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Sulforaphane (SFN) has shown potential as an antioxidant and anti-inflammatory agent. To improve its druggability, we developed new analgesic formulations with sulforaphane-loaded hyaluronic acid (HA)-poloxamer (PL) hydrogel. This study evaluated the pre-clinical safety and effectiveness of these formulations. Effectiveness was tested on Wistar rats divided into groups (n = 15) receiving (IM, 10 mg/kg) SFN formulations or control groups (without SFN). This study used a hind paw incision postoperative pain model to evaluate mechanical hypersensitivity with von Frey filaments. TNF-α, IL-1ß, substance P, and CGRP levels verified anti-inflammatory activity in the hind paw tissue. Histopathology of tissues surrounding the injection site was assessed after 2 and 7 days post-treatment. To corroborate drug safety, cell viability of 3T3 and RAW 264.7 cultures was assessed. Additionally, RAW 264.7 cultures primed with carrageenan evaluated nitric oxide (NO) levels. All animals exhibited post-incisional hypersensitivity, and F2 (PL 407/338 (18/2%) + HA 1% + SFN 0.1%) showed a longer analgesic effect (p < 0.05). F2 reduced TNF-α, IL-1ß, and CGRP levels (p < 0.05). Histopathological evaluation showed mild to moderate inflammatory reactions after the formulations' injections. F2 produced no significant difference in cell viability (p > 0.05) but reduced NO production (p < 0.05). Thus, our results highlight the biocompatibility and effectiveness of F2.
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This study aimed to investigate whether GSK-3 inhibition (CHIR99021) effectively promoted mineralization by cementoblasts (OCCM-30). OCCM-30 cells were used and treated with different concentrations of CHIR99021 (2.5, 5, and 10 mM). Experiments included proliferation and viability, cellular metabolic activity, gene expression, and mineral nodule formation by Xylene Orange at the experimental time points. In general, CHIR99021 did not significantly affect OCCM-30 viability and cell metabolism (MTT assay) (p > 0.05), but increased OCCM-30 proliferation at 2.5 mM on days 2 and 4 (p < 0.05). Data analysis further showed that inhibition of GSK-3 resulted in increased transcript levels of Axin2 in OCCM-30 cells starting as early as 4 h, and regulated the expression of key bone markers including alkaline phosphatase (Alp), runt-related transcription factor 2 (Runx-2), osteocalcin (Ocn), and osterix (Osx). In addition, CHIR99021 led to an enhanced mineral nodule formation in vitro under both osteogenic and non-osteogenic conditions as early as 5 days after treatment. Altogether, the results of the current study suggest that inhibition of GSK-3 has the potential to promote cementoblast differentiation leading to increased mineral deposition in vitro.
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Cemento Dentário , Quinase 3 da Glicogênio Sintase , Quinase 3 da Glicogênio Sintase/metabolismo , Quinase 3 da Glicogênio Sintase/farmacologia , Proliferação de Células , Osteocalcina/análise , Diferenciação Celular/fisiologiaRESUMO
OBJECTIVE: Semaphorin 4D (Sema4D) is a coupling factor expressed on osteoclasts that may hinder osteoblast differentiation. Since the leukocyte platelet-rich fibrin (L-PRF) membrane promotes growth factor concentration, this study aims to quantify the amount of Sema4D in L-PRF membranes, and analyze the impact of Sema4D on osteoblast cell function in vitro. DESIGN: Enzyme-linked immunosorbent assay (ELISA) was used to quantify the levels of Sema4D in both L-PRF and whole blood (serum). To analyze the impairment of Sema4D on osteoblasts, MC3T3-E1 cells were induced to osteogenic differentiation and exposed to Sema4D ranging from 10 to 500 ng/ml concentrations. The following parameters were assayed: 1) cell viability by MTT assay after 24, 48, and 72 h; 2) matrix mineralization by Alizarin Red staining after 14 days, 3) Runt-related transcription factor 2 (RUNX-2), osteocalcin (OCN), osteonectin (ONC), bone sialoprotein (BSP) and alkaline phosphatase (ALP) gene expression by qPCR. For all data, the significance level was set at 5%. RESULTS: The amount of Sema4D in the whole blood (serum) was higher than in L-PRF. Osteoblasts exposed to Sema4D at all tested concentrations exhibited a decrease in matrix mineralization formation as well in RUNX-2, OCN, ONC, BSP, and ALP gene expression (p < 0.05). CONCLUSION: The presence of Sema4D, a molecule known for suppressing osteoblast activity, diminishes within L-PRF, enhancing its ability to facilitate bone regeneration.
