Effects of Bifidobacterium animalis subsp. lactis HN019 on ligature-induced periodontitis in rats with experimental rheumatoid arthritis.

The purpose of this study was to evaluate the effects of systemic administration of the probiotic Bifidobacterium animalis subsp. lactis HN019 (HN019) on ligature-induced periodontitis in rats with experimental rheumatoid arthritis (RA). 28 rats were divided into four groups (n=7): RA (rheumatoid arthritis), RA/PROB (probiotic), RA/EP (experimental periodontitis) and RA/EP/PROB. From day zero, HN019 was added daily to the water of the PROB groups animals until the end of the experiment. From day seven, RA was induced. On day 28, in EP groups, ligatures were positioned around mandibular first molars and remained in position for 11 days, in order to induce periodontitis. The animals were euthanised on day 39. Microtomographic, histomorphometric, immunoenzymatic and microbiological analyses were performed. Data were statistically analysed (P<0.05). Group RA/EP/PROB presented reduced alveolar bone loss, tumour necrosis factor-α and interleukin (IL)-6 levels and increased IL-17 levels when compared with group RA/EP. There were no significant differences regarding connective tissue attachment level and IL-10 levels between groups RA/EP and RA/EP/PROB. Group RA/PROB showed decreased anti-citrullinated protein antibodies levels when compared with groups RA and RA/EP. Group RA/EP/PROB presented a higher rate of aerobic/anaerobic bacteria than group RA/EP. Systemic administration of HN019 promoted a protective effect against periodontal tissue destruction, decreasing both bone loss and inflammatory mediators and increasing the proportion of bacteria compatible with periodontal health, in rats with experimental RA and EP.

Impact of resveratrol on bone repair in rats exposed to cigarette smoke inhalation: histomorphometric and bone-related gene expression analysis.

This study investigated the effect of resveratrol on bone healing and its influence on the gene expression of bone-related markers in rats exposed to cigarette smoke. Two calvarial defects were created in each of 60 rats, which were assigned equally (n=20) to three groups: (1) resveratrol (10mg/kg)+smoke exposure (SMK+RESV); (2) placebo+smoke exposure (SMK+PLA); or (3) placebo+no smoke exposure (NS+PLA). Substances were administered daily for 30days following surgery. Smoke inhalation was started 7days before surgery and continued for 30days after surgery. One defect was processed for histomorphometric analysis and the other was used for mRNA quantification of bone-related gene expression by qPCR. The remaining defect was smaller in the SMK+RESV (2.27±0.61mm, P=0.0003) and NS+PLA (2.17±0.74mm, P=0.0005) groups than in the SMK+PLA group (3.12±0.47mm). Higher levels of Runx2 were observed in the NS+PLA group than in the smoke exposure groups (vs. SMK+PLA, P=0002; vs. SMK+RESV, P=0.052); levels of Lrp-5 were also higher in the no smoke exposure group (vs. SMK+RESV, P=0.009; vs. SMK+PLA, P=0.003). Resveratrol therapy decreased RANKL/OPG expression when compared to placebo (SMK+RESV vs. SMK+PLA, P=0.017). Dkk1 levels were decreased in the SMK+RESV group when compared to the SMK+PLA (P=0.006) and NS+PLA groups (P=0.005). In conclusion, resveratrol optimizes the repair of critical-sized bone defects, up-regulating the gene expression of important bone remodelling markers in rats exposed to cigarette smoke inhalation.

Periodontitis in Chédiak-Higashi Syndrome: An Altered Immunoinflammatory Response.

