatment of minocycline in experimental periodontitis of rats.

OBJECTIVE: The purpose of this study was to compare the effectiveness of minocycline on treating experimentally induced periodontitis in rats when administered either as a systemic subantimicrobial dose or as a topical ointment. DESIGN: Thirty-two adult male Sprague-Dawley rats in four experimental groups-(1) model group; (2) systemic subantimicrobial dose of minocycline (5mg/kg/day) treatment group; (3) topical subgingival dose of minocycline (2mg/animal/week) treatment group; (4) control group. Experimentally induced periodontitis-silk ligatures were placed around the crevices of the second molar teeth and the animals fed a 10% sucrose drink. Assessment was carried out at days 28 and 56 using a number of different visual, histological and ultrastructure approaches. (1) Visual assessment-tooth mobility, gingival index and alveolar bone loss. (2) Histological examination-monocyte infiltration and resorption lacunae with osteoclasts. (3) Transmission electron microscopy (TEM)-morphological transformation of fibroblasts and osteoclasts. The collected data were analysed for statistical significance using the analysis of variance statistical test. RESULTS: Minocycline significantly reduced tooth mobility, gingival index and alveolar bone loss when administered either systemically or as a topical ointment compared to the model group (P<0.01). However, the alveolar bone loss was significantly less (P<0.01 in the systemic treatment group compared to the local treatment group. Monocyte infiltration and resorption lacunae with osteoclasts were significantly less in the both treatment groups compared to the model group (P<0.01). The osteoclasts failed to form a ruffled border in the systemic treatment group. CONCLUSION: Topical treatment significantly reduces gingivitis while systemic treatment is beneficial in terms of inhibiting alveolar bone loss.

Efficacy of antibiotics against periodontopathogenic bacteria within epithelial cells: an in vitro study.

BACKGROUND: Periodontopathogenic bacteria can invade and survive within epithelial cells, but susceptibility of intracellular infection to antibiotics used in periodontitis treatment has not been studied to date. METHODS: KB cells were infected by Actinobacillus actinomycetemcomitans, strain NCTC 9710; Porphyromonas gingivalis, strains ATCC 33277 and JH16-1; or Streptococcus constellatus, strain J012b. After 2, 4, and 12 hours the bactericidal effect of antibiotics (clindamycin, doxycycline, metronidazole, and moxifloxacin) on intracellular microorganisms was tested at a concentration up to the 100-fold minimum inhibitory concentration (MIC) determined separately on planktonic bacteria. RESULTS: The P. gingivalis strains differed in their invasiveness and ATCC 33277 was 100-fold more invasive than JH16-1. Doxycycline and clindamycin at a concentration 10-fold MIC had no effect, but P. gingivalis intercellular infection was significantly reduced by metronidazole at 10-fold MIC after 2 and 4 hours. Moxifloxacin was effective, but a 100-fold MIC concentration was necessary to reduce P. gingivalis strains intracellular growth to 7% of the control. Other bacterial species grown inside the KB cells were more susceptible to antibiotics. Clindamycin at 10-fold MIC reduced the number of intracellular S. constellatus after 4 and 12 hours. This bacterium was eliminated by moxifloxacin at 50-fold MIC. Intracellular A. actinomycetemcomitans was killed by 10-fold MIC of doxycycline and moxifloxacin after 4 hours incubation. CONCLUSIONS: Moxifloxacin was the most efficient antibiotic to treat intracellular infection. However, taking into account the MIC values and the levels of antibiotics in gingival fluid, elimination of intracellular bacteria by antibiotics alone seems to be questionable.

ArF-193 excimer laser and Emdogain in the treatment of experimental periodontitis: an experimental study in rabbits.

OBJECTIVE: We aimed to investigate how the progress made on laser technology during the last ten years could overcome this obstacle and allow the use of lasers in periodontology, together with the application of a number of products permitting the regeneration of periodontal tissues. BACKGROUND DATA: The use of lasers in dentistry remains controversial, in spite of their increasing application in medical practice. The main reason for this discrepancy is the frequent report of damage to surrounding tissues and the dental pulp, due to the energy transfer, from the site of laser impact. METHODS: Experimental periodontitis was initiated in fifteen rabbits. Animals were divided into five equal groups. In the control group, no therapy was applied. The remaining four groups were treated with curettage or ArF 193 excimer laser, under conditions of strict control of frequency, fluency, and application, without or with the application of a periodontal healing product (Emdogain). Laser was applied by the use of a new, articulated arm for beam delivery. Pocket depth and microscopic analysis were performed three weeks after treatment. RESULTS: Our results show that all treatment groups decreased pocket depth significantly. ArF193 excimer laser does not produce any histological damage to the dental pulp, and facilitates periodontal regeneration. This result is highly facilitated by the application of Emdogain). CONCLUSIONS: The use of UV lasers, under a tight control of its energy, may be a valuable tool for the treatment of periodontal diseases, especially combined with the use of healing products. Further study is need to confirm these results.

Efectos clínicos e histológicos de la cocaína sobre el periodonto de protección / Clinical and histologic effects of cocaine on protecting periodontium

El consumo de drogas psicotrópicas es uno de los grandes problemas de salud en el mundo. La cocaína, en sus distintas presentaciones y vías de administración tales como la aplicación por frotación sobre la encía, se convirtió en especial interés para esta investigación, debido a que en nuestra actividad clínica hemos encontrado casos aislados de alteraciones periodontales en las cuales los pacientes aseguraron haber usado cocaína por frotación sobre la zona afectada. En este estudio se utilizaron 40 ratas de la cepa Wistar (20 experimentales y 20 control) durante 16 semanas. Se demostró que la cocaína produce alteraciones clínicas e histológicas en la encía, siendo la gingivitis crónica la patología más frecuente en un 70 por ciento de los casos, por lo cual se debe considerar en la evaluación clínica el consumo de drogas como factor de riesgo de la enfermedad periodontal (AU)

Ciliated fibroblasts in mouse connective tissues: a comparison between methods for estimating their frequency.

The frequency of ciliated fibroblasts in skin, gingiva, molar and incisor periodontal ligaments and incisor enamel-related periodontium of the mouse was estimated by straight counting or by methods based on the probability of observing a basal body in relation to other cell structures. Transmission electron microscopy of ultra-thin sections mounted in single slot grids was used. The results obtained with these methods differed, but indicated that periodontal ligament fibroblasts from molars or incisors generally had a higher ciliation index than the fibroblasts from skin and gingiva. These differences may not be real since the detection of cilia and/or centriolar structures seems to depend very much upon the plane of sectioning relative to the long axis of the fibroblasts, a situation which favours the more regularly arranged periodontal fibroblasts. This arrangement makes the periodontal tissues, particularly those of rodent incisors, a valuable model for studying ciliation in vivo because of the prompt response to experimental manipulation.