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Fibrina Rica em Plaquetas , Semaforinas , Diferenciação Celular/genética , Leucócitos/metabolismo , Osteoblastos , Osteocalcina/metabolismo , Osteogênese/genética , Fibrina Rica em Plaquetas/metabolismo , Semaforinas/farmacologia , Semaforinas/metabolismo , Animais , CamundongosRESUMO
Cell culture and invertebrate animal models reflect a significant evolution in scientific research by providing reliable evidence on the physiopathology of diseases, screening for new drugs, and toxicological tests while reducing the need for mammals. In this review, we discuss the progress and promise of alternative animal and non-animal methods in biomedical research, with a special focus on drug toxicity.
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Pesquisa Biomédica , Animais , Modelos Animais , MamíferosRESUMO
OBJECTIVE: In this study, we investigated the modulatory effects of PI3Kγ on IL-17A expression and the progression of experimental periodontitis in vivo. METHODS: Ligature-induced periodontitis was developed around the first molar of mice. Animals were treated with anti-mouse IL-17A or IPI-549 (PI3Kγ inhibitor). In addition, PI3Kγ-deficient mice (PI3Kγ-/-) were used in the study. Alveolar bone loss was measured and real-time PCR of Il17a and Rankl genes was performed. A bioinformatics analysis was carried out using the Gene Set Enrichment Analysis computational tool. RESULTS: Nine days after ligature placement, alveolar bone loss scores were significantly increased, with upregulation of Il17a and Rankl genes in the gingival tissues. Treatment with anti-mouse IL-17A (100 µg/mice) significantly attenuated alveolar bone loss. Mice with ligature-induced periodontitis treated with IPI-549 (3 mg/kg) or PI3Kγ-/- mice showed reduced alveolar bone loss and downregulation of Il17a and Rankl gene expression in the gingival tissues. Consistent with this, the bioinformatics analysis showed upregulation of IL17F, IL17A, IL17D, and STAT3 genes, as well as greater activation of IL-17 and PI3KCI pathways (upregulation of PIK3CG gene) in the gingival tissue of patients with periodontitis. CONCLUSION: PI3Kγ plays an important role in modulating IL-17A expression and alveolar bone loss in vivo and can be considered a promising pathway for the management of periodontal disease and the development of new therapies.
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Perda do Osso Alveolar , Periodontite , Animais , Camundongos , Perda do Osso Alveolar/tratamento farmacológico , Perda do Osso Alveolar/genética , Interleucina-17/genética , Interleucina-17/metabolismo , Periodontite/tratamento farmacológico , Periodontite/genética , Gengiva/metabolismo , Ligadura , Modelos Animais de DoençasRESUMO
Abstract This study aimed to investigate whether GSK-3 inhibition (CHIR99021) effectively promoted mineralization by cementoblasts (OCCM-30). OCCM-30 cells were used and treated with different concentrations of CHIR99021 (2.5, 5, and 10 mM). Experiments included proliferation and viability, cellular metabolic activity, gene expression, and mineral nodule formation by Xylene Orange at the experimental time points. In general, CHIR99021 did not significantly affect OCCM-30 viability and cell metabolism (MTT assay) (p > 0.05), but increased OCCM-30 proliferation at 2.5 mM on days 2 and 4 (p < 0.05). Data analysis further showed that inhibition of GSK-3 resulted in increased transcript levels of Axin2 in OCCM-30 cells starting as early as 4 h, and regulated the expression of key bone markers including alkaline phosphatase (Alp), runt-related transcription factor 2 (Runx-2), osteocalcin (Ocn), and osterix (Osx). In addition, CHIR99021 led to an enhanced mineral nodule formation in vitro under both osteogenic and non-osteogenic conditions as early as 5 days after treatment. Altogether, the results of the current study suggest that inhibition of GSK-3 has the potential to promote cementoblast differentiation leading to increased mineral deposition in vitro.