Chédiak-Higashi syndrome (CHS), a rare autosomal recessive disorder caused by mutations in the lysosomal trafficking regulator gene (LYST), is associated with aggressive periodontitis. It is suggested that LYST mutations affect the toll-like receptor (TLR)-mediated immunoinflammatory response, leading to frequent infections. This study sought to determine the periodontal status of patients with classic (severe) and atypical (milder) forms of CHS and the immunoregulatory functions of gingival fibroblasts in CHS patients. In contrast to aged-matched healthy controls, atypical (n = 4) and classic (n = 3) CHS patients presented with mild chronic periodontitis with no evidence of gingival ulceration, severe tooth mobility, or premature exfoliation of teeth. As a standard of care, all classic CHS patients had undergone bone marrow transplantation (BMT). Primary gingival fibroblasts obtained from atypical and BMT classic CHS patients displayed higher protein expression of TLR-2 (1.81-fold and 1.56-fold, respectively) and decreased expression of TLR-4 (-2.5-fold and -3.85-fold, respectively) at baseline when compared with healthy control gingival fibroblasts. When challenged with whole bacterial extract of Fusobacterium nucleatum, both atypical and classic CHS gingival fibroblasts failed to up-regulate TLR-2 and TLR-4 expression when compared with their respective untreated groups and control cells. Cytokine multiplex analysis following F. nucleatum challenge showed that atypical CHS gingival fibroblasts featured significantly increased cytokine expression (interleukin [IL]-2, IL-4, IL-5, IL-6, IL-10, IL-12, interferon-γ, tumor necrosis factor-α), whereas classic CHS cells featured similar/decreased cytokine expression when compared with treated control cells. Collectively, these results suggest that LYST mutations in CHS patients affect TLR-2 and TLR-4 expression/function, leading to dysregulated immunoinflammatory response, which in turn may influence the periodontal phenotype noted in CHS patients. Furthermore, our results suggest that atypical CHS patients and classic CHS patients who undergo BMT early in life are less susceptible to aggressive periodontitis and that hematopoietic cells play a critical role in mitigating the risk of aggressive periodontitis in CHS. Knowledge Transfer Statement: Results from this study can be used to create awareness among clinicians and researchers that not all CHS patients exhibit historically reported aggressive periodontitis, especially if they have atypical CHS disease or have received bone marrow transplantation. LYST mutations in CHS patients may affect TLR-2 and TLR-4 expression/function leading to dysregulated immunoinflammatory response, which in turn may influence the periodontal phenotype noted in CHS patients.

The impact of different torques for the insertion of immediately loaded implants on the peri-implant levels of angiogenesis- and bone-related markers.

The aim of this split-mouth, randomized, double-blind, controlled clinical trial was to evaluate the influence of different insertion torque values for dental implants on bone- and angiogenesis-related marker profiles. Eighteen edentulous patients received dental implants and fixed complete-arch mandibular prostheses. The implants (n=36) were assigned randomly to two groups: reduced torque (n=18), with insertion torque <30Ncm; and conventional torque (n=18), with insertion torque ≥30Ncm. Levels of vascular endothelial growth factor (VEGF), placental growth factor, bone morphogenetic protein 9 (BMP-9), periostin, osteoprotegerin (OPG), and tartrate-resistant acid phosphatase (TRAP) in the peri-implant fluid were quantified at 7, 14, 30, and 120days after implant placement. Inter-group comparisons showed that VEGF and OPG levels were higher in the low-level torque group than in the conventional torque group on days 7 and 30, respectively (P<0.05). BMP-9 and periostin levels were higher in the conventional group than in the low-level torque group on day 120, and TRAP was up-regulated around implants inserted with conventional torque when compared to those inserted with lower-level torque at all time points evaluated (P<0.05). In conclusion, the use of different levels of torque for implantation of immediately loaded implants significantly influenced the levels of bone- and angiogenesis-related markers during early peri-implant repair.

Transgenerational effects of a hypercaloric diet.

The effects of a maternal hypercaloric diet (HD) during puberty and early adulthood on neuroimmune aspects in offspring were investigated. In female rats of the F0 generation and male rats of the F1 generation, bodyweight (BW) gain, retroperitoneal fat (RPF) weight, the number of hypodermic adipocytes (HAs) and expression of glial fibrillary acidic protein (GFAP) were measured in hypothalamic astrocytes. On Postnatal Day 50, the F1 pups were challenged with lipopolysaccharide (LPS, 100µgkg-1, s.c.) or an equal volume of saline (S), and behaviour in the open field test was evaluated, as were plasma neuropeptide and cytokine concentrations. The maternal HD caused the female F0 rats to become overweight. The F1 offspring of dams fed the HD and challenged with saline (HDS group) exhibited increases in BW gain, RPF weight and in the number of large HAs and a decrease in GFAP immunoreactivity. F1 offspring of dams fed the HD and challenged with LPS (HDLPS group) exhibited decreases in BW gain, RPF weight and GFAP immunoreactivity, but no differences were observed in the number of larger and small HAs. Plasma tumour necrosis factor-α concentrations were high in the HDS and HDLPS groups. Thus, the maternal HD during puberty and early adulthood caused the F1 generation to become overweight despite the fact that they received a normocaloric diet. These results indicate a transgenerational effect of the HD that may occur, in part, through permanent changes in immune system programming. The attenuation of neuroinflammation biomarkers after LPS administration may have resulted in a decrease in the number of adipocytes, which, in turn, reduced cytokine, adipokine and chemokine levels, which are able to recruit inflammatory cells in adipose tissue.