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OBJECTIVE: To define the potential of polycaprolactone (PCL) scaffold for cementoblast delivery. BACKGROUND: Dental cementum is critical for tooth attachment and position, and its regenerative capabilities remain unpredictable. METHODS: PCL scaffolds were manufactured by the electrospinning technique at 10% and 20% (w/v) and seeded with cementoblasts (OCCM-30). Scaffolds were characterized for their morphology and biological performance by scanning electron microscopy (SEM), confocal and conventional histology, cytocompatibility (PrestoBlue assay), gene expression (type I collagen - Col1; bone sialoprotein - Bsp; runt-related transcription factor 2 - Runx-2; alkaline phosphatase - Alpl; osteopontin - Opn; osteocalcin - Ocn, osterix - Osx), and the potential to induce extracellular matrix deposition and mineralization in vitro. RESULTS: Overall, data analysis showed that PCL scaffolds allowed cell adhesion and proliferation, modulated the expression of key markers of cementoblasts, and led to enhanced extracellular matrix deposition and calcium deposition as compared to the control group. CONCLUSION: Altogether, our findings allow concluding that PCL scaffolds are a viable tool to culture OCCM-30 cells, leading to an increased potential to promote mineralization in vitro. Further studies should be designed in order to define the clinical relevance of cementoblast-loaded PCL scaffolds to promote new cementum formation.
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Materiais Biocompatíveis , Cemento Dentário , Diferenciação Celular , Sialoproteína de Ligação à Integrina/metabolismo , Poliésteres , Alicerces TeciduaisRESUMO
INTRODUCTION: Pain is a multidimensional experience involving the biological, psychological, and social dimensions of each individual. Particularly, the biological aspects of pain conditions are a response of the neuroimmunology system and the control of painful conditions is a worldwide challenge for researchers. Although years of investigation on pain experience and treatment exist, the high prevalence of chronic pain is still a fact. AREAS COVERED: Peroxisome proliferator-activated receptor-gamma (PPARγ) is a ligand-activated transcription factor belonging to the nuclear hormone receptor superfamily. It regulates several metabolic pathways, including lipid biosynthesis and glucose metabolism, when activated. However, PPARγ activation also has a critical immunomodulatory and neuroprotective effect. EXPERT OPINION: This review summarizes the evidence of synthetic or natural PPARγ ligands such as 15d-PGJ2, epoxyeicosatrienoic acids, thiazolidinediones, and specialized pro-resolving mediators, representing an interesting therapeutic tool for pain control.
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Imunomodulação , PPAR gama , Humanos , Imunomodulação/efeitos dos fármacos , Imunomodulação/fisiologia , Ligantes , PPAR gama/metabolismo , Dor , Prostaglandina D2/metabolismo , Tiazolidinedionas/uso terapêuticoRESUMO
Epoxyeicosatrienoic acids (EETs) are endogenous molecules that exerts effective antinociceptive and resolutive actions. However, because of their rapid metabolism by the soluble epoxide hydrolase (sEH), EETs are unable to remain bioavailable. Therefore, the aim of this study was to investigate whether local sEH inhibition could prevent inflammatory hyperalgesia in the temporomandibular joint (TMJ) of rats. For that, rats were pre-treated with an intra-TMJ injection of TPPU, followed by the noxious stimulus (1.5% of formalin intra-articular) to evaluate nociceptive behavior. Histological analysis was conducted to explore the inflammatory exudate and mast cell degranulation. Periarticular tissue over the TMJ was used to measure inflammatory lipids and cytokines/chemokine by Enzyme-Linked Immunosorbent Assay (ELISA). We demonstrated that peripheral pretreatment with TPPU prevents formalin-induced inflammatory hyperalgesia in the TMJ, and this effect is strictly local. Moreover, TPPU mitigates the leukocyte exudate in the TMJ, as well as inflammatory lipids mediators. Mast cell number and degranulation were abrogated by TPPU, and the inflammatory cytokine levels were decreased by TPPU. On the other hand, TPPU up-regulated the release of interleukin 10 (IL-10), an anti-inflammatory cytokine. We provide evidence that locally sEH by intra-TMJ injection of TPPU produces an antinociceptive and anti-inflammatory effect on rats' TMJ.
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Epóxido Hidrolases , Hiperalgesia , Analgésicos/uso terapêutico , Animais , Anti-Inflamatórios/uso terapêutico , Citocinas/metabolismo , Epóxido Hidrolases/metabolismo , Formaldeído/farmacologia , Hiperalgesia/induzido quimicamente , Hiperalgesia/tratamento farmacológico , Hiperalgesia/patologia , Lipídeos , Compostos de Fenilureia/toxicidade , Piperidinas/farmacologia , Ratos , Articulação Temporomandibular/metabolismo , Articulação Temporomandibular/patologiaRESUMO
Photobiomodulation (PBM) has been applied as a non-invasive technique for treating temporomandibular joint symptoms, especially on painful condition's relief, however the anti-inflammatory mechanism underlying the effect of PBM remains uncertain. This study aims to evaluate the mechanisms of action of PBM (808 nm) in a carrageenan-induced inflammation on temporomandibular joint (TMJ) of rats. In this study male Wistar rats were pre-treated with irradiation of a low-power diode laser for 15 s on TMJ (infra-red 808 nm, 100 mW, 50 J/cm2 and 1.5 J) 15 min prior an injection in the temporomandibular joint of carrageenan (100 µg/TMJ). 1 h after the TMJ treatments, the rats were terminally anesthetized for joint cavity wash and periarticular tissues collect. Samples analysis demonstrated that PBM inhibit leukocytes chemotaxis in the TMJ and significantly reduces amounts of TNF-α, IL-1ß and CINC-1. In addition, Western blotting analysis demonstrated that PBM significantly decreased the protein levels of P2X3 and P2X7 receptors in the periarticular tissues. On the other hand, PBM was able to increase protein level of IL-10 (anti-inflammatory cytokine). In summary, it is possible to suggest that PBM inhibit inflammatory chemotaxis, modulation the balance of the pro- and anti-inflammatory characteristics of inflammatory cells.
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Inflamação/terapia , Lasers Semicondutores/uso terapêutico , Terapia com Luz de Baixa Intensidade , Articulação Temporomandibular/efeitos da radiação , Animais , Carragenina/toxicidade , Movimento Celular/efeitos da radiação , Regulação para Baixo/efeitos da radiação , ELISPOT , Inflamação/induzido quimicamente , Interleucina-10/análise , Leucócitos/citologia , Leucócitos/metabolismo , Masculino , Ratos , Ratos Wistar , Receptores Purinérgicos P2X3/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Articulação Temporomandibular/metabolismo , Articulação Temporomandibular/patologia , Fator de Necrose Tumoral alfa/análiseRESUMO
Peripheral tramadol's delivery in the temporomandibular joint (TMJ) leads to significant analgesic outcomes and inflammatory process's resolvent actions. Mechanistically, these properties are apart from the opioid system. Nevertheless, the molecular mechanisms behind these effects are still unclear. Therefore, the present study investigated the hypothesis that adenosine A1 receptors are involved in the tramadol-induced analgesic and anti-inflammatory effects in the TMJ. Animals were pretreated with an intra-TMJ injection of DPCPX (antagonist of A1 receptor) or tramadol and subsequent nociceptive challenge with an intra-TMJ injection of 1.5% formalin. For over 45 min, the nociceptive behavior was quantitated, and by the end of this assessment, the animals were euthanized, and the periarticular tissue was collected. Lastly, an in vitro assay of BMDM (Bone Marrow-Derived Macrophages) was performed to investigate tramadol activity in macrophages. The intra-TMJ injection of tramadol ameliorates formalin-induced hypernociception along with inhibiting leukocyte migration. The tramadol's peripheral anti-inflammatory effect was mediated by the adenosine A1 receptor and was associated with increased protein expression of α2a-adrenoceptor in the periarticular tissues (p < 0.05: ANOVA, Tukey's test). Also, tramadol inhibits formalin-induced leukocyte migration and protein expression of P2X7 receptors in the periarticular tissue (p < 0.05); however, DPCPX did not alter this effect (p > 0.05). Moreover, DPCPX significantly reduced the protein expression of the M2 macrophage marker, MRC1. In BMDM, tramadol significantly reduces inflammatory cytokines release, and DPCPX abrogated this effect (p < 0.05). We identify tramadol's peripheral effect is mediated by adenosine A1 receptor, possibly expressed in macrophages in the TMJ tissue. We also determined an important discovery related to the activation of A1R/α2a receptors in the tramadol action.