Systemic treatment with resveratrol and/or curcumin reduces the progression of experimental periodontitis in rats.

BACKGROUND AND OBJECTIVE: Periodontitis is a chronic inflammatory disease of periodontal tissues that leads to the destruction of bone and other connective tissues. Resveratrol and curcumin are plant-derived substances with biological properties that may have immunomodulatory properties. This study investigated the effect of continuous administration of resveratrol and curcumin and the association of resveratrol and curcumin on the progression of experimental periodontitis in rats. MATERIAL AND METHODS: Forty Wistar rats were assigned randomly to the following groups: group 1, experimental periodontitis + placebo (PL) (n = 10); group 2, experimental periodontitis + resveratrol (RSV) (n = 10); group 3, experimental periodontitis + curcumin (C) (n = 10); and group 4, experimental periodontitis + resveratrol + curcumin (COMBI) (n = 10). Periodontitis was induced in rats by tying a silk suture, as a ligature, around one of the first molars. Daily administration of the placebo solution, 10 mg/kg of resveratrol, 100 mg/kg of curcumin or 10 mg/kg of resveratrol plus 100 mg/kg of curcumin was carried out from day 0 to day 30. At the end of the relevant experimental periods, rats were killed and the specimens obtained were processed for morphometric analysis of bone loss. Gingival tissues surrounding the first molar were collected for quantification of interleukin (IL)-1ß, IL-4, interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α) using a Luminex/MAGPIX assay. RESULTS: Intergroup comparisons of the morphometric outcomes revealed higher bone-loss values in the PL group (p < 0.05) when compared with RSV, C and COMBI groups. There was no difference in bone-loss values among RSV, C and COMBI groups (p > 0.05). The immunoenzymatic assay of the gingival tissue showed a lower concentration of IL-1ß in the COMBI group in comparison with the PL group (p < 0.05). Higher values of IL-4 were demonstrated in groups RSV, C and COMBI in comparison with the PL group (p < 0.05). Only RSV caused a reduction in the levels of IFN-γ (p < 0.05). There was no difference in the concentration of TNF-α amongst the four groups (p > 0.05). CONCLUSION: Resveratrol and curcumin are capable of reducing alveolar bone loss in an animal model of periodontitis. This occurred when these agents were added singly or in combination with one another, but there did not appear to be either synergistic or additive effects.

Impact of smoking on experimental gingivitis. A clinical, microbiological and immunological prospective study.

OBJECTIVE: The present study assessed the effect of smoking on clinical, microbiological and immunological parameters in an experimental gingivitis model. MATERIAL AND METHODS: Twenty-four healthy dental students were divided into two groups: smokers (n = 10); and nonsmokers (n = 14). Stents were used to prevent biofilm removal during brushing. Visible plaque index (VPI) and gingival bleeding index (GBI) were determined 5- on day -7 (running phase), baseline, 21 d (experimental gingivitis) and 28 d (resolution phase). Supragingival biofilm and gingival crevicular fluid were collected and assayed by checkerboard DNA-DNA hybridization and a multiplex analysis, respectively. Intragroup comparison was performed by Friedman and Dunn's multiple comparison tests, whereas the Mann-Whitney U-test was applied for intergroup analyses. RESULTS: Cessation of oral hygiene resulted in a significant increase in VPI, GBI and gingival crevicular fluid volume in both groups, which returned to baseline levels 7 d after oral hygiene was resumed. Smokers presented lower GBI than did nonsmokers (p < 0.05) at day 21. Smokers had higher total bacterial counts and higher proportions of red- and orange complex bacteria, as well as lower proportions of Actinomyces spp., and of purple- and yellow-complex bacteria (p < 0.05). Furthermore, the levels of key immune-regulatory cytokines, including interleukin (IL)-8, IL-17 and interferon-γ, were higher in smokers than in nonsmokers (p < 0.05). CONCLUSION: Smokers and nonsmokers developed gingival inflammation after supragingival biofilm accumulation, but smokers had less bleeding, higher proportions of periodontal pathogens and distinct host-response patterns during the course of experimental gingivitis.