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Agonistas do Receptor A1 de Adenosina/administração & dosagem , Artralgia/tratamento farmacológico , Receptor A1 de Adenosina/metabolismo , Receptores Adrenérgicos alfa 2/metabolismo , Tramadol/administração & dosagem , Analgésicos Opioides/administração & dosagem , Animais , Anti-Inflamatórios/administração & dosagem , Artralgia/induzido quimicamente , Artralgia/imunologia , Artralgia/patologia , Modelos Animais de Doenças , Formaldeído/administração & dosagem , Formaldeído/toxicidade , Humanos , Injeções Intra-Articulares , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Nociceptividade/efeitos dos fármacos , Ratos , Articulação Temporomandibular/efeitos dos fármacos , Articulação Temporomandibular/imunologia , Articulação Temporomandibular/patologia , Xantinas/administração & dosagem , Xantinas/toxicidadeRESUMO
BACKGROUND: P2X7 receptors are responsible for triggering inflammatory responses contributing to processes of pain in articular tissues. This study aimed to investigate whether the activation of the P2X7 receptor located in the temporomandibular joint (TMJ) tissues induces nociception through an inflammatory mechanisms and/or the activation of C-fibres (small-diameter primary afferents) of rats' TMJ. METHODS: The TMJ hypernociception induced by the activation of P2X7 receptor was assessed by measuring the behavioural nociceptive responses. After behavioural experiments, the animals were terminally anaesthetized and periarticular tissues were removed and homogenate for enzyme-linked immunosorbent assay, leukocyte infiltration and western blotting analysis. RESULTS: The nonselective P2X7 receptor agonist BzATP induced a dose-dependent TMJ nociception, which was blocked by the selective P2X7 receptor antagonist A-438079. The co-administration of the selective ß2-adrenoceptor antagonist (ICI-118,551) and the pre-treatment with cyclooxygenase inhibitor indomethacin or with the nonspecific selectin inhibitor Fucoidan significantly reduced BzATP-induced TMJ nociception. BzATP also induced an increase of pro-inflammatory cytokines TNFα, IL-1ß and CINC-1 levels, as well as leukocyte recruitment in TMJ tissue, effects that were reduced by A-438079. Moreover BzATP-induced TMJ nociception was inhibited in rats neonatal-treated with Capsaicin (depleting C-fibers). Finally, BzATP-induced an increase in TRPV1 expression in TMJ tissue. CONCLUSIONS: These findings suggest that P2X7 receptor activation in TMJ of rats induces nociceptive responses mediated by sympathomimetic amines, prostaglandins, leukocyte migration and increased levels of pro-inflammatory cytokines. Furthermore, the P2X7 receptor activation induces nociceptive responses dependent on the activation of the primary afferent nociceptors of rats' TMJ. SIGNIFICANCE: The activation of P2X7 receptors has an essential role in TMJ nociception and could be an interesting target to control the inflammatory pain in temporomandibular disorders.
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Nociceptividade , Transtornos da Articulação Temporomandibular , Animais , Dor , Ratos , Ratos Wistar , Articulação Temporomandibular , Transtornos da Articulação Temporomandibular/induzido quimicamenteRESUMO
PURPOSE: To investigate the effect of experimental traumatic occlusion (ETO) induced by metal crowns on alveolar bone loss. MATERIALS AND METHODS: Metal crowns were custom-made for the lower first molars with occlusal discrepancy of 0.4 and 0.7 mm from the maximum intercuspation. Thirty-six animals were randomly divided into three groups (n = 12 animals per group): 0.4-mm hyperocclusion group, 0.7-mm hyperocclusion group and the sham group (no metal crown). Twenty-eight days after crown cementation, the animals were euthanized and gingival tissue was collected to assess cytokine levels of IL-17, IL-6, and TNF-α using enzyme-linked immunosorbent assay (ELISA). Mandibles were stained with 1% methylene blue and alveolar bone levels were quantified. Western blotting was used to quantify the expression of receptor activator of nuclear factor κ B (RANK), and its ligand (RANKL), secreted osteoclastogenic factor of activated T cells (SOFAT) and TNF-α-converting enzyme (TACE). Also, mandibles were histologically processed and stained with hematoxylin and eosin, from which the presence of osteoclast-like cells, multinucleated cells containing ≥3 nuclei was counted at 100× magnification. The data were analyzed using one-way ANOVA and Tukey tests. RESULTS: Experimental occlusal trauma for 28 consecutive days significantly increased alveolar bone loss and multinucleated cell counts (p < 0.05). RANK, RANKL, SOFAT, TACE, IL-6, and TNF-α were significantly higher in gingival tissues of ETO groups (p < 0.05). IL-17 titers were unchanged among the groups (p > 0.05). CONCLUSION: Experimental traumatic occlusion activates and sustains bone resorption pathways in the periodontium inducing alveolar bone resorption. As the intensity of occlusal trauma increased, alternative osteoclastic pathways were activated, such as TACE and SOFAT.
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Perda do Osso Alveolar , Cimentação , Perda do Osso Alveolar/etiologia , Animais , Coroas , Osteoclastos , PeriodontoRESUMO
The objective of the present study was to evaluate discomfort and safety of microneedle (MN) insertion in several intraoral regions. A device was developed to standardize MN insertions. MNs were inserted in the following regions of the oral cavity: gingiva, palatine alveolar process, buccal mucosa, dorsum of the tongue and inner portion of the lower lip. Perforations from MNs post insertion were confirmed with topical gentian violet stain. Pain was evaluated in a randomized, double-blinded, crossover study in 30 volunteers. Each volunteer received a MN patch, a 30G hypodermic needle (positive control) and an identical MN patch with its needles laying flat in the plane of the patch (negative control). Adverse events were visually evaluated immediately after (0 h) and 24 h post MN application. The application device developed a consistent application force (10 N) and promoted perforation of all individual MNs on a patch. At all sites, insertion of the hypodermic needle promoted more pain when compared to the negative control (p < 0.001). Application of the MNs promoted less pain than the hypodermic needle (p < 0.05), but slightly more pain as compared to the negative control (p < 0.05) at all sites except the tongue, where the MN did not differ from the negative control (p > 0.05). Hypodermic needle caused bleeding at all insertion sites. In contrast, MNs did not cause bleeding at most sites except in some cases of insertion into the hard gingiva and the palatine alveolar process where tiny blood spots appeared immediately after MN application for few of the MNs on the patch. There were no cases of bleeding at 24 h post MN application. In conclusion, MNs can perforate different sites of the oral cavity in a safe and significantly less painful manner as compared to the 30G hypodermic needle. Thus, analogous to the skin, MN-based approaches could be an attractive approach for drug delivery in the oral cavity.