Impact of type 2 diabetes on the gene expression of bone-related factors at sites receiving dental implants.

This study evaluated the influence of type 2 diabetes mellitus (T2DM) on the gene expression of bone-related factors in alveolar bone tissue from sites designated to receive dental implants. Bone biopsies were harvested from sites of planned implants for 19 systemically healthy patients and 35 patients with T2DM (17 with better-controlled T2DM (glycated haemoglobin (HbA1c) levels ≤8%) and 18 with poorly controlled T2DM (HbA1c levels >8%)). The mRNA levels of tumour necrosis factor alpha, transforming growth factor beta, receptor activator of the nuclear factor kappa B ligand (RANKL), osteoprotegerin (OPG), runt-related transcription factor 2, alkaline phosphatase, bone sialoprotein (BSP), type I collagen (COL-I), and osteocalcin were evaluated by quantitative real-time polymerase chain reaction. T2DM up-regulates RANKL levels and the ratio of RANKL/OPG, whereas it down-regulates COL-I and BSP expression (P<0.05). Higher mRNA levels of RANKL/OPG were observed in the poorly controlled T2DM patients compared to those with better-controlled T2DM and systemically healthy patients (P<0.05). A lower amount of COL-I and BSP was detected in the biopsies from individuals with poorly controlled T2DM compared to systemically healthy patients (P<0.05). In conclusion, RANKL, RANKL/OPG, COL-I, and BSP are negatively affected in diabetics. Additionally, the patient's glycaemic status appears to modulate bone-related genes in a different manner.

Resveratrol improves bone repair by modulation of bone morphogenetic proteins and osteopontin gene expression in rats.

This study investigated the effect of resveratrol on bone healing and its influence on the gene expression of osteogenic markers. Two calvarial defects were created and one screw-shaped titanium implant was inserted in the tibia of rats that were assigned to daily administration of placebo (control group, n=15) or 10mg/kg of resveratrol (RESV group, n=15) for 30 days. The animals were then sacrificed. One of the calvarial defects was processed for histomorphometric analysis and the tissue relative to the other was collected for mRNA quantification of bone morphogenetic protein (BMP)-2, BMP-7, osteopontin (OPN), bone sialoprotein (BSP), osteoprotegrin (OPG), and receptor activator of NF-κB ligand (RANKL). Implants were removed by applying a counter-torque force. Histomorphometric analysis revealed higher remaining defect in the calvarial defects of the control group than the RESV group (P=0.026). Resveratrol increased the counter-torque values of implant removal when compared to control therapy (P=0.031). Gene expression analysis showed a higher expression of BMP-2 (P=0.011), BMP-7 (P=0.049), and OPN (P=0.002) genes in the RESV group than in the control group. In conclusion, resveratrol improved the repair of critical-sized bone defects and the biomechanical retention of implants. Indeed, this natural agent may up-regulate the gene expression of important osteogenic markers.

Release of bone markers in immediately loaded and nonloaded dental implants: a randomized clinical trial.

The aim of this study was to compare the release of bone markers during osseointegration of immediately loaded and nonloaded implants. Forty patients who were indicated for rehabilitation with dental implants randomly received either implant and prosthesis placement within 72 hours (group IM) or implant insertion and no prosthesis placement (group NL). Peri-implant crevicular fluid was collected immediately after implant insertion and 7, 15, 30, 60, 90, and 120 days after surgery and levels of osteoprotegerin, transforming growth factors, osteocalcin, osteopontin, and parathyroid hormone were evaluated using Luminex assay. Bleeding index and peri-implantar sulcus depth were also evaluated. The data were compared using statistical tests (α = 5%). No statistical difference was found regarding demographic and clinical parameters (p > .05). Transforming growth factors, osteoprotegerin, osteopontin, and parathyroid hormone presented an earlier release peak in group IM than in NL group (p < .05). Osteocalcin achieved higher levels in group IM versus group NL between 7 and 30 days of evaluation (p < .05). It may be concluded that earlier loading positively modulates bone mediators release around immediately loaded implants when compared with nonloaded dental implants.