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Agulhas , Pele , Administração Cutânea , Estudos Cross-Over , Sistemas de Liberação de Medicamentos , Humanos , Microinjeções , Boca , Dor/tratamento farmacológicoRESUMO
Analgesic mechanism of Botulinum toxin type A (BoNT/A) involves retrograde axonal transport to central nervous system, where it may interact with sensory neurons. Though, some authors suggested that BoNT/A antinociceptive action may also be associated with the inhibition intracellular factors and neuromodulators expressed by immune cells, especially by microglia. Antigen-induced arthritis in the temporomandibular joint (TMJ) of rats is signal by P2X7 receptor/Cathepsin S (CatS)/Fractalkine (FKN) microglia-activated pathway. Thus, we aimed to evaluate the possible modulatory effect of an intra-TMJ injection of BoNT/A on the P2X7/CatS/FKN microglia-activated pathway in the trigeminal subnucleus caudalis of rats with antigen-induced arthritis of the TMJ. A model of antigen-induced arthritis was used on Wistar rats (n = 40) by systemic injections of an emulsion containing complete Freund's adjuvant and methylated bovine serum albumin (mBSA) diluted in PBS. The arthritic condition was stablished by an intra-TMJ injection of mBSA (10 µg/TMJ/week) for 3 weeks. Then, animals were treated with an intra-TMJ injection of BoNT/A (onabotulinumtoxinA, Allergan®; 7U/kg) or vehicle saline. Animals were euthanized 24 h, 7 or 14 days after BoNT/A treatment and their trigeminal nucleus caudalis was harvested to evaluate the protein level of microglial purinergic P2X7 receptor and CX3 chemokine receptor 1 (CX3CR1) by Western blot, and to measure the protein level of microglial modulators CatS, FKN, and the pro-inflammatory cytokines tumor necrosis alfa (TNF-α) and interleukin 1ß (IL-1ß) by enzyme-linked immunosorbent assay (ELISA). The antigen-induced arthritis in the TMJ significantly increased the protein levels of P2X7, CatS, FKN, TNF-α and IL-1ß in the trigeminal subnucleus caudalis (P < 0.05). The intra-TMJ injection of BoNT/A reduced the protein levels of P2X7 in all time points tested. Additionally, BoNT/A significantly reduced the protein levels of CatS, FKN, and TNF-α 14 days after treatment. However, IL-1ß was significantly reduced just 24 h after the BoNT/A intra-TMJ treatment. Based on our results, we can suggest that the intra-TMJ injection of BoNT/A may promote a central effect by reducing the P2X7/CatS/FKN microglia-activated pathway in the trigeminal subnucleus caudalis.
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Toxinas Botulínicas Tipo A/uso terapêutico , Articulação Temporomandibular , Animais , Artrite Experimental/tratamento farmacológico , Quimiocina CX3CL1/metabolismo , Adjuvante de Freund , Injeções Intra-Articulares , Masculino , Microglia/metabolismo , Ratos , Ratos WistarRESUMO
Rheumatoid arthritis (RA) has an inflammatory milieu in the synovial compartment, which is regulated by a complex cytokine and chemokine network that induces continuously degenerative and inflammatory reactions. The secreted osteoclastogenic factor of activated T cells (SOFAT) is a unique cytokine and represents an alternative pathway for osteoclast activation. In this study, we examined whether SOFAT is able to induce joint pain and investigated the presence of SOFAT in a Collagen-induced Arthritis (CIA) model and in human subjects. Here, we found that an intra-articular stimulation with SOFAT (1, 10, 100, or 1,000 ng/10 µl) in the knee joint significantly decreases the mechanical threshold in the hind paw of mice (p < 0.05). Moreover, after a second injection of SOFAT, the mechanical threshold decrease was sustained for up to 8 days (p < 0.05). In the CIA model, the immunohistochemical assay of knee joint showed positivity stained for SOFAT, and the mRNA and protein expression of SOFAT were significantly higher in the affected-group (p < 0.05). Besides, the mRNA of RANKL, IL-1ß, IL-6, and IL-15 were significantly higher in the affected-group (p < 0.05). Finally, SOFAT was detected in the synovial fluid of RA patients, but not in OA patients (p < 0.05). In conclusion, SOFAT is up regulated in inflammatory milieu such as RA but not in non-inflammatory OA. SOFAT may be a novel molecule in the complex inflammatory phenotype of RA.
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Artrite Experimental/metabolismo , Artrite Reumatoide/metabolismo , Carbono-Oxigênio Liases/metabolismo , Citocinas/metabolismo , Articulações/fisiologia , Dor/metabolismo , Animais , Modelos Animais de Doenças , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos DBA , Osteoclastos/fisiologia , Osteogênese , Transdução de Sinais , Regulação para CimaRESUMO
Rheumatoid arthritis (RA) is characterized by chronic inflammation of the synovial tissue, joint dysfunction, and damage. Epoxyeicosatrienoic acids (EETs) are endogenous anti-inflammatory compounds, which are quickly converted by the soluble epoxide hydrolase (sEH) enzyme into a less active form with decreased biological effects. The inhibition of the sEH enzyme has been used as a strategy to lower nociception and inflammation. The goal of this study was to investigate whether the peripheral treatment with the sEH enzyme inhibitor 1- trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) could prevent the hypernociception and inflammation in the albumin-induced arthritis model in rats' temporomandibular joint (TMJ). After the induction of experimental arthritis, animals were assessed for nociceptive behavior test, leukocyte infiltration counts and histologic analysis, ELISA to quantify several cytokines and Western blotting. The peripheral pretreatment with TPPU inhibited the arthritis-induced TMJ hypernociception and leukocyte migration. Moreover, the local concentrations of proinflammatory cytokines were diminished by TPPU, while the anti-inflammatory cytokine interleukin-10 was up-regulated in the TMJ tissue. Finally, TPPU significantly decreased protein expression of iNOS, while did not alter the expression of MRC1. This study provides evidence that the peripheral administration of TPPU reduces hypernociception and inflammation in TMJ experimental arthritis.
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Analgésicos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Artrite Experimental/tratamento farmacológico , Epóxido Hidrolases/antagonistas & inibidores , Compostos de Fenilureia/uso terapêutico , Piperidinas/uso terapêutico , Articulação Temporomandibular/efeitos dos fármacos , Albuminas , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Artrite Experimental/imunologia , Artrite Experimental/patologia , Citocinas/imunologia , Masculino , Óxido Nítrico Sintase Tipo II/imunologia , Compostos de Fenilureia/farmacologia , Piperidinas/farmacologia , Ratos Wistar , Articulação Temporomandibular/imunologia , Articulação Temporomandibular/patologiaRESUMO
Although periodontitis is one of the commonest infectious inflammatory diseases in humans, the mechanisms involved with its immunopathology remain ill understood. Numerous molecules may induce inflammation and lead to bone resorption, secondary to activation of monocytes into osteoclasts. TACE (TNF-α converting enzyme) and DC-STAMP (dendritic cell-specific transmembrane protein) appear to play a role on bone resorption since TACE induces the release of sRANKL (soluble receptor activator of nuclear factor kappa-ß ligand) whereas DC-STAMP is a key factor in osteoclast induction. The present study evaluated the levels of TACE and DC-STAMP in patients with and without periodontitis. Twenty individuals were selected: 10 periodontally healthy participants undergoing gingivectomy for esthetic reasons and 10 diagnosed with periodontitis. Protein levels of such molecules in gingival tissue were established using Western blotting. Protein levels of both TACE and DC-STAMP were higher in the periodontitis group than in the control group (p<0.05; Student t-test). In conclusion, TACE and DC-STAMP protein levels are elevated in patients with periodontitis, favoring progression of bone resorption.
Assuntos
Proteína ADAM17 , Proteínas Adaptadoras de Transdução de Sinal , Reabsorção Óssea , Proteínas de Membrana , Periodontite , Proteína ADAM17/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Diferenciação Celular , Humanos , Proteínas de Membrana/metabolismo , OsteoclastosRESUMO
Epoxyeicosatrienoic acids (EET) and related epoxy fatty acids (EpFA) are endogenous anti-inflammatory compounds, which are converted by the soluble epoxide hydrolase (sEH) to dihydroxylethersatrienoic acids (DHETs) with lessened biological effects. Inhibition of sEH is used as a strategy to increase EET levels leading to lower inflammation. Rheumatoid arthritis is a chronic autoimmune disease that leads to destruction of joint tissues. This pathogenesis involves a complex interplay between the immune system, and environmental factors. Here, we investigate the effects of inhibiting sEH with 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) on a collagen-induced arthritis model. The treatment with TPPU ameliorates hyperalgesia, edema, and decreases the expression of important pro-inflammatory cytokines of Th1 and Th17 profiles, while increasing Treg cells. Considering the challenges to control RA, this study provides robust data supporting that inhibition of the sEH is a promising target to treat arthritis.